Engineering Materials

Rishabha Malviya
Sonali Sundram Editors

Engineered
Biomaterials
Synthesis and Applications
Engineering Materials
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Rishabha Malviya · Sonali Sundram
Editors

Engineered Biomaterials
Synthesis and Applications
Editors
Rishabha Malviya Sonali Sundram
Department of Pharmacy Department of Pharmacy
Galgotias University Galgotias University
Greater Noida, Uttar Pradesh, India Greater Noida, Uttar Pradesh, India

ISSN 1612-1317 ISSN 1868-1212 (electronic)

Engineering Materials
ISBN 978-981-99-6697-4 ISBN 978-981-99-6698-1 (eBook)
[Link]

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Dear Healthcare Professionals,
We are dedicating this book to you. Our Love
for profession shall live forever.

This book is also dedicated to friendship of

editors. We work together and edited a
fruitful book.
Foreword

Biomaterials are materials with novel properties that can enable them to come in
contact with the living tissue. They can be synthesised by a variety of processes and
engineered according to a specific use and application in order to mimic biological
phenomena or else be employed for some other process based on its use. There are
immense benefits of Biomaterials particularly in improving the quality of life bene-
fits apart from others. The progress in this field is dependent on interdisciplinary
research with scientists from diverse backgrounds like chemists, physicists, biolo-
gists, pharmaceutical scientists, medical doctors, engineers, material producers and
manufacturers etc. For medicinal purposes it can either be a diagnostic purpose or
therapeutic purpose. The main rationale behind writing this book is to cover all the
diverse aspects and the scientific knowledge associated with engineering of bioma-
terials as well as its utilisation in the healthcare field. This book attempts to explain
in detail the principles and foremost methods for the synthesis of biomaterials and
its application interconnected with prevention, mitigation, diagnosis and treatment
purposes. The readers of this book will get an idea about how biomaterials are
growing wider in terms of application as well as the increasing future opportunities
associated with them. This book can also serve as a useful source of information
for studying engineering biomaterials. Various methods for the synthesis and devel-
opment of biomaterials are provided in the contributed chapters. These methods
include those related to genetic engineering for the development of novel bioma-
terials, environmentally friendly approaches for the development of biomaterials
based on bone and tissue engineering, gold nanoparticles derived from microorgan-
isms as a synthetic method for developing biomaterials, and biomimetic approaches
for producing biomaterials.
Each chapter of this book highlights methods of synthesizing biomaterials and
how to apply them in healthcare. Not only the opportunities and advantages have
been covered but the possible challenges have been addressed so that the readers and
researchers get an idea about the obstacles and their means to overcome them.
It is my pleasure to present this forward for this book entitled Engineered Bioma-
terials: Synthetic Approach and Applications Edited by Dr. Rishabha Malviya.

vii
viii Foreword

Different chapters in this book have been contributed by the experts in their respective
fields.
I highly acknowledge all the authors as well as the editors for their endeavour and
efforts to compile this book and wish them all the very best.

Prof. Roop Krishen Khar

Director
B. S. Anangpuria Educational Institutes
Faridabad, India
Director
Formulators Koncept Pvt. Ltd.
New Delhi, India
Director
Unicure India Ltd.
New Delhi, India
Former Dean and HOD
Jamia Hamdard University
New Delhi, India
Preface

Biomaterial can be defined as a material which is engineered for interacting with the
biological system for the healthcare purpose in diagnosis, mitigation and treatment
motives. Engineering biomaterials means the study of biomaterials. Biomaterials
have a property of biocompatibility which makes them suitable as well as safe for
use. Methods for the synthesis of biomaterials and all the aspects associated with it
are covered in this book. In the initial chapter information is given about the biomate-
rials which are derived naturally, its advances as well as opportunities. Then various
techniques are given for synthesis and development of biomaterials which includes
techniques associated with genetic engineering for the development of novel bioma-
terials, green methods for the development of bone and tissue engineering based
biomaterials, gold nanoparticles from a microorganism—a synthetic approach for
development of biomaterials and the biomimetic approaches for manufacturing of
biomaterials. A detail description about genetically induced biomaterial advances in
medical science is given in one of the chapters. Nowadays biomaterials are applied
widely in healthcare applications which are growing wider along with time. Along
with the synthetic approaches, the applications of biomaterials are also covered in
this book. The usage of nano-sized biomaterials in drug delivery systems, recent
trends associated with it as well as the future opportunities are covered here. Usage
of biomaterials in implant devices, stimuli responsive materials in controlled release
of drug, advanced tissue engineering with novel engineered biomaterials, collagen
based nanomaterials for delivering drug, photo responsive material for 4D printing
in tissue engineering, surface modified biomaterials in developing medical devices,
nano-porous metal foams as versatile nanoplatforms for drug delivery applications
and its properties, recent progress as well as challenges related to it, development
of cardiovascular biomaterials from collagenous tissues, stimuli responsive material
in controlled release of drug, gold nanoparticles along with their clinical applica-
tions and morphological study of silver nanoparticles supported on ti1-xcexo2, their
synthesis and antimicrobial applications are covered in the chapters of this book. The
students as well as researchers reading this book will get a detailed idea about what
are biomaterials, how they are synthesized and how they can be applied in health-
care for medical purposes. The readers will find the information useful and precise.

ix
x Preface

Careful attention has been given by the authors to avoid making the information
complex so that readers do not get confused while reading the book. Authors have
tried to cover recent points linked to biomaterials so that latest information can be
conveyed through this book. We thank the authors for putting their efforts in writing
this book.

Greater Noida, India Rishabha Malviya

Sonali Sundram
Contents

Approach for the Synthesis of Biomaterials

Naturally Derived Biomaterials: Advances and Opportunities . . . . . . . . . 3
Ainil Hawa Jasni, Azlin Suhaida Azmi, Noor Illi Mohamad Puad,
Fathilah Ali, and Yusilawati Ahmad Nor
Different Techniques of Genetic Engineering Used
for the Development of Novel Biomaterials . . . . . . . . . . . . . . . . . . . . . . . . . . . 43
Aashveen Chhina, Vridhi Sachdeva, and Shubham Thakur
Green Methods for the Development of Bone and Tissue
Engineering-Based Biomaterials . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 73
Avipsa Hazra, Gowrav Baradwaj, A. S. Dhanu,
Gobianand Kuppannan, Malarvizhi Arthanari, and B. M. Kanthesh
Genetically Induced Biomaterial Advances in Medical Sciences . . . . . . . . 95
Eva Kaushik and Rohit Kaushik
Biomimetic Approaches for Biomaterials Development . . . . . . . . . . . . . . . . 125
Sudipta Choudhury, K. R. Arjun, M. N. Ramesh Bharadwaj,
M. Maghimaa, and Kanthesh M. Basalingappa
Plant-Based Biomaterials in Tissue Engineering and Drug Delivery
Systems . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 153
Azadeh Izadyari Aghmiuni, Arezoo Ghadi, and Elmira Azmoun
Gold Nanoparticles from a Microorganism: A Synthetic Approach . . . . . 199
Anil Thakur, Shubham Thakur, and Sonia Sharma

Applications of Biomaterials
Nanostructured Biomaterials in Drug Delivery . . . . . . . . . . . . . . . . . . . . . . . 233
İbrahim Mizan Kahyaoğlu, Erdi Can Aytar, Alper Durmaz,
and Selcan Karakuş

xi
xii Contents

Nanostructured Biomaterials in Drug Delivery: Current Trends

and Upcoming Possibilities . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 261
Pankaj Sharma
Advancement in Biomaterials in the Form of Implants . . . . . . . . . . . . . . . . 281
Riya Shivgotra, Bindu Soni, Manjot Kaur, and Shubham Thakur
Smart Biomaterials in Drug Delivery Applications . . . . . . . . . . . . . . . . . . . . 323
S. Giridhar Reddy and H. C. Ananda Murthy
Advanced Tissue Engineering with Novel Engineered Biomaterials . . . . . 361
Azadeh Izadyari Aghmiuni and Aref Gholami
An Insight into Collagen-Based Nano Biomaterials for Drug
Delivery Applications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 397
Amit Kumar Verma
Photo Responsive Material for 4D Printing in Tissue Engineering . . . . . . 429
Amisha, Shubham Thakur, and Amrinder Singh
Surface-Modified Biomaterials in Medical Device Development . . . . . . . . 465
Bindu Soni, Riya Shivgotra, Manjot Kaur, and Shubham Thakur
Nanoporous Materials as Versatile Nanoplatforms for Drug
Delivery Applications: Properties, Recent Progress, and Challenges . . . . 495
R. Abdel-Karim
Development of Cardiovascular Biomaterials From Collagenous
Tissues . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 521
Gowrav Baradwaj, Kshitija Aherkar, R. Mythreyi, T. S. Gopenath,
and Kanthesh M. Basalingappa
Stimuli-Responsive Material in Controlled Release of Drug . . . . . . . . . . . . 535
Karan Trehan, Muskaan Saini, and Shubham Thakur
Gold Nanoparticles: Clinical Applications . . . . . . . . . . . . . . . . . . . . . . . . . . . 563
Sheikdawood Parveen, T. Sathiyapriya, D. Tharani,
S. U. Mohammed Riyaz, Rakshi Anuja Dinesh,
Jayashree Shanmugam, K. Rajakumar, Dmitry Zherebtsov,
Manikandan Dhayalan, and Antony Stalin
Biocidal Effect of Copper Contained in a Mineral Tailing
on the Growth of Shewanella Putrefaciens . . . . . . . . . . . . . . . . . . . . . . . . . . . . 579
Hugo J. Marín-García, Ramiro Escudero-García,
Carlos Cortés-Penagos, and Ricardo Morales-Estrella
Contents xiii

Examining the Problems and Possibility of Immunological Control

for Engineered AAV as a CRISPR Vector and Other Genetic
Transfers . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 591
Asra Hamidi

Subject Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 615

About the Editors

Dr. Rishabha Malviya completed B. Pharmacy from

Uttar Pradesh Technical University and M. Phar-
macy (Pharmaceutics) from Gautam Buddha Technical
University, Lucknow Uttar Pradesh. His Ph.D. (Phar-
macy) work was in the area of Novel formulation devel-
opment techniques. He has 12 years of research experi-
ence and is presently working as Associate Professor
in the Department of Pharmacy, School of Medical
and Allied Sciences, Galgotias University for the past
eight years. His area of interest includes formulation
optimization, nanoformulation, targeted drug delivery,
localized drug delivery and characterization of natural
polymers as pharmaceutical excipients. He has authored
more than 200 research/review papers for national/
international journals of repute. He has 58 patents
(19 grants, 38 published, 1 filed) and publications in
reputed National and International journals with total
of more than 200 cumulative impact factor. He has also
received an Outstanding Reviewer award from Elsevier.
He has authored/edited/editing 48 books (Wiley, CRC
Press/Taylor and Francis, Springer, River Publisher, IOP
publishing and OMICS publication) and authored 31
book chapters. His name has been included in word’s
top 2% scientist list for the year 2020 and 2021 by Else-
vier BV and Stanford University. He is Reviewer/Editor/
Editorial board member of more than 50 national and
international journals of repute. He is the invited author
for Atlas of Science and pharma magazine dealing with
industry (B2B) “Ingredient south Asia Magazines”.

xv
xvi About the Editors

Prof. Sonali Sundram completed [Link]. and

[Link]. (Pharmacology) from AKTU, Lucknow. She
has worked as a research scientist in an ICMR project
at the King George’s Medical University, Lucknow
after which she joined BBDNIIT and currently working
in Galgotias University, Greater Noida. Her Ph.D.
(Pharmacy) work was in the area of Neurodegener-
ation and Nanoformulation. Her area of interest is
neurodegeneration, clinical research, artificial intelli-
gence. She has authored/edited/editing more than 15
books (Wiley, CRC Press/Taylor and Francis, IOP
publishing, Apple Academic Press/Taylor and Francis,
Springer Nature and River Publisher) She has attended
as well organized more than 15 national and interna-
tional seminar/conferences/workshops. She has more
than eight patents, national and international, to her
credit. She has published six SCI indexed manuscripts
(cumulative impact factor: 20.71) with reputed inter-
national publishers. She has delivered oral presenta-
tions in international conferences organized in different
European countries.
Approach for the Synthesis of Biomaterials
Naturally Derived Biomaterials:
Advances and Opportunities

Ainil Hawa Jasni, Azlin Suhaida Azmi, Noor Illi Mohamad Puad,
Fathilah Ali, and Yusilawati Ahmad Nor

Abstract Biomaterials are materials that have been formed from or created by
biological organisms such as plants, animals, bacteria, fungus, and other forms of life
are referred to as biologically derived materials. Biomaterials are normally designed
to interface with biological systems, for the treatment, augmentation, or replace-
ment of biological functions. Across billions of years, life has been composed of and
existed within these biomaterial molecules, monomers, and polymers. For instance,
biomaterials of polysaccharides are sugars or starch polymers. Cellulose is the most
ubiquitous and abundant polysaccharide. Polysaccharides are found in the tissues
of both trees and humans. Meanwhile, natural biomaterials are substances that are
derived from natural sources such as plants, animals, or minerals, and are used
in medical and healthcare applications. Examples of natural biomaterials include
collagen, chitosan, silk, cellulose, hyaluronic acid, and bone minerals such as hydrox-
yapatite. These materials are attractive in the field of regenerative medicine and tissue
engineering due to their biocompatibility and biodegradability. Additionally, some
natural biomaterials can mimic the physical and chemical properties of the body’s
natural tissues, making them ideal for use in implants and scaffolds. Recent advances
in the production of natural biomaterials include the development of more efficient
and scalable manufacturing processes, which has made them more widely available
and accessible for use in medical applications. In addition, advances in the under-
standing of the biological interactions between these materials and the body have
allowed for the development of new and improved medical devices and therapies.
The use of natural biomaterials also provides unique opportunities for customization
and personalization in medical treatment. For example, natural biomaterials such
as collagen and hyaluronic acid can be engineered to meet specific patient needs,
such as tissue repair and regeneration, wound healing, and drug delivery. Overall,
natural biomaterials have shown great promise in many fields. This chapter’s goal
is to give readers a quick introduction to naturally derived biomaterials and their
advances and opportunities. For example, recent developments in the production of
natural biomaterials have made them more widely available and accessible for use

A. H. Jasni (B) · A. S. Azmi · N. I. M. Puad · F. Ali · Y. A. Nor

Department of Chemical Engineering and Sustainability Kulliyyah of Engineering, International
Islamic University Malaysia, Jalan Gombak, 53100 Kuala Lumpur, Malaysia
e-mail: [Link]@[Link]

© The Author(s), under exclusive license to Springer Nature Singapore Pte Ltd. 2023 3
R. Malviya and S. Sundram (eds.), Engineered Biomaterials, Engineering Materials,
[Link]
4 A. H. Jasni et al.

in medical applications, and advances in the understanding of the biological interac-

tions between these materials and the body have allowed for the development of new
and improved medical devices and therapies. In the coming years, the adoption of
new advanced experimental methodologies, such as bioengineering approaches, will
alter the practice of medicine in the applications using natural derived biomaterials.
Tissue engineering, a multidisciplinary field of research involving the principles of
materials science, engineering, biological sciences, and medical research, is a clear
illustration of this.

Keywords Biocompatibility · Biomaterials · Natural biomaterials · Regenerative

medicine · Tissue engineering

1 Introduction

Natural biomaterials have developed into an increasingly popular area of study in

recent years. These materials are derived from natural sources, such as plants or
animals, and can be used for a variety of applications in the medical field. The fact
that naturally derived biomaterials are biocompatible, or do not have negative effects
when inserted into the body, is one of their main benefits. This makes them perfect for
application in tissue engineering, medical devices, and drug delivery systems. One
promising application of naturally derived biomaterials is in regenerative medicine.
Utilizing biological components, regenerative medicine aims to replace or repair
damaged tissues and organs.
Naturally derived biomaterials offer many advantages over synthetic materials in
this context, including better compatibility with living cells and tissues. For example,
researchers have developed scaffolds made from collagen, a protein found in skin
and connective tissues, which can support the growth of new tissues. Other naturally
derived biomaterials being investigated for regenerative medicine include chitosan,
silk fibroin, and hyaluronic acid.
Another exciting opportunity for naturally derived biomaterials is in the devel-
opment of sustainable materials. Many conventional materials used in the industry
are derived from non-renewable resources, which can have negative environmental
impacts. By contrast, naturally derived biomaterials can often be produced sustain-
ably and may even be biodegradable [1]. Overall, advances in naturally derived
biomaterials hold great promise for improving human health and reducing our impact
on the environment. Naturally derived biomaterials are offering exciting opportu-
nities when it comes to regenerative medicine. The ability to regenerate tissues
and organs using biological materials has been a long-standing goal for medical
researchers, and with the development of naturally derived biomaterials, this dream
is becoming a reality. One of the primary advantages of these materials over synthetic
ones is their compatibility with living cells and tissues. Additionally, advances in 3D
printing technology have made it possible to create complex structures that mimic
the architecture of natural tissues.
Naturally Derived Biomaterials: Advances and Opportunities 5

Naturally derived biomaterials also offer promising solutions for sustainable mate-
rials. Many conventional materials used in industry rely on non-renewable resources
and often result in negative environmental impacts. In contrast, biomaterials can be
produced sustainably and may even be biodegradable. By leveraging these materials,
we can reduce our impact on the environment while still producing high-quality prod-
ucts. Advances in naturally derived biomaterials represent an incredible opportunity
for improving human livelihoods while also protecting our environment. As research
continues to explore the full range of applications for these materials, we can look
forward to even more exciting discoveries and innovations in the years to come. It’s
clear that the possibilities are truly remarkable, and we should all be excited about
what the future holds for this cutting-edge field.

2 Definition of Naturally Derived Biomaterials

Naturally derived biomaterials are materials that are sourced from biological sources
such as plants, animals, and microorganisms [1]. These materials are usually biocom-
patible and biodegradable, meaning they can be safely used in living organisms
without causing harm and can eventually be broken down by biological processes.
Biomaterials are materials that interact with biological systems for various purposes,
for instance the delivery of drugs, medical devices, and tissue engineering [2]. Natu-
rally derived biomaterials, obtained from natural sources such as animals, plants,
and microorganisms [3] have undergone substantial research and are utilised in
biomedical applications for their biocompatibility, biodegradability, and bioactivity.

3 The Importance of Naturally Derived Biomaterials

in Various Applications

Here are some of the reasons why naturally derived biomaterials are important in
various applications (Table 1).

4 Scope of the Chapter

This chapter covers a variety of closely connected topics to the development, prop-
erties, and applications of biomaterials that are derived from natural sources. The
topics which are covered in this chapter include:
6 A. H. Jasni et al.

Table 1 The importance of naturally derived biomaterials in applications

Added values in naturally Definition and examples
derived biomaterials
1. Biocompatibility Naturally derived biomaterials are often biocompatible,
meaning they are not harmful to living tissues and can be used
in medical applications without causing adverse reactions. For
example, collagen and hyaluronic acid are naturally derived
biomaterials that are often utilised in tissue engineering and
regenerative medicine [4, 5]
2. Sustainability Many naturally derived biomaterials are renewable and
sustainable, making them environmentally friendly alternatives
to synthetic materials. For example, cellulose, a naturally
occurring polymer found in plants, can be used to make
biodegradable plastics [6] and packaging materials
3. Diversity Naturally derived biomaterials come in a wide variety of forms,
allowing for customization to specific applications. For
example, chitosan, a polysaccharide derived from crustacean
shells, can be modified to have different properties, and used in
wound dressings or drug delivery systems [7]
4. Biodegradability Naturally derived biomaterials can be designed to be
biodegradable, which is particularly important in applications
where long-term implantation is not desired. For example, silk
fibroin, a protein derived from silkworm cocoons, can be used as
a biodegradable scaffold in tissue engineering [8]
5. Antibacterial properties Some naturally derived biomaterials possess inherent
antibacterial properties, making them useful in medical
applications where preventing infection is critical [9]. For
example, silver nanoparticles can be incorporated into chitosan
to create a material which can be utilised in wound dressings
due to its antimicrobial qualities. Other than that, natural
antimicrobial peptides (AMPs) can be used in the treatment of
bacterial infections [9]

1. Overview of naturally derived biomaterials: This include an introduction to the

types of biomaterials that are commonly derived from natural sources. This
chapter offers a summary of the physical and chemical properties of these
biomaterials and their biological interactions.
2. Advances in the synthesis and processing of naturally derived biomaterials: This
section focuses on recent developments in the synthesis and processing of natural
biomaterials, including advances in biomaterials engineering and biotechnology.
This includes discussions of new methods for isolating and purifying natural
biomaterials, as well as approaches for modifying their chemical and physical
properties.
Naturally Derived Biomaterials: Advances and Opportunities 7

3. Applications of naturally derived biomaterials: This section explores the diverse

range of applications of natural biomaterials, including their use in wound
healing, tissue engineering, drug delivery, and medical devices. It also high-
lights the advantages of using natural biomaterials over synthetic alternatives,
such as their biodegradability, biocompatibility, and sustainability.
4. Challenges and opportunities in the field: This section examines some of the
current challenges and opportunities in the field of naturally derived biomate-
rials. This includes discussions of regulatory issues related to the use of natural
biomaterials in medical devices and pharmaceuticals, as well as challenges
related to scaling up production and optimizing biomaterial properties for specific
applications.

5 Classification of Naturally Derived Biomaterials

Naturally derived biomaterials can be categorised as several groups depending on

their chemical structure and origin. Here are some examples:
1. Proteins: Proteins are a diverse group of biomolecules that are composed of amino
acids [5]. They can be created with a variety of natural sources like animal tissues,
plants, and microorganisms. Among the naturally derived protein biomaterials
are collagen, gelatine, silk fibroin, and elastin.
2. Carbohydrates: Carbohydrates are a class of biomolecules that are made up of
carbon, hydrogen, and oxygen [6]. They can be obtained from numerous natural
sources, such as plants, algae, and microorganisms. Examples of naturally derived
carbohydrate biomaterials include chitosan, hyaluronic acid, and cellulose.
3. Lipids: Lipids are a class of biomolecules that are composed of fatty acids
and glycerol. They can come from a range of natural sources, such as animal
tissues and plant oils. Examples of naturally derived lipid biomaterials include
phospholipids, triglycerides, and waxes [7].
4. Nucleic acids: Nucleic acids are a class of biomolecules that are composed of
nucleotides [8]. There are numerous natural sources from which they can be
obtained, such as animal and plant tissues, as well as microorganisms. Exam-
ples of naturally derived nucleic acid biomaterials include DNA, RNA, and
oligonucleotides [9].
5. Extracellular matrix (ECM) components: The extracellular matrix consists of
an intricate network of proteins and carbs that provides structural support and
regulates cell behaviour in tissues [10]. ECM components can be extracted from
natural sources, including animal tissues or plant materials. Examples of naturally
derived ECM biomaterials include collagen, hyaluronic acid, and decellularized
tissues [11].
6. Bio-based polyester: Common polyester is not naturally derived; it is possible to
create bio-based polyesters by using sustainable resources, including plant-based
oils and sugars. Bio-based polyesters have a variety of prospective uses in the field
of biomaterials. For example, they can be used to create biodegradable sutures,
8 A. H. Jasni et al.

implants, and drug delivery systems. One advantage of bio-based polyesters is

that they can be designed to break down over time, reducing the risk of long-term
harm to the environment.
These biomaterials can be categorised further depending on their physical and
chemical characteristics. properties, such as their solubility, biodegradability, and
mechanical strength. This classification can help researchers select the appropriate
biomaterials for specific applications in tissue engineering, drug delivery, and other
biomedical fields.

6 Polysaccharides

Polysaccharides are lengthy chains of monosaccharide (simple sugar) molecules

that make up complex carbohydrates [12]. They are found in many natural sources
such as plants, animals, and microorganisms [13]. Polysaccharides play a variety
of roles in living organisms, such as energy storage, structural support, and cellular
communication [14]. Here are some examples of polysaccharides and their chemical,
physical properties, and functions.

6.1 Starch

Starch is a polysaccharide that acts as the main component for storing energy in
plants. Amylose and amylopectin, two different forms of glucose polymers, make up
the substance [15]. Amylopectin is a branching chain of glucose molecules, whereas
amylose is a linear chain [16]. Starch is a naturally occurring polymer composed
of glucose units that can be found in a variety of plant-based products, including
corn, wheat, rice, potatoes, and cassava [17]. It is frequently utilised as a thickening,
stabilizer, and gelling agent in the food sector because of its unique chemical and
physical properties [18].

6.1.1 Chemical Properties of Starch

Starch is insoluble in cold water but can be partially solubilized by heat or acid
hydrolysis, breaking down the starch into smaller glucose units. Starch has a high
molecular weight, which makes it viscous and difficult to dissolve [19] especially
in the electrospinning and membrane casting applications. Starch can be effectively
dissolved using ionic liquids, 4-methylmorpholine 4-oxide (NMMO), and dimethyl
sulfoxide (DMSO), with DMSO being the most cost-effective and soluble [20].
Raw starch must be chemically changed to make it more hydrophobic, stronger
Naturally Derived Biomaterials: Advances and Opportunities 9

film-forming abilities, and higher tensile resilience to broaden the applications of

starch.

6.1.2 Physical Properties of Starch

Starch has a white, odourless, and tasteless appearance [21]. Starch has a large
capacity to store water and can hold up to 20 times its weight in liquid [22]. Starch
can form a gel when heated in water, allowing it to thicken food products and increase
their viscosity [23]. Starch is a renewable and biodegradable resource, making it
an attractive material for use in various applications beyond food. Natural derived
biomaterials derived from starch can include:
1. Bioplastics: As an alternative, starch-based bioplastics can be used to replace
traditional petroleum-based plastics and can be used in packaging materials [24],
disposable utensils, and other single-use products
2. Adhesives: Starch-based adhesives are used in paper and cardboard production
and can replace synthetic adhesives [25].
3. Textiles: Starch-based materials can be used in textiles as a sizing agent, which
helps prevent shrinkage during washing and ironing [26].
4. Medicine: Starch can be used as a binder in tablet formulations and as a
disintegrant to help tablets dissolve in the body [27].
5. Agriculture: Materials made of starch can be utilised as a biodegradable substitute
to synthetic mulch films used in agriculture [28].

6.2 Glycogen

Glycogen is a polysaccharide that is the principal energy storage molecule in animals

[29]. It’s similar to starch. but is more highly branched and can be broken down
more rapidly to provide energy for cellular processes. Because of its unique features,
glycogen has sparked increased interest as a biomaterial in recent years. It is biocom-
patible, biodegradable, and readily available in large quantities, making it an attrac-
tive alternative to synthetic materials suitable for a variety of uses. One potential use
of glycogen is in drug delivery. Because glycogen is a natural product of the body
and in comparison, to synthetic materials, it is less likely to elicit an immunological
response or induce toxicity. Additionally, glycogen can be easily modified to allow
for controlled release of drugs. Consequently, it is an excellent option for targeted
medication delivery systems [30].
Glycogen also has the potential of a wound-healing material. It can be used to
create a scaffold for tissue regeneration [31], as well as to promote the growth of new
blood vessels. Because glycogen is highly branched, it can form a porous structure
that allows for the infiltration of cells and nutrients, aiding in the regeneration process.
For instance, Alkaline phosphatase (ALP) activity and gene expression investigations
10 A. H. Jasni et al.

revealed that collagen/glycogen/hydroxyapatite (HAP) components influenced the

differentiation of bone mesenchymal stem cells into osteoblasts or chondrocytes [31].
Overall, the use of glycogen as a biomaterial is still in its early stages, and more
research is needed to fully understand its potential. However, glycogen’s unique qual-
ities make it an intriguing contender for a variety of applications including medication
delivery, tissue engineering, and wound healing.

6.3 Peptidoglycan

Peptidoglycan is a polysaccharide found in bacterial cell walls [32]. For bacte-

rial cells, it offers structural stability and defence. It is made up of two sugars,
N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM), which are repeated
and connected by short peptides to form a mesh-like structure [33]. There has recently
been an increase in interest in using peptidoglycan as a biomaterial because of its
unique attributes. It is biocompatible, biodegradable, and can be easily isolated from
bacterial cell walls, making it an attractive alternative to a variety of applications for
synthetic materials.
One potential use of peptidoglycan is in tissue engineering. It can be used as a
scaffold for cell development, promoting the regeneration of the damaged tissue [34].
Additionally, peptidoglycan can be modified to provide growth factors or additional
bioactive compounds that can improve tissue regeneration [35]. Peptidoglycan also
has the potential as an antibacterial material [34]. It can be used to create coatings for
medical devices or implants, which can prevent bacterial colonization and biofilm
formation. Additionally, peptidoglycan can be modified to incorporate antimicrobial
peptides, which can further enhance its antibacterial properties.

6.4 Cellulose

Cellulose is a polysaccharide that makes up the structural component of plant cell

walls. It is composed of cross-linked, lengthy chains of glucose molecules. to form
a strong and rigid structure [36]. Cellulose is widely used as a biomaterial due to
its renewable nature, biocompatibility, and biodegradability [37]. Recent advance-
ments in the use of cellulose as a biomaterial have focused on developing new
processing techniques, modifying its chemical and physical properties, and exploring
its potential applications in various fields.
One recent advancement is the development of nanocellulose, which refers to
cellulose fibres that have been broken down into smaller nanoscale dimensions.
Nanocellulose has several special merits including high stiffness, strength, and
surface area, making it suitable for use in a variety of applications [38] including
packaging materials, medical devices, and drug delivery systems.
Naturally Derived Biomaterials: Advances and Opportunities 11

Another recent advancement is the development of cellulose-based hydrogels,

which are water-swollen networks of crosslinked cellulose fibres. These hydrogels
have shown promise in a range of applications, including tissue engineering, drug
delivery, and wound healing. The use of bacterial cellulose multifunction bandage
with potent antibacterial capabilities and wound healing for angiogenesis stimulators
had been reported by [39].
Cellulose has also been explored for use in 3D printing, where it can be used as
a feedstock material. By combining cellulose with other materials, such as chitin or
lignin, 3D printed structures with enhanced mechanical and physical properties have
been developed. For instance, the greatest capability was shown by methylcellulose
and wood glue, with the former having the benefit of being natural and producing a
bio-based composition. The result was a homogeneous paste with moderate adhesion
and viscosity, like the clay frequently used in 3D printing, that could be used with
a caulking gun and a syringe to create multi-layered extrusions that were stable and
smooth. Although it lacked the strength and rigidity of wood, it performed better than
other bio-based new materials currently being developed thanks to its mechanical
qualities, which put it on par with rigid polymer foams frequently used as insulation
boards [40].

6.5 Chitin and Chitosan

Insects, crustaceans, and other arthropods’ exoskeletons are made of the carbohy-
drate chitin It is also present in some fungi cell walls [41]. Chitin provides strength
and protection to these organisms. Chitin can be processed into nanofibers, nanopar-
ticles, and other nanostructures, which have special characteristics including a large
surface area, mechanical strength, and biocompatibility [42]. These nanomaterials
have potential applications in wound healing, tissue engineering, and drug delivery.
Another recent development in the use of chitin is when it comes to food packing
[43]. In the not-too-distant future, it has been reviewed by authors in [44] that a
significant amount of chitin and chitosan can be derived from mushrooms [44]. There
are bright prospects for the industrial use of fungus-sourced chitosan, especially
within the food and pharmaceutical industries, as a result of the comparisons made
between the physicochemical, functional, and biological traits of that substance and
that of marine chitin and chitosan [44].
Chitin has also been explored for its potential use in agriculture as a biopesticide.
Chitosan can act as a natural insecticide, disrupting the growth and development of
pests while being safe for humans and environment [45]. Additionally, chitin can be
used as a soil conditioner, enhancing soil fertility and plant growth [46]. It was found
that the exuviae of a particular insect species (mainly chitin) had enhanced bacteria
that grew after the topsoil underwent treatment with 10 g/kg in the rhizosphere of B.
oleracea. However, at a ratio of 1 g/kg, there were no appreciable changes in bacterial
abundance. The results are consistent with earlier research on chitin alterations, which
has shown to increase bacterial population abundance [47].
12 A. H. Jasni et al.

Chitin, a material present within the exoskeletons of crustaceans including crabs,

prawns and lobsters, is the source of chitosan, a naturally occurring polysaccharide
[48]. Fruits and vegetables have been coated with chitosan to prolong their shelf
life. The chitosan coating can slow down the rate of water loss and gas exchange,
thereby extending the freshness and quality of the produce [49]. Chitosan coatings
have been utilised to increase the implant surfaces’ antibacterial characteristics and
promote tissue integration [50]. Recent findings showed that the combination of
chitosan with enzymes and antimicrobial peptides enhances its antibacterial activity
and broadens its application in a variety of physiological circumstances. The combi-
nation of chitosan and polymers also produced coatings with improved antibacterial
and anti-adhesive qualities to reduce the incidence of implant-associated infections
[51].

6.6 Alginate

Brown seaweed is the source of the natural polymer known as alginate, such as
kelp [52]. This environmentally friendly and renewable biomaterial exhibits several
intriguing qualities, including good mild gelation when adding divalent cations (such
as Ca2+ ), biocompatibility, low toxicity, and cost effectiveness. High-molecular-
weight alginate’s poor solubility and high viscosity, the aqueous solution’s poly-
electrolyte nature, the lack of suitable organic solvents, the substantial amount of
intra- and intermolecular hydrogen bonds, continue to be problems in alginate alone
applications [53]. Thus, it must be coupled with other polymers to enhance its
values [54] that had developed natural polymer high-strength hydrogels using gelatin
and hydrazide alginate (HAlg), which were then crosslinked to mimic the struc-
ture of collagen and glycosaminoglycans (GAGs) within the extracellular matrix,
respectively. The Gelatin-HAlg-DN hydrogels exhibit excellent biodegradability and
swelling consistency in physiological conditions, as well as the capacity to facilitate
cell binding and proliferation. In a rat model with critical size bone defects, psoralen-
loaded gelatin-HAlg-DN hydrogels effectively induced bone regeneration offering
intriguing potential as tissue engineering scaffolds [54].

6.7 Hyaluronic Acid

A polymer present in the connective tissues called hyaluronic acid (HA) aids in
maintaining hydration and lubrication. It is also used in some cosmetic products
for its moisturizing properties [55]. Bioink additions include HA and multi-walled
carbon nanotubes (CNTs) in 3D bioprinting for fabrication of cartilage had been
reported [56]. HA acts as a molecule that is both structural and signalling. It possesses
extraordinary water-retention abilities that create a gel-like environment within the
tissue and give the entire structure flexibility for the structure [57].
Naturally Derived Biomaterials: Advances and Opportunities 13

6.8 Others

6.8.1 Inulin

Inulin a polysaccharide generated from plants, is a bioinspired, adaptable, and useful

biomaterial [58] In comparison to other biodegradable polysaccharides, inulin’s
distinctive and adaptable structure, stabilising and protecting properties, and capacity
to target specific organs make it an ideal drug delivery carrier. Each fructose unit
has three hydroxyl groups that act as an anchor for chemistry changes. Thus,
this enhances cellular bioavailability and absorption, and accomplishing medicines’
and biomolecules’ targeted, prolonged, and regulated release [58]. Inulin used via
solvent-casting technique was used to create sustainable water-based films for wound
healing [59] employed inulin (INL), a carbohydrate-based matrix, and polyvinyl
alcohol (PVA), a petroleum-free matrix. Due to its sustainability and antimicrobial
qualities, pumpkin powder made from vegetable waste was employed in the study
[59].

6.8.2 Xylan

Xylan is a fluoropolymer-based commercial coating that is mostly found in nonstick

cookware, footware and automotive coatings due to its water repellent characteristic
[60]. Click-chemistry techniques were used to create reactive xylan compounds that
can be crosslinked chemically and functionalized further. Quantitatively, functional
xylan carbamates (XCs) were created from xylan phenylcarbonates (XPCs) along
with magnitude of substitution (DS) with propargyl amine (PA) or 6-azidohexan-
1-amine (AA). Alkynyl-functionalized XCs were crosslinked with bisazide linkers
using copper-catalyzed 1,3-dipolar cycloaddition within an organic medium, and it
was demonstrated that the gelation process produced a reactive moiety plus effective
solubility in water for additional appropriate applications [61].

6.8.3 Proteins

Proteins are substantial, intricate molecules consisting of long chains of amino acids
[62]. They are found in every cell of every living organism and are involved in a variety
of functions, such as providing structural support, catalysing chemical reactions,
and transporting molecules within cells and throughout the body. A protein’s linear
sequence of amino acids makes up its primary structural component. These consistent
folding structures, alpha helices and beta sheets, make up a protein’s secondary
structure. The collection of curves and folds in a single linear chain of amino acids,
sometimes referred to as a polypeptide, is the tertiary structure of a protein. Last but
not least, macromolecules having multiple polypeptide chain structures or subunits
are known as proteins [63]. Figure 1 is the illustration each type of protein structure.
14 A. H. Jasni et al.

Fig. 1 Illustration of type of protein structure (Taken from open source, [Link]
bea.2021.100021)

Here are some examples of protein-based naturally derived biomaterials and their
functions:
• Structural proteins: Structural proteins, such as collagen, keratin and elastin,
provide support and shape to tissues and organs in the body [64].
• Transport proteins: Transport proteins, such as haemoglobin and albumin, bind
to and transport molecules, such as oxygen and nutrients, throughout the body.
For example, one of the most prevalent proteins in blood plasma, serum albumin
is crucial to all biological activities and has been used in several biomedical
procedures. Human albumin, bovine albumin, and ovalbumin, which are used to
manufacture biomaterials, are appropriate for use in bone regeneration because
they have the right microstructure, hydrophilicity, and biocompatibility [65].
• Contractile proteins: Actin and myosin are examples of contractile proteins that
cause muscles to contract and move.
In summary, proteins are necessary molecules that have several functions in living
things. Their unique properties, such as their ability to provide structural support,
and transport molecules, make them important in many material science applications,
such as food, medicine, and biotechnology.
Naturally Derived Biomaterials: Advances and Opportunities 15

Fig. 2 Collagen fibre structure, from fibrils and molecules to chains, together with amino acid
sequence of Hydroxyproline, Proline, and Glycine. (Created with biorender)

6.8.4 Collagen

The protein collagen, which is the most prevalent in the human body, gives numerous
tissues their structural support [66]. It has been frequently employed beyond the field
of tissue engineering owing to its remarkable biocompatibility, low immunogenicity,
and ability to support cell adhesion and proliferation. Collagen is made up of three
chains. The chains are linked to form a triple helix. Because glycine is the smallest
amino acid, it permits the chain to form a tight shape and endure stress [67]. Figure 2
is the structure of the collagen fibre.

6.8.5 Silk Fibroin

Silk is a naturally occurring protein fibre produced by silkworms. It has been used in
tissue engineering, implant coating and others due to its biocompatibility, biodegrad-
ability, and ability to form scaffolds with tuneable mechanical properties [68]. A
recent study reported the possibility of encouraging the development of bone-like
tissue from human dental pulp stromal cells (hDPSCs) both in in vitro as well as
in vivo within porous Bombyx Mori silk structures by employing the specific HDAC2
and 3 inhibitor MI192. Fabricated and evaluated were scaffolds made of 2 and 5
wt.% silk. In contrast to the 2 wt% silk scaffolds, the scaffolds with a weight of
5% exhibit larger internal lamellae, slower rates of expansion and deterioration, and
higher torsional moduli. The alkaline phosphatase activity of hDPSCs on a 5 wt%
16 A. H. Jasni et al.

Fig. 3 The technique used for fabrication silk scaffolds for biomedical applications (adapted from
open source, [Link]

silk scaffold was dramatically increased by pre-treatment with MI192 (ALP). Histo-
logical examination proved that, in contrast to the control group, the use of MI192-
pretreated hDPSCs-silk scaffolds improved bone extracellular matrix (ALP, Col1a,
and OCN) deposition and mineralization. After six weeks of subcutaneous implanta-
tion in mice, the MI192-pretreated hDPSCs-silk scaffold constructs increased vascu-
larization and extracellular matrix mineralization compared to the untreated control.
After six weeks of subcutaneous implantation in nude mice, the MI192-pretreated
hDPSCs-silk scaffold constructs increased vascularization and extracellular matrix
mineralization compared to the untreated control [69]. Figure 3 is the workflow of
silk fabrication and application.

6.8.6 Gelatin

Gelatin is a substantially hydrolyzed version of collagen with characteristics similar

to collagen. It can be easily modified to form hydrogels with tunable mechanical
properties, making it a suitable material for tissue engineering.

6.8.7 Elastin

Elastin materials can be derived from organic, recombinant, or synthetic sources.

Elastin can be recovered from elastin-rich tissues of animals by hydrolyzing a portion
of the peptide linkages in the insoluble elastin [70].
Naturally Derived Biomaterials: Advances and Opportunities 17

6.9 Others

6.9.1 Fibrin

Fibrin is a naturally occurring protein that aids in the formation of clots and wound
healing. It is utilized in tissue engineering to form hydrogels that can support cell
growth and migration [71].

6.9.2 Lipids

Lipids are a wide class of naturally occurring macromolecules present in plants and
animals [72]. They are distinguished by their hydrophobic characteristics and are
used in a number of biomaterials research applications. One major use of lipids
is in the creation of liposomes. Liposomes are spherical vesicles consisting of a
bilayer of phospholipids that can be used to encapsulate drugs and other therapeutic
molecules for targeted drug delivery [73]. The inside of the liposome is formed by
the hydrophilic portions of the phospholipids, which contain the drug, while the
hydrophilic head groups form the outer shell, allowing the liposome to interact with
biological systems [74].
Lipids are also used in the creation of biodegradable polymers. Lipid-based poly-
mers can be designed to degrade in vivo, making them suitable for use in the fields of
tissue engineering and drug delivery [75]. They are additionally employed to produce
micro- and nanoparticle for targeted medication delivery [76]. Another use of lipids is
the formation of lipid membranes. Lipid membranes are used in biomimetic systems
to mimic the properties of natural cell membranes [77]. They can be used for a
variety of purposes, including water treatment, biological sensors, and drug detection
[78]. Lipids are additionally useful to create hydrogels. Hydrogels comprise three-
dimensional, water-swollen polymer networks that can be useful for drug delivery,
wound care, and tissue engineering. Lipids can be incorporated into hydrogels to
provide mechanical support and improve cell adhesion and proliferation [79].
Overall, lipids are a versatile biomaterial with many potential applications in
biomaterial science. Their hydrophobic properties and ability to self-assemble make
them appealing candidates for drug delivery, tissue engineering, and biomimetic
system.

6.9.3 Fatty Acids

Fatty acids are naturally occurring biomolecules that are commonly found in plant
and animal tissues. They are composed of long hydrocarbon chains with a carboxyl
group at one end, making them amphipathic like phospholipids [72]. Fatty acids have
many uses in biomaterial science due to their biocompatibility, biodegradability, and
versatility.
18 A. H. Jasni et al.

One major use of fatty acids is in the creation of bio-based polymers. Fatty acids
can be used as monomers to create polymers such as polyesters, polyamides, and
polyurethanes. These polymers are used for drug delivery, tissue engineering, and
exterior coatings thar are only a few of the potential applications [80]. Hydrogels are
also manufactured using fatty acids. Hydrogels are cross-linked networks of liquid-
swollen polymers which are able to be utilized for the administration of drugs, wound
healing, and other applications. tissue engineering. Fatty acids can be incorporated
into hydrogels to provide mechanical support and improve cell adhesion and prolif-
eration. A recent study showed that hydrogenated phosphatidylcholine and oleic acid
were used to create a highly hydrated hydrogel (95% water) for potential biomedical
uses [81].
Another application of fatty acids is in the production of surfactants. Surfactants
are molecules that can reduce the surface tension of liquids and increase their ability to
mix with other substances. Fatty acids can be used to create natural surfactants that are
biodegradable and non-toxic, making them suitable for use in personal care products,
detergents, and other industrial applications. It has been known that oleochemical-
based surfactants outperform petroleum-based ones in terms of biocompatibility
and biodegradability [82]. Sorbitan esters are commercially known as “Span” in
the contemporary industrial sector, specifically Span 80, a biodegradable surfactant
made from oleic acid and sorbitol-based sugar alcohol. These esters are commonly
used in the development and formulation of water-in-oil emulsions due to their rela-
tive hydrophobicity. Sorbitan esters are widely derivatized by combining with ethy-
lene oxide to form sorbitan ester ethoxylates, which enhances their hydrophilicity.
Polyethoxylated sorbitan monoesters, popularly known as “Tween” in the industry,
are surfactants that are ideal for creating oil-in-water emulsions [83, 84].
Overall, fatty acids are a versatile biomaterial with many potential applications
in biomaterials science. Their biocompatibility, biodegradability, and versatility
make them an attractive option for the development of sustainable and biologically
compatible materials.

6.9.4 Phospholipids

Phospholipids are naturally occurring biomolecules that are found in cell membranes
and other biological structures. They are amphipathic, meaning that they have both
hydrophilic (water-loving) and hydrophobic (water-repelling) regions [85]. This
unique property makes phospholipids useful in a variety of applications in biomaterial
science.
One major use of phospholipids is in drug delivery systems. Phospholipid-based
liposomes can be used to encapsulate drugs and other therapeutic molecules, allowing
for targeted delivery to specific tissues or cells. The liposome’s outer shell is formed
by the hydrophilic head regions of the phospholipids, while the hydrophobic tails
form the interior, where the drug is contained. The liposome can then be designed to be
released by the reaction to certain stimuli, such as changes in pH or temperature, and
the medication is released. The optimum phospholipid for managing the release of
Naturally Derived Biomaterials: Advances and Opportunities 19

the anaesthetic bupivacaine (BUP) medication from liposomal depots and controlling
the regulated aggregation of the liposomes is phosphatidylglycerol (PG) [86].
Phospholipids are also used in the creation of biomimetic membranes, which are
synthetic structures designed to mimic the properties of natural cell membranes.
These membranes have a wide range of uses, including water filtration, biosensors,
and drug screening. The amphipathic nature of phospholipids allows them to form
bilayers, which can be used as the foundation for these biomimetic membranes. The
best exemplar of this is a study done by [87] where the cell membrane phospho-
lipids are functions as dentin’s mineralization sites and potential raw materials for
bottom-up strategies aiming for quicker and more intricate production of dentin-like
structures.
Phospholipids are also used in tissue engineering applications. They can be added
to scaffolds to offer structural support while also promoting cell adhesion and prolif-
eration. Because of their improved blood compatibility and cytocompatibility, elec-
trospun poly (lactic-co-glycolic acid) (PLGA) membranes were a suitable case in
point [88]. Overall, phospholipids’ distinct features make them flexible biomaterials
with numerous uses in pharmaceutical delivery, tissue engineering, and biomimetic
membrane design.

6.9.5 Waxes

Wax is a simple lipid that is generated by esterifying a long-chain alcohol with a fatty
acid. Alcohol can have 12–32 carbon atoms. Waxes are found in nature as coatings
on plants and stems. The wax protects the plant from excessive water loss [89]. In
practise, cellulose combined with a trace amount of plant-derived wax (nonacosane-
10-ol and nonacosane-5, 10-diol) has a higher mechanical strength and modulus of
elasticity [90] than synthetic plastic. The diffusion coefficients of oxygen, nitrogen,
and water molecules through this material were found to be at least 50% lower than
those found in polyethylene. It is an environmentally friendly, long-lasting packaging
material with outstanding mechanical, thermal, and barrier properties [90].

7 Synthesis and Modification of Naturally Derived

Biomaterials

The synthesis of naturally derived biomaterials involves the extraction, purification,

and modification of the natural source to obtain a suitable material for the intended
application. The following approaches can be used for the synthesis of naturally
derived biomaterials:
1. Extraction: The first step in the synthesis of naturally derived biomaterials
is the extraction of the material from its natural source [91]. The extraction
method varies depending on the source material and the intended application.
20 A. H. Jasni et al.

For example, collagen can be extracted from animal tissues using acid or enzy-
matic digestion [92], while cellulose can be extracted from plants using chemical
or mechanical methods [93].
2. Purification: After extraction, the material is usually purified to remove impurities
and obtain a pure form of the material. Purification methods include filtration,
centrifugation, and chromatography [94].
3. Modification: Modification of naturally derived biomaterials is frequently
required to modify their qualities and adapt them to certain uses. Modification
can be achieved through chemical or physical methods. For example, cross-
linking of collagen can improve its mechanical strength and stability, while
surface modification of cellulose can improve its biocompatibility [95].
4. Characterization: Characterization of naturally derived biomaterials is essen-
tial to determine their properties and suitability for the intended application.
Characterization methods include spectroscopy, microscopy, and mechanical
testing.
5. Quantification: Quantification is the act of assigning a numerical value to a
measurement of anything, or counting the quanta of whatever is being measured.
Quantification creates a defined method of measurement that permits statistical
operations and mathematical computations to be performed [96].
6. Pilot production: A pilot study, pilot project, pilot test, pilot production or pilot
experiment is a small-scale preliminary study undertaken prior to the performance
of a full-scale research project to evaluate feasibility, duration, cost, adverse
occurrences, and to enhance the study design [97]. Figure 4 is the summary of
biomaterials synthesis workflow.

Fig. 4 Workflow and techniques in biomaterials synthesis

Naturally Derived Biomaterials: Advances and Opportunities 21

8 Chemical Modification

Chemical modification of naturally derived biomaterials is a crucial step in devel-

oping biomaterials with improved properties and functionality [98]. The chemical
modification can be performed on different biomaterials such as proteins, polysac-
charides, and lipids. Here are some examples of chemical modifications of naturally
derived biomaterials:
1. Crosslinking: Crosslinking is the development of covalent connections between
polymer chains in a biomaterial, resulting in improved mechanical properties
and stability. Crosslinking can be achieved using various crosslinkers, such
as glutaraldehyde, genipin, and formaldehyde [99]. Other than that hydrogel
formation involves the crosslinking of a hydrophilic polymer to form a three-
dimensional network, resulting in improved mechanical properties and biocom-
patibility [100]. Hydrogels can be formed from various biomaterials such as
alginate, chitosan, and gelatin.
2. Acylation: Acylation involves the introduction of acyl groups into the biomaterial,
resulting in improved solubility, stability, and biocompatibility. For example,
acylation of chitosan can improve its solubility in organic solvents, making it
easier to process for various applications [101].
3. Grafting: Grafting involves attaching a functional group or a polymer onto the
surface of the biomaterial, resulting in improved properties such as biocompat-
ibility and hydrophilicity. Grafting can be achieved using various methods such
as surface-initiated polymerization, click chemistry, and plasma treatment [102].
4. Glycosylation: Glycosylation involves the addition of sugar moieties onto the
biomaterial, resulting in improved bioactivity and biocompatibility. For example,
glycosylation of proteins such as collagen and elastin can improve their interac-
tions with cells and tissues. A case of this was the use of glycosylation in biomate-
rial engineering by adding saccharides to the catalogue of construction blocks to
embellish it with adhesion properties. The use of a Campylobacter jejuni glyco-
sylation circuit to de novo glycosylate the Bacillus subtilis amyloid-like biofilm
protein TasA was reported as a unique biomaterial engineering technique for
increasing the adhesiveness of TasA fibrils [103].
In conclusion, chemical modification of naturally derived biomaterials can
improve their properties and functionality, making them suitable for various biomed-
ical applications. The method of modification used is determined on the biomaterial
and the desired application as well.
22 A. H. Jasni et al.

9 Physical Modification

Physical modification of biomaterials refers to the changes made to the physical

properties of a material without altering its chemical composition [104]. This can
include changes to the surface topography, roughness, stiffness, and shape of the
material. Some common physical modifications of biomaterials include:
1. Surface modification: This involves changing the topography and roughness
of the surface of a material to improve its interaction with cells and tissues.
Surface modification techniques can include plasma treatment, micro- and
nano-patterning, and coatings [105].
2. Mechanical modification: This involves altering the stiffness and elasticity of
a material to mimic the mechanical properties of natural tissues. This can
be achieved through various methods of mechanical modification such as
mechanical stretching, compression, and shear forces [106].
3. Shape modification: This involves changing the shape and size of a material
to better fit a specific application or tissue. Techniques can include moulding,
cutting, and 3D printing [105].
4. Porosity modification: This entails altering a material’s pore size and shape to
promote cell infiltration and tissue ingrowth. This can be accomplished using
a variety of procedures such as salt leaching, solvent casting, and gas foaming
[107].
Physical modification of biomaterials can greatly enhance their biocompatibility,
bioactivity, and mechanical properties, making them better ideal for a wide range
of biomedical applications, including tissue engineering, medication delivery, and
medical devices.

10 Enzymatic Modification

Enzymatic modification of biomaterials refers to the surface modification with

enzymes or bulk properties of biomaterials without changing their chemical compo-
sition [108]. Enzymes are biological catalysts that can selectively modify the surface
functional groups of biomaterials to add new functions, improve biocompatibility,
and enhance the ability of the material to interact with cells and tissues. Some
common enzymatic modifications of biomaterials include:
1. Surface functionalization: This entails using enzymes to change the outermost
layer of a biomaterial in order to introduce new functional groups that can facili-
tate cell adhesion and proliferation. For example, enzymes like horseradish perox-
idase [109] and tyrosinase can be used to generate reactive oxygen species [110]
that can covalently bind proteins and peptides to the surface of the biomaterial.
Naturally Derived Biomaterials: Advances and Opportunities 23

2. Degradation: This involves the use of enzymes to selectively degrade the bioma-
terials to control the rate of release of bioactive molecules, drugs, or cells from
the biomaterial. For example, enzymes like metalloproteinase [111] collagenase,
hyaluronidase, and chondroitinase can be used to selectively degrade collagen,
hyaluronic acid, and chondroitin sulphate, respectively [112].
3. Cross-linking: This involves the use of enzymes to cross-link biomaterials to
enhance their mechanical and structural properties. For example, enzymes like
transglutaminase can be used to cross-link proteins, such as gelatin and collagen,
to form stable hydrogels [113].
4. Surface cleaning: This involves the use of enzymes to remove contaminants and
impurities from the surface of the biomaterial to improve its biocompatibility.
For example, enzymes like lipase and protease can be used to remove lipids and
proteins from the surface of the biomaterial [114].
Enzymatic modification of biomaterials is an intriguing approach for improving
biocompatibility and performance for a variety of purposes, namely drug delivery,
tissue engineering, and medical devices.

11 Applications of Naturally Derived Biomaterials

11.1 Medical and Biomedical Applications

Because of their capacity for biocompatibility, biodegradability, and propensity to

replicate the extracellular matrix (ECM) of natural tissues, naturally produced bioma-
terials have been extensively researched for use in tissue engineering. Because of
their biocompatibility, biodegradability, and ability to package and transport medi-
cations to specific tissues, naturally produced biomaterials have showed consider-
able promise in drug delivery. Some naturally generated biomaterials that have been
employed in biomedical applications are listed in Table 2.
Naturally derived biomaterials offer many advantages over synthetic materials
in tissue engineering, drug delivery and implant coating, including their capacity
to replicate the ECM of living tissues and promote cell adhesion and prolifera-
tion. However, there are also some limitations, such as batch-to-batch variability,
difficulty in controlling their mechanical properties, and potential immunogenicity.
Overall, naturally derived biomaterials hold great promise for the development of
safer products for these constructs that can more closely resemble natural tissues.

12 Industrial Applications

Naturally derived biomaterials have several industrial applications, including:

24 A. H. Jasni et al.

Table 2 Recent discoveries of naturally produced biomaterials used in biomedical applications

No. Type of biomaterials Findings Study
Tissue engineering
1 Fibrin: • A wound dressing consisting of [115]
– Is a naturally occurring protein that helps sodium carboxymethylcellulose
the body’s blood clot and repair wounds (Hcel® NaT), fibrin, and
– It has been used in tissue engineering to in vitro-seeded dermal fibroblasts
support cell growth and migration • Fibrin promotes healthy cell
adhesion and spreading, and aids
in cell migration and subsequent
proliferation in a wound
2 HA+Chitosan • Degradable hydrogel from N, [116]
O-Carboxymethyl Chitosan
(NOCC), Aldehyde-Hyaluronic
Acid, and the addition of Allium
sativum (garlic oil)
• A breakthrough for preventing
intraperitoneal adhesion following
surgery
3 Chitosan+collagen type 1, hyaluronic acid, • Improved cartilage and bone cell [117]
Poly(L-lacticacid)/gelatin/β-tricalcium lesions repair when mesenchymal
phosphate, gamma-poly [glutamic acid] stem cells (MSCs) are combined
polyelectrolyte/titanium alloy, modified with chitosan-based scaffolds
Poly(L-Lactide-co-Epsilon-Caprolactone), • The scaffold’s synergistic effects
calcium phosphate, β-glycerophosphate on the proliferation and
hydrogel/calcium phosphate cement differentiation of MSCs into bone
(CPC), and CPC-Chitosan-RGD and cartilage tissue
Drug delivery
1 Albumin • Flexible hollow human serum [118]
albumin (HHSA) loaded with the
chemotherapy medication
doxorubicin (DOX) and the
photosensitizer chlorin e6 (Ce6)
for synergistic cancer treatment
• Stronger therapeutic effects
without radiation development of
4T1 breast tumours
• Surprisingly, remarkable
biocompatibility has been
demonstrated both in vitro and
in vivo
(continued)
Naturally Derived Biomaterials: Advances and Opportunities 25

Table 2 (continued)
No. Type of biomaterials Findings Study
2 Collagen • Collagen nanostructures using the [119]
scales of fish were produced for
the very first time using
desolvation techniques
• According to the histological and
macroscopical study, the synthetic
fish scale’s collagen nanomaterials
aided in the healing process with
little toxicity compared to the
saline group
3 Alginate-chitosan+HA • Alginate-chitosan microbeads and [120]
thermo-responsive hyaluronic
acid-poly(N-isopropylacrylamide)
(HA-pNIPAM) hydrogels have
been created as bioresorbable local
bacteriophage (Staphylococcus
aureus phage ISP and
Pseudomonas aeruginosa phage
LUZ19) delivery systems to
permit both delayed and quick
phage release
• Both phages were gradually
released hydrogels HA-pNIPAM
during the course of 21 days. The
short-tailed LUZ19 was
continuously released from the
microbeads for 21 days, whereas
the long-tailed ISP released in
bursts at first and then at a
declining rate over time
4 Fibrin • Research on the drug release and [121]
antibacterial properties of
platelet-rich fibrin (PRF), a natural
vehicle for the administration of
antibiotics
• Following oral surgery, using
antibiotic-loaded PRF may reduce
the risk of post-operative infection,
replace or enhance systemic
antibiotic therapy, and maintain
the healing properties of PRF
(continued)
26 A. H. Jasni et al.

Table 2 (continued)
No. Type of biomaterials Findings Study
5 Cellulose • Using an in-situ co-precipitation [122]
approach, Magnetic iron oxide
(MIO)-incorporated waste tissue
sheets (WTP) and sugarcane
bagasse (SCB) were incorporated
to make WTP/MIO and SCB/MIO
nanocomposite particles (NCPs)
• By including MIO-NPs into the
cellulose matrix, the swelling
capacity, drug loading capacity,
and drug release time were all
improved
Implant coatings
1 Laminin • A poly (D, L-lactide) [123]
(PDLLA)-Laminin 332 (LN332)
hybrid film that displayed timed
release of LN332for up to 28 days
was successfully created using a
layer-by-layer manufacturing
process
• It accelerated gingival
mesenchymal stem cells’ epithelial
growth and promoted their
adhesion, dissemination, and
proliferation
2 Hydroxyapatite • 10 implants featuring 10 implants [124]
having a dual acid-etching (DAA)
surface and a nanostructured
hydroxyapatite coating (HAnano).
was given to ten lambs (aged 2–4)
• Low-density bone in sheep after
28 days with HAnano enhances
bone development as compared to
DAA surface
3 HA • Tannic acid (TA) and hyaluronic [125]
acid (HYA) use is necessary to
lower the rate of corrosion of
implants made of magnesium
alloys
• The magnesium alloy’s corrosion
rate was lowered by the TA-HYA
coating from 7.379 mm/year to
0.204 mm/year in in vitro
corrosion testing employing Tafel
polarisation
(continued)
Naturally Derived Biomaterials: Advances and Opportunities 27

Table 2 (continued)
No. Type of biomaterials Findings Study
4 Silk • FibroFix Cartilage P™ [126]
(FibroFix™)—commercial brand
name
• Substantial bone repair and strong
implant tissue integration, which,
in our opinion, will speed up the
healing process (promote tissue
regeneration)
5 Collagen • An innovative injectable medicinal [127]
product on primary cultures of
human gingival fibroblasts (hGF),
dental SKIN BioRegulation (Guna
S.p.A., Milan, Italy) was put to the
test
• It is made of type I collagen of
porcine origin
• Significant improvement in 48-h
viability of hGF-grown cells (p =
0.05) and 24-h wound healing (p
= 0.001)

1. Food industry: Natural polymers with gelling, thickening, and stabilising

qualities, like cellulose, chitosan, and alginate, are utilised as food additives.
2. Agriculture: Natural biopolymers such as starch and cellulose are used as plant
growth promoters and soil conditioners.
3. Textile industry: Natural polymers such as silk, cotton, and wool are used to
make textiles.
4. Packaging industry Biodegradable packaging materials are created using natural
polymers like cellulose, chitosan, and starch.
5. Cosmetic industry: Natural polymers with moisturising and anti-aging qualities,
like collagen, elastin, and hyaluronic acid, are employed in cosmetic products.
6. Medical industry: Medical uses for naturally generated biomaterials include
tissue engineering, medication delivery, and wound healing. Examples include
collagen, chitosan, and hyaluronic acid.
7. Construction industry: Building materials are produced using natural biopoly-
mers like cellulose and lignin for insulation, cement composite, and adhesives.
The use of naturally derived biomaterials in these industries offers several advan-
tages over synthetic materials, including their biodegradability, biocompatibility,
and renewability. Their use in industry is also in keeping with the rising demand for
environmentally friendly and sustainable materials. Table 3 summarizes the recent
discoveries within industrial applications. Composite made up of Oyster shell is
shown in Fig. 5.
28 A. H. Jasni et al.

Table 3 Recent industrial applications of naturally derived biomaterials

No. Sample type Findings References
Food industry
1. Polylactic acid (PLA) • Bilayer films were [128]
developed that included a
PLA layer with a layer
made of washed
cottonseed meal (CSM)
• The elongation at break
has decreased while the
Young’s modulus and
tensile strength have both
increased
Agriculture industry
2. Chitosan • Hydrogels made of [129]
chitosan that have
embedded mineral
fertilisers were developed
• It was discovered that the
experimental group’s
seedling survival rate was
40% greater than the
control group
3. Chitosan • Vegan chitosan from [130]
Cunninghamella
echinulate was extracted
• It has been utilised as an
alternative to chemical
treatments to manage
plant diseases
Textile industry
4. Bacterial cellulose • Adapted microbial [131]
weaving process via
Komagataeibacter
rhaeticus bacteria
• The incredibly tiny
nanocellulose fibres are
eight times stiffer and
stronger than steel
(continued)
Naturally Derived Biomaterials: Advances and Opportunities 29

Table 3 (continued)
No. Sample type Findings References
5. Cellulose • Bio sequins fabricated [132]
from bioplastic derived
from tree woods namely
‘Bio Iridescent’
• Traditional sequins are
produced of polyvinyl
chloride (PVC), a
polyester film, often
known as Mylar, which
poses serious
environmental and health
dangers by creating
hazardous, bioaccumulate
compounds, including
hormone disruptors and
carcinogens like
phthalates
Packaging industry
6. Glycol • monoethylene glycols [133]
(meg) and monopropylene
glycols (mpg) from woods
• a genuine greener option
for making recyclable pet,
polyesters, and industrial
liquids
Cosmetic industry
7. Glycol • A fully environmentally [133]
friendly option for the
manufacture of carrier
liquids. the secret to
providing a generation of
resins, detergents,
de-icing agents, and
cosmetics that is
sustainable and renewable
8. PLA+Polyhydroxyalkanoates (PHA) • The cosmetic case is made [134]
of polylactic acid (PLA)
and amorphous PHA
technology for a private
brand, ‘Wakemake’
• An alternative substitute
of acrylonitrile butadiene
styrene (ABS) casing
(continued)
30 A. H. Jasni et al.

Table 3 (continued)
No. Sample type Findings References
Medical industry
9. HA • HA-coated gold [135]
nanoparticles (AuNP-HA)
synthesised in one pot.
Usage as a drug
(Sulfasalazine) (SSZ),
delivery vehicle
• AuNP-HA exhibits a
promising material as a
modern medicine delivery
method with
characteristics of
controlled release
10. Collagen+Glycogen+Hydroxyapatite • Composite hydrogels [31]
functions as bone repair
scaffold
• When cultivated on the
composite hydrogels,
bone mesenchymal stem
cells displayed favourable
cell adhesion, vitality, and
proliferation
Construction industry
11. Cellulose • Food leftovers are [136]
combined with water and
seasoning, then heated up
and pressed into a mould.
It is called ‘Fabula’
• Has a bending strength
four times more than that
of concrete and is
primarily made of
vacuum-dried, ground-up
fruit peels and coffee
grounds
12. Alginate+Oyster shells • ‘Ostra’-A biomaterial [136]
resembling ceramic and
cement that is created
from used oyster shells
and seaweed extract
• Sourced from local
seafood restaurants in Italy
Naturally Derived Biomaterials: Advances and Opportunities 31

Fig. 5 Composite made of

oyster shell [137]

13 Challenges and Future Opportunities

Naturally derived biomaterials hold great promise for a variety of commercial and
biological uses. However, there are also several concerns that must be completely
addressed to realize their potential.

13.1 Challenges in the Use of Naturally Derived Biomaterials

Some of the challenges and future opportunities associated with naturally derived
biomaterials are:
• Standardization and quality control: Naturally derived biomaterials can have
batch-to-batch variability, which can affect their properties and performance.
Standardization and quality control measures need to be developed to ensure
consistent quality of naturally derived biomaterials [138].
• Cost-effectiveness: Naturally derived biomaterials can be expensive to produce
and process, which can limit their use in industrial applications [139]. Improve-
ments in production efficiency and processing methods can help to reduce costs
and increase the scalability of naturally derived biomaterials.
• Mechanical and chemical properties: The mechanical and chemical properties
of naturally derived biomaterials can vary widely, which can make it difficult to
control their properties for specific applications. Advances in processing methods
and functionalization techniques can help to overcome these limitations [140].
32 A. H. Jasni et al.

• Biodegradability and stability: The biodegradability of naturally derived bioma-

terials can be both an advantage and a challenge, as it can affect their stability and
longevity in applications where long-term stability is required. Research on stabi-
lizing naturally derived biomaterials while maintaining their biodegradability can
help to overcome this challenge.
• Biocompatibility: While naturally derived biomaterials are generally biocompat-
ible, some may cause an immune response or other adverse reactions in certain
individuals. This is to better comprehend the biocompatibility of naturally derived
biomaterials and to create new strategies to overcome any potential limitations
[141].

13.2 Future Opportunities in Research and Development

Advances in biotechnology and material science are likely to drive new opportu-
nities for the development of naturally derived biomaterials. For example, genetic
engineering and synthetic biology approaches can be used to tailor the natural bioma-
terials’ characteristics for particular uses, while utilising novel processing techniques
that can help to overcome challenges related to scalability and processing efficiency.
Genetic engineering allows for the modification of an organism’s DNA to produce
specific traits or characteristics that can be beneficial for certain applications [142].
For example, scientists can engineer bacteria to produce large quantities of proteins
or other biomolecules that can be used in medical or industrial settings. Similarly,
genetic engineering can be used to modify the properties of natural biomaterials such
as silk or collagen, allowing them to be tailored for specific applications. Synthetic
biology, on the other hand, involves creating new biological systems and designing
existing ones or organisms with desired properties or functions [143]. This approach
can be used to create entirely new biomaterials that do not exist in nature, such as
biofuels or biodegradable plastics.
Both genetic engineering and synthetic biology can be used to improve the prop-
erties of natural biomaterials for specific applications. For example, scientists can
use genetic engineering to modify the properties of silk to make it more elastic or
stronger. Synthetic biology can also be used to create new biomaterials with unique
properties, such as self-healing materials or materials that respond to changes in their
environment.
In addition to genetic engineering and synthetic biology, novel processing tech-
niques can also be used to enhance the functionality and performance of natural
biomaterials. For example, scientists can use electrospinning or 3D printing to
construct scaffolds for delivery of drugs or tissue engineering. These methods enable
fine-grained control over the composition and characteristics of the biomaterials,
enabling them to be tailored for specific applications.
The capacity of natural biomaterials to combine with nanotechnology is perhaps
the most fascinating part of them. By combining the unique properties of natural mate-
rials with the precision of nanotechnology, researchers can create smart biomaterials
Naturally Derived Biomaterials: Advances and Opportunities 33

that having the capacity to revolutionize a vast area of industries. The possibilities
are truly endless.

13.3 What Are Among the Crucial Areas for Biomaterials

Research in the Future?

1. Immunomodulation is the process of adjusting the immune reaction to a specific

level. Immunomodulating biomaterials could aid in the fight against common
chronic illnesses like type 1 diabetes, an autoimmune condition in which the
body’s defences kill the insulin-producing cells within the pancreas. Injectable
synthetic biomaterial that treated type 1 diabetes in non-obese diabetic mice
was recently developed by researchers. This is a significant step in creating a
sustainable platform to help control the disease’s effects [144].
2. Injectable biomaterials are indeed being employed more frequently to deliver
therapeutic agents such drugs, genetic material, and proteins. They provide
focused distribution while preventing immune system absorption, opening the
possibility of treating a number of illnesses. Injectable biomaterials that are now
being researched and made from both artificial and naturally present materials
might someday be utilised to treat heart attacks, cancer, and bone abnormalities
[144].
3. Supramolecular biomaterials, which are collections of molecules that go beyond
the capabilities of individual molecules, have the capacity to feel and react,
making them perfect materials for treating disease or injury. Supramolecular
biomaterials that imitate natural biological signalling or that can be switched
either on or off regarding physiological cues are being investigated by researchers
[144].
4. Quorum sensing in synthetic biology which are needed in areas like bioterrorism,
combat, food and water safety, therapeutic properties, and fuel reliability are
pertinent to work on exploiting the indigenous microbial communication mech-
anism involving the signalling molecules Autoinducer-2 (AI-2) [145]. It has
been suggested that the signal molecule AI-2, which is created by the enzyme S-
Ribosyl homocysteinase (LuxS), is present in many bacterial species, and enables
interspecies communication [145]. To create a reliable and sensitive biosensing
device, the researchers use synthetic biology to design a bacterial sensor and
rewire the communication system of bacteria. It would be possible to create
more advanced and/or enhanced systems for a larger range of applications with
the aid of additional theoretical studies in genome circuits [146].
5. Photonic biomaterials: new fields including neuromorphic photonics, biodegrad-
able photonics, and Artificial Intelligence (AI) design are currently the subject of
progressive investigation. Combined, these new inventions—from an all-optical
internet to quantum communications—will help further transform the world
today [147].
34 A. H. Jasni et al.

6. Long-lived biomaterial-based fuel cells and batteries for wearable and

implantable electronics or high-power microbial fuel cells. Without any commer-
cial success, biofuel cells and biobatteries have only ever been considered labo-
ratory marvels. The primary difficulty is because historically, research has been
conducted by scientists and research organizations, and this decentralized method
encourages an imbalanced and deceptive image of biofuel cells that suggests itself
more to science fiction than an instant answer [148].
7. Food biopolymers for cultured food production: Due to the rise of societal
and commercial interest, cultured meat has gradually become well-known.
The cultured meat method comprises the laboratory cultivation of designed
muscle tissue as opposed to the traditional meat manufacturing method, which
relies on animals. Unfortunately, bioengineers have limited experience in engi-
neering muscle tissue for its post-mortem features, which largely determine how
consumers define meat quality. Until now, they have only built tissues to fulfil
biological objectives. Furthermore, the technical viability and industrial scala-
bility of current tissue engineering technologies for the manufacture of cultured
meat face basic hurdles. Thus, new explorations are needed in this area [149].
8. Grave to cradle concept of recycling waste as beneficial biomaterials for multiple
application. Reprocessing liquid and solid waste is an emerging research area for
the development of novel biomaterials with potential industrial applications as
wastes are abundant and cheap. This concept is aligned with the bio circular
economy model and is sought globally.

14 Conclusion

In summary, while there are challenges associated with naturally derived bioma-
terials, ongoing research and development efforts hold significant promise for the
future of these materials in an extensive array of manufacturing and biomedical
industries. Overall, this chapter delivers a summary of the synthesis of natural bioma-
terials and their possible applications, emphasizing the need for continued research
in this field. Regardless of the approach used, sustainability is always an important
consideration when synthesizing biomaterials. By utilizing renewable resources and
minimizing environmental impact, researchers can create more sustainable bioma-
terials that benefit both the users and the planet. This is particularly significant given
the growing concern about environment variation besides the requirement for more
eco-friendly solutions in all areas of life. As more research is conducted, we will
undoubtedly discover even more applications for these incredible materials, making
a positive impact on human health and our planet’s future.
Naturally Derived Biomaterials: Advances and Opportunities 35

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A. H. Jasni a former graduate of Monash University (MU) and

National Defence University of Malaysia (NDUM). She just
completed her Ph.D. in Engineering at Faculty of Engineering,
International Islamic University Malaysia. She obtained her
Bachelor of Science in Biotechnology from MU and Master of
Science in Biology from NDUM. She had been working under
the area of nanotechnology, material science, biotechnology
in military applications including microbiology, biosensor,
proteomic, and electrospinning since 2013.

A. S. Azmi a former graduate of the Widener University, Penn-

sylvania, USA started her career as a Chemist at First Malaysia
Coating Sdn. Bhd. on April 1999. On September 1999, she
received an offer from UniversitiTeknologi PETRONAS (UTP)
and working as trainee lecturer. She continued her study in MSc
in Process Integration at University of Manchester Institute of
Science and Technology (UMIST), Manchester, UK in 2001 for
one year. Early 2002 she was back to UTP and being appointed
and served as Lecturer until October 2006. On November 2006,
she moved to International Islamic University Malaysia (IIUM)
as lecturer before she pursued her study at University of Malaya
(UM), Malaysia on December 2007. She obtained her Ph.D.
in Bioprocess in Chemical Engineering on September 2012.
Currently she is attached as an Associate Professor to the
Department of Chemical Engineering and Sustainability, Inter-
national Islamic University Malaysia, Kuala Lumpur.
Naturally Derived Biomaterials: Advances and Opportunities 41

N. I. M. Puad graduated with B. Eng (Biochemical-

Biotechnology) (Honors) from International Islamic University
Malaysia (IIUM) in 2007. She was then appointed as an Assis-
tant Lecturer at the Calatrava Department of Biotechnology
Engineering, IIUM in the same year. Later in 2011, she obtained
her Ph.D. in Chemical Engineering and Analytical Science from
The University of Manchester, UK. Her research interest
is mainly on Plant Cell Culture Technology, Flux Balance
Analysis, Kinetic Modelling and Simulation, Plant Secondary
Metabolite and Natural Products, Bioprocess and Renewable
Energy. Presently, she is an Assistant Professor at the Depart-
ment of Chemical Engineering and Sustainability, Faculty of
Engineering, International Islamic University Malaysia (IIUM).

F. Ali is an Assistant Professor at the Department of Biotech-

nology Engineering, Faculty of Engineering, International
Islamic University Malaysia (IIUM), Gombak 50728, Kuala
Lumpur. She obtained her Ph.D. in Chemical and Biomolec-
ular Engineering from Korea Advanced Institute of Technology
(KAIST) in 2013. Her research interest is in the area of Polymer
Blend and Composites, Polymer Nanocomposites, Synthesis
of polymers, block copolymers and polyurethane, Synthesis
of Nanoparticles using natural sources, Polymers in Sensors,
Polymers for packaging materials, Functional Polymers for
lithography, Polymerization of biopolymer from the monomers
produced from fermentation process, and Separation and
Purification Techniques. She has nearly two years’ experience
in teaching on Process Plant and Design, Biopharmaceutical
Engineering, Seminar for undergraduates and Fluid Mechanics.
Dr. Fathilah also serves as a reviewer for polymer related
journals. She published 11 papers in National and International
conferences, 6 papers in SCOPUS and ISI journals and 2
book chapters. She was secretary for Nano Research Group
and currently the Deputy Dean of Student Affairs (January
2015–present).

Y. A. Nor obtained her qualifications of Chemical and Biolog-

ical Science (Nanotechnology) for her Doctor of Philosophy
from University of Queensland and she acquired her Biotech-
nology Engineering Masters Degree from the International
Islamic University Malaysia (IIUM). She is now working as an
Assistant Professor at the Department of Chemical Engineering
and Sustainability, Faculty of Engineering IIUM.
Different Techniques of Genetic
Engineering Used for the Development
of Novel Biomaterials

Aashveen Chhina, Vridhi Sachdeva, and Shubham Thakur

Abstract Biomaterials are garnering huge success in the modern era of unforeseen
disease conditions. They find their utility in specifically interacting with biolog-
ical systems for therapeutic or diagnostic purposes. They help patients recuperate
from disease or injury by restoring tissue function thereby, sometimes also leading
to regeneration. Some of the biomaterials are living (microorganisms based) and
can be prepared through microencapsulation, 3D printing, coating, and spinning by
which they can be inculcated into matrices. On the other hand, microbes can even
produce their own matrix which can be genetically engineered to have control over
the responses of the body. Genetic engineering and biotechnological approaches are
an ideal amalgamation of systems for preparing biomaterials that handle accuracy
and complexity quite conveniently. The biomaterials generated by the use of geneti-
cally engineered techniques have proven to be a boon for the regenerative medicines
market, gene delivery systems, tissue regeneration platforms, and controlled drug
delivery systems. In the current scenario, researchers have focused their attention
on molecular evolution for the preparation of advanced designs of genetically engi-
neered biomaterials and the development of inert biomaterials that can be properly
integrated into devices. The advancements and research carried out in genetic engi-
neering continue to assist researchers to gain insights into a specific change in the
structure of employed molecules for the preparation of biomaterials and the way it
will affect the development of biomaterials and their function in the longer run. The
manufacturing sector of biomaterials is a domain that has significantly evolved and
is still being researched.

Keywords Genetic engineering · Biomaterials · Recombinant DNA technology ·

Genetically engineered biomaterials · CRISPR- Cas9 · mRNA

A. Chhina · V. Sachdeva · S. Thakur (B)

Department of Pharmaceutical Sciences, Guru Nanak Dev University, Amritsar, Punjab 143005,
India
e-mail: [Link]@[Link]

© The Author(s), under exclusive license to Springer Nature Singapore Pte Ltd. 2023 43
R. Malviya and S. Sundram (eds.), Engineered Biomaterials, Engineering Materials,
[Link]
44 A. Chhina et al.

Abbreviations

DNA Deoxyribonucleic Acid

GF Growth Factors
ECM Extracellular Matrix
GR and CNT Graphene and Carbon Nanotubes
ELP Elastin-like Proteins
SLP Silk-like Proteins
SELP Silk-elastin-like Proteins
HDP Host Defense Peptides
CPP Cell-penetrating Peptides
AMP Antimicrobial Peptides
mRNA Messenger RNA
SDS Sodium Dodecyl Sulfate
CDI Carbodiimide
HMDC Hexamethylene Diamine Carbamate
EtO Ethylene Oxide
PAA Peracetic Acid
GI Gamma Irradiation
dECM Decellularized Extracellular Matrix
PBS Phosphate Buffer Saline
VEGF Vascular Endothelial Growth Factor
bFGF Basic Fibroblast Growth Factor
BMP Bone Morphogenetic Proteins
TENG Triboelectric Nanogenerator
PENG Piezoelectric Nanogenerator
PyENG Pyroelectric Nanogenerator
Bio TENG, Biodegradable Nanogenerator
PTFE Polytetraflouroethylene
AFR Aniline Formaldehyde Resin
S-TENG Starch Triboelectric Nanogenerator
CD-TENG Chitosan-diatom-based biomaterial TENG
TDD Transdermal Drug Delivery
HAP Hydroxyapatite
CRISPR/CAS 9 CRISPR, Associated Protein 9
HFG Hepatocyte Growth Factor
sgRNA Single Guide RNA
PLGA Polylactic Glycolic Acid
GCE Genetic Code Expansion
HDACi HDAC Inhibitors
CAR T-cell Chimeric Antigen Receptor T-cell
Different Techniques of Genetic Engineering Used for the Development … 45

1 Introduction

The main goal of medical research aims to convert multifaceted engineering science
along with the biological science into procedures for the substitution, rejuvenate, cell
repair, tissue repair, or organs in order to restore compromised function. Biomaterials
are any substances, whether organic or inorganic, living or dead, typically composed
of several parts that engage with the body system. In medical uses, such intelligent
materials are frequently used to supplement or take the place of a natural function.
To state it in a more simplified manner, a biomaterial is a substance that has been
developed to take on a shape that, either on its own or as part of a complex system,
is utilized to regulate interactions with living systems and has the potential aid in the
growth of any curative or clinical method. Smart biomaterials are used in medicinal
uses to brace, improve, and restore a biological functionality or tissue that’s damaged.
They can be natural or synthetic. Contrary to what the term “biomaterial” might
imply, a biomaterial need not be biological or composed of materials linked to life.
The actual substance may be made of metal, plastic, or different types of composites,
but it may also be bioinspired and derived from natural sources. They are intended
to interact with the body or live in a biological environment. Biomaterial sciences
deal with investigations regarding biomaterials. They have expanded steadily and
significantly during the course of their existence as a result of various enterprises
making considerable financial expenditures in the development of new stuff. Aspects
of tissue engineering, biology, chemistry, and materials science are all included in
the discipline of biomaterials science.
Due to the incorporation of genetic engineering, cell-based biomaterials have
attracted attention from all over the globe for their potential use in a variety of
medical disciplines. These engineered biomaterials can change shape or mechanical
characteristics, release bioactive ions, and actively discharge chemicals as a result of
physiological triggers. All of these material-induced activities may result in native
cells and influence the immune system These designed biomaterials can be utilized
in the deposition of extracellular matrix (ECM) to aid in the repair of tissues, and
enhance cell adhesion and proliferation by incorporating biophysical and biochem-
ical signals [1]. Evaluation of the short- and long-lasting cellular and when creating
the next generation of bio-responsive materials, understanding how materials interact
at the molecular level is essential as material capabilities increase [2].
Medications, genetic materials, and proteins are all delivered therapeutically
through the use of injectable biomaterials. They provide the opportunity to address
a range of conditions by delivering medications precisely and preventing immune
system uptake. Injectable biomaterials with both synthetic and organically derived
components are being investigated for use in managing heart attacks, cancer, and
bone defects. Widespread chronic illnesses like type 1 diabetes, an autoimmune
disease in which the body’s defenses attack pancreatic cells that produce insulin,
may be combated with the aid of immunomodulating biomaterials. An injectable
synthetic biomaterial recently created by researchers reversed type 1 diabetes in
non-obese diabetic mice. This is a significant move toward creating a biodegradable
46 A. Chhina et al.

platform to help manage the disease’s effects. Applications for genetically engi-
neered materials include the delivery of drug, nanoparticle coatings, carriers that are
macromolecular, and hydrogels. Engineering of the tissue has a significant influence
on novel cutting-edge approaches to gene engineering. Studies supported various
approaches to obtaining, creating, and using these materials while highlighting the
evolved functions and properties.

2 Biomaterials: Structural and Functional Roles

Biomaterials are more frequently integrated into devices than they are used as
simple materials in medical uses. Supramolecular biomaterials, which are collec-
tions of molecules that go beyond the capabilities of individual molecules, have the
capacity to detect and react, making them perfect materials for treating disease or
injury. Supramolecular biomaterials that can respond to physiological signals by
activating or deactivating or that mimic biological signaling are being investigated
by researchers.
The method of creating biomaterials has changed over time. Many of the bioma-
terials that are used in medicine today were not intended to be used in medicine;
instead, they were discovered by clinicians to help treat patients. Thus, cellulose
acetate, a common plastic, was used to create the first dialysis conduit. Initial
vascular grafts employed polymers like Dacron, which were taken from fabrics.
Initially, polyurethanes of industrial quality were used as the basis for the mate-
rials for artificial hearts. These resources made it possible to handle critical medical
issues. However, they also brought about challenges. Platelets and the complement
system may be activated by dialysis tubing; vascular grafts made of dacron that are at
least 6 mm thick can be used; otherwise, biological processes at the blood-material
and tissue-material interfaces could result in occlusion; additionally, blood-material
interactions could cause clot development in an artificial heart, which would raise
the risk of stroke and other complications [3]. Recent studies are based on drug and
cell transporters made of a variety of biomaterials. In tissue engineering, it may be
necessary to combine medications, cells, and an appropriate carrier with a tailored
degradation profile, specialized macroscopic features, and specific biological cues to
promote healthy tissue growth [4]. Biological activity can be added to biomaterials
by incorporating oligopeptides that promote binding.
Traditional biomaterials like polytetrafluoroethylene, silicone rubber, or polyethy-
lene are implicitly recognized by cells in vivo. Non-specific protein adsorption
from bodily secretions occurs on the surface of surfaces, and adsorbed homolo-
gous adhesive proteins encourage adherence of the cell by binding to cell surface
receptors. The explicit power of adhesion of cells on intelligent materials is possible
by preventing nonspecific protein adhesion on the material surface and utilizing
adhesion-promoting peptides that are unique to particular cell types [5]. Short primary
segments of these peptides are derived from the domains that are receptor bound with
the adhesion proteins. The tri-peptide sequence RGD is the adhesion peptide that is
Different Techniques of Genetic Engineering Used for the Development … 47

Fig. 1 The scope of biomaterials in various fields

most frequently investigated [6]. Figure 1 illustrates the scope of biomaterials in

various fields. Over the last ten years, significant advancements have been made
in our knowledge of how cells interact with biomaterials [7, 8]. There is growing
proof that biomaterials do more than just act as a scaffold that is temporary since
they facilitate the coordination of tissue morphogenesis and cell adhesion [9, 10].
Biocompatibility is a frequently employed notion involving determining the viability
of biomaterials in therapeutic implementation for tissue repair and recovery. Biocom-
patibility, however, reveals some limitations in the ability to describe and define
some crucial characteristics of superior biomaterials should have as immunology
and biology science advance.
A specific biomaterial must enable long-term use in any application without being
rejected by the host organism or losing its intended purpose. The design and selection
of biomaterials must take a variety of properties into consideration to achieve this.
Although these characteristics can be broken down into mechanical, physical, molec-
ular, and biological groups, in reality, these divisions are frequently entwined. The
ideal properties of biomaterials are always evaluated in the context of their particular
use because the requirements for various biomaterials are frequently different. To
prevent disruption, biomaterials must take into consideration the structure and oper-
ation of the tissues and systems in the vicinity. According to an implantation region
and possibly even the patient’s medical history, the properties of an ideal bioma-
terial may vary. Bifunctionality is required in biomaterials [11]. This implies that
they must have the necessary qualities to perform their particular role, as implants
or other types of devices. It might restore a diseased or injured tissue’s lost function-
ality, completely replace that function, or be used to make a diagnosis. A sustainable
48 A. Chhina et al.

Table 1 Applications of biomaterials

[Link] Nature of biomaterial Organ Application
system
1 Ceramics (Bioactive ceramics, Bones, Joint replacement, heart valve, dental
non-oxide bioinert ceramics, glass heart, implant
ceramics) joints,
teeth
2 Metals and alloys (Zinc, Heart, Stents, orthopedic screws, dental implant,
magnesium, copper, titanium, bones, cochlear replacement
platinum, high-entropy alloys) teeth,
ears
3 Polymers (Poly (α-hydroxyesters), Skin, Surgical sutures, skin repair templates,
polylactic acid, polyglycolic acid eyes, ocular implants, corneal bandage, cartilage,
and their co-polymers (PLGA, bones, contact lenses, bone cement, heart–lung
polycaprolactone, silicones, heart, machine, blood vessel prosthesis, artificial
polyurethane) kidney kidney
4 Genetically engineered Bones, Elastin-like-polypeptides based
biomaterials (Collagen, silk, skin Hydroxyapatite matrices, tissue
gelatin, chitin, cellulose, glucose) regeneration

biomaterial is a relationship due to which we have a great opportunity to build inno-

vative sustainable development strategies in the upcoming years owing to the synergy
between biomaterials and renewable natural resources. Significant efforts and steps
are being made now to create and design materials using sustainable resources that
will eventually replace conventional materials. We are currently at a critical juncture
where we require sustainable energy, which is only made feasible by advancements
in green technologies [12]. The applications of various types of biomaterials have
been listed in Table 1.

3 Need for Genetic Engineering of Biomaterials

Genetically engineered biomaterials have been a topic of research ever since recom-
binant DNA technology has come into play. Such biomaterials can be of animal or
plant origin with properties of strength, firmness, and ability to adapt to the environ-
ment. Where genetically engineered techniques aid in developing new, relevant, and
functional biomaterials, there they also help in developing a better version of already
existing biomaterials along with enhancing chemical and physical qualities leading
to an increase in the applicability of biomaterials which have a significant influ-
ence on biomedical applications such vaccine production, engineering of tissue and
regeneration, drug administration, and zoological interpretations, etc. In the creation
of hydrogels, development of films, formulation of matrices, development of phar-
maceuticals, as well as several physiological and anatomical implants, the use of
Different Techniques of Genetic Engineering Used for the Development … 49

genetically altered proteins can provide good quality materials with excellent multi-
functional properties, broadening the range of applications and need for developing
better genetically engineered biomaterials [13]. Moreover, electrical functions can
also be inculcated into the biomaterials through the use of technology which helps
in excellent antibacterial activities which could be in vitro or in vivo.
Multi-stage moderation of smart biomaterials, which includes gene alteration
(e.g., modifying gene sequence, molecular weight, and chain size), chemical conju-
gation (e.g., adding light-sensitive molecules, oil-loving and/or water-loving side
chains), and mesoscopic blending (e.g., doping with metallic nanoparticles, GR, and
CNT), allows for the integration of biochemical and electrical capabilities into the
material system based on biopolymers [14]. To circumvent the limitations of viral
vectors for NK cell engineering, attempts have been made to substitute nanoparticles
for viral vectors as they did not show any efficacy due to a transduction efficiency of
less than 20%. Due to the distinct physicochemical features of genetic modification,
nanoparticles can add imaging modality to the vector while avoiding many of the
possible safety problems associated with viral vectors [15].
Living-engineered biomaterials that have microorganisms involved have the
capacity to respond in complicated ways to environmental cues, and they can be
genetically modified to enable user control of behavior and the incorporation of
a variety of inputs. The altered microbes can either produce their matrix, as in
biofilms, or they can be integrated into matrices utilizing numerous innovations,
including coating, 3D printing, spinning, and microencapsulation [16]. In addition,
the use of biomaterials in clinical environments may be hampered by a few drawbacks
that have been observed. For instance, a significant hurdle impeding effective tissue
repair and regeneration is biomaterial-mediated inflammation; as a result, biocompat-
ibility is being continuously researched and enhanced which can be fulfilled through
the process of genetic engineering [17]. Various sorts of immunomodulatory genes
were introduced for alteration to stop biomaterials from being immunologically
rejected. Transmembrane proteins, cytokines, and genes genes-producing all have
immunomodulatory effects and other proteins have been proven in several investiga-
tions and these genes acted in a variety of ways to exert influence that is suppressive
or agitative on the immune system elements [18].
Live cells can now be incorporated into structures and allowed to flourish owing to
developments in 3D bioprinting. Also, chimeric biomaterials can be 3D printed more
accurately and intricately. The conventional biomaterials used as non-conventional
biomaterials can be produced with more precision and finer qualities and hence
can also help in saving cost [19]. Consequently, there are many uses for intelligent
biomaterials, ranging from health (such as engineering of the tissues, and delivery
of the drugs, and biosensors) to more newly researched environmental uses like
ecosystem restoration (e.g., coral reefs and environmental remediation) [20]. The
development of physicochemical features of protein-based drug delivery devices
that are accurately calibrated is made possible by genetic engineering approaches,
offering a higher degree of customization than is possible with synthetic polymers.
Elastin-like proteins (ELP), silk-like proteins (SLP), and silk-elastin-like proteins
(SELP) due to their inbuilt physical and chemical attributes and the simplicity of
50 A. Chhina et al.

engineering made possible by recombinant DNA technology, a particular set of

substitutes for creating drug delivery systems [21].

4 Techniques of Genetic Engineering

4.1 Scaffolding

As a result of the variety of biomaterial amenities, including the adaptable

nature of protein-engineered materials can be independently tailored, offering a
desirable option to harvested natural scaffolds or synthetic polymeric scaffolds.
Different modular peptide domains are created by encoding specific molecular
sequences in a DNA plasmid, transfecting the plasmid into a target organism,
expressing the plasmid, and purifying the resulting biopolymer definition make up
protein-engineered biomaterials [22]. By combining a variety of peptide sequences
allowing in a single, modular biomaterial, the scaffolds can be created to imitate
many properties of the natural extracellular matrix, including cell adhesion, cell
signaling, elasticity, and biodegradability. The usefulness and scope of protein-
engineered biomaterials have recently been increased by the inclusion of functional
elements that are uncommon in the extracellular matrix.
Each functional peptide module’s location and density can be precisely adjusted
thanks to the genetically encoded DNA sequence being used as a template to create
these protein-based materials, allowing the independent tailoring of numerous mate-
rial properties. For each engineered biomaterial, a precise DNA template needs to
be made for the sake of completely realizing the potential of this design approach
at molecular level. Therefore, meticulous control of DNA template design is essen-
tial for enabling the adaptability of biomaterials made of modified proteins. For the
prevention of DNA recombination when generating extreme repetition of amino acid
patterns, a variety of codons must be used [23]. The smart biomaterial is engineered
through protein in which a DNA template is created and it is then turned into a
recombinant plasmid. The chosen host cell is transfected with the plasmid, which
causes the chosen host cell to translate the genetic information involved with the
desired protein. A protein-engineered scaffold is created once the protein has been
processed after being refined in step five. The resulting scaffold is examined using
methods involving in vivo and in vitro techniques, which reveal data on how to
enhance the scaffold’s characteristics and lead to remodeling the biomaterial and
altering the encoding template’s DNA. Figure 2 elucidates the steps involved in the
scaffolding technique.
Different Techniques of Genetic Engineering Used for the Development … 51

Fig. 2 The steps involved in the scaffolding technique

4.2 Employment of Sequences

Both organic and synthetic nanoengineered biomaterials that are polymeric are
employed in biomedical utilization like targeted and smart drug delivery with
prolonged and controlled release. Natural polymeric nanocomposites are frequently
employed in medicine because they are biodegradable, nearly biocompatible, and not
poisonous. Applications for synthetic biopolymer nanocomposites are numerous. By
combining with biomolecules, they can be altered to become more biocompatible
and harmless to the body [24].
Sequencing in the genetic engineering of biomaterials is one of the significant tech-
niques as the properties of biomaterials depend on the sequences that are employed
in the biomaterials which is an emerging topic for research recently. Examples
of synthetic methods for sequence, control include iterative methods where each
monomer unit is added sequentially, regulated polymerization techniques using both
chain and step-growth mechanisms, and bioinspired and supported methodologies
like molecular imprinting and templated syntheses. In the case of genetic engi-
neering of biomaterials. Biomaterials made of particular peptidomimetics and statis-
tical copolymers have both been made using the comparatively simple process of
sequence selection known as “bioabstraction,” or replicating minimum functional
DNA or protein sequences. Synthetic analogs have been produced using this tech-
nique which are biologically important compounds, including host defense peptides
(HDPs), cell-penetrating peptides (CPPs), and antimicrobial peptides (AMPs) [25].

4.3 Recombinant DNA Technology

Protein-based biomaterials are created using recombinant DNA technology, or

genetic engineering. Because of precise control and the modification of the DNA
structure that encodes the protein structure, it is possible to alter the basic structure,
52 A. Chhina et al.

layout, and length of the protein biomaterial’s chain. This quickly developing tech-
nology has given rise to effective tools for creating unique smart biomaterials with
preset 3D structures [26]. However, several issues need to be resolved to synthesize
protein chains using genetic engineering techniques: Repetitive DNA sequences may
be altered and deleted in various ways, sequences of messenger RNA (mRNA) neces-
sary for a certain codon for a particular amino acid residue. The host cell may break
down the target protein before sufficient levels are synthesized, reducing the yield.
Biomaterials that have been genetically modified have many benefits. By carefully
regulating the length, stereochemistry, and 3D structure of the polymer chain, it is
possible to create materials with specific, distinctive characteristics.

4.4 Decellularization

Recently, novel methods based on genetic changes have been developed, including
decellularized biomaterials. Decellularization is the elimination of cells and
constituents involving the nuclear ones, of the tissue, using a variety of chemi-
cals while conserving the ultrastructure of the collagen, elastin, microfibrils, proteo-
glycans, glycosaminoglycans (GAGs), and other growth factor components found
in the extracellular matrix [27]. Decellularization seeks to reduce the immuno-
genicity, calcification-inducing potential, and cytotoxicity of native extracellular
matrix (ECM) by removing the majority of cells from it with the aid of physical,
chemical, and biological methods [28]. Extracellular matrix (ECM)-based bioma-
terials now have new prospects in the transfusion of organs and tissue regrowth in
preclinical and clinical settings because of advances in decellularization techniques.
The resultant dECM scaffold should ideally maintain its functional content and three-
dimensional (3D) structure for tissue restoration purposes. Before dECM scaffold
transplantation, it was important to recellularize the dECM with a particular cell
type to increase cell survival and reduce immunological rejection [29]. Steps in the
decellularization are given in Fig. 3.
Decullarization occurs either through physical methods such as freeze-thaw,
hydrostatic pressure, ultrahydrostatic pressure, and oscillation, chemical methods
like hypertonic/hypotonic solutions, detergents namely sodium dodecyl sulfate
(SDS), alcohols or triton X-100 and enzymatic methods such as RNases, DNases,
and trypsin which aids in separating a tissue’s extracellular matrix (ECM) from the
cells that reside there, leaving behind an ECM scaffold that can be employed in arti-
ficial organ and tissue regeneration [29]. Crosslinking after decellularization occurs
through a number of crosslinking agents which can be physical, chemical, or natural.
Thermal dehydrogenation and photo-oxidation crosslinking are the two basic types of
physical crosslinking. Thermal dehydrogenation was frequently utilized in the begin-
ning and now, this approach is frequently employed as a backup strategy because
of the difficult crosslinking conditions. The crosslinking conditions are difficult to
manage using the photo-oxidation approach. This makes it more frequently employed
in the decellularization of tumor tissue, which is typically done for disease modeling
Different Techniques of Genetic Engineering Used for the Development … 53

Fig. 3 Steps involved in the decellularization process

purposes. The dECM of the tumor was crosslinked by Lü et al. using photo-oxidation
mediated by methylene blue [14]. This was the first study to show whether photo-
oxidative crosslinking affected the tumor dECM scaffold’s structural, physical, and
biological characteristics. Epoxy compounds, carbodiimide (CDI), glutaraldehyde,
and hexamethylene diamine carbamate (HMDC) are examples of crosslinkers which
are employed in chemical crosslinking but have huge side effects due to which better
crosslinking agents need to be used [29].
Sterilization being the third step is the most crucial step when decellularization is
involved as the applicability of different biomaterials is put off due to limited steril-
ization methods for a period that is more than expected. The structure of the ECM
will inevitably change as a result of current sterilization techniques, such as exposing
the material to radiations such as ethylene oxide, gamma, or electron beam, and this
could negatively impact the biomaterial’s mechanical properties [29]. With respect
to the list provided above, a more suitable option is provided by the development
of supercritical carbon dioxide. [30]. The impact of three widely used sterilizing
techniques, including ethylene oxide (EtO), peracetic acid (PAA), and gamma irra-
diation (GI), on material qualities, was examined by Matuska et al. In this procedure,
they studied the early cellular contacts and discovered that, unlike GI and EtO, PAA
showed advantages in cell adhesion but did not significantly alter the structure of the
cell. According to this, the choice of sterilizing technique is important for modifying
the properties of biomaterials in a way that promotes cellular adherence and is highly
relevant to the use of biomaterials in vivo [31]. There is currently no sterilization
method that can completely stop the incidence of undesirable consequences [32].
54 A. Chhina et al.

Consequently, selecting the best sterilizing process is a crucial step for using decel-
lularized scaffolds in the future for therapeutic applications, and more research is
still required.
Future uses of the decellularized matrix-based materials will depend on the final
step of preservation in the process of decellularization as well. The manufactured
decellularized scaffolds were now typically stored in PBS alongside antibiotics and
antimycotics at a temperature of 4°C for a brief length of time, on standby, or at
20 °C for long-term preservation [29, 33]. The degradation of active substances and
structural damage, however, seems to be unavoidable at the moment and require
urgently more work. The ideal characteristics of a tissue for repair in wound repair
include strong bioactivity, adequate mechanical characteristics, excellent degrada-
tion, and outstanding biocompatibility are all desirable characteristics in a scaffold
for tissue repair. Decellularized ECM (dECM) is a three-dimensional natural scaf-
fold produced from autologous, allogeneic, and xenogenic tissues that retains its
intrinsic tissue structure despite having had the cellular components removed. The
dECM is promising in translational medicine [34] because of exceptional physio-
logical activity, high biocompatibility, lack of immunogenicity, and use as a diverse
source of basic materials [35].
New advancements in the biomaterial’s domain in the realms of regenerative
medicine and engineering of tissue, dECM-based smart biomaterials have had great
success. Also, scientists were inspired to create a novel standard for the application
of dECM biomaterials, such as constructing a safe and biocompatible method for
delivering therapeutic agents like drugs and cells. For instance, dECM can be ground
into nano- or microparticle size and used as a therapeutic delivery equipment to repair
anastomoses [29]. Clinically, both Axogen’s Avance nerve transplant and AlloDerm,
an acellular skin matrix from Biohorizon is a regenerative tissue matrix intended to
hasten skin regeneration. are used to treat wounded nerves and are two examples of
emerging clinical applications of dECM. DECM can also be utilized to make bioink-
related hydrogels. Of the inks now on the market for 3D bioprinting, bioinks based on
dECM exhibit the highest level of biomimetic capability. The building block for 3D
printing a functioning human organ is made of dECM bioinks in conjunction with the
recent development of bioprinters. It is fascinating that multiple studies have been
published employing ECM hydrogels synthesized from almost every organ where
dECM bioinks are employed for bioprinting. ECM hydrogels could be employed in
minimally invasive deliveries as an easily administered pre-gel viscous solution into
a patient utilizing a catheter or syringe [36].

4.5 Physical Adsorption

Combining protein-based biomaterials with bioactive molecules can improve their

properties and increase either in live conditions or the laboratory. Surfaces generated
from proteins can be changed by chemical, physical, or encapsulating procedures.
Using these methods, proprotein-based materials can be functionalized with various
Different Techniques of Genetic Engineering Used for the Development … 55

antimicrobials and growth factors are two examples of biologically active chemi-
cals that can improve cellular and tissue reactions [27]. When introducing bioactive
compounds, such as growth hormones or extracellular matrix proteins, through dip
coating, to the surface of scaffolds physical adsorption, a straightforward immobi-
lization technique, is commonly used. Topography of surface, functional groups,
pH, wettability, and electrical charge of the substance are just a few examples of the
physical and chemical characteristics that affect adsorption efficiency. Since many
nanomaterials are hydrophobic, techniques are required to increase their wettability
and turn them into hydrophilic materials. Chemical interactions between carbon and
non-carbon atoms are broken down using physical techniques like ion bombard-
ment, UV light, and plasma modification. As a result, oxygen reacts with unsatu-
rated bonds and radicals to promote hydrophilicity and reactivity toward substances
that are biological. Since they contain a large number of reactive chemical groups
(hydroxyl, carboxyl, and amide), natural polymers have the benefit of being less
hydrophobic and able to interact with physiological molecules. Materials composed
of proteins, such as collagen and silk, have been customized by the adsorption of
biologically active compounds, such as vascular endothelial growth factor (VEGF)
[37], basic fibroblast growth factor (bFGF) [38], and bone morphogenetic proteins
(BMPs) [39], and pharmaceuticals like antibiotics, and heparin [40].
Van der Waals, hydrogen, and hydrophobic interactions serve a purpose in adsorp-
tion while they are electrostatic in nature, external factors can affect how stable the
adsorbed molecules’ bond is relatively weak or moderate. In this manner, uncon-
trolled release of the immobilized species can be brought on by changes in the pH,
strength of ions, and the number of adsorbed species in the surrounding media. For
instance, large doses of bone morphogenetic proteins (BMPs) in order to trigger the
correct osteogenic response, substances that diffuse away from the fracture location
are required. The BMP-2 release profile from collagen sponges exhibits an early
spike within the first ten minutes, during which 30% of the BMP-2 is lost by the
messenger carrier, and is thereafter sluggish over the course of the next 3–5 days.
Clinical conditions like soft tissue hematomas, ectopic bone growth, and receding
bones may result from this original burst release [41].

4.6 Triboelectric Nanogenerators (TENG)

TENG is one of three types of nanogenerators in which the concepts of tribo-

electrification and electrostatic induction work together to transform mechanical
energy, including that from very low frequencies to electrical power. In the “tri-
boelectric effect,” the charges are transferred during mutual contact between mate-
rials having varying triboelectric polarities (distinct capacities for electron binding).
TENG makes use of this phenomenon by using an external force to separate two mate-
rials which further creates a potential difference hence, creating an electric field [42].
Another type is piezoelectric nanogenerators (PENGs), which function by interacting
56 A. Chhina et al.

between the mechanical and electrical states linearly while responding to mechan-
ical stress and are made of crystalline materials lacking inversion symmetry [43].
Pyroelectric nanogenerators (PyENGs), which transform variations in heat energy
into electrical energy, come in third [44]. Recently, it was shown that TENGs and
PyENGs can work together by harnessing the human body’s mechanical and thermal
energy to generate electrical signals [45]. Nonetheless, triboelectric nanogenerators
are frequently chosen in comparison to their counterparts piezoelectric nanogenera-
tors or pyroelectric nanogenerators due to their conditions of operation for regions of
low frequency/amplitude, ecosystem quality, more extensive resources, and material
selection [46].
Polymers, such as polyvinylidene fluoride, polytetrafluoroethylene, poly-
dimethylsiloxane, and others, make up the majority of triboelectric nanogenerators
because they all include the element fluorine, which has a great affinity for elec-
trons. Although these polymers can produce significant triboelectricity, they can be
challenging to handle and decompose, and their accumulation and disposal could be
hazardous to both human health and the environment. The ecologically friendly and
biodegradable TENGs (bio-TENGs) will create smart portable tools to address these
issues, setting a new standard for the establishment of a smart and green society.
TENGs in which the triboelectric layer components are partially or entirely made of
natural substances or their constituents are classified as bio-TENGs. Firstly, bioma-
terials having high biocompatibility and biodegradability are obtained from natural
species and they can be decomposed by microorganisms after being disposed of
in nature. Additionally, because TENGs have an innate ability to generate green
energy from their environment and biologic processes, these intelligent bio-TENGs
can be used to power autonomous driving sensors or smart electronics. Finally, there
are numerous sources of biological materials, ranging from marine products like
shellfish and seaweed to agricultural by-products like straw, rice husks, bark, and
bagasse [47]. The majority of biodegradable polymers have low toxicity and strong
biocompatibility, which makes BD-TENGs effective in biomedical applications. The
implantations for medical devices should be removed promptly when their purpose
has been served in the human body. Biodegradable devices are typically employed
to treat disorders of the heart and there is no requirement of additional surgery,
in contrast to non-degradable devices that can only be removed through procedures,
which causes subsequent injury to the patients [48]. They are also employed in tissue
regeneration.
For the current triboelectric series to be expanded, new triboelectric materials
must be developed. The design of positive and negative materials for placement
in the triboelectric series is under the limelight. The synthesis of novel materials
containing significantly more accurate halogenic elements is the main focus of tech-
niques for creating novel tribo-negative materials. As an illustration, Lee et al. stated
that fluorinated polymeric sulfur outperformed commercial PTFE, which is ranked
the worst in the series of triboelectric materials. A disadvantage associated with tribo-
negative materials is their insulation. The creation of materials that are tribo-positive
is inspired by the emergence of the highest positive charges in polymers comprising
nitrogen and oxygen with pyridine amide, amine, or hydroxyl groups. Based on this
Different Techniques of Genetic Engineering Used for the Development … 57

hypothesis, Zhao et al. prepared an aniline formaldehyde resin (AFR) with sufficient
mechanical strength and controlled microstructures for TENG device applications
[43].
By capturing high entropy energy, TENGs offer a fresh approach to the problem
of supply of energy. Nevertheless, flexible, humidity-resistant, and affordable TENG
must meet strict standards for wearable electronic devices. Here, a straightforward
and environmentally benign process was used to create the multipurpose wheat starch
TENG (S-TENG). The S-TENG opens the door for the mass production of multifunc-
tional biomaterials-based TENG as well as the real-world use of wearable electronics
and self-powered sensors [49].
There is a reported way for easily improving the quality of a chitosan-diatom-
based biomaterial TENG (CD-TENG) by controlling the weight ratio of the diatom
frustule, which can increase the density of charge in the chitosan-diatom composite
film [50]. This is useful to apply in wearable devices that might be easily attached to
human skin and will not be associated with any adverse implications. Nanocarriers
are stimuli-responsive biopolymers that are commonly used in invasive and trans-
dermal drug delivery (TDD) systems that are stimulated and regulated by TENGs
power supply. Other common drug-delivery agents include nanoparticles, micronee-
dles, and liposomes. TENG is therefore utilized to deliver drugs with organelles and
biomaterials for offering a target-oriented controlled drug release [51]. Paper-based
cellulose TENGs, nano cellulose TENGs, and micro/nano composite TENGs are
other advancements in biomaterials based on the triboelectric nanogenerator prin-
ciple. The advantages of cellulose as a naturally occurring polymer are cost effective-
ness, processability, good mechanical flexibility, biocompatibility, and biodegrad-
ability. Additionally, it may display a special combination of chemical, structural,
dielectric, and optical characteristics. These benefits may transform cellulose-based
functional materials into appealing substrates or TENG elements [52].
The creation of a durable, stable encapsulating covering is necessary for the
commercialization of biomaterial TENG devices as clinical implants since it will
shield the device from the hostile in vivo environment. This is because overcoming
the immune system’s response to a foreign body is challenging, and fibroblast growth
may eventually cause the implanted TENGs to respond to motion stimuli less effec-
tively [53]. To construct well-designed TENG sensors, researchers will eventually
need to use fabric, shape-memory polymers made of latex or rubber, hydrogel, and
other cutting-edge functional materials. In addition, significant progress in the area
of flexible electrodes is required to increase the wearability of TENGs both inside
and outside the living systems. Hard and soft materials, including carbonaceous
nanoparticles, polymeric substances, 2D materials, inorganic and ceramic materials,
and polymers, are used to create today’s naturally flexible electrodes [54]. The TENG-
based healthcare system benefits greatly from TENG-based biomaterials, which have
a bright future in terms of consistency, sustainability, comfort, and adaptability.
58 A. Chhina et al.

4.7 Elastin like Polypeptides (ELPs)-Based Hydroxyapatite

Composites

Organic matrix macromolecules are essential for improving the mechanical char-
acteristics of biomineralized compounds like bone and teeth in nature. Because
there is still a lot to learn about how natural matrix components work, creating
artificial matrix counterparts is both exciting and difficult. Biomimetic matrices can
be created utilizing genetically produced elastin-like polypeptides (ELPs) instead
of natural components for the design of mechanically robust ELP-hydroxyapatite
(HAP) composites. ELPs with clearly defined backbone charge distributions can
be made by intermittently including negative, positive, or neutral side chains or
HAP-binding octa glutamic acid patterns at one or both protein termini. ELPs have
sequence-specific properties that affect how they interact with ions, bind HAP, and
spread HAP nanoparticles. With ELPs that bind with HAP, calcium phosphate cement
can be further improved to provide materials with enhanced mechanical strength,
injectability, and anti-washout properties [55].
Elastin-like polypeptides possess some adaptable properties which enable them
to form different nanostructures, including nanoparticles, nanofibers, and nanocom-
posites [56, 57]. When utilized in drug delivery, these nanotechnologies increase
the effectiveness of the therapy because the nanostructure can target the molecule,
increasing its stability and increasing contact surface. This could boost these struc-
tures’ activity, which is essential for maintaining their thermodynamic properties
[58, 59].

4.8 CRISPR-Cas 9

Streptococcus thermophilus, a bacterium frequently employed in the dairy industry,

was utilized to identify the CRISPR/Cas mechanism system for the initial time in
2007. By functioning as a highly effective “antiviral system” to prevent viral infec-
tions, this system serves as an adaptive immunological mechanism that combats
this gram-negative bacterium. Later, it was found that prokaryotes, which comprise
84% of archaea and 45% of bacteria, are the organisms where CRISPR is currently
most common. The term “CRISPR/Cas system” refers to the locus, which is perpet-
ually active in the presence of CRISPR-associated (Cas) genes which generate the
endonuclease enzyme. CRISPR’s operons of Cas genes, spacers, and repetitions are
its essential components [60]. Table 2 describes the biomaterial vectors along with
their Cas9 cargoes with their applications and advantages.
Since viral predation poses a constant threat to bacteria, they have developed a
range of defense mechanisms, including CRISPR/Cas systems. CRISPR-Cas 9 oper-
ates by modifying DNA. The nuclease protein (Cas9) of the Streptococcus pyogenes
Type II-A system addresses a particular sequence of DNA under the direction of an
Different Techniques of Genetic Engineering Used for the Development … 59

Table 2 Biomaterial vectors along with their Cas9 cargoes: Applications and Advantages [63]
Cas9 Cargo Delivery System Application Advantages
DNA Palmstearin derived Analyzing The PS-Lips cationic lipid
fatty acyl chains nanoparticle molecular nanocarrier system’s
architecture effect on liposomes successfully
Cas9 plasmid delivery carried genome-editing
and characterization of tools containing CRISPR/
delivery to HEK-293 Cas9 encoded pDNA
cells
DNA Turbofect Identification of Nf-1 Effective pain
pain-related management
therapeutic targets
mRNA Extracellular vesicles Targeting in MOLM13 In bothhuman cells and
derived RBCs cells xenograft animal models,
RNA medication delivery
with RBCEVs exhibits
highly strong microRNA
inhibition and
CRISPR-Cas9 genome
editing with no
detectableharm
Protein Nanoclews involving Disruption of EGFP in Enhanced delivery of
yarn-like DNA U2OS-EGFP cells CRISPR-Cas9
in vitro and in vivo
DNA Liposome-Exosome Lipofectamine 2000 Exosome-liposome hybrid
hybrids and nanoparticles are
HEK293FT-derived promising for in vivo gene
exosome performing editing since they can
targeting in transport the
mesenchymal stem CRISPR-Cas9 system to
cells MSCs
mRNA Zwitter-ionic lipids Lung cancer cell Improvement in safety
amino delivery and in vivo and effective usage of
targeting gene editing
Protein-tagged Arginine coated Works by targeting The method assures the
NLS and glutamic cationic gold Hela cells and development of
acid nanoparticles disrupting signals for “weaponized”
(ArgNPs) cancer survival macrophages for cancer
immunotherapy due to the
enhanced attack and
destruction of cancer cells
mRNAwith Cholesterol,DSPC In TTR amydoilosis Helps in reducing
modified sgRNA lipid, PEG-2000 for gene targeting Transthyretin (TTR)
DMG and LP01 lipid serum protein
concurrently with in vivo
genome-editing levels that
are clinically significant
(continued)
60 A. Chhina et al.

Table 2 (continued)
Cas9 Cargo Delivery System Application Advantages
Highly negative Lipofectamine 2000, Delivery which is When compared to DNA
GFP fused protein RNAiMAX in vivo to mice cochlea transfection, delivery of
unaltered complexes
resulted in genomic
modifications of up to
80%
DNA Cationic polypeptide Disrupting GFP and The system provides a
made with surface targeting in HeLa flexible gene-editing
PEGylation tumor-bearing mice platform for biological
research and therapeutic
applications, enabling
multiplex gene knock-out,
gene knock-in, and gene
activation in vitro and in
vivo

associated RNA sequence, is the system that is used for the majority of CRISPR appli-
cations in bacteria. This method causes an incision in the designated area, allowing for
DNA editing. The DNA that recognizes the protospacer is bound by the Cas9-sgRNA
complex, which leads to a slow unraveling of the DNA. The CRISPR-Cas9 tech-
nology has the advantage of enabling researchers to more precisely control how the
CRISPR/Cas systems are activated by targeting transcriptional activators to specific
genomic regions. The latest advances in the interaction of Porphyromonas gingivalis
(P. gingivalis), Enterococcus faecalis (E. faecalis), and Streptococcus mutans (S.
mutans) with bacterial functions and CRISPR/Cas systems are relevant to dentistry
[61].
Delivering the CRISPR-Cas complex to particular cells or tissues is challenging
and laborious because its components must transcend tissue and cell membrane
barriers to enter the nucleus, where they can affect the nuclear genome. The two
broadly used physical techniques for introducing the CRISPR-Cas complex into
cells are electroporation and single-cell microinjection. These techniques are crucial
for editing genes in developing embryos and creating transgenic animals [62].
Due to their adaptability, biocompatibility, and rising transfection effectiveness,
biomaterials are increasingly being used as non-viral vectors. Capacity restrictions
are not an issue, and nanoparticles can be further tailored to enhance nuclear transport
and tissue selectivity. The capacity of biomaterials to be tuned offers considerable
potential for enhancing CRISPR-Cas9 delivery for in vivo gene editing. Since there
are practically no restrictions on how biomaterials can be customized, this gives phys-
ical and viral delivery a particular advantage. This is due to the fact that several organ
systems along with their types of cells will eventually need gene editing methods
that are specially tailored to their microenvironments [63]. By incorporating various
biomaterials into them, Cas9 can be supplied in the form of DNA, plasmids, mRNA,
and proteins.
Different Techniques of Genetic Engineering Used for the Development … 61

4.9 3D Printing

The methods of genetic engineering are utilized to extract and mix particular DNA
from diverse materials to create chimera DNA. When the chimeric DNA is sent back
with the help of plasmids to bacterial cells, it can then be duplicated. The survival of
cells carrying chimeric DNA can be backed by bioprinting techniques like inkjet-,
extrusion-, and laser-based printing. They can design chimeric biomaterials for 3D
printing in a controlled and cost-effective way. Recombinant DNA technology allows
engineers to employ chimeric biomaterials in a number of biological research and
development applications by taking their desirable traits from nonhuman origin. The
creation of fake CAR T cells for use in adoptive cell therapy for the treatment of
cancer is the most well-known application of chimeric biomaterials. Particularly,
Fc-chimeric proteins may be utilized for the identification of desired stem cells
and as test compounds in advanced 3D printers. Chimeric organoids that mimic the
tumor microenvironment can be designed by bioprinting cancer cells and healthy
mammary epithelial cells. Furthermore, chimeric biomaterials can be included in
drug delivery systems to improve the therapeutic potency of previously accessible
proteins and drugs. Recently, attempts have been made to combine chimera biomate-
rials with biopolymers [19]. Despite being incredibly effective in CAR T-cell therapy
as a customized approach to cancer treatment, chimeric biomaterials can potentially
have unfavorable side effects and flaws in patients. Additionally, because of its high
cost and drawn-out approach, CAR T-cell treatment is not commonly available.
The ethical ramifications of research using human organoids and/or the introduction
and integration of chimera material into a host mammal are also complex. Sound
ethical principles must therefore be upheld as chimeric biomaterials are increasingly
used in modern science. It is necessary to harvest DNA from numerous genomes in
order to create chimeric biomaterials, which calls for the employment of additional
3D printing settings that allow high cell survival and density. Across all printing
techniques, there is a definite trade-off between cell survival, resolution, and price.
Chimeric biomaterials are designed using recombinant DNA technology, which
reconstructs genes from many species into desired effects by programming them
at specified gene sequences. Chimeric polymers have a much greater degree of
control over the amino acid sequences than their chemically synthesized equiva-
lents, which is much greater than what is possible with solution or solid-phase peptide
synthesis. Incorporating stimuli-responsive motifs into a chimeric polymer’s back-
bone allows for accurate phase changes in response to changing temperature, pH,
and ionic strength. Additionally, this addition might enhance interactions between
an implant and already-existing cells. Chimeric polymer-based biodegradable struc-
tures can also maintain their temporal and spatial precision so that breakdown only
happens in the presence of specific enzymes. This can be achieved through the addi-
tion of amino acids which are prone to breakdown [64]. Engineers could advance
the creation of treatments that address a variety of unresolved medical issues that
have plagued society since the dawn of humankind by leveraging desirable properties
from other sources.
62 A. Chhina et al.

4.9.1 Epigenetic Modifications/Omics-Based Approaches

Recent advances in biology technology, notably in the domain of omics approaches,

have enabled functional high throughput reading of the cell’s genome, epigenome,
transcriptome, proteome, and metabolome. The capacity of omics-based techniques
in forecasting the biological response to designed biomaterials is the primary check-
point in developing the next generation of biomaterials, and it has the potential to
revolutionize biomedical research and replace the traditional “trial and error” method.
These omics-based tools can also be applied to the study of the in vivo behavior of
biomaterials, a field that is based on semi-quantitative imaging-based techniques
instead of precise computational techniques [65].
In order to make hydrogels whose mechanical characteristics depend on calcium
concentration, calmodulin, a protein that changes conformation in order to bind
calcium ions, was used and is one of the biomaterials that is prepared through a
protein [2]. Successfully combining hydrogel encapsulation and mRNA transfection
opens up new avenues for the prospective clinical delivery of mRNA-engineered cell
products. Primarily, improved cell attachment is necessary for significant secretion
of the IVT-mRNA encoded protein, a recently discovered synergism between mRNA
transfection of cells and their microenvironment. The increased secretion of indige-
nous HGF and mRNA-overexpressed VEGF in collagen hydrogels indicates that
material-mediated effects can further modulate the cells’ secretome in concert with
IVT-mRNA overexpression [2]. Depending on the method of nuclease delivery, gene
editing needs different regulatory provisions than other gene therapy medications.
The location where nuclease genes are inserted into the genome, how the genes are
delivered, how they are administered, and what kinds of gene editing nucleases and
sgRNA are used can all affect the level of risk and the categories of risk factors are
one of the challenges when a gene needs to be modified [66].
The most extensively researched epigenetic modifications are DNA methyla-
tion and histone protein alterations, that are directly involved in regulation of the
gene expression levels and defining the characteristics of the cell. Methylation of
DNA alters gene transcription by preventing transcription factors from attaching to
the DNA. The same is true for post-translational histone modifications, which are
crucial for determining chromatin structure and regulating DNA expression. Enzy-
matic proteins that are involved in these epigenetic changes can also reverse them.
These changes are practical because they can be undone, which opens the door to
using them for in situ tissue regeneration. In fact, chromatin structure and epige-
netic processes in cells have been regulated by nanomaterials [67]. In a recent study,
epigenetic regulators were employed to stimulate osteogenic differentiation in human
MSCs113. These regulators included gemcitabine, decitabine, I-CBP112, chidamide,
and SIRT1/2 inhibitor IV. Notably, stem cells obtained from elderly adult donors
showed a fivefold increase in osteogenic efficacy when treated with gemcitabine and
decitabine. This research emphasizes the function of the nucleosome remodeling and
epigenetics in cellular differentiation as well as their potential application to in situ
tissue regeneration [68]. The surface organization and material energy of biomate-
rials influence the epigenetic patterns of cells, which can subsequently influence the
Different Techniques of Genetic Engineering Used for the Development … 63

gene expression of cells that come into contact with the material. Substances in tissue
engineering that may affect epigenetic pathways are titanium, silica, PLGA, bioglass,
and ceramics [69]. Another organic material that has undergone epigenetic function-
alization is silk. James et al. examined an anti-sense-miRNA-214 silk device utilizing
surface coating, which led to a continuous release of miRNA inhibitors up to 7 days
in vitro and increased the expression of osteogenic genes in the human mesenchymal
stem cells implanted on these devices [70]. These findings would suggest that this
unique technique could be helpful for localized bone tissue creation and for promoting
osteogenesis at the implant surface. Collagen sponges and macroporous biphasic
calcium phosphate scaffolds combined with HDAC inhibitors (HDACi) caused the
creation of woven bone and newly produced bone at the scaffold’s interface, which is
another epigenetic mechanism for collagen functionalization that has been explored
[71]. Although Cre recombinases are widely utilized, optogenetics has been found to
be beneficial for a variety of phylogenetic histone deacetylases, methyltransferases,
acetyltransferase inhibitors, and proteins that recruit these enzymes. Included in this
group of proteins are KYP, TgSET8, NUE, PHF19, Sin3a, Sirt3, NcoR, HDAC8,
RPD3, and Sir2a [72].
Genetic code expansion (GCE), a related approach, is comparable to it. Next-
generation protein engineering, in particular GCE, enables heterologous expression
of proteins with NCAA/UNAA inclusion in proteins in both prokaryotic and eukary-
otic organisms [73]. GCE promotes biomedical and tissue engineering research by
developing innovative protein-based biopolymers with broad structural and func-
tional flexibility. Silk, elastin, collagen, and fibrin are used to create a variety of
biomaterials that are created using the GCE process. The ability to enhance the
production of biomaterial proteins by GCE is expected to usher in a new era of tissue
engineering and biotechnology. According to recent advancements in tissue engi-
neering, customized tissue constructs with anisotropic architecture, heterogeneous
cells, and composite nano/biomaterials are being created for regenerating functional
tissue and organotypic tissue models [74]. Because they enable precise control over
the loading of bioactive moieties, microstructure, and handling, biomaterials with
spatiotemporal variations are essential for 3D manufacturing. GCE improves the
possibility of producing biomaterials made of growth factors and congener proteins
that act as brokers for interactions between cells and the matrix. GCE presents a
unique opportunity for the creation of bioinspired proteins and customized biomate-
rials that show potential for use in precision and personalized medicine. GCE creates
protein biomaterials with all-inclusive qualities to ensure enhanced interactions
between the components of the tissue engineering trio [74].
64 A. Chhina et al.

5 Pre-Clinical and Clinical Relevance of Genetically

Engineered Biomaterials

Biomaterials can serve a variety of purposes in cartilage repair and regeneration,

including mechanical support systems (such as visco supplementation, defect plugs),
delivery vehicles for the delivery of therapeutic molecules as well as scaffolds for cell-
driven repair of tissues. Hydrogels, 3D polymer networks respond viscoelastically
and are the optimal materials for cartilage repair since they are high in water content,
like healthy cartilage [75]. Drug delivery, the development of cell-rich bioengineered
devices, and tissue and immunological engineering are some possible applications
for smart materials [76]. The mechanical properties of hydrogels can be reversibly,
wavelength-specifically, dose- and space-controlled tuned by incorporating a protein
under optical control into polymeric polymeric materials. These light-controlled poly
(ethylene glycol) (PEG) hydrogels were first made successful, according to a PhoCl
protein, which raised the potential of their practical implementation [77].
Biomaterials that are appropriate for human use and of neurosurgical grade may
provide a feasible replacement. In particular, the neurosurgical grade using the bioma-
terial Duragen PlusTM matrix that has been approved by regulators and is mostly
utilized in duraplasty treatments. Type I bovine collagen is used to make it. Here, it
permits fibroblast percolation to restore and heal the dura mater after an intraoperative
breach, and it is said to be resorbed after six to eight weeks. According to reports, the
substance is conformable, clean, and biocompatible as well as lacking immuno-
genicity, cytotoxicity, and pyrogenicity. It has been demonstrated that Duragen
PlusTM, which has been transplanted into the nervous system, stimulates neural
cell growth, which might involve that of rat cortical neurons, without producing any
adverse side effects [78].
dECM-based biomaterials enhance the advancement of more precise delivery of
a certain drug at the intended region and facilitate chemical component distribution
as carriers. Decellularized ECM biomaterials have also been investigated for growth
factor delivery due to the intrinsic qualities of ECM and its natural interactions with
various growth factors. For instance, the decellularized matrix containing hydrogel
was designed to load bFGF to treat spinal cord injury since ECM is essential in
directing growth factor signaling in vivo for bone regeneration [79]. The delivery
of biological cues using dECM-based material is advantageous because it increases
in vivo utilization and effectiveness. As a number of ECM goods (AlloDerm, Life-
Cell Corp.), as well as injectable ECM materials (ACell’s CytalTM Wound Matrix,
MicroMatrix® , and CorMatrix® ), have already passed the commercialization stage,
injectable ECM hydrogels may soon follow this trend. It is crucial to remember that
musculoskeletal dECM biomaterials have had a tremendous impact on clinic tissue
repair all over the world. Applications for dECM biomaterials in preclinical studies
have included urethral reconstruction, bone tissue healing, and muscle regeneration
[79, 80].
Biomaterials can be used in a variety of ways to induce immunological tolerance
because they offer reliable control over immune cells’ interactions with specific
Different Techniques of Genetic Engineering Used for the Development … 65

antigens. They can therefore make excellent use of the immune system’s numerous
internal regulating mechanisms. One way to promote tolerance is through inducing
energy or deletion of self-reactive lymphocytes. Additionally, targeting helper T cell
activation toward the regulatory pathway can help prevent the activation of other T
cells’ undesired effector functions.
Biomaterials allow for engineered solutions to complex biological issues, as was
the case with vaccine development, and they hold hope for the future of our ability to
effectively treat autoimmune diseases and aberrant immune activity [81]. Macroscale
scaffolds based on biomaterials can be utilized to disseminate immunomodula-
tory elements like proteins (like cytokines and antibodies), oligonucleotides (such
silencing RNA and plasmid DNA), and scavengers that eliminate or reroute harmful
substances (like ROS) locally. Future biomaterials-based cell carriers and niches
may be created to monitor certain immune responses for diagnostic, investigation
and observation, and clinical study purposes together with their use for therapeutic
goals. For instance, the delivery of a pancreatic-islet lysate to gather, separate, and
analyze the repertoire of T cells that are diabetogenic has been investigated using
a PLGA-based microporous scaffold. Similar to this, a locally applied adjuvant-
and antigen-loaded alginate microneedle patches can attract T cells indicating active
systemic responses. [82].
By combining a suitable biomaterial (which is an essential part of the extracel-
lular matrix involving the airway system) using airway organoids harboring disease-
relevant cell types, it is possible to construct a structure like the tissues, with favor-
able functional and morphological attributes. Lung-on-a-chip systems also need to be
enhanced and optimized in order to replicate and imitate the critical pulmonary tissue
microenvironment conditions. This could aid in the discovery of innovative medica-
tions and therapeutics for challenging lung diseases like the newer SARS-CoV-2. A
promising method for determining potential side effects of recently created SARS-
CoV-2 medications and vaccinations is to combine lung-on-a-chip technology with
heart, liver, and kidney organs-on-a-chip systems [83]. An acellular dermal matrix
created by the usage of skin from the human deceased when mixed fat grafts in skin
graft and breast reconstruction surgeries, to enhance the localization and survival of
transplanted fat cells procedures to improve the localization and survival of trans-
planted fat cells. In the commercially available product Epigraft, collagen has been
used to enable the organization of composite cell types within tissue grafts, such
as skin grafts. Lately, cardiomyocytes derived from stem cells and collagen sheet-
laden epicardium cells have been used in combination for myocardial tissue healing
following infarction [84].
Hence it is believed that biomaterials will eventually be used for more sophisti-
cated purposes, such as signal transducers that trigger cellular roles. Early viability
studies in animals suggest that remote control of cell activity is possible using external
actuators including ultrasonic, magnetic fields, and electronic inputs. These external
actuators generate forces that the body can withstand [1]. These technologies are
in their nascent stages, so advancements in the production of external signals will
be required for propelling their development, and therefore, genetically modified
biomaterials will reach heights in the near future.
66 A. Chhina et al.

6 The Future of Genetically Engineered Biomaterials

Over the past fifteen years, there has been a lot of research into genetically modified
biomaterials, and CRISPR has already begun to play a significant part in the advance-
ment of this field. With the advancement of manufacturing techniques that produce
genetically engineered biomaterials, new ways have been paved for harnessing this
potential of the manufacturing sector and formulating genetically engineered bioma-
terials from it. Scalable production has provided hope for the synthesis of genetically
engineered nanofibrous materials. With features like biocompatibility, biodegrad-
ability, prospective energy collection, thermal, optical, and carcinogenic detection
potentialities, among others, these methodologies may arise in the not too-distant
future. Intelligent and genetically generated biomaterials and medicine delivery
systems have created tremendous strides in the last few years. It is being tested
to create new materials with unique properties. Researchers presently concentrate
mostly on designs found in nature, such as spider silk motifs, in order to develop
materials with outstanding mechanical properties. The design of novel materials, not
present in nature, that are based on an understanding of the relationship between
the structure of protein-based materials and biorecognition, self-assembly, and char-
acteristics has the most potential, though [1]. Bio adaptability would be the best
factor for selecting the optimal biomaterials that prove to be useful in certain tissue
repair. Biomaterials with exceptional bio adaptability must possess some essential
qualities, such as adaptable components that mimic cell response and tissue recon-
struction, matched mechanical properties with natural tissues, suitable surfaces with
advantageous tissue reaction, and biomimetic multi-level structures.
Tissue engineering and regenerative medicine, two techniques that are now in
development, will someday transform medical implants. Tissue engineering will
make it possible to create functional, living alternatives for many tissues and organs.
Tissue engineering will benefit from developments in biodegradable polymers, fast
prototyping, drug administration, cell culture, stem cell procedures, angiogenesis,
and biomimetic approaches for building extracellular, matrix-like biomimetics. The
potential applications of clever systems and micro- and nanofabrication are almost
endless. Future in vitro systems will be built on a sophisticated knowledge of how
biological systems engage with synthetic surfaces. These novel technologies will
also be carefully and morally addressed because they have the potential to alter
expectations for humans. Another area of study that will improve the functionality
of implants is lowering the risk of infection. This can be assisted by antibacterial
surface coatings, non-fouling surfaces, and antibiotic-controlled release mechanisms.
The development of devices that function as both a device and a drug will also be
made possible by advancements in controlled drug release in general.
Different Techniques of Genetic Engineering Used for the Development … 67

7 Conclusion

This chapter offers a comprehensive overview of the biomaterials industry. It is

meant to give the reader a vantage position from which they can start to situate all
the subthemes within the context of the bigger picture. To reiterate a crucial point,
the study of biomaterials may be the one that is most interdisciplinary. As a result,
in order to be proficient in their area, biomaterials scientists need to be knowledge-
able in a wide range of science, technology, engineering, and medical specialties.
Being involved in a mentally challenging project that advances our knowledge of
fundamental sciences and also helps to alleviate human suffering is the reward for
mastering this volume of material. The journey of biomaterials can surely be consid-
ered a success from the significance they hold for contemporary medical treatments,
the market’s economic growth, and steady development of the industry over the past
50 years. The body will require fewer synthetic elements as a result of regenera-
tive medicine. We anticipate the need for genetically engineered biomaterials to last
well into this century and that many uses will continue to call for synthetic mate-
rials. Modern biomaterial development necessitates the design of materials that are
bioinert and at the same time, also engage with and even react to their surrounding
living environment. Biological cues that may communicate cell adhesion, migra-
tion, proliferation, differentiation, or even apoptosis must be provided by bioactive
materials. The materials can communicate with the nearby biological environment
in this way to convey their messages, for instance, by sending a signal of growth to
encourage tissue repair. Bioactive substances with built-in responses to stimuli may
respond to nearby physiological activity, such as by releasing a drug or breaking
down to respond to a biological stimulus. Future genetically engineered intelligent
biomaterials will depend heavily on biomaterials’ capacity to detect changes in their
surroundings or biological demand.
Conclusively, this is the stage where our knowledge of cellular biological sciences
and biochemistry allows us to incorporate particular biologic functions into geneti-
cally engineered biomaterials. As new information is discovered, it can be immedi-
ately incorporated into a biomaterial to govern specific amino acid, lipid, or carbohy-
drate sequences that regulate cell differentiation, immune responses, or other biolog-
ical phenomena. Thus, a key activity in the future will be the integration of materials
science and cell biology knowledge to create a new class of materials that can truly
facilitate desired medical outcomes.

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Aashveen has a master’s degree in pharmaceutical sciences and

a keen interest in developing novel drug delivery systems. She
has worked on projects involving medical writing skills involving
the knowledge of polymer sciences and how they work in various
delivery systems along with their roles. Some of these articles
have been published and some are in the process of publishing.
72 A. Chhina et al.

Vridhi is a Formulation Scientist with experience in the

Research & Development sector of the pharmaceutical industry.
Having a Master’s degree in Pharmaceutical Sciences, she works
on novel drug delivery systems to enhance active pharmaceutical
ingredients’ therapeutic potential and reduce the associated side
effects. She is actively involved in the publication of scientific
and health-related articles.

Shubham Thakur completed his undergraduate and postgrad-

uate studies in pharmacy at Guru Nanak Dev University in
Amritsar in 2016 and 2018 respectively. Presently, he serves as
an Indian Council of Medical Research-Senior Research Fellow
and is pursuing a Ph.D. in the Department of Pharmaceutical
Sciences at Guru Nanak Dev University, Amritsar. His research
focuses on the advancement of novel drug delivery systems,
aiming to enhance the kinetics, efficacy, and safety of existing
drugs. Specifically, his current project involves developing a safe
oral formulation for pregnant women. Till now he has published
25 articles and 5 book chapters in Scopus indexed journals and
books with h-index of 5.
Green Methods for the Development
of Bone and Tissue Engineering-Based
Biomaterials

Avipsa Hazra , Gowrav Baradwaj , A. S. Dhanu ,

Gobianand Kuppannan , Malarvizhi Arthanari , and B. M. Kanthesh

Abstract Bioengineering of bones and tissues includes the culmination of principles

of bioactive compounds, scaffoldings, and developing materials for the treatment of
body cells and organs that are required to be replaced due to damage of some kind.
However, synthetic materials used are known to cause extensive pollution and thus to
develop methods with no or lesser pollution multiple green manufacturing technolo-
gies are being designed. Due to their outstanding biodegradability and biocompati-
bility in biological media, green materials are proven to be a useful factor in medicine.
Though this technology is still in its cradle stage, there have been multiple kinds of
materials like chitosan, collagen, alginate, and corals that are being implemented
for manufacturing composite for hard tissue implants such as artificial bone, bone
cement and for the construction of multiple-generation biomaterials. This chapter
will cover the various materials and methodologies that are already in practice or are
being developed to regenerate bones and tissues, keeping in sight the safety of the
environment and to give us a pollution free world.

Keywords Bioengineering · Biomaterials · Scaffolds · Bone tissue engineering

Abbreviations

TE Tissue Engineering
BE Bone Engineering

A. Hazra · G. Baradwaj · A. S. Dhanu · B. M. Kanthesh (B)

Division of Molecular Biology, School of Life Sciences, JSS Academy of Higher Education and
Research, Mysuru, Karnataka 570015, India
e-mail: kantheshmb@[Link]
G. Kuppannan
Department of Microbiology, Noorul Islam College of Dental Sciences, Aralummoodu,
Thiruvananthapuram, Kerala 695123, India
M. Arthanari
Vivekanandha College of Arts and Sciences for Women (Autonomous), Elayampalayam,
Tiruchengode, Tamil Nadu 637205, India

© The Author(s), under exclusive license to Springer Nature Singapore Pte Ltd. 2023 73
R. Malviya and S. Sundram (eds.), Engineered Biomaterials, Engineering Materials,
[Link]
74 A. Hazra et al.

PGA Polyglycolic Acid

PEG Polyethylene Glycol
PCL Polycaprolactone
3D Three-dimensional
BTE Bone Tissue Engineering
ECM Extracellular Matrix
ALP Alkaline Phosphatase
GlcNAc N-acetyl Glucosamine
GAGs Glycosaminoglycans
PMMA Polymethyl methacrylate
PLLA Poly-L-Lactic acid
MSC Mesenchymal Stem Cell
DNA Deoxyribonucleic Acid
HAP Hydroxyapatite
HFIP Hexafluoro-2-propanol
GHG Greenhouse Gases
HEMA 2-Hydroxyethyl methacrylate
PEO Polyethylene oxide
PVA Polyvinyl alcohol

1 Introduction

With an increase in the rates of diseases affecting various organs of the human body,
and the rate expected to double by the end of 2030, the need for the ability to replace
any such organ has become imperative [1]. It has led to the expansion of the field of
medicine known as regenerative medicine, which involves tissue engineering (TE),
regeneration of bones, and self-healing through activation of the host’s immune
system [2]. It is a multidisciplinary discipline that integrates physics concepts with
principles derived from life science to provide clinical treatment strategies. The
concept of regenerative medicine emerged in the early 1990s, and since then, efforts
have been made to strengthen research in the field.
Although bone bioengineering is a type of tissue engineering, it is considered
distinct from tissue engineering of other organs in today’s context, with bone engi-
neering being one of the most developed arenas. The creation of bio-active artificial
cells and tissues for the substitution and regeneration of faulty tissues or organs is the
focus of tissue engineering [3]. This includes isolation of suitable cells, development
of compatible scaffolds, and implantation of the molecules into the tissues to alter
functions or initiate repair of damaged organs and tissues [2]. Bone engineering (BE)
involves the synergistic effect of engineered biomaterials with the host’s recovery
factors to regenerate bone tissues. It appears to have no side effects so far and allows
for healing of bone fractures and other flaws using the host’s cells [4]. However, with
Green Methods for the Development of Bone and Tissue … 75

the progress and improvement in the field, there has been a distinct rate of pollution
because of it and has led to the environmental depletion.
The raw materials being used, the chemicals being depleted, and the destruction of
faulty end products have led to soil erosion as well as major water pollution. It has also
been known to affect the health of individuals being involved in the process which
has now led to the thought of developing a process that is responsible toward human
health and environment- green practice. Green practices involve the implementation
of resource- and environmentally conscious methods throughout the entire lifecycle,
from design to construction, operation, maintenance, renovation, and destruction [5].
The traditional considerations of economy, usefulness, durability, and comfort are
expanded and are now being complemented with green practices.
The primary concept of green practices was presented by Anastas and Warner who
put forward the foundation based on 12 principles, which revolves around reducing
waste, hazardous products and derivatives, balancing atom economy and energy
efficiency, developing designs that make use of catalysis and based on the usage of
benign compounds and renewable sources, and always having a detailed plan about
degradation of the product, pollution control, and accident prevention. Thus, in the
light of 2023, today scientists from all fields are working on improving the production
of biomaterials such that it keeps the environment stress free [6].

2 Usual Methods and Their Issues

To develop a system which was economically sustainable, environmental wellness

had often got a backseat and so has been the case with the initial bioengineering
methods. The prime concern for the methodologies was the disposal of waste—as
most of it is unusable, they have often been disposed of into landfills. Alongside the
foul odor and land wastage, the waste has even been recorded to yield pathogens
[7]. Even though theoretically the use of nanoparticles to maintain cell viability
seemed like a sublime idea, over the years the use of it in medicine and under human
biology has posed multiple adverse effects due to the variation in manufacturing,
particle size, chemical composition, and materials involved in the production of
the nanoparticles [8–10]. Poly-glycolic acid (PGA), lactic acid, polyethylene glycol
(PEG), and polycaprolactone (PCL) were used as the sources to produce synthetic
polymers for the fabrication of scaffolds for tissue engineering [11–14]. However, the
biological and mechanical functioning of the said chemicals proved to be weakened
when used singly in tissue engineering and demanded reinforced materials for the
increase of the functioning properties [10].
Usually, scaffolds in tissue engineering have been created and produced using
methods such as freeze-drying, salt leaching, gas forming, and phase separation;
however, the precision of this method to develop complex structures and fit the
internal architecture is insufficient. Owing to these issues, production of scaffolds
with the use of 3D printing came into play as it was able to mold materials into
the desired shape [15–17]. Nevertheless, the use of 3D printers is limited because
76 A. Hazra et al.

not every substance can be modified in them, reducing the range of sources for
producing scaffolds in tissue engineering [18]. Among each of the other tissue engi-
neering processes, bone regeneration is the most critical one as it is founded on
numerous principles of molecular and cellular biology, biological development, and
structural biology, all of which are monitored by biomechanics and bioengineering.
This is because bone tissues are responsible for dynamics of the body and must be
adjusted according to its different locations. The most difficult aspect of bone tissue
development is retaining vascularization of blood vessels by making the scaffolding
porous and integrating it with the host system. With current synthetic mechanisms,
only the periphery of a scaffolding is suitable to be used as an implantation thus
limiting its use. Owing to these reasons, the synthetic mechanisms for production of
bone tissues are not widely in use [19].
Usually, the current strategies are based on either the cellular or source material
mechanism, thus for the usage to be widespread all the factors must be collaborated.
Bone engineering is also an expensive process due to it being patient-dependent
and poses health issues due to erroneous cell isolations, seeding, culturing, and
implantation [20]. Methods need to be established such that all the factors can be
culminated, and they impose no adverse effects on human health [4]. Moreover,
the growth in the field of tissue regeneration has been quite stagnant over the past
two decades. The clinical translations of the basic research for bone, cartilage, and
ligament regeneration have met a roadblock ever since the involvement of stem
cells had been implemented. Though the synthetic polymers at one time seemed
like the most promising solution to tissue engineering, in the current scenario there
seems to be an unresolved failure at the translation stage for application of the
methods [4, 21]. Also, for the current approaches to be effective a proper cell for the
source must be determined for the model to be designed. However, the separation of
human cells and culturing it in vitro such that its sensitivity is maintained, and its
differentiation capacity is not reduced is a tedious task. Also, for some cells to work
they need to be converted into their embryonic forms. Furthermore, the different
methods are influenced by the tissue origin, the patient’s age, and the individual’s
health condition. Also, the environment around the cell must be replicated exactly
to establish the correct behavior of the polymer of the scaffold. Thus is the need of
designing materials from natural sources which can be used as scaffolds by exploiting
their chemical, physical, and biological properties [22–29]. These natural fibers can
also be combined with synthetic materials depending on the need of the tissue at the
site [30].
Another issue that crops up while developing regenerated tissues is the innervation
of those tissues. For the structures to attain complete functionality after implantation,
the structures need to get consistent with other growing tissues in the region [31].
These requirements of innervations have been seen to be very critical for regions like
bladders and small intestines [32]. This association not only develops an interlink
with the nervous system but also rejuvenates the tissue regenerated [33, 34]. For
the bioengineering of peripheral nerves, an important criterion is the inculcation of
neurotrophic factors which would ultimately lead to synapse formation, even muscle
Green Methods for the Development of Bone and Tissue … 77

tissues require electric impulse for stimulation. These are critical steps and are diffi-
cult to implement as well [35, 36]. Some grafted organs require neovascularization
to occur, such that the branches developed after morphogenesis can help in accep-
tance of the implantation, but it is more than often seen to be not there with synthetic
polymers. However, in other cases the synthetic materials sometimes exceed the
functionality rate of native tissues, leading to some complications [37, 38]. Overall
many trials with synthetic fibers have been postponed owing to the financial factors
as some of the synthetic fibers which are best suited for human use have a high cost
[39].

3 Development of Green Methods and Its Types

The idea of developing environmentally friendly techniques for bone and tissue
engineering arose because of the dangers that synthetic techniques posed to the
human body and the environment [40]. This resulted in the creation of engineered
biomaterials and scaffolds composed of natural polymers that offer little to no risk to
the environment or human health. Green approaches involve using healthy, non-toxic
materials and procedures to lessen the hazardous and damaging effects of scientific
study. The reduction of pollution’s harmful impacts on the environment and the
human body is a key objective of green technologies and practices [41]. The goal
of bone tissue engineering (BTE) is to create new, functional bone tissues using
knowledge of the structure of bones, bone mechanics, and tissue building. To put it
another way, if you want to efficiently regenerate as well as repair bone, you must
first understand the biology of bone and how it develops. The ideas for biologically
active chemicals, scaffolds, engineering, and material production are also combined
to build, repair, or treat injured tissues or organs [42–44].
Because of their inherent bioactivity, including cell adhesion and proliferation,
as well as the fact that they are harmless to native, healthy tissue, natural biomate-
rials are critical for tissue engineering. Biomaterials made from natural substances
include those made from glycosaminoglycans, cellulose, chitin, collagen, chitosan,
elastin, alginate, fibrinogen, dextran, gelatin, laminin, and silk. Among these mate-
rials, collagen, fibrinogen, glycosaminoglycans, elastin, and laminin are mimics of
original extracellular matrix (ECM) components. Extracellular matrix is crucial for
tissue regeneration because it supports cells physically and controls cellular processes
including growth, proliferation, differentiation, and homeostasis, enabling the body
to repair tiny flaws in cells, tissues, and organs [45, 46].
The human body is unable to regenerate deficiencies because of a shortage of
extracellular matrix (ECM) to make up large defects. Natural biomaterials were
created in this manner to be employed in applications involving tissue engineering.
These natural biomaterials are employed in hydrogels, that additionally serve as an
excellent tool for tissue engineering. Hydrogels are capable of being made from
both synthetic and naturally occurring biomaterials. Artificial polymers include poly
78 A. Hazra et al.

HEMA, PEO, and PVA. Naturally occurring biomaterials used in hydrogel prepa-
ration include collagen, chitosan, gelatin, fibrin, alginate, agarose, and many more
[47].

3.1 Collagen

In mammals, the protein collagen is a particularly abundant structural protein in

the extracellular space. It accounts for approximately 30% of total protein weight.
These are fibrous proteins which are major building block components in most of
the animals (invertebrates, vertebrates) and sponges [44]. The chemical makeup,
microscopic patterns, and mechanical strength of collagen fibrils all work together
to preserve the structural integrity of each connecting tissue. Collagen, a component
of the ECM, is found naturally in human tissues such as bone, skin, ligaments, and
other types of connective tissue. The first type of collagen fibrils is relatively homo-
geneous in size, ranging from 50 to 500 nm [48–50]. Collagen is among the most
used biomaterial for nanofibrous scaffolds, and several recent research articles have
reported on the use of collagen and collagen-based composites in bone tissue engi-
neering. According to some studies, the collagen/chitosan/hydroxyapatite composite
is extremely effective in promoting osteogenic differentiation of mesenchymal stem
cells [51].
Collagen is the triple helix structure made up of 2 α1 chains and one α2 chains, each
chain weighing approximately about 100 kDa. Among the 29 types of collagens, type
I is the most used for bone tissue engineering. The connective tissues of the vertebrae
almost all contain collagen type I, while tendon and bone contain the most. The ability
of this collagen to produce insoluble fibers also gives tissues a high tensile strength
and stability [52]. Numerous discoveries proved that type I collagen can be isolated
from different animal species and tissues. The most common source of its extraction
is from cattle, which could have concerns including spreading bovine spongiform
encephalopathy. Human recombinant collagen, on the other hand, has been used in
tissue engineering processes [53]. It is possible to change the mechanical properties
and collagen rate of degradation by adjusting various characteristics, including the
amount of cross-linking.
Collagen is currently reported to be used in a variety of scaffolds for bone tissue
engineering. Freeze drying is one of the most common and widely used processes
for producing collagen scaffolds, in which acetic acid-based collagen suspensions
are made and collagen slurries are allowed to develop prior to the freeze-drying
procedure [54]. Another method included mineralization of collagen fibrils. This
method included precipitation of collagen and their mineralization using amorphous
calcium. These fibrils are then subjected to freeze drying to produce scaffolds.
Type I collagen is known to be an important component of bone mineraliza-
tion since it is known to be secreted by osteoblasts during ossification. As a result,
when collagen scaffolding is used in vivo, improved physiological integration with
the surrounding tissue is possible. Several commercial products used in bone tissue
Green Methods for the Development of Bone and Tissue … 79

Fig. 1 Development of Scaffolds from Collagen

engineering are based on type I collagen. Collagen/cellulose scaffolds were shown

to support cellular attachment and the phenotypic preservation of cultivated human
osteoblasts in vitro. High levels of alkaline phosphatase (ALP), an osteogenic
enzyme, were also observed [55, 56]. Development of scaffold from collagen is
shown in flowchart in Fig. 1.

3.2 Chitosan

After cellulose, chitosan is the most abundant biopolymer. It is frequently found

in invertebrates as shells or cuticles in crustaceans or insects. It is also found in
some yeast and algae cell walls [56]. It is a deacetylated derivative from chitin. It
is a polysaccharide that is made up of N-glucosamine and N-acetyl glucosamine
(GlcNAc). It has a deacetylation degree of 30–95% and a molecular weight range of
300–1000 kDa [57, 58]. Chitosan is undissolved in aqueous solutions above the pH of
7. Nonetheless, free amino groups become protonated in weak acids, and the molecule
becomes completely soluble below a pH of 5. Chitosan is a promising biomaterial in
tissue engineering due to its antimicrobial properties and induced minimal foreign
body response. Another important reason for using chitosan in tissue engineering
is its structural similarity to GAGs, as it contains glucosamine, a key component
of ECM. One of GAG’s characteristics is that it has several specialized interactions
with growth factors, receptors, and adhesion proteins. Chitosan’s similar structure
may also have comparable bioactivities. Most importantly, chitosan is cationic in
nature, which aids in the retention or accumulation of molecules due to interac-
tions with anionic GAGs and proteoglycans [59]. Chitosan is also appealing as a
80 A. Hazra et al.

material for bone scaffolds because it promotes osteoblast cell adhesion and prolif-
eration as well as the development of mineralized bone matrix. Because chitosan
is a polymer composed of amino acids and polysaccharides, it contains breakable
glycosidic bonds. In vivochitosan degradation is typically carried out by the enzyme
lysozyme [60].
Chitosan has been combined with a variety of delivery components, including algi-
nate, hydroxyapatite, hyaluronic acid, calcium phosphate, PMMA, poly-l-lactic acid
(PLLA), and growth factors, for potential use in orthopedics. Chitosan, in general,
offers a wide range of options for cell-based tissue creation. Chitosan can be made
into 3D highly interconnected, porous structures with a high degree of porosity using
a variety of technologies, allowing cells to proliferate and move nutrients. Porous
scaffolds, for example, can be created by lyophilizing a frozen chitosan solution in
acetic acid [61]. Chitosan and acetic acid may phase split and form ice crystals when
frozen. The scaffold develops holes as a result of the sublimation of acetic acid ice
crystals during the freeze-drying process. Another method is to use porogens (salts,
sugar) in conjunction with chitosan in a melt-based technique [62].
Rapid prototyping technology can also be used to create chitosan scaffolds. This
is a computer-aided method [63]. The use of chitosan-based scaffolds that mimic
some bone characteristics can improve osteoblast adhesion and proliferation. For
example, chitosan scaffolds combined with bioactive ceramics can impart osteo-
conductive and even osteo-inductive properties to the final structure, directing bone
development engineering [64]. Chitosan, in combination with hydroxyapatite and
tricalcium phosphates, has shown promising results in mesenchymal stem cell (MSC)
adhesion, proliferation, and differentiation, as well as intriguing results in bone tissue
engineering [65]. Chitosan has been used to improve the biological and mechanical
properties of marine-derived polymers in order to provide an alternative to collagen
derived from animal sources. When combined with polyesters, chitosan increased cell
viability and adhesion [62]. Development of scaffolds from chitosan is in flowchart
shown in Fig. 2.

3.3 Gelatin

Gelatin is a naturally occurring biopolymer created when animal collagen from skin,
bones, and tendons is partially hydrolyzed in an acid or alkaline solution, and the
structure is very similar to collagen. It is comprised of various proteins and amino
acids. Regardless of how collagen is hydrolyzed to create gelatin, both polymers
include up to 20 distinct amino acids in varying amounts in their basic structure.
This basic structure provides a three-amino-acid detection sequence for integrin-
mediated adhesion of cells. The properties of gelatin are determined by the source
of collagen, the type of collagen, and the method of collagen conversion to gelatin.
Various methods are implemented for the creation of gelatin scaffolds. One such
process includes electrospinning. The production of polyelectrolyte colloidal sol
makes it very challenging to electro-spun gelatin using water as the solvent.
Green Methods for the Development of Bone and Tissue … 81

Fig. 2 Development of
Scaffolds from Chitosan

The silky gelatin nanofibers were created using a variety of polar organic solvents,
including trifluoroethanol and hexafluoro isopropanol [66]. Biomaterials made of
gelatin are widely used in tissue engineering. Gelatin scaffolds demonstrated high
DNA, GAG, and collagen contents and enabled the development of chondrogenic
cells. Gelatin used with chitosan was used in skin tissue engineering [67]. Much
interest has been shown in electro-spun nanofibrous scaffolds as prospective plat-
forms for bone repair and regeneration. Degradable polymers, whether they are
synthetic or natural, were used as a starting point for research into the advanta-
geous properties of nanofibrous matrices. Many synthetic polymers’ hydrophobic
properties restrict cell affinities and early cell adhesion. Another method to increase
hydrophilicity and, consequently, cell compatibility in polymers is to combine
synthetic and natural polymers. This is done using a composite of polycaprolactone-
gelatin scaffolds. Gelatin-apatite composite was also used since it mimicked the
properties of bone ECM [68].

3.4 Silk Fibroin

Silk fibroin has been extensively researched for tissue engineering applications due
to its superior biological and mechanical strength, as well as biocompatibility and
biodegradability.
Silk is made up of fibrous protein (SF) and sericin. Silk is produced by a variety
of arthropods, including spiders and silkworms with the most used silk produced
by the silkworm Bombyx mori. SF has been shown to be very biocompatible. After
observing its attachment to fibroblast cells and growth on SF matrices, its role as a
biomaterial was evaluated. Silk fibroin can be easily separated from sericin by using
a steaming alkaline or surfacting solution (degumming). The resulting fibers have a
82 A. Hazra et al.

Fig. 3 Development of Scaffolds from Silk Fibroin

diameter of 10–25 mm and are composed of a hydrophobic heavy chain (350 kDa)
and a hydrophilic light chain (25 kDa).
Depending on the type of scaffold required, silk fibroin can be created using
a variety of techniques. One such technique is salt leaching using NaCl particles
for scaffolds with spherical pores and for scaffolds with lamellar pores a process
involving freezing, and lyophilizing was used which resulted in the formation of beta
sheets. Another technique included electrospinning which produced fibers with low
tensile strength [69]. Silk fibroin is a very useful biomaterial in tissue engineering.
By making the fibers stiffer, it was seen to increase osteogenic induction. Silk used
along with calcium phosphate in engineering of bone showed increased stability, and
bioactivity with no toxicity. An osteo-inductive composite was created by combining
silk and HAp, the important mineral seen in bone, to replicate the composition of
bone. HAp improved the scaffolds’ compressive strength as well as the ability of
cells to proliferate and produce bone. Other biofunctional substances for restoration
or therapy of tissues of bone are titanium dioxide (TiO2 ) nanoparticle and natural
silk fibroin-based nanocomposite scaffolds [70, 71]. The development of scaffolds
from silk fibroin in flowchart is shown in Fig. 3.

3.5 Starch

Starch is a polysaccharide that plants and algae produce as an energy-storing

compound in a variety of granules. Amylose and amylopectin are the two subunits of
starch. Native starch includes waxy starch, which contains little amylose, regular
Green Methods for the Development of Bone and Tissue … 83

starch, which contains 15–30% amylose, and higher content of amylose, which
contains more than 50% amylose. Crystallinity can range from 15% in high-amylose
starch to 50% in waxy starch [72]. Starch’s brittleness prevents it from being used in
most situations. As a result, starch must be modified or combined with other polymers
to improve its properties.
Starch scaffolds can be produced by traditional casting in solvent procedures. In
this procedure, the starch is dissolved in chloroform and cast into a mold. Cellu-
lose and starch were combined by leaching with salt and freeze-drying methods
in a different approach. Wet spinning method is also used to create scaffolds of
desired pore size which include mixing of starch with other polymers. Chitosan-starch
microparticles can be obtained from water in oil emulsification techniques. Starch
can also be used along with HAp and chitosan which is processed by co-precipitation
[73]. Several studies on bone tissue engineering have used starch-based scaffolds.
When inserted into a critical-sized defect, scaffolds made of polycaprolactone, starch,
and stem cells boosted the repair of bone. In comparison to scaffolds loaded with
no differentiated cells, scaffolds loaded with osteoblastic cells produced more new
bone. Modified starch scaffolds and polycaprolactone with silanol group showed
improved osteogenic differentiation and bone healing. 3D scaffolds promoted adhe-
sion and recruitment of cells and promoted tissue regeneration [74]. The development
of scaffolds from starch in Flowchart is shown in Fig. 4.

Fig. 4 Development of Scaffolds from Starch

84 A. Hazra et al.

4 Advantages and Disadvantages of Green Methods

The goal of numerous studies is to create techniques that are cost-effective, low-
energy, or sustainable to produce various tissues while decreasing the consumption
of harmful ingredients. One of the safest ways to create tissues and organs is to apply
natural substances taken from plants or animal bones [75]. Synthetic materials exhibit
physicochemical properties that may be regulated during the production process.
They are also robust, affordable, dependable, frequently easily electro spinnable,
and strong. Their lack of cell-recognition sites, however, results in a low affinity for
cell attachment.
Thus, natural polymers are favored because they resemble the macromolecular
components of the human body [76]. The main benefits of bio-based and biodegrad-
able polymers include reducing GHG emissions, utilizing local resources, reusing
by products, and preserving fossil fuels. Without any prior processing, biopolymers
print beautifully and have good scent and fat barriers. Polymers made of starch offer
excellent surface finishing and antistatic characteristics. Naturally occurring biopoly-
mers (collagen, chitosan, starch, silk) have a better role in cell adhesion, proliferation,
and less cytotoxicity, hence are better materials for scaffold preparation for tissue
engineering. Polymers made from natural materials outperform semi-synthetic or
synthetic polymers in replicating the ECM and interacting with tissues, owing to
their high resemblance to the tissue environment [77].
Despite having the above said advantages, natural polymers do have some disad-
vantages, such as often high production costs (collagen, hyaluronic acid). Its complex
structural and chemical makeup, as well as macromolecular complexity, architec-
ture and shape, unpredictable hydration rate, resource limitations, and potential for
microbial deterioration are further disadvantages that may limit their use in tissue
engineering. Furthermore, because of their low stability, the rate of disintegration
and catabolization is high of some spontaneously created scaffolds is higher than the
pace of regeneration of the host tissue. Certain natural polymers, like cellulose and
chitosan, have limited mechanical characteristics and some have poor processability
e.g., polypeptides. Synthetic biopolymers have a few advantages over natural poly-
mers, such as a tunable and engineerable hydrophilic/hydrophobic ratio, a faster rate
of breakdown, and better mechanical properties. Another disadvantage of natural
biopolymers is degradation, so it can’t be used for a long period of time [78].

5 Challenges and Future Scope for Green Methods

Though there has been overwhelming progress in the field of green bioengineering,
some constraints have always been present. The acquisition of the natural sources and
its behavior has always been a point of criticality. Current focus is being shifted to
use perennial plants as source material so that the restriction due to seasonal changes
can be avoided [79]. This would be helpful as the production would not be limited
Green Methods for the Development of Bone and Tissue … 85

by time frames. Also, plant sources are being tried to be extracted from plants which
have broader availability, that it is found in multiple regions to make the production
uniform [80, 81]. Also, to make the environment a safer place and reduce economical
stress, waste utilization has cropped up as a concept over the years, where agricultural
wastes are being worked upon to lead to the conformation of different scaffolding
frameworks [82].
Also, currently most production processes require high energy, which differs
depending on the source material that would be extracted. One of the current aims
is to develop methodologies or extract forms that can be molded at less than 100 °C
[83, 84]. Despite a lot of challenges in the field, the major challenges that need to be
combated include, developing the natural polymer in a way that it imitates all struc-
tural, functional, and morphological properties of the said tissue, and can also show
protein adsorption as well as inter- and intra-cellular interactions. Also, they must be
suitable for drug delivery as well as having a window for chemical and biomechanical
modifications [85, 86]. Thus, though the research in the field has made remarkable
discoveries with respect to the use of natural polymers, they are now trying to make
it more feasible and extraction and development to be easier as well as affordable
[87].

6 Conclusion

Current research in this field has dealt with the extraction, development, and modifica-
tion of natural polymers to produce particulars with concerns to bone and engineering
of tissue. The use of polysaccharides and hydrogels has given a new dimension to
discovery in this field and has contributed toward reducing environmental damage
and quite a bit of the economic load. These new compositions have been seen to have
better congregation with human tissues and thus can work toward the replacement of
faulty ones. The progression of them into organ transplantation remains to be envis-
aged into. The multiple methodologies that have been developed for regeneration are
still left to be optimally standardized and its reach to be increased. The novel strate-
gies and newly constructed biomaterials have played a pivotal role in turning engi-
neered templates for tissue replacement, into fully functional independent systems.
They possess all characteristics including biocompatibility, conductivity of impulses,
cellular interactions, and integration with host’s native cells. Further research aims
at investing the natural polymers into other complicated surgeries and formulations
of degradable, injectable, and mechanically stronger components for restoration of
functionality of various organs.
86 A. Hazra et al.

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A. Hazra is a postgraduate student under Dr. Kanthesh BM

an Associate Professor and Coordinator Division of Molecular
Biology, in JSS Academy of Higher Education & Research. She
is currently pursuing her master’s in Molecular Biology. Before
joining JSS, she was in St. Xavier’s College, Mumbai for her
undergraduate where she got a double major in Microbiology
and Biochemistry. She has also worked as a research trainee
at ACTREC, Tata Memorial Centre in Bose Lab, an Integrated
Biophysics and Structural Biology Lab. After completing her
postgraduate studies, Avipsa wants to pursue Ph.D. and continue
with research in the field of Genetics and Cancer. She is a
research-oriented individual who is always looking to expand
her reach of science.

G. Baradwaj is a postgraduate student under Dr. Kanthesh BM,

an Associate Professor and Coordinator of Division of Molec-
ular biology, in JSS Academy of Higher Education & Research.
He is currently pursuing his master’s in Molecular biology. He
has completed his undergraduate in NITTE University Centre
for Science Education and Research, Mangaluru in Biomed-
ical Science Hons. After completion of postgraduate studies,
Gowrav wants to pursue his Ph.D. and continue with research
in the field of Stem cell research and cancer.
Green Methods for the Development of Bone and Tissue … 91

A. S. Dhanu is a Research Scholar under Dr. Kanthesh M

Basalingappa, an Associate Professor and course coordinator at
Division of Molecular Biology, School of Life Sciences, JSS
Academy of Higher Education & Research-Mysore. Her current
research interest is oriented towards Traditional medicine in
Triple negative Breast cancer. In the year 2020, she graduated
from JSS College of Arts, Commerce, and Science—Mysore
with a UG degree in Biochemistry, Microbiology, and Biotech-
nology. In the year 2022, she was awarded a postgraduate
degree, an [Link]. in Biotechnology from Shivagangotri, Davan-
gere University—Davangere. The author is a gold medalist,
holds the first rank from Davangere University, Shivagangotri,
Karnataka, in the academic year 2022. The author is currently
pursuing her Ph.D. at Division of Molecular Biology, School of
Life Sciences, JSS Academy of Higher Education & Research—
Mysore. She is a young Molecular Biology researcher with a
passion for invention and innovation.

G. Kuppannan is a Reader of Microbiology at the Noorul

Islam College of Dental Sciences in Aralumoodu, Thiruvan-
thapuram, India, which is affiliated with the Kerala Univer-
sity of Health Sciences in Thrissur. He is focusing his research
interests in the fields of microbiology, bacteriology, virology,
stem cell research, and innovative medication creation using
natural resources. He is now working on an environmental and
soil microbiology as main research project that will lead to an
epidemiological investigation of organic and inorganic fertilisers
used in diverse crop cultivation soil samples. With the current
research’s integrated inquiry seeking to create the bioconsortium
utilising a substantial strains for all crop sorts. Dr. Gobianand
Kuppannan earned his doctorate from the University of Madras
in 2007, and then worked as a Senior Researcher in the Depart-
ment of Medical Microbiology at the University of Malaya in
Kuala Lumpur, Malaysia (2007–2009), before being awarded
as a Researcher by the National Institute of Animal Sciences,
Central Government of South Korea (2009–2012). He joined
the Department of Microbiology, K. S. Rangasamy College of
Arts and Science, Tiruchengode, as an Assistant Professor with
a research [Link] awarded as Senior Researcher by the
Malaysian Agricultural Research and Development Institute in
connection Universiti Malaya (2007). In 2009, He was awarded
as Post-Doctoral research fellow by the Central Government of
South Korea. He has been devoted to academics and research
for the last 20 years, and he has produced 23 research articles
and two review papers during his career. He is a Life Member
of the Indian Association of Applied Microbiology (IAAM), the
Indian Association of Biomedical Scientists (IABMS), and the
Association of Microbiologists of India (AMI). He contributes
as an editor, technical editor, associate editor, and reviewer for
a number of prestigious journals for its publications.
92 A. Hazra et al.

M. Arthanari is serving as a Head and Assistant Professor,

Department of Microbiology, Vivekanandha College of Arts
and Sciences for Women (Autonomous), Elayampalayam,
Tiruchengode, Namakkal (DT), Tamilnadu, India, affiliated
under Peiyar University, Salem. Her field of research is mainly
bacteriology and virology. Currently her research focuses on
gut microbiome associated with diabetes and its complications,
mood swinging and behavioral changes. She is mainly working
on biodegradation of crude oil spills on marine environment,
also she studied phytochemical compounds extracted from
macroalgae and worked on their applications. She is aiming her
research interest that focusing on environmental degradation of
oil spills and concentrating to elucidate, identify and elucidate
the bioactive compounds from macroalgal. Dr. Malarvizhi
Arthanari rearmed her doctorate from Bharathiyar University
in the year of 2014. She is serving as active teaching faculty
in Vivekanandha College of Arts and Sciences for women
(Autonomous), Elayampalayam, Tiruchengode, Namakkal
(DT), Tamilnadu for more than 16 years of experience. She is
actively involved in academic responsibilities, and she is serving
as Board of Studies Member in reputed Universities/Higher
Education Institutes. She has published 22 research/review
manuscripts in reputed journals. She is actively connecting and
occupied with the Scientific associations/bodies with her allo-
cated responsibilities with very sincere and punctual. She has
the role in her present institution as Mentor and Coordinator for
various activities in which she is regularly monitoring, serving,
and recording to the respective responsibilities to bring it to
the maximum height. She is supervising two scholars for their
Doctoral Degree, registered with Periyar University.

B. M. Kanthesh is an Associate Professor of Molecular Biology

at the School of Life Sciences, JSS Academy of Higher Educa-
tion and Research, Mysuru, India. His goal of research is to
determine the role of RNA Binding proteins in tumor progres-
sion and metastasis. Post-transcriptional regulation of gene
expression by RNA binding protein is a crucial mechanism in
regulating the timing and the amount of expression of genes.
Growing evidence indicate that the alteration of the expression
and function of RNA binding proteins could potentially play a
role in inflammation and cancer. Dr. Kanthesh BM did is Ph.D.
from University of Madras (2005), He also did postdoctoral
research at the University Malaya, Kuala Lumpur, Malaysia
(2007–2009); West Virginia University, Morgantown, USA
(2090–2011) and University of Oklahoma Health Sciences,
Oklahoma, USA (2011-2014). He received Malaysia Presti-
gious Bio-Malaysia Gold medal Award (2008). For his research
area is Arbovirus infections, in that they done patented work
on “Early detection of BK virus using molecular methods”. He
also Received Dr. Wilson Aruni “Best Research Mentor and
Teacher Gold medal Award” from the Indian Association of
Applied Microbiology (IAAM) (2018). He has been engaged in
teaching and research in Microbiology and Molecular Biology
Green Methods for the Development of Bone and Tissue … 93

for the past 22 years. He has published over 95 original research

papers, 15 book chapters, and 15 review articles. He is also
Professional and Scientific Memberships in, American Asso-
ciation for Cancer Research (AACR), Life Member of Indian
Association of Applied Microbiology (IAAM), Life Member of
Indian Association of Biomedical Scientists (IABMS), Indian
Association of Medical Microbiologist (IAMM). He Received
Fellowship Award from Indian Association of Applied Micro-
biology (FIAAM). At present he is a having collaboration with
Royal Research Foundation, a research institute in India.
Genetically Induced Biomaterial
Advances in Medical Sciences

Eva Kaushik and Rohit Kaushik

Abstract Protein-based monomers are among the most alluring options for
achieving improved results with cutting-edge biomaterials since recent advances
in bioartificial and microbiology technologies enable the very complicated, accurate
design, and fabrication of protein-based biomaterials. These sequences are easily
enhanced with bioactive motifs that improve their functionalities, material-host inter-
actions, and basic biological requirements since their sequences are built from archi-
tectural protein-based modules. These polypeptides have gained increasing interest
for usage in biomedical activities such as cell cultures, bioengineering, separation,
and purification design is vital, as well as controlled drug administration because of
their versatility, self-assembly behavior, cued, and biocompatibility. The biopolymers
discussed in this piece are biomimetic materials and genetic algorithms created from
elastin-derived proteins. These genetically programmed polymers’ design, manufac-
turing, and characterization as well as their potential uses in regenerative medicine,
bioinformatics, targeted medication delivery, and stem cell therapy will also be
covered.

Keywords Extracellular matrix · Tissue engineering · Regenerative medicine ·

Stem cell · Biomaterial · Protein engineering

1 Introduction

Medical science, biology, chemistry, materials science, and engineering are all inter-
related fields in the study of biomaterials. It has been said that “biopolymer is utilized
to create systems that substitute an element or a function that the body performs
in a secure, dependable, cost-effective, and physiologically reasonable way.” An

E. Kaushik (B)
Information Technology and Engineering Department, Dr. Akhilesh Das Gupta Institute of
Technology & Management, New Delhi, India
e-mail: Kaushikeva0026@[Link]
R. Kaushik
CS and Mathematics Department, Data Analytics Program, University of Illinois, Illinois, USA

© The Author(s), under exclusive license to Springer Nature Singapore Pte Ltd. 2023 95
R. Malviya and S. Sundram (eds.), Engineered Biomaterials, Engineering Materials,
[Link]
96 E. Kaushik and R. Kaushik

unworkable resource utilized in a pharmaceutical drug that is meant to engage with

biochemical processes is a biological material. Biomaterials are widely employed in
medicine, and some are even used in medical equipment. Dental implants, artificial
hip joints, intraocular lenses, replacement heart valves, vascular grafts, kidney dial-
ysis, and heart–lung machines are only a few examples of medical technology. All
three types of materials—metals, ceramics, and polymers—are utilized as biomate-
rials [1]. Metallic nanomaterials must be utilized in load-bearing applications and
have enough fatigue strength to withstand regular activities such as walking and
chewing to be employed in implant placement and prosthetic hip joints. Ceramic
biomaterials are often used for applications like joint surfaces in bones and teeth and
bone adhesive interface in implants due to their wear resistance and hardness. Poly-
meric materials, which are frequently used for their suppleness and stability, have
also been used to generate low-friction articulated surfaces. Engineers must have a
thorough understanding of biomaterials since they must choose their materials and
create their designs in accordance with the needs of the biomaterials. The choice
of material and design is crucial because either one gone wrong could result in a
person’s demise [2]. Metals are used in load-bearing applications, but because most
metals react with water and the human body contains a lot of water, when metals
interact with human tissue, they form oxide layers. Ceramic coating is used on the
surface of metals to prevent this because ceramic does not interact with body cells.
Medical device design utilizing biomaterials is extremely constrained and requires
great accuracy and precision because the design must fit with the following body
parts and engage with human body parts to carry out the desired operation. As a
result, without knowledge of biomaterials, engineers who create or build biomate-
rial devices are utterly helpless. In the field of biomedicine, novel nanotechnology
with cutting-edge and complex capabilities has found specific applications and can
be particularly significant in fields like controlled drug delivery tissue repair and
bioengineering in general. Classes of biomaterials are shown in Fig. 1.
For specialized use as a carrier for the dispersal of bioactive chemicals in
controlled medication delivery, new biomaterials are required. Only a small portion of
treatments administered really reach the sites for which they were intended, making
improving the effectiveness of chemotherapy medications a key area of research.
Standard chemotherapeutic drugs’ negative impacts on cells beyond the target cells
limit their efficacy [3, 4]. Because they are created to lessen the noxious effect and
enhance the solubility, consistency, pharmacokinetics, and pharmacodynamics of
conventional pharmaceuticals, these novel types of drugs can therefore offer signif-
icant benefits about the toxic metabolites and degenerative diseases brought on by
the distribution of the free drug. By responding to specific cues, efficient pharmaceu-
tical systems are developed to control the dosage of therapies and their associated
side effects. This reduces the dispersion of medications into target tissues. Innova-
tive biomaterials with the ability to organize supramolecular at higher orders and
self-assemble in a manner that is bio-responsive will therefore be required. Physical
aid to the healing process, proactive engagement with biochemical processes, and
preservation or enhancement of tissue function are all expected from these tools. The
innovation of these systems frequently coincides with an improved understanding
Genetically Induced Biomaterial Advances in Medical Sciences 97

Fig. 1 Biomaterials classification

of the mechanisms dictating the body’s systems and advances in genomic science,
allowing the use of refined methods that take inspiration from the natural world to
mimic the mechanical and biological attributes of the tumor site while ensuring secu-
rity. Depending on the purpose, biomedical technologies for muscle restoration and
medication development could be produced or natural, degradable, or not. Protein-
based galactose, which was inspired by naturally occurring proteins discovered in the
ECM components, stands out as a leading candidate for use as essential components
of cutting-edge medical technology and improved replacements in tissue regenera-
tion because their characteristics are determined by the physicochemical character-
istics and loop of constituent monomers. Genetic manipulation still offers significant
control over the component organic chemicals used to make delivery, transportation,
and skin substitutes, despite the significant advances made in the field of polyelec-
trolyte materials in terms of advancements in the effectiveness of polymeric materials
research methods and lower polydispersity’s [5]. According to sequence-structure
interactions, the length and concentration of genetically created drug carriers affect
a variety of macro, micro, and nanoscale characteristics. Protein polymers made
of elastin-like polypeptides are used as well-developed genetic engineering drug
carriers as well as personalized supports for tissue restoration. Polymers known as
elastin-like peptides (ELPs) have brief pentapeptide repetitions that mimic those in
real elastin. They are influenced biologically. The most frequent recurrence of any of
98 E. Kaushik and R. Kaushik

these amino acid residues is VPGXG, where X can be any other important ingredient
than L-proline. The term “elastin-like recombinases” (ELRs), a recent invention, is
already in use to describe genetically modified ELPs. With absolute control over
the sequence of amino acids and the possibility to include a range of functions and
bioactive sequences, ELRs can be made utilizing DNA technology. Because of their
biological and thermal properties, such as biomaterials, degradability, and behavior
that is responsive to temperatures and environment, ELRs, which are microbiologi-
cally biomaterials, present an appealing alternative for the development of innovative
biomedical devices [6, 7]. ELRs also can self-assemble sterically over a temperature
slowly referred to as the low ambient temperatures (Tt), which causes a conforma-
tional reorganization at the cellular scale and the reversible development of steric
effects. The inverse thermal transformation (ITT) of ELRs is the name given to
this characteristic. Numerous elements, such as variations in temperature, pH, light,
ionic imbalances, etc., can worsen the polymer backbone. Furthermore, because it
is controlled by the molecular size and make-up of the recombinant, the activity
that responds to stimuli can be altered. They are ideal materials for the majority of
materials technology and bioscience applications, particularly for controlled tissue
engineering and drug delivery because of their metabolic behavior, flexible mechan-
ical characteristics, heat sensitivity, and capacity for self-assembly. In this study,
we explore the application of ELRs as cutting-edge drug delivery systems to safely
deliver therapeutic drugs to targeted areas. ELRs’ diverse topologies, sequencing,
and assembly enable the creation of specialized pharmacological devices such as
glucose molecules, nanoparticles, and three-dimensional assemblages. It will also
be discussed how monomeric ELRs can be used as aqueous delivery methods for
certain peptides that act as knh or cell-penetrating sections. Such systems are suit-
able for systemic distribution since they are designed to self-assemble in response to
environmental cues. They are very beneficial in tumor tissue therapy because of their
thermoresponsive activity. ELRs can self-assemble similarly to how they can produce
nanoparticles (NPs). Thus, the usage of bases and hydrogels as a 3D framework for
drug-delivery devices as well as the applications of NPs as drug nanomaterials will
be discussed [8]. Utilization of NPs as drug solid lipid nanoparticles when deliv-
ering therapeutic approaches, proteins, or signaling molecules to the skin’s surface,
or those arrangements that can internalize the medication, bioactive constituents,
or even DNA to maximize absorption and persistence effects. Problems with inad-
equate drug absorption loading can be solved by building macroscopic scaffolds
using chemical and physical ELR chain bridging, which also enables controllable
localization of the reactive drug release region and rate.

1.1 Regenerative Medicine and Tissue Engineering

Tissue engineered and tissue regeneration are very beneficial methods for the preser-
vation, repair, or enhancement of functional body materials or complete systems
after they have exhausted their viability. The development of support systems that
Genetically Induced Biomaterial Advances in Medical Sciences 99

are “allowed to exist” when applied directly to injured tissues but are unable to
recreate on their own has progressed in recent years in the field of nanomaterials
for tissue regeneration. Systems that can interact with the organism and reproduce
biological environments are being created using contemporary technology. To build
functional tissues, these diverse fields of nanomaterials research are progressively
combining the use of cutting-edge scaffolds, the right cells, and bioactive chemi-
cals [9, 10]. For the construction of a functional scaffold, a few components have
been identified as being crucial, including the following: Factors include the needed
three-dimensional graphene, its porosity, its unique material performance, its water
sorption and permeable, epithelial connection via chemical signals, its hydrophilicity
and permeation, and its length of residence before host tissue remodeling.
Elastin-like scaffolds have been created with the goal to mimic some of the char-
acteristics of the ECM, including the architectural surface features, appropriate firm-
ness and porosity, improvement of ligand binding and viability, sensitivity to cell
signaling, processability, and cued to variations in their environment, among others.
The skeletal system, the vascular system, as well as the rejuvenation of soft tissues
are among the areas of the body where elastin has been designed to obtain specialized
support. These methods are covered in this study. The following discussion examines
a few recent cases.

1.2 Cardiovascular System Regeneration

One of the main challenges in tissue healing is managing vascularization. In this

context, artificial tissue-engineered scaffolds must allow for appropriate oxygen and
nutrient transport as well as the expulsion of catabolic waste. Therefore, it is essen-
tial that the scaffold develop an effective network of capillaries and blood vessels.
ELR Hydrogels offer a viscous, bioactive substance that is both biocompatible and
bioactive despite the wide variety of polymers that are now available. Human adipose
tissue-derived SVF cells are transplanted in a variety of ELR hyaluronan, from non-
functionalized composite (NF-hydrogels) to a dendrimer gelatin incorporating two
cell adherence integrin-mediated motifs, to promote in vitro vascularization (REDV is
specific for endothelial cells). The SEM photos of a non-functional system revealed
that there was little cell attachment on the flat, elongated parallel granules of the
RGD/REDV agarose. The anti-systems had poor cell adherence, while the RGD/
REDV polymers had flat, elongated symmetry cells that were well-organized [11].
Moreover, endothelial cells gathered in the RGD/REDV hydrogel but not in the
non-functionalized hydrogel, according to CD-31 labeling. Notably, cells contained
in NF-hydrogels produced 79% more anti-angiogenic factors than proangiogenic
factors, including FGF or IL-8, which were each up 21%. One of these molecules
that prevent angiogenesis is vasohibin, along with IGFBP-3, prolactin, depositors,
and others. One of the more powerful proangiogenic agents, VEGF, was studied for its
release pattern. It of released, and it was found that while it was maintained within the
polymeric and was not liberated in the case of NF hyaluronic acid, it was liberated into
100 E. Kaushik and R. Kaushik

the precipitation of RGD/REDV hydrogels. RGD/REDV hydrogels sustained blood

vessel penetration and were penetrable by host vascularization, according to an in vivo
experiment performed on naked rats. NF-hydrogels also showed a decrease in host
cell penetration in conjunction with not displaying any additional vascularization. As
a result of the VGVAPG sequence’s ability to attract monocytes and its vulnerability
to proteolysis, RGD/RED hydrogels break down, allowing fibroblasts and endothe-
lial cells to populate the matrix. Even though vascular repair is challenging, some
research groups are trying to speed up the procedure by stimulating small-caliber
vascular regeneration. ELR-based scaffolds were implanted in the rat’s abdomen. To
investigate the healing of blood vascular tissue, an ELR-RGD was selected. After
the temperature approaches 12 °C, the VPGIG-based hydrophobic sequence of the
ELR begins to assemble into nanomaterials up to 10 m in diameter [12, 13].
In vitro, studies showed that HUVEC adhesion to the scaffold was boosted by the
RGD motif, not just the ELR by itself. In the in vivo testing, rats that got the ELR-
RGD scaffold showed evidence of arterial tissue regeneration, but rats that obtained
an ELR scaffold did not. One of the most frequent negative effects of many illnesses is
intracranial hemorrhage (ICH), which results in brain edema and harms brain tissue.
It is true that patients with ICH have high rates of morbidity and mortality. Cardiac
tissue engineering (CTE) and their transplantation is shown in Table 1.
To address the severe symptoms of ICH, it is crucial to create both novel thera-
peutic approaches and new materials. In this instance, an ELR with RGD has been
recommended to manage intracerebral hemorrhage while it is still acute. The effec-
tiveness of the medication was evaluated in vivo tests by measuring the thrombus
diameter and the inflammatory response. 6 h after the intracranial hemorrhage, the
edema mass was lower in the group that received RGD-ELR as opposed to those
getting RGD alone [14]. Moreover, it protected the damaged blood vessels to stop the
subsequent loss of blood components, especially IgG. Von Willebrand factor (vWF),

Table 1 Cardiac tissue engineering (CTE) and their transplantation

Cells CTE strategy Scaffold Transplantation
Fetal rat CM Classical Gelatine mesh Avascular graft
Hesc-cm + human Classical Poly-1-lactic acid + Vascularized graft
umbilical vein EC + Matrigel
embryonic fibroblasts
Neonatal rat CM EHT Collagen type 1 from Avascular graft
rat tails + Matrigel
Neonatal rat CM + Myocardial micro and Cell generated Vascularized graft
human umbilical vein macro tissue
EC
Neonatal rat CM + rat Decellularized heart Decellularized rat heart Organ
aortic EC scaffold
Fetal rat CM Classical 3D porous alginate Avascular graft
Neonatal rat CM In vivo flow through Fibrin Gel Flap
pedicle chamber
Genetically Induced Biomaterial Advances in Medical Sciences 101

a vital part of the coagulation system required for platelet adherence to collagen,
was detected using immunohistochemistry testing. Additionally, the immunological
response brought on by post-ICH activated microglia was dramatically diminished
by the RGD-containing ELR therapy, as well as vWF overexpression.

1.3 Skeletal System

These substances constitute a significant portion of the bone and ligaments that
make up the bones and joints, of the body’s skeleton. Upon injury or in response
to ongoing modifying stimuli, the mature bone does have the innate capacity to
repair. Despite this ability to regenerate, some conditions or diseases that affect
how efficiently the body can heal itself, such as inflammation brought on by the
severity of the injury or oncogenic or pathologic bone problems, may impede the
process. Tissue-engineered supports are necessary for these circumstances because
of their osteogenic, osteoinduction, and osteogenic components, which provide struc-
tural functioning and promote bone healing. Due to their amphiphilic characteristics,
which enable them to attach to thermo-responsive hydrogels in physiological circum-
stances, two distinct ELRs have been produced. The domains from self-aware fibrin
typically contain the Arg-Gly-Asp (RGD) themes in addition to the osteoblastic and
osteogenic bone technologist and technology protein-2 (BMP-2) motif. Elastase-
sensitive action scenes are included in the segments of these ELRs to investigate the
rate of degradation of the ELR molecules. While the RGD sequencing helped host
cells adhere to the substrate and increase cell survival, the BMP-2 component encour-
aged osteoblast development. The deterioration of the artificial scaffold’s organic
composition occurred concurrently with bone regeneration, which was used in this
work to show how host bone remodeling affects how rapidly the scaffold dissolves.
In vivo experiments on female Zealand white bunnies revealed hematologic marrow
and fibroblast layer infiltration and multiplication, suggesting a large potential for
bone graft recovery and outstanding biocompatibility. The V40C2 ELR block, a
pentapeptide made of valine and disulfide, was previously combined with human
recombinant BMP-2 to increase the osteoblastic and osteoinductive capabilities of
the ELR scaffold. At 37 °C, this ELR block is identified [15, 16]. The main goal
was to maintain the bioactivity of BMP-2 in the fusion protein. It was demonstrated
that despite the protein’s diminished biocompatibility, however, neither ELR gene
product nor the commercially available pure BMP-2 lost any of their cytocompat-
ibility. BMP-V40C2 showed stronger calcium deposition than the control during a
21-day treatment regimen, as well as increased mRNA concentrations of the tran-
scription factors Osterix (OSX), which is only expressed in osteoblasts. The produc-
tion of artificial bone tissue utilizing hydrogel hydroxyapatite (Si-HA) scaffolding
and ELRs is an additional intriguing method. Due to its osteoblasts, osteogenic, and
accessibility, which encourage bone development and osseointegration, HA—a vital
element of vertebrate bone and teeth—is widely utilized in orthopedic, dental, and
cranial operations. The 3D degradable scaffolds employed in this investigation were
102 E. Kaushik and R. Kaushik

coated with two bioactive ELRs that were produced and analyzed. It has been demon-
strated that these ELRs mediate biochemical mechanisms at the interface of minerals
and have a strong affinity for calcium phosphorus, much like HA. The production
of the osteoblast markers OC and ALP, however, showed that the Si-HA-SNA15-
containing scaffold was particularly effective at directing the BMSCs toward the
osteoblastic lineage when osteoblast differentiation was assessed [17]. All studied
scaffolds allowed cells to colonize and proliferate, but the functionalized ELR gener-
ated a biochemical pathway that markedly enhanced the scaffolds’ ability to stimulate
osteoconduction. Unlike bone, some tissues, like enamel, cannot regenerate. An ELR,
a tiny protein that stimulates enamel remineralization, contains statherin. In contrast
to the untreated substrates, the resultant STNA15-ELR formed mineral platelets after
8 days of growth in a mineralization solution at 37 °C. ELRs may therefore be crucial
for preserving the mineral’s stability. On day 8, SEM pictures verified the presence
of a sodium fluoride and fluorapatite combination [18]. Fluorapatite was also visible
in TEM images. This exploratory study might lead to the creation of a useful enamel
biomineralization substance. One of the most challenging academic problems has
long been developing the finest biomechanically feasible skeletal scaffolding for
tissue engineering. To produce a hydrogel with sufficient wear resistance over time,
cysteines and the domain peptide were included in this case. After 21 days in culture,
the human mesenchymal stem cells (hMSCs) on the biomaterials appeared to still
be alive, proving that contraction rather than erosion was the cause of the loss of
volume.
These hMSCs also had higher levels of osteogenesis markers such as osteopontin
and osteocalcin than hMSCs that were placed on glass slides. Chondrocytes were
put to the gel after 28 days of growth and displayed 86% more vitality, respectable
architecture, and elevated activity of alkaline phosphatase than the control [19, 20].
As we mentioned at the beginning, one of the most intriguing applications of elastin-
like particles is the creation of scaffolding that mimics biological structures.
The final design of a 3D spheroid using an ELR chemically coupled to
polyethyleneimine (PEI) was compared with that of a 2D monolayer culture, even
though 2D cultures cannot replicate the physiological 3D environment. To create
3D microspheres as a 3D model of culture, these scientists plated hybrid fat-derived
stem cells (hASCs) on normal styrene coated with the ions with a positive charge
ELR-PEI. Viability tests were performed after both had been grown in differentiation
media, and they revealed the same number of cells as for the 2D layer. Scaffolds or
scaffolding for glycosidic bonds are also produced as composites to regulate bone
regeneration because of the advantages mentioned above. The mechanical proper-
ties of the scaffolds can be improved by the stiffness and flexibility of ELRs [21].
Four distinct composites with various ELR/collagen ratios were examined in this
experiment. The composite with the highest ELR/collagen content (18 mg/mL) had
the highest levels of ALP activity, OCN transcription, and stiffer matrix even though
all these composites contained hASCs that were proliferating and differentiating.
Composites constructed entirely of chondro had substantially less matrix and took
Genetically Induced Biomaterial Advances in Medical Sciences 103

longer to mature than those with lower ELR/collagen ratios. Currently, one of the
most prevalent causes of discomfort and incapacity in the elderly is the degeneration
of joint cartilage.
Due to collagen’s perivascular nature, it is challenging to identify articular struc-
tures that are harmed because of trauma or aging joints because collagen has a limited
capacity to rebuild [22, 23].
Like this, hydrogels are intriguing scaffolds because of their mechanical prop-
erties and ease of manipulation, especially those that can self-gelate under physi-
ological conditions and may precisely integrate a device in situ. Available internal
and external ELR-HA hydrogel foundations have been created to compare scaffolds
with varying component ratios. The stiffness of the supports remained consistent for
all the ratios that were taken into consideration. When HA concentration in chon-
drocytes increased, there was a dose-dependent rise in the transcription of cartilage
marker genes like aggrecan, SOX9, and type 2 collagen. The importance of these
chondrogenic indicators must be stressed because chondrocytes are the only cells
present in the knees that have the charge of controlling the extracellular matrix. The
hydrogel’s chondrocytes also showed increasing levels of type 1 collagen, a marker
of an undesirable fibrous connective tissue, and metallopeptidase-13, a marker of
matrix disintegration and remodeling, when the concentration of HA was reduced
[24]. Moreover, chondrogenic multiplication was boosted in biomaterials with lower
HA concentrations, whereas sGAG (sulfated glycosaminoglycan) deposition was
promoted in hydrogels with higher HA contents. This system (5% HAhydrogel) was
shown to be the best scaffold since it had the highest peak of cartilage indicators,
the largest sGAG deposit, the least amount of cellular proliferation, and the fewest
of type 1 collagen and fibrous tissue phenotypic development.

2 Advanced Medicine Delivery System from ELRs

The various ELR architectures are examined in this chapter as the building blocks
for numerous drug delivery systems. In these subchapters, devices spanning between
monomeric ELRs to amphoteric ELRs that can create nanoparticles have been
discussed. The chapter is concluded with macroscale ELRs for the delivery of drugs
in the forms of depots or hydrogels.

2.1 Drug Nanocarriers Made of Monomeric ELRs

Drug nanocarriers made of monomeric ELRs have been suggested as prospective

drug carriers because they improve the size and durability of medical molecules,
which are typically tiny and have short half-lives because they promote quick
glomerular filtration. Simple elastin-based polypeptides called monomeric elastin-
like recombinases can be modified and used as drugs [25]. As a result, they can be
104 E. Kaushik and R. Kaushik

used as solvent delivery systems when connected to biopharmaceuticals or as phar-

macokinetics boosters when incorporated into fusion proteins that make medications.
A monomeric ELR carrier provides unique benefits over the free drug by improving
the pharmacokinetics and drug load pharmacokinetic profile. Incorporating target
peptides, such as knh or cell-penetrating motifs, as well as reactive areas useful for
the strong covalent conjugation of medications is also made possible by genetically
designing elastin-like recombinases. ELRs can be engineered to self-assemble in
response to external stimulation, and they can be employed to deliver therapeutic
medications to injured tissue both systemically and locally. Beneficial clinical drug
delivery methods include monomeric ELRs coupled to drug-delivery molecules with
known therapeutic potential [26, 27]. A drug is joined to a macromolecule during the
thermoplastic coupling process; the most popular options are proteins and synthetic
substances. The insertion of sensitive residues that can be used to further compound
other medications, like the hydroxyl group in cysteines or the amine groups in lysines,
is made possible by the recombinant origin of ELRs.
If the loading is a protein, it is possible to create the ELR-load complex through
genetic engineering. The most common disease in the world is cancer, however, the
majority of therapeutic drugs employed for treating it have molecular masses of less
than 500 Da, are hydrophilic, have significant toxicities, have poor pharmacokinetic,
and have low therapeutic indices. The therapeutic index is the ratio between the
lowest dose necessary to achieve the desired result in 50% of a society and the dose
necessary to kill 50% of a community (effective dose). Interaction among triad is
shown in Fig. 2.
Additionally, several chemotherapy medications with anti-cancer effects,
including Paclitaxel, Doxorubicin, Camptothecin, or Melphalan, have a significant
drawback known as toxicity in normal healthy cells. Future uses of the thermal capac-
ities of ELRs for therapeutic hypothermia have a decent probability of becoming
successful. ELRs are anti-cancer medication delivery devices [28]. Increasing the
local, regional, or bodily temperature above the physiological level (37 °C) to roughly
42 °C is known as hyperthermia. Under a specific transition temperature (Tt), elastin-
like recombinamers are disordered and water-soluble, but as the temperature rises
over this Tt, they amass and collect. This property makes them intrigue as drug
delivery vehicles. Because ELRs are liquid at 37 °C, the body temperature, can
move freely. These compounds become intractable when they circulate in tumor

Fig. 2 Interaction among

the triad resulting in the
variation in functional,
survival, and aesthetic
qualities
Genetically Induced Biomaterial Advances in Medical Sciences 105

tissue and concentrate in the tumor, even though hypothermia is characterized as

the substance being warmer than the transition temperature. This suggests that using
the phase change of these polypeptides could facilitate the localized drug delivery to
tissues (Tt). The plasma membrane is one of the most challenging problems a system
for drug delivery must solve. The plasma membrane, a permeable barrier found on
living cells, is essential to their existence and ability to operate. Consequently, the
main challenge to assuring intracellular drug administration is getting a bioactive
molecule across the plasma membrane. Small compounds can pass right through the
lipid bilayer of this membrane, while protein-based therapies must first penetrate the
membrane via mobile transport. To increase the effectiveness of a medicinal agent’s
entry, cell-penetrating peptides (CPPs), among the most well-liked and effective
vectors for improving cellular absorption, could be used. Surface functionalization
of the ELR with aCPP can facilitate non-specific cell viability of the load via path-
ways, simplifying the cell survival of ELRs. CPPs are 5–30 peptide polypeptides that
can pass through cell membranes. As a result, a variety of CPPs have been included
in the bioactive of ELR drugs to enhance the targeted distribution of anticancer drugs
in the tumor [29, 30]. CPPs are also used to reduce the rapid breakdown and insuf-
ficient infiltration of tumor cells by these drugs. Frankel and Green, who discovered
that the HIV activator of transcription (TAT) peptide can traverse cell membranes
and was effectively absorbed in vitro, reported the first CPP in 1988.

2.2 Nanoparticles Based on ELR as Pharmaceutical Delivery

The utilization of nanoparticles as having to cut drug delivery systems is covered in

this chapter along with a variety of ELR-based architectures that can produce them.
We discuss a variety of devices, including triblock copolymers, cell-specific ligands,
hybrid diblock ELRs, and ELRs-blocks.

2.3 Amphiphilic ELRs Nanoparticles

Amphiphilic ELR-based diblocks can self-assemble to produce structures like

nanoparticles, suggesting that there are several ways to induce or reinforce their self-
assembly depending on the polymer design. The hydrophobic portion of the particle
has also received several bioactive domains that are accessible at the subatomic
particle shell. ELRs can release drugs under regulated conditions in reaction to
pathological changes in temperature because of their thermosensitivity. Anticancer
therapy benefits from heat generation at the tumor site because ELR-based methods
are efficient at creating well-organized nanoparticles at the infection site. The pH
differential between normal tissue and the cancerous environment is another factor
that aids in the therapy of cancer. Callahan and colleagues created an ELR block
copolymer (ELRBC), specifically [VG7A8]-80/[VH4]-100, to create nanoparticles
106 E. Kaushik and R. Kaushik

with a pH-responsive component that can also be stabilized with Zn 2 + ions [31,
32]. When compared to ELRBCs made of [VG7A8]-64/[V]-120, a non-pH-sensitive
ELR block copolymer, in vivo penetration, and distribution in tumors. The solid
tumor tissues usually have extracellular pH ranges of 6.2–6.9 as opposed to the
normal tissue’s circulatory pH range of 7.2–7.4, therefore the ionic intensity ELR-
based polyamide microspheres break down at this lower pH. The 60 pentapeptide
repeats in the diblock, which include the hydroxyl group as a guest residue, form a
hydrophobic block, whereas the 60 core protein residues, which contain glutamine
and glycine in a 1:1 ratio, form a hydrophilic block. Then, Silaffin R5 and the
hydrophilic region of the ELR were genetically connected. When phosphorus ions
are present in phosphate-buffered saline, the ELR-R5 is crosslinked, resulting in the
formation of silica nanoparticles. Afterward, silicic acid is added to the solution of
the made-up templates to make the silica polycondensation. Many researchers have
used the strain-promoted azide-alkyne cycloaddition (SPAAC) method to enhance
the formation procedure by connecting the chain at the particle’s outer shell. To
ensure peripheral crosslinking, a clearly defined ELR was created by joining the more
hydrophobic block to a lysine-rich area. The arginine amine groups were converted
into azide units using the diazo-transfer method, and the NPs were subsequently stabi-
lized through exergonic crosslinking using the chemical bis-cyclooctyne. Additional
approaches involve employing membrane transporters made of polymer nanoparti-
cles and genetically altered drug receptors after the prescription has been coupled to
the receptor [33]. The basis of this innovative encapsulation method, developed by
MacKay and colleagues, is the high selectivity between a given molecular medication
and its protein sequence targets attached to the corona of protein polymer nanopar-
ticles. The drug is administered gradually once it has been securely attached to the
carrier. Rapamycin (Rapa), an immunosuppressive macrolide antibiotic, is applied
over stents to prevent organ transplant rejection. Rapa’s anti-proliferative qualities,
constrained dispersion, low oral absorption, and quick systemic clearance made it
the ideal candidate for these studies. 23 days after the initial dose, the rodents lost
over 15% of their entire body mass, demonstrating the high toxicity of free Rapa.
The drug for the prostate xenograft mouse model showed superior antitumor action,
showing notably lower cytotoxicity in contrast to free Rapa, without exhibiting any
symptoms of behavioral issues or body weight loss. Eventually, diverse proteins and
small molecules will be encapsulated, targeted, and released using this technique.

2.4 Modern Techniques for the Synthesis of ELR

Nanoparticles

More and more sophisticated devices have been developed that use ELR-based tech-
nologies to recognize complex NPs that can transport a variety of small molecules
to improve regulated medication delivery systems. One of the best ways to incor-
porate bits into macro material is by self-assembly, which can transform nanoscale
Genetically Induced Biomaterial Advances in Medical Sciences 107

into devices with unique capabilities. Coworkers created the H6-ELR-CP triblock
polymer in this situation, which consists of a polypeptide block with thermal respon-
siveness, a hex histidine block with the potential to bind ionic species, and the capsid
peptides of the cowpea erythema mottle virus, for whom the self-assembly may
be pH-dependent [34, 35]. New nanocarriers built on recombinamers that mimic
elastin require inventive ways. Active targeting abilities can also be added thanks
to the customization of these ELRs with fibroblast ligands. To facilitate active cell-
targeting, ELR with a polymeric acid tail (ELR-D) and then added epidermal growth
factor EGF (ELR-D-E). The resultant nanoparticles have been shown to have a
specific biological interaction, and in vitro results have been encouraging using A549
cells, a normal lung cancer epithelium line that pushes the limits of the EGF recep-
tors. It looked at how to distribute suicide genes using a plasmid that contains the
Rna for the toxin PAP-S.
With excellent selectivity for MCF-7 cancer cell lines as opposed to a MUC-1-
negative tumor line, this technique effectively transported the plasmid into tumor
cells while protecting normal human cells. This aptamer-ELR vector’s ability to
transfect cells, as demonstrated by micropinocytosis uptake, opens the door for the
use of ELRs in suicidal genome therapy to cure cancer and further opens the door
for their use in the biomedical sector.

2.5 In-Situ Therapeutic Depots

One form of treatment where a prolonged local treatment of the tumor is required
for success is anticancer therapy. Therapeutic drugs must navigate several biological
barriers on their way to the tumor to be successful, and systemic medicines usually
have high rates of toxicity as a side effect. Thank goodness, localized anticancer
therapy exposure and reduced drug storage inside the tumor can be achieved through
intertumoral injection, minimizing negative side effects. An enticing alternative to
local delivery is the creation of a biocompatible substance that may be injected as a
medication delivery deposit at room temperature. The thermosensitive ELR sequence
in this instance carried a biopolymer into a tumor when soluble, and because of air
temp coacervation, produced a depot in situ [36]. Intertumoral radiation therapy has
a very good response rate in solid tumors, especially aggressive prostate cancers
(brachytherapy). By introducing radioactive “seeds” into or close to the tumor, this
technique gives the tumor a significant dosage of radiation while limiting unneeded
exposure to the neighboring healthy tissues. Despite the method’s overall benefits,
existing seed implantations are permanent and can only be used in the treatment of
low- or intermediate-grade malignancies. They are ineffective for treating high-grade
tumors. The development of polymeric devices that function in conjunction with
irradiation has undergone several attempts. It is preferable to avoid the problems with
conventional brachytherapy. Avoiding the issues with traditional brachytherapy is
preferred-based systems that have recently been taken into consideration for topically
applied delivery, particularly in the context of creating new capabilities considering
108 E. Kaushik and R. Kaushik

the prevalence of eye-related disorders. Lacritin is closely connected to a group of

contemporary elements that can enhance corneal mitogen action and encourage tear
protein secretion (Lacrt) [37, 38]. To create a monomer that can create a base on inter
cache at temperatures close to and protect Lacrt-mediated cell-signaling pathways,
Lacrt was fused to an ELR made up of 96 repeats of the pentapeptide VPGVG. In a
mouse model based on inter defect, the therapeutic efficacy of the extended infusion
of IL-1a and/or tumor necrosis factor was assessed. IL-1 inhibition significantly
reduces PTA-related indicators like synovial inflammation, bone repair, and chondro
degradation, according to in vivo research. Yet, lowering TNF-, whether alone or
in conjunction with IL1Ra, impairs bone structure and hastens articular cartilage
degeneration.
This novel method dramatically minimizes clearance from the joint space while
boosting the impacts of IL-1a and decreasing injection dose and frequency by
combining xELR with IL-1a for continual intra-articular administration.

2.6 Hydrogels

These highly hydrated hydrophilic, three-dimensional, insoluble viscoelastic poly-

mers can swell when in contact with liquid but do not dissolve it. Physical substances
that are activated by a range of inputs, such as pH, temperature, or even sonar waves,
can create these networks. Cross-linking during chemical processes can also produce
them. These nanogels are structurally stable due to crosslinking or physical interac-
tion between polymer strands. Composites can get over some delivery restrictions
because they can be placed close to the area of issue and distribute drugs locally for
a long time. The window for reducing systemic effects just at target site is expanded
by this restricted exposure. Enzymatically cross-linked ELRs are one of the most
cutting-edge components of hyaluronan delivery systems for drugs because they
could release bioactive compounds in response to specific environmental stimuli.
As was already mentioned, elastolytic activity has been connected to pathological
disorders like atherosclerosis, cystic fibrosis, and chronic wounds. Crosslinking was
made possible by the addition of microbial transglutaminases. After being subjected
to P. aeruginosa or human granulocytes, the final hydrogels emitted the recombinant
green fluorescent protein (eGFP) that was added during the gelation process over
a longer period (PMNs). Similar results were obtained once an electron transport
system was added [39]. ELRs are a potential biopolymer for making genuine pass
depots or biodegradable under specific circumstances because of their adaptability.
On the other hand, noncovalent cross-linking frequently leads to insufficient struc-
tural and mechanical stability, rendering them unsuitable for a variety of applications.
An injectable, repairable, dimeric crucifix silicone gel with properties halfway in the
range of coacervates and more durable chemical hydrogels was created to increase
the results in the applicability of these biomaterials. These results demonstrate the
effectiveness of nanomaterials for the in vitro gradual release of a reference protein
as well as the injectable biomaterial’s transport properties. The composite’s two
Genetically Induced Biomaterial Advances in Medical Sciences 109

components interacted secondarily, causing the ELR-collagen combo hydrogels to

release doxycycline more rapidly than gelatin hydrogels. Four bacterial strains that
are frequently found in clinical settings were used in bioassays to determine the
cytocompatibility of the doxycycline discharged from the hydrogels.
Staphylococcus aureus multidrug testing was carried out using the function.
Knowing that all hyaluronic acid produced doxycycline, which was effective against
all but four of the bacterial strains tested, it was demonstrated that the hydrogels had
no impact on the medicine’s bioactivity. Combining ELR with collagen hydrogels
can hasten the healing of wounds by reducing postoperative infection and extending
the duration of an antibiotic’s local site of action.

3 Background Related Work

[1] Frappier et al. [40]; Atomically small material alterations enable a variety of
nanoelectronics devices, such as resonant tunneling diodes (RTDs), quantum well-
integrating photodetectors (QWIPs), subatomic good lasers, and heterojunction junc-
tion effect transistors (HFETs). For the development and creation of such devices,
a fundamental knowledge of electron propagation in such wavelengths is required.
Based on a basic nonequilibrium electron transport theory, NEMO is a flexible tool
for designing and studying quantum devices. EMO was linked with the parallel
processing evolutionary algorithm application PGAPACK to modify the properties
of the structural materials to match a certain set of experimental data. A numer-
ical experiment is done to generate structural parameters like layer thicknesses and
doping levels to evaluate an experimental current–voltage characteristic. We find
that the basic and doping characteristics of the heNEMO model parameters closely
reflect the genetic algorithm. Synthesis is achievable with such precise agreement
between theory and technique are known.
[2] Schloss et al. [41]; In order to create new kinds of allergy vaccinations, DNA
Fragment was used in allergens research. Details on the genetic sequencing and
makeup were made available. One technique that is widely employed in the creation
of antimicrobial peptides that meet certain T- or B-cell epitopes. A new technique
for developing hypersensitive vaccinations that can provide a better and much less
replication of the epitopes is genetic engineering allergen synthesis. To show how
well these hypo-allergens reduce allergenicity, numerous stimulating skin and nasal
testing have been employed. The vaccines’ capacity to generate anti-allergen IgG
antibodies and sustain T fibroblast activity have both frequently been shown, despite
the diminished immunoglobulin E (IgE)-binding reactivity. The main hypoallergenic
have been polypeptide segments and tetrameric birch allergen structures.
[3] Olorunniji et al. [42]; Atomic size differences in the materials enable devices like
resonance tunneling diodes (RTDs), quantum well-infrared photodetectors (QWIPs),
classical good lasers, and heterojunction field effect transistors (HFETs). Such
heterostructure devices call for an in-depth knowledge of electron transport in
110 E. Kaushik and R. Kaushik

quantum states for their design and optimization. This problem is addressed by
the overall design and analysis tool for nanodevices known as the Nanoelectronics
Modeling Tool (NEMO). NEMO was used with PGAPACK, a library of parallel
processing neural network methods, to improve structural and material properties.
The quasi-band effects in the longitudinal and transverse dimensions as well as the
Wkb charge personality are included in the electron transport calculations displayed
here.
[4] Tang [43]; A popular biocompatible substance for stem-cell cartilage healing
is elastin-like polypeptides (ELPs), a class of artificial polypeptides with special
characteristics. The pentapeptide Val-Pro-Gly-Xaa-Gly makes up ELPs, and Xaa
can be any other amino acid besides Pro that passes through an inverted temperature
phase change. They can dissolve in water when it is below their transition temperature
(T t). Yet, when the temperature of water surpasses their T t, they congeal. This study
evaluates the rheological characteristics of an uncross-linked ELP below and above
its T t as well as the effectiveness of ELP in triggering chondrogenesis in vitro.
Recombinant DNA techniques were used to create an ELP with a T t of 35 °C.

4 Application

4.1 Peptide-Protein Interactions

Under conditions of kinetic and thermodynamic equilibrium, the intrinsic self-

recognition and processes of biomolecules enable the formation of organized
structures just at the nanoscale or even at the macroscopic scale. Peptides and
peptidomimetics are significant because they may allow for the rational dissec-
tion of biological pathways to produce new medications, materials, catalysts, and
other products. Van der Waals forces, hydrogen bonds, and electrostatic attraction
are a few instances of interactions that regulate self-recognition or self-assembly
processes. They also produce secondary structures like -helix, -sheet, and polypro-
line II helix, which are necessary for many biological functions. Here, we give some
recent and noteworthy examples of how design has been successfully used to create
functional structural themes. These investigations are crucial for comprehending
the fundamental relationships governing biological operations and the emergence
of numerous diseases. The types of secondary configurations that peptides take
on during self-assembly play a key role in the properties of biomaterials, such as
the contacts, encapsulating, non-covalent contacts, or covalent interactions that are
eventually useful for drug delivery applications [40].
The definition of the chemical self-assembly process is the bidirectional and
spontaneous grouping of some substances (small or macromolecules, peptides, or
proteins) into a well-defined and stable configuration while in thermodynamic equi-
librium. Both healthy and pathological processes result in the formation of several
Genetically Induced Biomaterial Advances in Medical Sciences 111

beneficial sub-cellular structures. A few examples include amyloids linked to disor-

ders including Alzheimer’s, Parkinson’s, diabetes, amyotrophic lateral sclerosis,
hemoglobin, and membrane transport channels. In the field of protein interaction
(PPI), where the connection may be connected to a conformational change that initi-
ates a biological pathway, the supramolecular association created by low intramolec-
ular pressures is an essential mechanism both in chemical and biological recogni-
tion. For instance, the growth of fiber-like structures is driven by hydrogen bonds
or interactions. If the dissociation energy is compared, hydrogen bonds only require
10–65 kJ/mol for homolytic breakdown while covalent bonds take 100–400 kJ/mol.
Multiple interaction sites are frequently combined throughout the assembly process.
Many organic compounds have been found to be capable of forming supramolec-
ular polymers under specific conditions [41, 42]. The fibrillation process is likely
the most remarkable biological example of peptide self-assembling, such as amel-
ogenesis in Alzheimer’s disease. Peptides represent a different class of identity and
self-recognition molecules. Moreover, recently, peptidomimetics. Since the early
1990s, researchers have been studying self-assembling peptides. Self-assembling
peptides can be used to create novel biological materials, coatings for surfaces and
semiconductors, as well as a new group of antibiotics to prevent and treat antibiotic
resistance. A self-assembling peptide (SAP) system uses organic or synthetic scaf-
folding that can display a variety of cell-interactive constituents in spatially defined
networks. They do this by using biomolecules self-assembly or naturally occurring
organized aggregates. Hydrophobic interactions, hydrogen bonds, van der Waals
forces, and electrostatic contacts all contribute to the stability and loss of energy
of protein self-assembled structures. The length of the peptides, the percentage of
hydrophobicity, the I amino acid sequence, and the duration of the self-assembling
process are all factors that influence the self-assembling processes. To be used in
a variety of biomedical fields, these innovative SAP-based materials usually make
hydrogels that may mimic the extracellular matrix and have synthesized scaffolds
that may contain different cell-interactive components. Comparison of peptide array
is shown in Table 2.

4.2 Silk Protein Sericin in Biomaterial

A natural polymeric protein called silk sericin is derived from the Bombyx mori silk-
worm. Sericin can be retrieved and used in other ways during the many processes used
to make raw silk and textiles. Additionally, sericin recovery lessens the impact of silk
production on the environment. Sericin protein has advantages due to its characteris-
tics. The protein is UV-resistant, antimicrobial, and resistant to oxidation [43]. It also
readily collects and releases moisture. To create products with better qualities, sericin
proteins can be polymerized, cross-linked, and combined with other biomolecules,
notably synthetic polymers. Additionally, the protein can be used to enhance or
coat both natural and synthetic fibers, fabrics, and products. Bioplastics, biomedical
112 E. Kaushik and R. Kaushik

Table 2 Peptide array comparison

Product Technology Support Scanning
Pepscan PepChip Standard Fmoc Glass slides Standard
microarray SPPS screening with
antibody-based
detection
LC Sciences PepArray Photolithography Chips Microfluidic
micro-array Chip using system
microfluidics
PEPperPRINT PEPperCHIP SPPS amino acids Chips/glass slides Antibody-based
microarray scanned with the detection and
electrical field in-house services
produced via laser for staining
printer
JPT PepSpot Standard Fmoc Cellulose sheets Electrotransfer
Microarray SPPS using the blotting to PVDF
SPOT synthesis membrane
technique
Intavis Celluspots SC2 Technique Glass slides Antibody-based
microarray detection directly
on an array

components, articles-forming polymers, functional membranes, fibers, and textiles

can all be made from sericin-modified components and sericin composites.

4.3 Modulation of Bone Cells’ Responsiveness to Plasma

Treatment with Starch-Based Biomaterials

Investigated were the effects of o2 radio wave glow discharge (rfGD) on the interfaces
of various starch-based biomaterials (SBB) as well as the effects of protein adsorp-
tion on controlling bone-cell activity. Both simple and complicated protein systems
used fibronectin, vitronectin, and bovine serum albumin. Surfaces coated with RfGD
demonstrated greater hydrophilicity and interface energy as compared to untreated
SBB. In cornstarch-based biodegradable polymeric blends, polycaprolactone (SPCL;
30/70 weight percent), poly (ethylene alcohol (SEVA-C; 50/50 weight percent),
and methylcellulose (SCA; 50/50 weight percent) were investigated. The maximum
degree of change was seen in SCA and SCA reinforced with 10% fluorapatite (HA)
after rfGD treatment [44]. On SCA, it was discovered that cell adhesion and multipli-
cation were increased compared to untreated surfaces, and the plasma modification
had no effect on SCA + 10%HA. BSA, FN, and VN single solutions enhanced cell
adherence to SCA surfaces, and a similar impact was observed for ternary systems.
Moreover, when compared to the untreated surfaces, SEVA-C treated with blood
shows better adhesion and proliferation. MG63 cell multiplication was clearly aided
Genetically Induced Biomaterial Advances in Medical Sciences 113

by plasma modification, even though adherence to treated and uncontrolled SPCL

was roughly equal. It was demonstrated that the variance in MG63 cells’ morphology
on SEVA-C surfaces was primarily controlled by the peptide system, whereas it was
primarily regulated by the treatments on SPCL surfaces.

4.4 Tissue Engineering

In vivo tissue-engineered uses elastin-like biomaterials, however novel ELRs are

often developed and enhanced to work better. For instance, the interaction and preser-
vation of mouse pluripotent cells were achieved using four distinct constructs. Most
of these inventions were composed of the ELR sequence (APGVGV) 12, the cell-
adhesion sequence (RGD), as well as the CBP peptide, a lawfully binding peptide
(iPSCs). The best method for encouraging mouse iPSCs to adhere to and develop on
untreated culture plates was determined to be the ELR construct (ERE-CBP), which
contains both the lower perceived and the CBP sequence. Additionally, it was shown
that CBP peptide enhanced the RGD gene’s capacity to permit cellular adhesion [43].
Additionally, compared to ERE alone, ERE-CBP showed much-increased expres-
sion of the embryonic marker Oct3/4 and alkaline phosphatase activities. There were
no obvious differences between the construction without RGD and ERE-CBP. Some
application of tissue engineering is shown in Fig. 3.
In opposition to CBP itself, which could not bind cells but successfully sustained
pluripotency, RGD showed its ability to increase cell attachment but not pluripotency
maintenance. The ERE-CBP study demonstrated how important it is for RGD and
CBP to coexist to preserve pluripotency. This unique recombinant ECM protein may
be best used in future in vivo tissue-engineered studies. One of tissue engineering’s
primary goals is likely to be the establishment of an extracellular environment that
is similar to the natural one. The growth and cell adhesion of fetal mouse fibroblasts
were examined in one study using GPG, or fiber of an ELR that had a double-
hydrophobic triblock structure (NIH-3T3). The scientists developed three different
hydrophilic structures with the RGD motif known to enhance cell adhesion proper-
ties: GPG, GPG-KAAK, and GPG-KAAK-GRGDS. They are all possibly suitable
for coating polystyrene surfaces after being cleaned with D-PBS, and over 90% of
these nanostructures remained intact. A noteworthy technique in the field of tissue
engineering is the development of cytocompatibility adhesives [45, 45]. Unexpect-
edly, mELY16 outperformed glass coverslips in terms of adsorption capacity. Both
constructions (ELY and mELY16) outperformed BSA and commercial sealant in
terms of good adhesion (>2 MPa under dry conditions and 0.24 MPa under wet
conditions, respectively) This new biomaterial could therefore have a significant
impact on regenerative medicine and tissue engineering.
114 E. Kaushik and R. Kaushik

Fig. 3 Applications of tissue engineering

5 Case Study

5.1 Elastin-Based Biomaterials that Have Undergone

Genetic Modification

One of the important features that distinguish the genetic algorithm from other algo-
rithms like evolutionary algorithms is that its main search operators are crossover and
mutation. The two main genetic operators utilized in genetic algorithms are crossover
and mutation. The objective of the current study was to investigate the impacts of
genetic algorithm operators such as mutation and crossover (P c & P), population
size (n), and the number of iterations (I) used to forecast the biological material
extruder’s lowest hardness (N). The second-order polynomial regression equation
developed for the extrudate characteristic hardness in terms of independent factors
such as temperature, screw speed, the fish proportion of the feed, and feed moisture
content was employed as the objective function in the GA analysis [47]. A simple
genetic algorithm (SGA) based on crossover and mutation operators was used in the
current study. A rank-based fitness selection algorithm was built into a C language
program for an SGA. The maximum number of iterations and inhabitants was set to
100. It was discovered that a medium community of 50, iterations of the algorithm,
and the lowest hardness values were feasible. According to the Pareto charts, P c
Genetically Induced Biomaterial Advances in Medical Sciences 115

Table 3 Biomaterial state for

Biomaterial Physical State
myocardium tissue
engineering Natural
Hyaluronan Solid
Alginate mesh Solid
Collagen Liquid/gel
Synthetic
PGA and copolymer with PLA Solid
PPS Solid

was found to have a greater impact when the population was below 50, while P m
was crucial when the population was below ten. The threshold values for a crossover
chance of less than 50% and a mutation probability of less than 5%. Physical state
of biomaterial for myocardium tissue engineering is shown in Table 3.

5.2 Engineered Biomaterials to Enhance Stem Cell-Based

Cardiac Tissue Engineering and Therapy

One of the biggest causes of death in the globe is cardiovascular disease. After a
myocardial infarction, cardiac tissue containing deceased or damaged cardiac cells
downstream of the occluded channel does not recover because adult cardiac cells
have a restricted capacity for proliferating. The heart becomes feeble when non-
functional fibrotic scar tissue takes the place of the original cardiac tissue [48].
Researchers are looking into the possibility of stem cells to regenerate damaged
cardiac tissue because host cardiac cells have a restricted capacity for proliferation.
This project has made tremendous progress. Currently, there is no agreement on the
ideal stem cell type, matrix material, or microenvironmental stimuli for cells. This
article provides a summary of the various biofunctional compounds and bioactive
matrices that, when combined with stem cells, have demonstrated promise for the
renewal and reinforcement of cardiac tissue. Engineered biomaterials are also used
in cardiac tissue engineering, which uses stem cells and biomaterial scaffolds to
generate tissues in vitro that can be used for drug testing or eventual implantation. This
study highlights the advantages of repairing injured myocardium with biomaterials
and stem cells and gives a quick overview of the characteristics of these polymers
that make them such useful instruments in the field.
116 E. Kaushik and R. Kaushik

5.3 Recombinant Cellulose Crosslinking Protein

Cellulose-binding domains have been discovered in a variety of cellulases and

proteins that lack hydrolytic function. The ability to alter the surfaces and mechan-
ical properties of paper by a cellulose-binding domain was examined. Two cellulose-
binding domains from Clostridium cellulovorans were combined to create a polypep-
tide that crosslinks cellulose (CCP). The mechanical characteristics of the synthetic
polyfunctional cellulose-binding proteins were studied when it was submerged in
Whatman cellulose filter paper after being expressed in E. coli. The extracted protein
improved the tensile strength, fragility, Young’s modulus, and energy to break off the
treated paper. Furthermore, it was demonstrated that filter paper improved in water
repellency following treatment with cellulose crosslinking protein. For the creation
of a new class of paper-modifying materials, it is intriguing that cellulose-binding
motifs can bind to cellulose under a variety of environmental conditions without the
necessity for chemical interactions [49, 50].

6 Challenges

• Bio Ceramic Challenge

The long-term durability of tissue-engineered constructions after implantation

depends on obtaining and sustaining a blood supply. Third-generation bioactive,
resorbable composites have been employed in studies to improve the vasculariza-
tion of a soft tissue construct made from regenerated tissue. To find the appropriate
concentration of bioactive particles, the production of vascular endothelial factors
was boosted while cell proliferation was observed using a rat fibroblast model. Adult
rats were given subcutaneous implantation of samples of the biodegradable, active
composite mesh. Embryonic stem cells and other cells were introduced into the
composite meshes.
• Stem Cell Engineering
Stem cell therapy, a revolutionary twenty-first-century approach to healthcare, is
built on the foundation of stem cells, which have the capacity for self-renewal and
the potential for multiple lineages. The ability of cells to differentiate into a range
of body tissues is widely known. The challenge is in getting stem cells to differ-
entiate into the required lineage, obtaining higher purity populations of the desired
cell phenotype, confirming that the cell line has no cancer-causing potential, and
then implanting the selected cell bloodline in a way that ensures the organisms will
multiply and replace or enhance the function of pathology or aging tissues. These
cells can be collected, grown, and, if necessary, integrated into a tissue construction
before being administered utilizing minimally invasive methods to the location that
requires healing [51]. The tissue at the location will regenerate because of the acti-
vated stem cells. There are issues with accessibility, infrasound (in the marrow, there
Genetically Induced Biomaterial Advances in Medical Sciences 117

is one cell type for every 100,000 cells), and diminished developmental capacity with
aging for some stem cell types. The control of stem cell development is a potential
use of mechanistically customized bioactive materials that may also serve as the
rationale for stem cell-based therapy.
• Bio Ceramics: Control of Infection
Due to bacterial adherence to biomaterials, which causes biomaterial-centered infec-
tion and insufficient tissue integration, several medical devices have a short lifespan.
Globally, the problem of the spread of bacteria resistant to common antibiotics is
getting worse. The increased incidence of chronic foot and limb lesions, which
frequently require amputation, is a substantial barrier. The bacteria and inflammation
that lead to cellular dedifferentiation must be controlled locally. AgBG concentra-
tions of 0.05−0.20 mg per microliter of culture medium suppress the development
of these bacteria. AgBG has a quick antibacterial effect in addition to being bacterio-
static. At AgBG concentrations of 10 mg ml1, a full bactericidal action was evoked
within the first hours of incubation. 45S5 Both Bioglass and BG had no impact on
the viability or proliferation of bacteria. Only the leaching of Ag+ ions from the glass
matrix is responsible for AgBG’s antibacterial activity [52]. Analytical measurements
exclude any involvement of pH changes, ionic strength changes, or the dissolution
of other ionic species from the biomaterials in AgBG-mediated bacterial death. The
patterns of Ag+ dissolution from AgBG in the addition and exclusion of bacteria
lend credence to the notion that the microbes are storing silver. According to XRD
patterns, FTIR spectra, and ICP data, the Ag-doped gel glass exhibits the same bioac-
tive behavior as bioactive gel glasses that have Approval from the FDA for use in
bone repair.
• Predicted in vitro testing of the biomaterials’ and nanoparticles’ toxicity and
biocompatibility
For both ethical and budgetary reasons, society is worried about the existing reliance
on in vivo testing on animals to assess the safety of novel biomaterials, TE structures,
and nanoparticles. The development of reliable and cost-effective predictive in vitro
experiments based on human cells is a major challenge for the twenty-first century.
There are a variety of issues with in vitro research that need to be resolved to ensure
relevance to prospective in vivo applications [53]. A mature cell phenotypic in culture
must, first and foremost, be representative of the same form of human cells in vivo.
Cell culture research currently frequently uses eternal cell lines, which are unable
to express the intricate protein arrays necessary for mature phenotypes. Third, while
testing, the mature cell phenotype in the cultured cells must be preserved. The drug
being tested might not be toxic enough to solely kill the cells; instead, it could cause
de-differentiation and impair the tissues’ ability to repair. These demand monitoring
of cell morphologies, preferably in situ. Fourth, the in vitro testing should contain
details regarding the molecular biological alterations that the chemical causes in
the cells. Fifth, the in vitro tests ought to be able to do statistical analysis to discern
between minute alterations in the cell population. Sixth, it’s important to take pricing
and usability into account.
118 E. Kaushik and R. Kaushik

7 Future Scope

Again, a biological basis to produce a generation of biomaterials provides the scien-

tific foundation for the protein engineering of scaffolding for TE and for in situ organ
regeneration and repair, preferably via minimally invasive surgical procedures. Every
one of these cutting-edge approaches has significant financial advantages and could
assist in resolving the problems related to caring for an aging population. A new class
of marker nanoparticles that are uniquely adapted for specific patients and illness
scenarios should be conceivable [54]. With the increased accessibility of predictive
analytics approaches, innovative plans for universal healthcare can be developed.
Until recently, this idea would have seemed ludicrous. It was difficult to envisage a
material that living tissues wouldn’t trash just 40 years ago. Millions of people have
benefited from this therapeutic reality, and it most likely will inspire new ideas in
the years to come.

8 Conclusion

It has also been studied how to distribute therapeutic drugs safely and effectively
to certain places, as well as a variety of ELR-based architectures and their prospec-
tive application as superior drug delivery systems. Whether utilized with transgenic
biopharma in a liquid delivery mechanism or as a pharmacokinetics enhancer, either
via chemical linkage or by creating fusion proteins of the drug, monomeric elastin-
like proteins that are implicated have been explored. These polypeptides can increase
the half-life of medications when utilized as therapeutic systems. Genetic engineering
also makes it possible for better cellular reception of nanostructures with therapeutic
functions in damaged tissues by inserting targeting peptides like knh or fibroblast
domains. Additionally, the right amino acid selection enables the existence of reactive
groups that are beneficial for the covalent connections formed during the conjugation
of molecules with proven therapeutic efficacy, with drugs and peptides being among
the most popular choices. ELRs can be designed to identify in response to external
stimuli, making them suited for systemic and local delivery of therapeutic drugs to
the injured tissue. ELRs that are available at body temperature after thermorespon-
sive transition have been demonstrated to accumulate in cancerous tissue because of
local overheating. In fact, by utilizing the porosity and perfusion of the vasculature,
therapeutic hyperthermia has improved the local distribution of ELR prodrugs into
tumor tissues. According to the diverse device structures, the biological applications
of NPs—in particular, ELR-based nanoparticles having signaling pathways that can
interact with organisms on their surfaces—have been studied. When used to heal
brain injury, and persistent skin damage, or to promote bone regeneration, several
of these have shown positive results. Lacritin nanomaterials are just one type of
NP that have been employed as therapeutic agents and are coupled to proteins that
really are visible on their surface. They support the integrity of the epithelium and
Genetically Induced Biomaterial Advances in Medical Sciences 119

corneal tissue healing. Last but just not least, studies on 3D ELR have been conducted.
These include depots, hydrogels, and macromolecular carriers. Because they provide
highly localized treatment, these are particularly promising for anticancer therapy. To
create injectable depots for venous distribution, ELRs can collaborate. This enhances
therapy options, localized anticancer medicine exposure, and drug stores in the
tumor. As macromolecular carriers, fusion-based ELR depots can carry curcumin
or glucagon-like peptide-1, which represents a substantial development in the treat-
ment of t2dm and pro therapy, etc. [55, 56]. The benefit of utilizing biomaterials that
have been chemically and structurally copolymerized is their ability to be positioned
close to the area of interest and disperse embedded drugs over a long period of time
locally, minimizing any negative effects. Because they enable the regulated release
of several bioactive compounds like hormones, antibodies, signaling pathways, and
other therapeutic biomolecules with possible biological uses, hydrogels containing
a domain sensitive to proteolytic disintegration have drawn particular interest. In
addition, the use of ELR-based 3D, notably hydrogels, has been investigated for
tissue-engineered with an emphasis on skeletal regeneration. Numerous strategies
have been examined, including the utilization of physiological conditions that repli-
cate the affected body part, such as the musculoskeletal or circulatory systems, or
the use of special scaffolds created from amphiphilic ELRs to restore soft tissues.
Because they can introduce various bioactive patterns into the specifically provided
sequencing or because they possess the appropriate cells for the regeneration of func-
tional tissues, these creative and biomimetic scaffolds are very significant for tissue
engineering. Studies have demonstrated that ELRs are excellent candidates for tissue
engineering because of the synergy that occurs when they are combined with bicom-
ponent scaffolds like ELR-collagen, ELR-silicon replacement perovskite, or ELR-
hyaluronic acid hydrogel scaffolds. For bone reconstructive surgery, new methods
based on precisely defined combinations of osteogenic materials, osteoinductive
chemicals, and/or osteoblastic cells are highly desired. The objective is to effectively
reproduce the original tissue without going through the bone harvesting process [57,
58]. To incorporate all the aforementioned qualities, a carefully controlled hybrid
material must yet be developed. Further developments require cooperation between
experts from a variety of disciplines, including stem cell biology genetics, diagnostic
devices, material sciences, medicine release, synthetic biology, and surgery.

9 Discussion

Materials that have potential for use in biomedical and therapeutic applications are
referred to as biomaterials [59]. They could be any or all the following, depending
on the application:
• Bioactive: They actively interact with bodily systems. This is mostly considered
in applications for tissue repair.
120 E. Kaushik and R. Kaushik

• Bioinert: This phrase implies that the substance in question does not affect or
upset the body system.
• Bioresolvable/Biodegradable: To prevent environmental pollution, the material
used for clinical or medicinal usage must be easily decomposed after use.
• Biocompatible: The material must be appropriate for its use, which is the most
crucial requirement of all [60, 60]. The material that is best for a certain application
will have mechanical characteristics that are like those of the real body component
that is being used or replaced.
• Mechanical Properties: The mechanical qualities of a biomaterial reveal the
molecule’s biocompatibility with the body system by describing the material’s
toughness, strength, and ductility.

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Genetically Induced Biomaterial Advances in Medical Sciences 123

Eva Kaushik is an IT professional with extensive experi-

ence in research. As a researcher, she is well-versed in varied
domains, including fintech, haptics, astrophysics, financial
economics, bioinformatics, and genomics. She has published
research manuscripts, book chapters, and articles in renowned
journals supporting her interest. She has been a founding
pioneer at Dexigner, a co-founder at FiCord community, and
even augmented the FinTech niche via various initiatives. Her
belief in hard work and perfection has led her to prominent
positions. She has served as chairperson of IEEE ADGITM, an
educational activist in IEEE USA, a core committee member
of the IEEE QT3 Series, an editorial coordinator at IEEE
WIE DS, and a public speaker at YPLO. She was chosen by
Microsoft for the start-up program “Binance Build for Bharat”
and received the IEEE WIE Affinity Group Award. She is
serving as an advisor at FrontForumFocus (a non-profit organi-
zation). She has volunteered with MHRD, WWF, IEEE MOVE
India, and the World Youth Alliance. Along with this, she is
spreading cognition about technology at different platforms for
the advancement of society. Precisely, being a woman of serious
potential, she believes in achieving accuracy and perfection.

Rohit Kaushik is a graduate student at the University of

Illinois, majoring in data analytics. His research interests lie
in the fields of machine learning, mathematical computation,
statistical analysis, artificial intelligence, and data mining. His
previous research has focused on the integration of technology
to diminish societal issues, leading to betterment. Rohit has
enormous potential and a burning desire to accomplish his
pursuits, along with a creative mind filled with new ideas.
Adding new heights to his research, he has been selected
by the Illinois Department of Public Health (IDPH) to research
the motor vehicle data linkage project in collaboration with
the Illinois Department of Transportation (IDOT). He has been
awarded the title of Director of Technology at “DREAMtorous,”
a business networking conclave and brand of Global Conflux
of Stalwarts (GCS). His dynamic personality leads him to new
opportunities, like getting invited as a guest speaker at the IEEE
Student Branch (2021) to deliver a session on data science,
machine learning, and deep learning. He has amazing analytical
and management skills, that directed him to serve as a commu-
nity manager at non-profit communities to support and enhance
their development using precise statistical data insights.
Biomimetic Approaches for Biomaterials
Development

Sudipta Choudhury, K. R. Arjun, M. N. Ramesh Bharadwaj, M. Maghimaa,

and Kanthesh M. Basalingappa

Abstract In recent times, the concept of biomimetics has gained widespread accep-
tance across different industries. This interdisciplinary field combines principles
from biology, engineering, and chemistry to develop materials, devices, or artifi-
cial systems that imitate biological processes. In the medical field, Biomaterials
play an important part in the recovery process by restoring function and speeding
up healing. These components, which might be natural or synthetic, are utilized to
preserve, enhance, or replace damaged tissues or biological processes. Additionally,
biomimetic technology has highlighted the importance of organelle attachment and
detachment in an organism’s ability to adapt to its environment. Tissue engineering’s
major goal is to make successful tissue grafts that can replace or repair damaged or
deteriorated tissues and organs. Currently, there are ongoing pre-clinical studies and
clinical applications of engineered tissues such as bones, cartilage, skin, skeletal
muscles, blood vessels, and bladder. In this chapter, the interconnection between
regenerative biology and engineering is examined, with emphasis on the utilization
of biomimetics in tissue engineering and the creation of functional tissue transplants
for regenerative medicine.

Abbreviation

BME Biomedical engineering

BID Biologically inspired design
ECM extracellular matrices
3D 3-dimensional
M meniscus

S. Choudhury · K. R. Arjun · M. N. R. Bharadwaj · K. M. Basalingappa (B)

Division of Molecular Biology, School of Life Sciences, JSS Academy of Higher Education and
Research, Mysuru, Karnataka, India
e-mail: kantheshmb@[Link]
M. Maghimaa
Department of Microbiology, Muthayammal College of Arts and Science, Rasipuram, Namakkal,
Tamil Nadu, India

© The Author(s), under exclusive license to Springer Nature Singapore Pte Ltd. 2023 125
R. Malviya and S. Sundram (eds.), Engineered Biomaterials, Engineering Materials,
[Link]
126 S. Choudhury et al.

T tendon
L ligament
TEN tissue-engineered nerve
CRISPR clustered regularly interspaced short palindromic repeats
Cas9 clustered regularly interspaced short palindromic repeats-associated
9
ZIF-8 zeolitic imidazolate framework
ELPs Elastin-like polypeptides
PGA poly-glycolic acid
PLA poly-lactic acid
OATS osteochondral autograft transfer system
NYSERDA New York State Energy Research and Development Authority
BIONIS Biomimetics Network for Industrial Sustainability
PLMC Poly D, L-lactide-co-trimethylene carbonate

1 Introduction

Human design has been “inspired by nature” for over three thousand years [1]. The
phrase “learning from nature” can be referred to in a variety of ways, and as a
systematic methodology, it is still in its infancy as a subject of study, particularly in
engineering design. Biomimetics covers a wide range of study areas, has an impact
on a number of application areas, and is thought to significantly improve the quality
of life on the scientific, social, and economic levels [2]. The majority of important
findings, however, have remained in their respective fields because of the large and
dispersed nature of research topics. According to studies, individual parties rather
than institutions have tended to practice biomimetics [3].
Nature has long served as an inspiration for the creation of useful materials and
systems [4]. Biomimetics involves drawing inspiration, adapting, or deriving design
ideas from nature. It is a process of developing innovative technologies by studying
biological systems. Through biomimetics, concepts from the field of biology are
applied to engineering, enabling the development of technologies that benefit from
millions of years of development evolution in natural biological systems as a result of
natural selection [2]. It denotes the investigation and imitation of natural processes,
systems, and approaches [5].
Biomimetics is a relatively recent topic of study that entails applying biological
scientific concepts and functions to engineering, design, chemistry, electronics, and
other disciplines [1]. Biologists, physicists, chemists, material scientists, engineers,
and other professionals combine to create biomimetics. Exploring the functions,
structures, and principles of several natural things, as well as engineering and engi-
neering materials and techniques with commercial uses, are all part of this multidis-
ciplinary discipline [6]. In 1974, the term “biomimetics” first appeared in print in
Webster’s dictionary [7].
Biomimetic Approaches for Biomaterials Development 127

In biological components, two forms of polymers, namely proteins and polysac-

charides (mostly composed of six-carbon sugars or hexoses), are commonly found.
Interestingly, When the densities of biological materials are evaluated, their mechan-
ical characteristics are equivalent to those of man-made materials [7]. Biomimetics
can provide valuable insights for the design of composite materials, drawing inspi-
ration from various natural examples such as the honeycomb beehive structure,
The fiber arrangement incorporates wood, spider silks, nacre, bones, and hedgehog
quills. Biomimetics explores solutions to human issues by studying nature’s models,
systems, processes, and components [8]. When using biomimetic membranes, the
immobilization of biorecognition molecules may not significantly affect the affinity
constant between the molecule and target analyte. This is because biomimetic
membranes imitate a natural environment that can contain numerous biorecognition
molecules [9].
Biomimeticists have traditionally concentrated on duplicating or copying biosys-
tems with synthetic components and traditional processes, deriving inspiration from
biological architecture and functions. However, recent advances in molecular and
nanoscale engineering in the physical sciences and molecular biology have enabled
biomimetics to be applied at the molecular level. Molecular biomimetics is a hybrid
approach that combines synthetic nanoscale structures with natural molecular
tools [4].

2 Terminology

2.1 Bioinspiration

“Based on observations of biological systems, an innovative method was developed”

[3]. Bioinspiration is utilized as a means to develop intricate and advanced engi-
neering models across different length scales, with the aim of harnessing the vast
information available to address the urgent challenges confronting humanity [10].

2.2 Biomimicry

The principles of sustainable development can be approached through the application

of interdisciplinary design techniques and philosophical frameworks that draw inspi-
ration from nature as a model for addressing social, environmental, and economic
concerns [3]. The convergence of biology and engineering, which encompasses
biomimicry, is a prevailing trend that is fuelling innovation in the twenty-first century.
This convergence presents opportunities for the transfer of knowledge across different
domains, the emergence of novel areas of expertise, and the transportation of physical
128 S. Choudhury et al.

materials [11]. The field of engineered biomimicry is rapidly evolving and involves
various disciplines, including biology, materials science, and manufacturing, making
it a challenging area of study [12].

2.3 Biomimetics

The collaboration among diverse fields such as biology, technology, and other innova-
tive areas has the goal of addressing practical issues by analyzing biological systems,
creating models of them, and transferring and implementing these models to discover
solutions [3]. “Biomimetics” refers to the imitation of life or nature. It comes from
the Greek word bio mimesis [6].

2.4 Bionics

The term “bionics” was coined by Jack Steele of the US Air Force Medical Division in
1960 by combining the words “biology” and “technics”. Bionics refers to a technical
domain that aims to replicate, enhance, or replace biological functions with elec-
tronic and/or mechanical equivalents [13]. In addition, there is a growing interest
in researching the improved regenerative abilities of bionic tissue-engineered
nerve (TEN) grafts, with a specific focus on the bionics of their structure and
components [14].

2.5 Biomedicine

Biomedicine is a medical field that applies biological and biochemical concepts to

medical research or practice. It is distinctive within a culture because it incorporates
specialized knowledge and unique practices based on that knowledge. Biomedicine
also involves a hierarchical division of labor and specific principles or guidelines
for social and professional interactions. The human body is the primary focus of
biomedicine [15].

2.6 Biomedical Engineering (BME)

The application of engineering principles to the medical field is the focus of biomed-
ical engineering, which is an engineering discipline that has gained increasing accep-
tance within the field [16]. Another fast-developing field of biomedical engineering
is tissue engineering [17].
Biomimetic Approaches for Biomaterials Development 129

2.7 Biologically Inspired Design (BID)

BID is a prominent trend in contemporary design that involves the incorporation of

biological functions and mechanisms to tackle human-related problems. Though
not mandatory, BID is often associated with engineering because it deals with
design issues akin to those encountered by engineers in various domains, such
as mechanical and aerospace engineering, electrical and computer engineering,
chemical engineering, and biomedical engineering [18].

2.8 Biomechatronics

“Biomechatronics refers to the utilization of medical and biological expertise to

enhance mechatronic procedures and products” [19]. Unlike biomimetics, biomecha-
tronics has a strong basis in engineering. Although it incorporates biomimetics, which
is a significant part of Bio4Eng (biology for engineering) for human–machine inter-
action, it is not restricted to it and is considered a related field [20]. Interlink between
Biology and Biomimetic approaches is shown in Fig. 1.

Fig. 1 Interlink between biology and biomimetic approaches (Made via powerpoint)
130 S. Choudhury et al.

3 Background to Biomimetics

Biomimetics holds the potential for application in various biological research

domains, including ecology, zoology, botany, molecular biology, and other related
subfields. Research areas with high promise include structure and systems design,
learning and memory, self-assembly and self-repair, perception and sensory systems,
movement and locomotion, control structures and self-regulation, and new physio-
logically active materials. At the nanoscale level, biomimetic research is expected
to have positive implications for component miniaturization, energy efficiency, and
self-configuration. The development of biomimetic robots that can operate in various
settings, including water, land, and air, is a significant area of attention. Additionally,
modeling brain systems, known as neuromimetics, is a crucial field of study, partic-
ularly for comprehending the foundation of intelligent behavior. The integration of
neuromimetic controllers in hardware, called “neuromorphic,” and in the control
frameworks of robots, referred to as neurorobotics, are emerging research fields in
the domain of biomimetics [2].
During the 1950s, Otto Schmitt introduced the term “biomimetics” to differentiate
between biological and engineering/physics approaches to science, which he referred
to as “biomimetics” and “biophysics,” respectively. Schmitt is also recognized as the
founder of biomedical engineering, which encompasses biomaterials and is closely
linked to biomimetics. The term “bionics,” coined by Jack Steele of the US Air Force,
pertains to the process of reproducing and modifying natural concepts [2].
According to the Wenzel and Cassie theories, superhydrophobic surfaces may
be generated by biomimetic materials with extremely hydrophobic qualities, which
need rough surfaces with low surface energy [21]. Many biological surfaces in nature,
such as lotus leaves, are superhydrophobic [22], wings of a butterfly [23], feet of a
water strider [24], and the petals of a rose. Superhydrophobic and superoleophilic
biomimetic materials are also utilized to extract oil from water.
An innovative electrochemical method has been devised to fabricate biomimetic
graphene on stainless steel surfaces, and this approach has proven successful in sepa-
rating oil and water. By combining biomimetic structures with graphene, biomimetic
graphene can be created, which could result in a range of devices with diverse prop-
erties based on graphene, potentially giving birth to a new graphene scientific field.
The surface and structure of biomimetic graphene mimic those found in nature [8].
Molecular biomimicry is an intriguing field that involves the precise and robust
implementation of synthetic techniques to replicate, alter, enhance, and analyze
biological systems. Synthetic membranes and ion channels, artificial photosyn-
thesis, synthetic enzymes, and, more recently, synthetic cellular life systems are
some of the current areas of molecular biomimetic research. Additionally, subfields
in materials research have expanded over the last few decades to include bioengi-
neered and hybrid-polymer materials, adhesives, biomineralization, and imitations
of extracellular matrix in terms of function, surface, and morphology [25].
Biomimetic Approaches for Biomaterials Development 131

4 Biological Materials

The body is a chemical laboratory that converts natural chemicals into energy,
construction materials, trash, and a variety of multifunctional structures [26]. Humans
have long recognized natural materials as sources of food, clothing, comfort, and
other essentials [27]. Despite their tiny size, animals and insects may create huge
quantities of natural products such as fur, leather, honey, wax, milk, and silk to suit
human requirements [5]. Due to their closely regulated structure and better-oriented
connections, many biological materials are less dense than synthetic materials with
equivalent qualities [7].
For centuries, natural materials have been utilized due to their appealing char-
acteristics like strength, durability, and beauty. In order to replicate these benefits,
attempts have been made to produce artificial imitations of these materials in any
quantity required. Natural materials possess unique features such as self-replication,
self-healing, reconfigurability, multifunctionality, and chemical equilibrium. Unlike
man-made materials that are typically processed through heating and pressuriza-
tion, natural materials are created under ambient conditions. Materials derived from
biological sources result in minimal waste and pollution, and the final product is
usually biodegradable and recyclable by nature. Mastering the handling of these
materials will expand our range of material options and enable us to produce more
environmentally friendly and recyclable materials [5].
Biomaterials play a crucial role in wound healing by acting as carriers for protein
and gene delivery, as well as providing scaffolds for cell attachment, growth, and
differentiation. They have been utilized to transport growth factors, small organic
compounds, and genes for the treatment of various diseases and wound healing [28].
Spider with their incredible webs is shown in Fig. 2.

5 The Two Approaches of Biomimetics

Recent advancements in biomedical research have been greatly aided by biomimetic

methods [29]. Generally speaking, there are two ways to practice biomimetics:
Problem-driven or solution-oriented. Both problem-driven and solution-based
methodologies have distinct starting points and design features (Figs. 3 and 4) [3].

5.1 The Solution-Based Approach

The solution-based approach involves using the knowledge of biological systems to

design and develop technology that replicates the functions of these systems. To do
so, a thorough understanding of the biological system is necessary in order to identify
132 S. Choudhury et al.

Fig. 2 Spiders are impressive “manufacturing engineers” of the natural world, able to create mate-
rials with remarkable efficiency, as evidenced by their incredible webs [5] The silk material used
in the spider’s web is five times stronger than steel, when compared by weight. (Original picture
captured by author)

Fig. 3 The biologically inspired design employs two analogical processes: a An analogy based on
a solution

Fig. 4 The biologically inspired design employs two analogical processes: b analogy based on an
issue

potential design solutions. The principles learned through fundamental research can
then be applied to technology. The transfer of scientific knowledge to industry is
closely tied to this approach [3] (Fig. 3).
Biomimetic Approaches for Biomaterials Development 133

The subsequent steps in the design process that is driven by solutions and inspired
by biology [30]:
Step 1: Recognition of Biological Solutions
Designers start with a certain biological solution in mind [30]
Step 2: Definition of Biological Solution
Step 3: Extraction of the Principle
Step 4: Reframing the Problem
Creators are compelled to reinterpret their work in order to assess how consumers
will appreciate the biological purpose attained [30].
Step 5: Look for a Problem
In contrast to search in the biological world, which entails searching through
some finite space of biological answers, it may encompass inventing whole new
challenges [30].
Step 6: Difficulties Identification
Step 7: Implementation of the Principle

5.2 The Problem-Driven Approach

The problem-driven approach in biomimetic development centers around identi-

fying real-world issues and finding solutions to them, with the first step being the
identification of a problem. By recognizing biological systems that accomplish a
certain function or mechanism, it is feasible to integrate new or improved functions
into technology by transferring and abstracting these principles. Despite the fact
that the problem-solving method and problem-driven approach are closely related,
they have inherent differences, and a detailed analysis is necessary to gain a deeper
understanding of each process [3].
In biologically inspired design that is problem-driven, the process follows a
pattern of non-linearity and dynamics, where the results of later stages can impact
the outcomes of earlier stages. This pattern enables iterative loops of feedback and
improvement [30].
Step 1: Formulation and definition of a problem
After being pushed to find or construct an issue to remedy, designers were asked to
define their problem as a function [30].
Step 2: Problem Reframing
Designers reframed their problems in biological terms in order to find biological
analogies for solutions [30].
134 S. Choudhury et al.

Step 3: Search for Biological Solutions

If the problem is clearly mentioned as “staying cool,” alter the limitations to
“thermoregulation” to broaden the search range [30].
Multifunctionality: Look for animals or systems that can solve many problems at
once [30].
Champion Adapters: Locate a system or organism that can withstand the most severe
form of the issue under investigation [30].
Step 4: Definition of Biological Solution
Initially, designers uncovered biological system structures and surface mechanisms
related to the reframed function [30].
Step 5: Extraction of the Principle
The identification of significant principles from a solution required a description
that minimized specific structural and environmental limitations once a proper
understanding of it was achieved [30].
Step 6: Application of the Principle
Once a thorough comprehension of a solution had been achieved, it was necessary to
provide a description that eliminated as many particular structural and environmental
constraints as feasible to identify significant principles. This procedure requires a
translation from one domain area, such as biology, to another, such as mechanical
engineering [30] (Fig. 4).

6 Biomimetic Materials Applications

6.1 Bioinspired Materials for Tissue Engineering

Tissue engineering enables the creation of biological substitutes for diseased and
damaged tissues using a number of techniques [31]. To attain the desired outcome,
numerous approaches are being investigated in musculoskeletal tissue engineering.
Among these, implanting 3D synthetic polymeric scaffolds at the site of tissue
damage is a widely used method. These scaffolds provide a framework that facili-
tates cell attachment, proliferation, and extracellular matrix (ECM) synthesis [32],
to accelerate tissue regeneration [33]. Many researchers have created scaffolds using
standard approaches such as phase separation salt leaching [34, 35].
The discovery of biomimetic scaffolds that mimic the architecture of the extra-
cellular matrix (ECM) has recently emerged as a promising strategy for tissue regen-
eration. To fabricate these 3D biomimetic scaffolds, advanced techniques such as
electrospinning and rapid prototyping have been employed. These scaffolds possess
Biomimetic Approaches for Biomaterials Development 135

a hierarchical 3D organization and are composed of collagen as the organic compo-

nent and carbonated hydroxyapatite nanoparticles as the inorganic component. The
human skeletal system is predominantly composed of bone, which serves as a biolog-
ical composite [8]. However, bone tissue engineering makes treating harvested and
injured tissues much easier [36].

6.2 Bioinspired Components Used in Genome Technologies

The CRISPR (clustered regularly interspaced short palindromic repeats)-associated

9 (Cas9) gene-editing technology is widely employed in plants, model organisms,
bacteria, and human cells (8). Cas9 allows for the disruption of a specific gene at
a specific site and the activation of the endogenous repair process by inserting a
specific piece of DNA, which can change the underlying cause of genetic illnesses
[37]. CRISPR/Cas9 delivery systems are based on physical or viral techniques
with limited applicability [38]. Ongoing research is being conducted to develop
novel delivery methods for CRISPR/Cas9, which entails investigating synthetic
delivery vehicles and metal-organic frameworks. Some examples of these include
DNA nanoclews, gold nanowires, gold nanoparticles, and the zeolitic imidazolate
framework [ZIF-8] [8].
Biomimetic nanoparticles, which blend synthetic cores with natural cell
membranes, are commonly used in cancer therapy and immunization. Specifically,
CC-ZIFs (zeolitic imidazolate frameworks enclosing CRISPR/Cas9) are nanopar-
ticles coated with cancer cell membranes, enhancing the precision of cell-targeted
gene editing by utilizing the homotypic binding events of tumor cells. Consequently,
the creation of CC-ZIF metal-organic frameworks represents a major advancement
in genome editing technologies, particularly in the domain of cell-specific cancer
treatment [8].

6.3 Bioinspired Components for Ultrasound Imaging

Compared to other imaging techniques like single-photon emission tomography,

positron emission tomography, and magnetic resonance imaging, ultrasound imaging
is generally known for producing images of lower quality. This is owing in part to
the input of echogenicities from adjacent tissues, which might result in poor picture
contrast [39]. When circulation proteins are adsorbed onto a material, it can alter its
physicochemical properties, including surface charge, surface composition, and size.
This process can affect the material’s cellular uptake, circulation duration, toxicity,
and tendency to agglomerate.
Changes in biological environments can negatively impact the ability of particles
to reach their targets effectively, resulting in suboptimal outcomes. A new tech-
nique called biomimetic coating or cloaking has been developed to address this
136 S. Choudhury et al.

problem. This technique involves using cell membranes obtained from lymphocytes,
macrophages, thrombocytes, and other sources to overcome the challenges related
to protein adsorption [8]. The biological composition of the cell membrane surface
is crucial in controlling the adsorption of biological components on particles, which
affects the circulation time, immune-mediated degradation, and biocompatibility of
the particles [40].

6.4 Bioinspired Materials Used in Actuators

Stretchable and flexible pressure sensors rely on biomimetic materials with micro-
and nano-hierarchical structures to work. These sensors have high sensitivity,
stability, and a wide pressure range [8]. Stimuli-responsive actuators, which can
transform diverse forms of energy such as thermal, optical, and electrical energy into
mechanical energy, are another application of biomimetic materials. These actu-
ators are highly versatile and can perform intricate mechanical movements in a
precise sequence, making them useful for bionic devices, artificial muscles, and intel-
ligent robots [41]. In recent times, there has been a growing focus on multi-stimulus
actuators instead of single-stimulus actuators [41]. Bilayer actuators are an attrac-
tive candidate for biomimetic applications because of their outstanding mechanical
characteristics and significant photo-induced stress [8].

7 Tissue Engineering

One of the most successful approaches for replacing or repairing damaged skin
and other biological components is tissue engineering [42]. In the artificial tissue
regeneration process, both in vivo and in vitro procedures are utilized [43]. It offers a
significant treatment alternative by combining the patient’s own cells with a scaffold
for bone healing [42]. The purpose of biomimetic tissue engineering approaches is
to use biomaterials to actively drive tissue repair and regeneration [44]. To guarantee
effective regeneration of the target tissue, the biomaterials used in tissue-engineered
constructs (TECs) and drug delivery must fulfill particular biological and mechanical
parameters [45].
Biomimetic materials that mimic the characteristics of the natural extracellular
matrix (ECM) have been created with the help of recent advancements in biomaterials
and tissue engineering [46]