ASEPTIC

BEHAVIOUR AND
PRACTICES
P R E S E N T E D B Y : PA L A S H C H D A S
Version 1.0
LEARNING OBJECTIVE

• Basics of Microbiology
• Concepts of Sterile/Aseptic Processing
• Aseptic processing
• Aseptic Behaviour

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CLASSIFICATION OF INFECTIOUS
AGENTS
• Bacteria – survive on appropriate media, stain gram-positive or -negative
• Viruses – obbligate intracellular parasites which only replicate intracellularly (DNA, RNA)
• Fungi – non-motile filamentous, branching strands of connected cells
• Metazoa – multicellular animals (e.g.parasites) with complicated life cycles often involving
several hosts
• Protozoa – single cell organisms with a well-defined nucleus
• Rickettsia – very small bacteria spread by ticks
• Prions – unique proteins lacking genetic molecules
• Chlamydia – bacteria lacking cell walls

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CLASSIFICATION OF INFECTIOUS
AGENTS
Bacteria Fungi

Bacteriophage

Avian Flu Giardia Ameba

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Viruses Palash Chandra Das
Protozoans 4
SIZE COMPARISON OF MICROBES

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BACTERIAL CELL STRUCTURE

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TAXONOMY OF BACTERIA

Domain
Phylum
Class
Order
Family
Genus
Species

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TAXONOMY OF BACTERIA

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MICROBIOLOGICAL CLASSIFICATION
OF INFECTIOUS DISEASES
• Bacteria are classified by their Gram stain characteristics.
• Gram staining is the application of a crystal violet dye to a culture of bacteria. Bacteria that
retain the color of the dye are called Gram positive; bacteria that don't are Gram negative.
– The Gram stain attaches to peptidoglycan in the bacterial cell wall.
• In Gram-negative bacteria, the peptidoglycan layer is protected by an outer membrane.

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MICROBIOLOGICAL CLASSIFICATION
OF INFECTIOUS DISEASES

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CLASSIFICATION OF BACTERIA

Gram + cocci Gram - bacilli

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CLASSIFICATION OF BACTERIA

Gram - Spirochete Gram + bacilli

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MICROBIOLOGICAL CLASSIFICATION
OF INFECTIOUS DISEASES

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UNDERSTANDING CONTAMINATION

• Definitions
• Contamination: introduction of undesired impurities (chemical, microbiological, or foreign
matter), at different stages of manufacturing during receipt, production, sampling, packaging,
storage or transport
• Cross Contamination: a contamination of starting material intermediate product or
finished product with another starting material or a product during production. A Typical
source of cross contamination is Premises, equipment, people, production process, clothing,
utilities and services.
• Mix-up: A mistake that results from taking one thing to be another. A wrong action
attributable to bad judgment or ignorance or inattention

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UNDERSTANDING CONTAMINATION
Bacteria

Viable Fungus

Other Microbes

Contamination Dust

API / Excipients
Non-viable
Skin flakes, Hair,
sweat

Gases / smoke

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UNDERSTANDING CONTAMINATION
Human

Practices Air

Contamination
Sources

Material Water
Equipment

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DIFFERENCE IN TERMINAL STERILIZER
AND ASEPTIC PROCESSING
Terminal sterilization Aseptic processing

▪ Product containers are filled and sealed ▪ Drug product, container, and closure
under high-quality environmental are subject to sterilization separately,
conditions designed to minimize and then brought together.
contamination, but not to guarantee
sterility. ▪ Because there is no process to
sterilize the product in its final
▪ Product in its final container is subject container, it is critical that containers
to a sterilization process such as heat or be filled and sealed in an extremely
irradiation. high –quality environment.

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 ASEPTIC PROCESSING

• Aseptic Processing is the processing of drug

components ( drug product, containers, excipients,
etc.) in a manner that makes impossible of
microbiological contamination of the final sealed
product

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CONCEPTS OF STERILE/ASEPTIC
PROCESSING
• Sterile: Free from living organism
• Aseptic: Absence of pathogenic microorganism or technique used to prevent microbial and
particulate contamination
• Aseptic Processing (John W Levchuk, CBER, FDA): The product and all of its contact parts
are sterilized separately and brought together under exposed conditions where, if not properly
controlled, could result in contamination

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CONCEPTS OF STERILE/ASEPTIC
PROCESSING Nested Manufacturing Zones (Diagrammatic Product: Out-Flow Not Shown

External Areas
Street, Offices, Restaurant

Transition Zone
Brings People, Materials etc., from External Areas to the
Manufacturing Areas in a ‘Controlled’ Manner

Clean Area
Provides a Protective Envelope to Minimise the
Challenge to the Critical Areas

Critical Processing
Remove Raw
People Change Change Area Compounding
Outers Materials
e.g. Point of Fill

Sterilise

Remove
Outers

Container Closures

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CONCEPTS OF STERILE/ASEPTIC
PROCESSING

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CONCEPTS OF STERILE/ASEPTIC
PROCESSING
• Parenteral Drugs:
➢ Injectable drugs bypass the body’s natural defences so must be sterile
• Terminal Sterilisation:
– Sterilise drug in final container (after manufacture and packaging)
– Preferred method of sterilisation
❖ Moist Heat Sterilisation
❖ Gamma Radiation
❖ E-Beam
❖ Microwave
➢ Many drugs not suitable for this method of sterilisation
• Aseptic Processing:
➢ Individual components are sterilised separately and brought together in the final form in a sterile environment.

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 CONCEPTS OF STERILE/ASEPTIC
PROCESSING
• Clean Room
– A room designed, maintained and controlled to prevent particulate and microbiological
contamination of drug products

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 CONCEPTS OF STERILE/ASEPTIC
PROCESSING
• An area with a defined particulate and microbiological cleanliness standard, usually consisting
of Laminar Air Flow protection.
• A clean zone is designed to maintain asepsis or sterility of the critical activity, product, test, or
material contained within it.

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 CONCEPTS OF STERILE/ASEPTIC
PROCESSING
• Critical activity
• an activity that involves close proximity or manipulation of exposed product, raw materials or
sterile equipment and components
• Laminar Air Flow (LAF)
• Uni-Directional Air Flow (UDAF) at a velocity sufficient to uniformly sweep particulates away
from the Clean zone

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ASEPTIC PROCESSING:

where the drug product, container, and closure are subject to sterilisation processes separately,
as appropriate, and then brought together

• ‘Any manual or mechanical manipulation of the sterilised drug, components, containers and
closures prior to, or during, aseptic assembly poses a risk of contamination and thus
necessitates careful control’

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ASEPTIC PROCESSING:

• Poor cGMP conditions…can ultimately pose a life threatening health risk to a patient’
• ‘Even successfully qualified systems can be compromised by:
poor personnel activities
poor operational activities
poor maintenance activities’

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ASEPTIC PROCESSING:

• ‘It is essential that operators involved in aseptic manipulations adhere to the basic principles of aseptic technique at all times…’
• Appropriate training should include:
– cleanroom behaviour
– aseptic technique
– microbiology
– gowning
– patient safety hazard posed by a non-sterile product
• Personnel training should be updated regularly
• Supervisors should routinely evaluate operators
• Standards of behaviour and aseptic technique must be transferable to all relevant situations
– this will be observed by an auditor/inspector
– if in doubt guidance must be sought

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ASEPTIC PROCESSING:

• Everyone entering cleanrooms/zones must:

– be trained and competent, or
– must be ‘closely’ supervised
• Everyone has a responsibility to:
– be familiar/compliant with expected standards
– report substandard behaviour (e.g. gowning)
– report damaged room fabric, equipment, gowns
Any persons who pose an unacceptable risk to the product, process or facility must be excluded
immediately and retrained

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ASEPTIC PROCESSING:

• Items entering cleanrooms

– avoid material that cannot be effectively sanitised
– avoid material that sheds excessive particles
– Work areas must be kept clean, neat and tidy to reduce cross-contamination/mix-ups
– Items within cleanrooms must not obstruct HVAC/LAF vents
– Standing water must be cleaned up
• Status labelling
– e.g. cleaned, sanitised, disinfected or sterilised
– soiled disposable equipment must be discarded
– soiled reusable equipment must be removed

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 ASEPTIC BEHAVIOUR:

• Good practice:
– gowns/PPE changed if damaged, wet or used for long durations
– check yours and others regularly
– target max duration = 4 hours
 Avoid rapid movements
– creates particles
– disturbs air flows
 Avoid aerosol production
  personnel =  contamination

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 ASEPTIC BEHAVIOUR:

• Aseptic technique must always be used wherever applicable

• Fresh sterilised gloves must be worn immediately before a critical activity and regularly
sanitised
– but do not use disinfectant spray near product, components, raw
materials or environment monitoring equipment

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ASEPTIC BEHAVIOUR:

Minimise spread of contamination during critical activities:

• avoid touching your person or other people
• avoid touching human contact sites such as:
• pens bin handles
• keyboards paperwork
• keypads desks
• doors plugs/switches
• chairs any unclean equipment
• telephones containers (disinfectant cans?!)
•if you do make contact - sanitise gloves

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 ASEPTIC BEHAVIOUR:

• Sterile/sanitised implements must be used for handling product. If sterile their sterility must
be maintained throughout the activity
• Minimise particles when opening autoclave bags

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 ASEPTIC BEHAVIOUR:

• Items dropped on the floor

– floors are relatively dirty areas
– items must NOT be picked up during a critical activity unless under exceptional
circumstances
• if really needed the item must be thoroughly sanitised/sterilized and the operators
gloves changed if they are completing the activity
• Touching floors must be avoided at all times
– if having to kneel, sit or lie on the floor, either
• disinfect the floor before and after
• place a sanitised/sterile sheet on the floor first
• Aseptic items must not be placed on the floor, or any other unsterile/unsanitised surface
– place on a suitable surface, or
– place on a sterile, or disinfectant wipe
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ASEPTIC BEHAVIOUR:

To avoid contamination and disruption of air flow:

• Control movements within, and adjacent to, cleanzones
• Talking must be avoided within, or adjacent to, cleanzones
• Interventions must be performed slowly and deliberately
• Covers, curtains and doors surrounding cleanzones must be opened with minimal
distance/force
• Aseptic technique must always be used regardless of any other protection present

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ASEPTIC BEHAVIOUR:

• ‘First Air’ principle:

– items being protected by air flow must receive clean ‘first air’ - so all operator manipulations must
be done downstream of air flow

• Aseptic manipulations and operator gloves must be restricted to

within the protected zone during a critical activity

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 ASEPTIC BEHAVIOUR:

Operation of cleanzone:
• Workareas must be clean, neat and tidy.
• Items present within the cleanzone must be minimised and located
so as not to disrupt or reduce air flow protection
 Cleanzone airflows must be allowed to operate for at least 10 mins*
prior to use
 Cleanzone work areas must be cleaned prior to use

*Need to demonstrate during qualification

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ASEPTIC BEHAVIOUR:

Gloves must be sanitised at regular intervals, but especially when related to a critical
activity:
Before leaving the final change
Entering the filling room/critical zone
Before opening clean zone screens/doors
Before handling container/closure bags
Before handling a sterilised implement/surface
After touching any part of ones self or others
After touching a human contact site
After touching microbiological media

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 ASEPTIC BEHAVIOUR:

Use of disinfectant sprays

avoid excessive particles
spray away from environment monitoring equipment
including particle counters (use Safe Zones in filling
area/isolator)
Use a jet spray for sanitising gloves
• deliver a pool into the palm of the hand
• spread across all surfaces
• allow to dry before commencing

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ASEPTIC BEHAVIOUR:

Filling rooms
must not be used as short cuts, even when not in use
must not be used to store items not required for filling activity in progress
must have personnel numbers minimised
• Long 24” outer gloves are mandatory at all times
• Goggles to be worn during critical activities

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 ASEPTIC BEHAVIOUR:

• When working in clean zones the operator must hold gloved hands:
– at working height
– at/above waste height
– under protective air flow
– avoiding touching anything unsterile
• Aseptic technique must be demonstrated at all times

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COMMON BAD PRACTICES – CLEAN ROOM
BEHAVIOR
Not wearing Safety Goggles
•Eyebrows / eye lashes can contaminate product

Improper gowning
•Dirt / dust from street clothes can contaminate product

Carrying unclean objects into cleanroom

•The lint from paper / cardboard can contaminate products

Carrying personal belongings to cleanroom

•They can fall and contaminate products

Wearing make-up in clean room

•Items like body spray can contaminate through aerosol
•Make up like nail paint can also contaminate product

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CONCLUSIONS

• Contamination control is a continuous battle

• User behavior has a critical impact on contamination
• Follow good hygiene practices
• Proper procedures must be followed at all times
• Any automatic control to process should not disable
• Do proper maintenance of machine
• Avoid occasional break down, do proper cleaning or testing after rectification
• Think about your actions! Be an active participant

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