World Journal of Microbiology and Biotechnology, 2023
Onion skins, actually recycled as organic fertilizers, could be used as a substrate in environmen... more Onion skins, actually recycled as organic fertilizers, could be used as a substrate in environmental-friendly bioprocesses to recover high-value bioactive compounds and food ingredients. In this work, a bioprospecting method was carried out including 94 bacterial and 45 yeast strains from several agri-food and environmental niches to verify their ability to grow on onion skins as unique nutrients source. Red and yellow onion skins were assessed by newly selected starter-driven liquid submerged fermentation assays mainly aimed at the release and modification of polyphenols through microbial activities. Fermented onion skins were also investigated as a inexpensive favourable source of microbial enzymes (amylases, proteases, lipases, esterases, cellulases, xylanases). In red onion skins, the treatment with Lactiplantibacillus plantarum TB 11-32 produced a slight increase of bioactive compounds in terms of total phenolics, whereas with the yeast strain Zygosaccharomyces mrakii CL 30 − 29 the quercetin aglycone content increased of about 25% of the initial raw material. In yellow onion skins inoculated, the highest content of phenolic compounds was detected with the yeast strain Saccharomyces cerevisiae En SC, while quercetin aglycone increased of about 60% of the initial raw material in presence of the bacterial strain L. plantarum C 180 − 34. In conclusion, the proposed microbial pre-treatment method can be a potential strategy to re-use onion skins as a fermentation substrate, and as a first step in the development of a biorefinery process to produce value-added products from onion by-products.
Golden thistle (Scolymus hispanicus L.) is a wild edible plant belonging to Asteraceae family, wi... more Golden thistle (Scolymus hispanicus L.) is a wild edible plant belonging to Asteraceae family, with a great potential for food applications. The aim of this study was to identify the best cooking procedure able to provide a high-quality, ready-to-use product. For this purpose, leaf midribs (the most used edible part of the plant) were cooked by boiling, steaming, and ‘sous vide’, and the cooked products were compared for their phenolic content and composition, antioxidant activity, sugar and inorganic ion content, organoleptic characteristics, and microbial safety, this latter also during storage. In general, boiling caused a decrease in the value of these parameters, despite being the best product for taste and overall acceptability. On the contrary, steaming and ‘sous vide’ resulted in the best treatments to preserve antioxidant activity, total phenols, and chlorogenic acid. In particular, in ‘sous vide’ cooked samples, a significant increase in the value of these parameters and a remarkable decrease in nitrate content were found. Moreover, ‘sous vide’ resulted in the best treatment also regarding microbial safety during shelf life; actually, after 15 days of storage at 8 °C, Enterobacteriaceae and mesophilic aerobic bacteria were not detectable in ‘sous vide’ samples. These results contributed to increase the knowledge of a wild edible plant with high nutritional properties and promoting its consumption by obtaining a ready-to-use product with good organoleptic characteristics and endowed with a long period of shelf life.
The increasing demand for natural, safe, and sustainable food preservation methods drove research... more The increasing demand for natural, safe, and sustainable food preservation methods drove research towards the use of plant antimicrobials as an alternative to synthetic preservatives. This review article comprehensively discussed the potential applications of plant extracts, essential oils, and their compounds as antimicrobial agents in the food industry. The antimicrobial properties of several plant-derived substances against foodborne pathogens and spoilage microorganisms, along with their modes of action, factors affecting their efficacy, and potential negative sensory impacts, were presented. The review highlighted the synergistic or additive effects displayed by combinations of plant antimicrobials, as well as the successful integration of plant extracts with food technologies ensuring an improved hurdle effect, which can enhance food safety and shelf life. The review likewise emphasized the need for further research in fields such as mode of action, optimized formulations, sensory properties, safety assessment, regulatory aspects, eco-friendly production methods, and consumer education. By addressing these gaps, plant antimicrobials can pave the way for more effective, safe, and sustainable food preservation strategies in the future.
In this study, the effect of selected Lactobacillus acidophilus ATCC 4356, Limosilactobacillus fe... more In this study, the effect of selected Lactobacillus acidophilus ATCC 4356, Limosilactobacillus fermentum DSM 20052, and Lacticaseibacillus paracasei subsp. paracasei DSM 20312 strains on the sensory characteristics, and protein and amino acid content of fermented water extracts derived from lupin, pea, and bean grains is reported. Even though all strains were able to grow over 7 log cfu mL −1 and to decrease pH in the range of −0.52 to −1.25 within 24 h, the release of an unpleasant ferricsulfurous off-odor from the fermented bean water extract prohibited further characterization. Lupin and pea grain-based beverages underwent an in-depth sensory evaluation using a simplified checkall-that-apply (CATA) method, finding new and appreciable sensory notes such as cooked ham, almonds, and sandalwood. Fermented lupin water extract showed higher total protein content (on average, 0.93 mg mL −1) in comparison to that of pea grains (on average, 0.08 mg mL −1), and a free amino acid content (on average, 3.9 mg mL −1) close to that of cow milk. The concentrations of these nutrients decreased during refrigerated storage, when the lactic acid bacteria load was always higher than 7 log cfu mL −1. The results of this study indicated that lactic fermentation improves the sensory characteristics of these innovative legume-based beverages, which sustained high loads of viable lactobacilli up to the end of cold storage.
In this work, the development, analytical characterization and bioactivity of zeolite-thymol comp... more In this work, the development, analytical characterization and bioactivity of zeolite-thymol composites, obtained using wet, semi-dry and dry processes, were carried out in order to obtain sustainable and powerful antimicrobial additives. FT-IR, XRD, DSC, TGA, SEM and B.E.T. analyses were carried out to gain comprehensive information on the chemical-physical, thermal, and morphological features of the composites. GC-MS analyses allowed quantifying the active molecule loaded in the zeolite, released by the functionalized composites and its stability over time. Among the three procedures, the dry approach allowed to reach the highest thymol loading content and efficiency (49.8 AE 1.6% and 99.6 AE 1.2%, respectively), as well as the highest composite specific surface area value, feature which promises the best interaction between the surface of the composite and the bacterial population. Therefore, the bioactive surface of composites obtained by this solvent-free method was assayed for its antimicrobial activity against four microbial strains belonging to Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa and Candida albicans species. The higher antimicrobial activity produced by the solvent-free composite in comparison with that of pure thymol, at the same thymol concentration, was ascribed to the large interfacial contact between the composite and the bacterial target. This feature, together with its enhanced storage stability, suggested that this composite could be employed as effective additives for the development of antimicrobial biointerfaces for food, home and personal care applications.
Wine pomace has attracted the attention of the food industry, due to its high content in bioactiv... more Wine pomace has attracted the attention of the food industry, due to its high content in bioactive compounds, and its multiple healthy activities. In this work, whole and separated skin pomaces from fermented (red) and un-fermented (white) grape by-products were characterized for their antioxidant and antimicrobial activities in order to exploit them as functional food ingredient. Antioxidant activity, measured by both ORAC and TEAC assays, was higher in whole than in skin pomace extracts. The characterization of phenolic composition in whole and skin pomace extracts confirmed the peculiarity of some compounds such as anthocyanins (107.84 + 10.3 mg/g TP) in red skin pomace and a great amount of flavanols (80.73 + 4.04 mg/g TP) in white skin pomace. Whole and skin pomace extracts displayed the same antibacterial activity at 250 µg gallic acid equivalents (GAE)/mL. Red and white skin pomace extracts showed a Minimum Inhibitory Concentration (MIC) of 31.25–62.5 GAE/mL against Staphylococcus aureus and Enterococcus faecalis. Pseudomonas spp. were more sensitive to red skin pomace extracts rather than white skin pomace extracts. Given these results, both red and white pomace extracts could be exploited for future application in food, pharmaceutical and cosmetic industry.
Hydrogel formulations (masks or patches, without tissue support) represent the new frontier for c... more Hydrogel formulations (masks or patches, without tissue support) represent the new frontier for customizable skin beauty and health. The employment of these materials is becoming popular in wound dressing, to speed up the healing process while protecting the affected area, as well as to provide a moisturizing reservoir, control the inflammatory process and the onset of bacterial development. Most of these hydrogels are acrylic-based at present, not biodegradable and potentially toxic, due to acrylic monomers residues. In this work, we selected a new class of cellulose-derived and biodegradable hydrogel films to incorporate and convey an active compound for dermatological issues. Films were obtained from a combination of different polysaccharides and clays, and berberine hydrochloride, a polyphenolic molecule showing anti-inflammatory, immunomodulatory, antibacterial and antioxidant properties, was chosen and then embedded in the hydrogel films. These innovative hydrogel-based systems were characterized in terms of water uptake profile, in vitro cytocompatibility and skin permeation kinetics by Franz diffusion cell. Berberine permeation fitted well to Korsmeyer–Peppas kinetic model and achieved a release higher than 100 µg/cm2 within 24 h. The latter study, exploiting a reliable skin model membrane, together with the biological assessment, gained insights into the most promising formulation for future investigations. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY)
Current trends in food packaging systems are oriented to biodegradable materials, especially with... more Current trends in food packaging systems are oriented to biodegradable materials, especially with enhanced features. The latter are often provided by the combination of natural biopolymers, clays and bioactive molecules. This review summarises recent developments in the production of biopolymers-clay hybrid materials for food packaging. The main production methods of bio-composites and the improvement of their physicochemical properties due to the clay addition are discussed. The improvement of the film properties is explained with a focus on the molecular interactions between biopolymer matrix, clay, and bioactive compounds. Recent researches on their food application as active and intelligent packaging are summerized. This review shows that the application of biodegradable bio-composites as packaging material is a promising green option to ensure food quality and safety. The research on the chemical properties of tailored and tunable biopolymers could lead to new functional films, suitable for the industrial scale-up.
The aim of this work was to evaluate the antifungal activity in vapor phase of thymol, p-cymene, ... more The aim of this work was to evaluate the antifungal activity in vapor phase of thymol, p-cymene, and γ-terpinene, the red thyme essential oil compounds (RTOCs). The Minimum Inhibitory Concentration (MIC) of RTOCs was determined against postharvest spoilage fungi of the genera Botrytis, Penicillium, Alternaria, and Monilinia, by measuring the reduction of the fungal biomass after exposure for 72 h at 25 • C. Thymol showed the lowest MIC (7.0 µg/L), followed by γ-terpinene (28.4 µg/L) and p-cymene (40.0 µg/L). In the case of P. digitatum ITEM 9569, resistant to commercial RTO, a better evaluation of interactions among RTOCs was performed using the checkerboard assay and the calculation of the Fractional Inhibitory Concentration Index (FICI). During incubation, changes in the RTOCs concentration were measured by GC-MS analysis. A synergistic effect between thymol (0.013 ± 0.003 L/L) and γ-terpinene (0.990 ± 0.030 L/L) (FICI = 0.50) in binary combinations, and between p-cymene (0.700 ± 0.010 L/L) and γ-terpinene (0.290 ± 0.010 L/L) in presence of thymol (0.008 ± 0.001 L/L) (FICI = 0.19), in ternary combinations was found. The synergistic effect against the strain P. digitatum ITEM 9569 suggests that different combinations among RTOCs could be defined to control fungal strains causing different food spoilage phenomena.
The aim of this work has been to evaluate the combined effect of modified atmosphere packaging (M... more The aim of this work has been to evaluate the combined effect of modified atmosphere packaging (MAP), and a gaseous ozone pre-treatment on the main microbiological and chemical parameters of strawberries, raspberries and blueberries during cold storage. The three types of small fruit were exposed to 13 mg m − 3 of gaseous ozone (about 6 ppm) for 16 h at 1 • C (± 0.5 • C) and then stored for 15 d under MAP (10 kPa O2 and 40 kPa CO2) or in air at 4 • C. The combined technology determined a significant (P ≤ 0.05) reduction of the yeast and mould counts, compared to samples stored in air and MAP without any pre-treatment. The main quality parameters of the three types of fruit were not affected by combined ozone pre-treatment and MAP storage. The combined effects of ozone and MAP led to a better preservation of the total and individual anthocyanins in the blueberries during cold storage. In conclusion, the exposure to gaseous ozone before packaging followed by storage under MAP could be a useful technological approach to extend the postharvest storage of small berry fruit.
This work has been aimed at studying the effect of red thyme oil (RTO, Thymus vulgaris L.) on the... more This work has been aimed at studying the effect of red thyme oil (RTO, Thymus vulgaris L.) on the shelf-life and Penicillium decay of oranges during cold storage. RTO vapours significantly reduced (P ≤ 0.05) the percentage of infected wounds, the external growth area and the production of spores in inoculated orange fruit stored for 12 days at 7°C in a polypropylene film selected for its appropriate permeability. Among the RTO compounds, p-cymene and thymol were the most abundant in packed boxes at the end of cold storage. The RTO vapours did not affect the main quality parameters of the oranges, or the taste and odour of the juice. The results have shown that an active packaging, using RTO vapours, could be employed, by the citrus industry, to extend the shelf-life of oranges for fresh market use and juice processing.
Background The application of emerging technologies for the food preservation is a relevant topic... more Background The application of emerging technologies for the food preservation is a relevant topic for the industry; these technologies are highly efficient to reduce the growth of spoilage microorganisms as well as the production of undesirable microbial metabolites in foods and the processing environment. Spoilage caused by the yeast metabolic activity affects the wine marketability. These novel technologies can be easily implemented during dynamic and customized winemaking processes, in order to control the microbial contaminations and to preserve the wine quality. Scope and approach During the last two decades, the Brettanomyces spp. contamination in the bottled wines and the winery environment has attracted interest due to production of the volatile phenol compounds, the high stress resistance and the peculiar metabolic features. The use of the emerging technologies, such as high hydrostatic pressure, ultrasounds, pulsed electric fields, UV irradiation, microwaves, ozone and electrolyzed water, novel finishing agents, and biotechnological approaches resulted useful to reduce the contamination levels and the production of off-flavours. In addition, the application of the emerging technologies allowed the reduction of sulphur dioxide, considering its allergenic properties and the raise of Brettanomyces spp. resistance to this compound. The effect of these technologies on nutritional and sensory properties of wines is also discussed. Key findings and conclusions This review highlights some useful novel approaches to control Brettanomyces spp. during winemaking, allowing the producers to follow the “hurdle concept” and to minimize wine sensory changes.
Red chicory is a leafy vegetable, currently used for the preparation of ready-to-eat salads that ... more Red chicory is a leafy vegetable, currently used for the preparation of ready-to-eat salads that undergo heavy losses during cold storage mainly due to microbial soft rot development. In this work, chicory was stored at (1°C) in a passive refrigeration chamber with low doses of gaseous ozone (0.1-1 ppm) applied during 21 days in order to reduce postharvest losses. Conventional refrigeration in absence of gaseous ozone was used as the reference storage method. In addition, in order to evaluate the antimicrobial effect a challenge test was performed inoculating sound red chicory heads with a specific spoilage microbiota, accounting approximately 8 log cfu mL-1 bacteria and 7 log cfu mL-1 yeasts and moulds, responsible for soft rot development under cold storage. The results showed that soft rot developed from day 8, especially in the inoculated samples, even though the leaf appearance was unchanged. At the end of cold storage, non-inoculated red chicory heads stored under conventional conditions lost all characteristics related to vegetable fresh appearance; on the contrary, colour brightness and tissue turgidity were retained in samples stored under passive refrigeration. Oxidative discoloration of leaf margins was recorded in red chicory heads stored under 1 ppm ozone. At the day 21, microbial counts of spoiled vegetable regardless the different cold storage conditions did not show any significant difference. Conversely, as expected, evident difference in soft rot severity was recorded between inoculated and non-inoculated samples. Passive refrigeration determined a lower amount of food waste in comparison with conventional refrigeration. Control samples, inoculated or not, showed 100% of losses whereas those under passive refrigeration produced 40.09±4.24 and 90.69±12.03% of food waste for inoculated and non-inoculated samples, respectively. A further reduction of vegetable waste (ranging from 10 to 30%) was registered when passive refrigeration was associated with gaseous ozone treatment at 0.1 ppm. Higher ozone doses caused no significant additional improvements. Passive refrigeration alone was able to reduce significantly weight losses and preserve the quality of chicory up to 21 days. In addition, the combined application of gaseous ozone and passive refrigeration resulted a valid approach to counteract soft rot development on red chicory heads and to reduce the amount of food waste. However, ozone exposure needs to be carefully calibrated to preserve vegetable tissues from oxidative damages caused by this gas.
The present study was designed to evaluate possible sugar-based trophic interactions between acet... more The present study was designed to evaluate possible sugar-based trophic interactions between acetic acid bacteria (AAB) and non-Saccharomyces yeasts (NSY) involved in table grape sour rot, a disease in which berries spoilage is caused by the accumulation of several microbial metabolites. Acetobacter syzygii LMG 21419 (As) and Candida zemplinina CBS 9494 (Cz), a simplified AAB-NSY association responsible for table grape sour rot, grew differently in a minimal medium (YP) supplemented with glucose, ethanol, acetic and gluconic acid under monoculture conditions. In As-Cz co-culture media, after 24 h of incubation, As showed high relative abundance in YP-ethanol, whereas Cz was the dominant strain in YP-glucose medium. Co-culture in YP-glucose showed that glucose was converted into ethanol by Cz that, in turn, promoted the growth of As population. Gluconic acid was the main bacterial metabolite from glucose in monoculture, whereas acetic acid putatively derived from ethanol oxidation was found only in co-culture. However, gluconic acid showed inhibitory effect against As whereas acetic acid mainly inhibited Cz. Negative effects of both metabolites were mitigated in the glucose-supplemented medium. The results suggest a possible metabolic-based temporal succession between AAB and NSY during grape sour rot development. At the begin of sour rot, low glucose concentration promotes NSY producing ethanol, then, the AAB could take advantage from the oxidation of ethanol into acetic acid, becoming the dominant microbial sour rot population during the late stages of the process.
Ready-to-eat asparagus (Asparagus officinalis L.) is a tasty food with excellent nutraceutical pr... more Ready-to-eat asparagus (Asparagus officinalis L.) is a tasty food with excellent nutraceutical properties. In order to realize a new ready-to-eat product, in this study asparagus's spears were cooked by blanching or microwave, and then dehydrated until they reached a weight loss of 25%, and packaged in air or in modified atmosphere (30% CO 2 + 70% N 2). Sensorial, physico-chemical, biochemical, and microbiological parameters were evaluated during a 30 days storage period at 4 °C. The microwave cooking proved to be the most effective method to preserve green colour, improving the overall acceptability of the product. Moreover, the storage in the absence of O 2 and in the presence of high CO 2 percentage was the most effective method to preserve phytochemical composition, total antioxidant capacity, and hygienic quality. In conclusion, asparagus spears cooked by microwave , semi-dried, packaged in modified atmosphere and stored at 4 °C for 30 days retained their quality and sensorial properties.
Washing water used for processing fruits and vegetables can convey spoilage fungi and bacteria. T... more Washing water used for processing fruits and vegetables can convey spoilage fungi and bacteria. The common procedure to reduce microbial contamination involves the use of chlorine based compounds. Recently, electrolyzed water (EW) has been evaluated as an alternative measure in controlling microbial spoilage contamination occurring during washing steps. This work reviews results related to the application of EW for controlling microbial viability responsible for decay development during storage period. EW produced with sodium bicarbonate as electrolyte reduced Penicillium spp. population in water and, consequently, green mould decay in citrus fruits; the use of sodium chloride in EW production inactivated spores of Fusarium sp. in water and reduced pineapple decay during storage at 12°C for 20 days as well as controlled yeast and mould population in date fruit up to six months of cold storage. EW was also found effective in controlling spoilage bacteria on ready-to-eat produce. Pseudomonas fluorescens, Pantoea agglomerans, and Rhanella aquatilis were undetectable in electrolyzed process water amended with sodium chloride although similar treatment slightly reduced Erwinia carotovora load inoculated onto lettuce. EW at low free chlorine concentration reduced viability of Pseudomonas spp. and psychrotrophic bacteria in both simulated and industrial washing water. EW treatment of fresh cut lettuce dipped in microbial contaminated water reduced Pseudomonas spp. of about 1 log cfu g-1 delaying spoilage symptoms that occurred early in untreated vegetables. These results demonstrate that the use of EW can control spoilage microorganisms in washing water, reduce cross-contamination phenomena and delay fruit and vegetable decay.
In the present study, we evaluated the antimicrobial activity of neutral electrolyzed water (NEW)... more In the present study, we evaluated the antimicrobial activity of neutral electrolyzed water (NEW) against 14 strains of spoilage Pseudomonas of fresh cut vegetables under cold storage. The NEW, produced from solutions of potassium and sodium chloride, and sodium bicarbonate developed up to 4000 mg/L of free chlorine, depending on the salt and relative concentration used. The antimicrobial effect of the NEW was evaluated against different bacterial strains at 10 5 cells/ml, with different combinations of free chlorine concentration/contact time; all concentrations above 100 mg/L, regardless of the salt used, were found to be bactericidal already after 2 min. When catalogna chicory and lettuce leaves were dipped for 5 min in diluted NEW, microbial loads of mesophilic bacteria and Enterobacteriaceae were reduced on average of 1.7 log cfu/g. In addition, when lettuce leaves were dipped in a cellular suspension of the spoiler Pseu-domonas chicorii I3C strain, diluted NEW was able to reduce Pseudomonas population of about 1.0 log cfu/ g. Thanks to its high antimicrobial activity against spoilage microorganisms, and low cost of operation, the application of cycles of electrolysis to the washing water looks as an effective tool in controlling fresh cut vegetable microbial spoilage contamination occurring during washing steps.
This work aims at studying the efficacy of low doses of gaseous ozone in postharvest control of t... more This work aims at studying the efficacy of low doses of gaseous ozone in postharvest control of the table grape sour rot, a disease generally attributed to a consortium of non-Saccharomyces yeasts (NSY) and acetic acid bacteria (AAB). Sour rot incidence of wounded berries, inoculated with 8 NSYstrains, or 7 AAB, or 56 yeast-bacterium associations, was monitored at 25 °C up to six days. Sour rot incidence in wounded berries inoculated with yeast-bacterium associations resulted higher than in berries inoculated with one single NSY or AAB strain. Among all NSY-AAB associations, the yeast-bacterium association composed of Candida zemplinina CBS 9494 (Cz) and Acetobacter syzygii LMG 21419 (As) showed the highest prevalence of sour rot; thus, after preliminary in vitro assays, this simplified As-Cz microbial consortium was inoculated in wounded berries that were stored at 4 °C for ten days under ozone (2.14 mg m−3) or in air. At the end of cold storage, no berries showed sour-rot symptoms although ozonation mainly affected As viable cell count. After additional 12 days at 25 °C, the sour rot index of inoculated As-Cz berries previously cold-stored under ozone or in air accounted for 22.6 ± 3.7% and 66.7 ± 4.5%, respectively. Molecular analyses of dominant AAB and NSY populations of both sound and rotten berries during post-refrigeration period revealed the appearance of new strains mainly belonging to Gluconobacter albidus and Hanseniaspora uvarum species, respectively. Cold ozonation resulted an effective approach to extend the shelf-life of table grapes also after cold storage.
The microbial content of plant tissues has been reported to cause the spoilage of ca. 30% of chlo... more The microbial content of plant tissues has been reported to cause the spoilage of ca. 30% of chlorine-disinfected fresh vegetables during cold storage. The aim of this work was to evaluate the efficacy of antimicrobial peptides in controlling microbial vegetable spoilage under cold storage conditions. A total of 48 bacterial isolates were collected from ready-to-eat (RTE) vegetables and identified as belonging to Acinetobacter calcoaceticus, Aeromonas media, Pseudomonas cichorii, Pseudomonas fluorescens, Pseudomonas jessenii, Pseudomonas koreensis, Pseudomonas putida, Pseudomonas simiae and Pseudomonas viridiflava species. Reddish or brownish pigmentation was found when Pseudomonas strains were inoculated in wounds on leaves of Iceberg and Trocadero lettuce and escarole chicory throughout cold storage. Bovine lactoferrin (BLF) and its hydrolysates (LFHs) produced by pepsin, papain and rennin, were assayed in vitro against four Pseudomonas spp. strains selected for their heavy spoiling ability. As the pepsin-LFH showed the strongest antimicrobial effect, subsequent experiments were carried out using the peptide lactoferricin B (LfcinB), well known to be responsible for its antimicrobial activity. LfcinB significantly reduced (P ≤ 0.05) spoilage by a mean of 36% caused by three out of four inoculated spoiler pseudomonads on RTE lettuce leaves after six days of cold storage. The reduction in the extent of spoilage was unrelated to viable cell density in the inoculated wounds. This is the first paper providing direct evidence regarding the application of an antimicrobial peptide to control microbial spoilage affecting RTE leafy vegetables during cold storage.
World Journal of Microbiology and Biotechnology, 2023
Onion skins, actually recycled as organic fertilizers, could be used as a substrate in environmen... more Onion skins, actually recycled as organic fertilizers, could be used as a substrate in environmental-friendly bioprocesses to recover high-value bioactive compounds and food ingredients. In this work, a bioprospecting method was carried out including 94 bacterial and 45 yeast strains from several agri-food and environmental niches to verify their ability to grow on onion skins as unique nutrients source. Red and yellow onion skins were assessed by newly selected starter-driven liquid submerged fermentation assays mainly aimed at the release and modification of polyphenols through microbial activities. Fermented onion skins were also investigated as a inexpensive favourable source of microbial enzymes (amylases, proteases, lipases, esterases, cellulases, xylanases). In red onion skins, the treatment with Lactiplantibacillus plantarum TB 11-32 produced a slight increase of bioactive compounds in terms of total phenolics, whereas with the yeast strain Zygosaccharomyces mrakii CL 30 − 29 the quercetin aglycone content increased of about 25% of the initial raw material. In yellow onion skins inoculated, the highest content of phenolic compounds was detected with the yeast strain Saccharomyces cerevisiae En SC, while quercetin aglycone increased of about 60% of the initial raw material in presence of the bacterial strain L. plantarum C 180 − 34. In conclusion, the proposed microbial pre-treatment method can be a potential strategy to re-use onion skins as a fermentation substrate, and as a first step in the development of a biorefinery process to produce value-added products from onion by-products.
Golden thistle (Scolymus hispanicus L.) is a wild edible plant belonging to Asteraceae family, wi... more Golden thistle (Scolymus hispanicus L.) is a wild edible plant belonging to Asteraceae family, with a great potential for food applications. The aim of this study was to identify the best cooking procedure able to provide a high-quality, ready-to-use product. For this purpose, leaf midribs (the most used edible part of the plant) were cooked by boiling, steaming, and ‘sous vide’, and the cooked products were compared for their phenolic content and composition, antioxidant activity, sugar and inorganic ion content, organoleptic characteristics, and microbial safety, this latter also during storage. In general, boiling caused a decrease in the value of these parameters, despite being the best product for taste and overall acceptability. On the contrary, steaming and ‘sous vide’ resulted in the best treatments to preserve antioxidant activity, total phenols, and chlorogenic acid. In particular, in ‘sous vide’ cooked samples, a significant increase in the value of these parameters and a remarkable decrease in nitrate content were found. Moreover, ‘sous vide’ resulted in the best treatment also regarding microbial safety during shelf life; actually, after 15 days of storage at 8 °C, Enterobacteriaceae and mesophilic aerobic bacteria were not detectable in ‘sous vide’ samples. These results contributed to increase the knowledge of a wild edible plant with high nutritional properties and promoting its consumption by obtaining a ready-to-use product with good organoleptic characteristics and endowed with a long period of shelf life.
The increasing demand for natural, safe, and sustainable food preservation methods drove research... more The increasing demand for natural, safe, and sustainable food preservation methods drove research towards the use of plant antimicrobials as an alternative to synthetic preservatives. This review article comprehensively discussed the potential applications of plant extracts, essential oils, and their compounds as antimicrobial agents in the food industry. The antimicrobial properties of several plant-derived substances against foodborne pathogens and spoilage microorganisms, along with their modes of action, factors affecting their efficacy, and potential negative sensory impacts, were presented. The review highlighted the synergistic or additive effects displayed by combinations of plant antimicrobials, as well as the successful integration of plant extracts with food technologies ensuring an improved hurdle effect, which can enhance food safety and shelf life. The review likewise emphasized the need for further research in fields such as mode of action, optimized formulations, sensory properties, safety assessment, regulatory aspects, eco-friendly production methods, and consumer education. By addressing these gaps, plant antimicrobials can pave the way for more effective, safe, and sustainable food preservation strategies in the future.
In this study, the effect of selected Lactobacillus acidophilus ATCC 4356, Limosilactobacillus fe... more In this study, the effect of selected Lactobacillus acidophilus ATCC 4356, Limosilactobacillus fermentum DSM 20052, and Lacticaseibacillus paracasei subsp. paracasei DSM 20312 strains on the sensory characteristics, and protein and amino acid content of fermented water extracts derived from lupin, pea, and bean grains is reported. Even though all strains were able to grow over 7 log cfu mL −1 and to decrease pH in the range of −0.52 to −1.25 within 24 h, the release of an unpleasant ferricsulfurous off-odor from the fermented bean water extract prohibited further characterization. Lupin and pea grain-based beverages underwent an in-depth sensory evaluation using a simplified checkall-that-apply (CATA) method, finding new and appreciable sensory notes such as cooked ham, almonds, and sandalwood. Fermented lupin water extract showed higher total protein content (on average, 0.93 mg mL −1) in comparison to that of pea grains (on average, 0.08 mg mL −1), and a free amino acid content (on average, 3.9 mg mL −1) close to that of cow milk. The concentrations of these nutrients decreased during refrigerated storage, when the lactic acid bacteria load was always higher than 7 log cfu mL −1. The results of this study indicated that lactic fermentation improves the sensory characteristics of these innovative legume-based beverages, which sustained high loads of viable lactobacilli up to the end of cold storage.
In this work, the development, analytical characterization and bioactivity of zeolite-thymol comp... more In this work, the development, analytical characterization and bioactivity of zeolite-thymol composites, obtained using wet, semi-dry and dry processes, were carried out in order to obtain sustainable and powerful antimicrobial additives. FT-IR, XRD, DSC, TGA, SEM and B.E.T. analyses were carried out to gain comprehensive information on the chemical-physical, thermal, and morphological features of the composites. GC-MS analyses allowed quantifying the active molecule loaded in the zeolite, released by the functionalized composites and its stability over time. Among the three procedures, the dry approach allowed to reach the highest thymol loading content and efficiency (49.8 AE 1.6% and 99.6 AE 1.2%, respectively), as well as the highest composite specific surface area value, feature which promises the best interaction between the surface of the composite and the bacterial population. Therefore, the bioactive surface of composites obtained by this solvent-free method was assayed for its antimicrobial activity against four microbial strains belonging to Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa and Candida albicans species. The higher antimicrobial activity produced by the solvent-free composite in comparison with that of pure thymol, at the same thymol concentration, was ascribed to the large interfacial contact between the composite and the bacterial target. This feature, together with its enhanced storage stability, suggested that this composite could be employed as effective additives for the development of antimicrobial biointerfaces for food, home and personal care applications.
Wine pomace has attracted the attention of the food industry, due to its high content in bioactiv... more Wine pomace has attracted the attention of the food industry, due to its high content in bioactive compounds, and its multiple healthy activities. In this work, whole and separated skin pomaces from fermented (red) and un-fermented (white) grape by-products were characterized for their antioxidant and antimicrobial activities in order to exploit them as functional food ingredient. Antioxidant activity, measured by both ORAC and TEAC assays, was higher in whole than in skin pomace extracts. The characterization of phenolic composition in whole and skin pomace extracts confirmed the peculiarity of some compounds such as anthocyanins (107.84 + 10.3 mg/g TP) in red skin pomace and a great amount of flavanols (80.73 + 4.04 mg/g TP) in white skin pomace. Whole and skin pomace extracts displayed the same antibacterial activity at 250 µg gallic acid equivalents (GAE)/mL. Red and white skin pomace extracts showed a Minimum Inhibitory Concentration (MIC) of 31.25–62.5 GAE/mL against Staphylococcus aureus and Enterococcus faecalis. Pseudomonas spp. were more sensitive to red skin pomace extracts rather than white skin pomace extracts. Given these results, both red and white pomace extracts could be exploited for future application in food, pharmaceutical and cosmetic industry.
Hydrogel formulations (masks or patches, without tissue support) represent the new frontier for c... more Hydrogel formulations (masks or patches, without tissue support) represent the new frontier for customizable skin beauty and health. The employment of these materials is becoming popular in wound dressing, to speed up the healing process while protecting the affected area, as well as to provide a moisturizing reservoir, control the inflammatory process and the onset of bacterial development. Most of these hydrogels are acrylic-based at present, not biodegradable and potentially toxic, due to acrylic monomers residues. In this work, we selected a new class of cellulose-derived and biodegradable hydrogel films to incorporate and convey an active compound for dermatological issues. Films were obtained from a combination of different polysaccharides and clays, and berberine hydrochloride, a polyphenolic molecule showing anti-inflammatory, immunomodulatory, antibacterial and antioxidant properties, was chosen and then embedded in the hydrogel films. These innovative hydrogel-based systems were characterized in terms of water uptake profile, in vitro cytocompatibility and skin permeation kinetics by Franz diffusion cell. Berberine permeation fitted well to Korsmeyer–Peppas kinetic model and achieved a release higher than 100 µg/cm2 within 24 h. The latter study, exploiting a reliable skin model membrane, together with the biological assessment, gained insights into the most promising formulation for future investigations. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY)
Current trends in food packaging systems are oriented to biodegradable materials, especially with... more Current trends in food packaging systems are oriented to biodegradable materials, especially with enhanced features. The latter are often provided by the combination of natural biopolymers, clays and bioactive molecules. This review summarises recent developments in the production of biopolymers-clay hybrid materials for food packaging. The main production methods of bio-composites and the improvement of their physicochemical properties due to the clay addition are discussed. The improvement of the film properties is explained with a focus on the molecular interactions between biopolymer matrix, clay, and bioactive compounds. Recent researches on their food application as active and intelligent packaging are summerized. This review shows that the application of biodegradable bio-composites as packaging material is a promising green option to ensure food quality and safety. The research on the chemical properties of tailored and tunable biopolymers could lead to new functional films, suitable for the industrial scale-up.
The aim of this work was to evaluate the antifungal activity in vapor phase of thymol, p-cymene, ... more The aim of this work was to evaluate the antifungal activity in vapor phase of thymol, p-cymene, and γ-terpinene, the red thyme essential oil compounds (RTOCs). The Minimum Inhibitory Concentration (MIC) of RTOCs was determined against postharvest spoilage fungi of the genera Botrytis, Penicillium, Alternaria, and Monilinia, by measuring the reduction of the fungal biomass after exposure for 72 h at 25 • C. Thymol showed the lowest MIC (7.0 µg/L), followed by γ-terpinene (28.4 µg/L) and p-cymene (40.0 µg/L). In the case of P. digitatum ITEM 9569, resistant to commercial RTO, a better evaluation of interactions among RTOCs was performed using the checkerboard assay and the calculation of the Fractional Inhibitory Concentration Index (FICI). During incubation, changes in the RTOCs concentration were measured by GC-MS analysis. A synergistic effect between thymol (0.013 ± 0.003 L/L) and γ-terpinene (0.990 ± 0.030 L/L) (FICI = 0.50) in binary combinations, and between p-cymene (0.700 ± 0.010 L/L) and γ-terpinene (0.290 ± 0.010 L/L) in presence of thymol (0.008 ± 0.001 L/L) (FICI = 0.19), in ternary combinations was found. The synergistic effect against the strain P. digitatum ITEM 9569 suggests that different combinations among RTOCs could be defined to control fungal strains causing different food spoilage phenomena.
The aim of this work has been to evaluate the combined effect of modified atmosphere packaging (M... more The aim of this work has been to evaluate the combined effect of modified atmosphere packaging (MAP), and a gaseous ozone pre-treatment on the main microbiological and chemical parameters of strawberries, raspberries and blueberries during cold storage. The three types of small fruit were exposed to 13 mg m − 3 of gaseous ozone (about 6 ppm) for 16 h at 1 • C (± 0.5 • C) and then stored for 15 d under MAP (10 kPa O2 and 40 kPa CO2) or in air at 4 • C. The combined technology determined a significant (P ≤ 0.05) reduction of the yeast and mould counts, compared to samples stored in air and MAP without any pre-treatment. The main quality parameters of the three types of fruit were not affected by combined ozone pre-treatment and MAP storage. The combined effects of ozone and MAP led to a better preservation of the total and individual anthocyanins in the blueberries during cold storage. In conclusion, the exposure to gaseous ozone before packaging followed by storage under MAP could be a useful technological approach to extend the postharvest storage of small berry fruit.
This work has been aimed at studying the effect of red thyme oil (RTO, Thymus vulgaris L.) on the... more This work has been aimed at studying the effect of red thyme oil (RTO, Thymus vulgaris L.) on the shelf-life and Penicillium decay of oranges during cold storage. RTO vapours significantly reduced (P ≤ 0.05) the percentage of infected wounds, the external growth area and the production of spores in inoculated orange fruit stored for 12 days at 7°C in a polypropylene film selected for its appropriate permeability. Among the RTO compounds, p-cymene and thymol were the most abundant in packed boxes at the end of cold storage. The RTO vapours did not affect the main quality parameters of the oranges, or the taste and odour of the juice. The results have shown that an active packaging, using RTO vapours, could be employed, by the citrus industry, to extend the shelf-life of oranges for fresh market use and juice processing.
Background The application of emerging technologies for the food preservation is a relevant topic... more Background The application of emerging technologies for the food preservation is a relevant topic for the industry; these technologies are highly efficient to reduce the growth of spoilage microorganisms as well as the production of undesirable microbial metabolites in foods and the processing environment. Spoilage caused by the yeast metabolic activity affects the wine marketability. These novel technologies can be easily implemented during dynamic and customized winemaking processes, in order to control the microbial contaminations and to preserve the wine quality. Scope and approach During the last two decades, the Brettanomyces spp. contamination in the bottled wines and the winery environment has attracted interest due to production of the volatile phenol compounds, the high stress resistance and the peculiar metabolic features. The use of the emerging technologies, such as high hydrostatic pressure, ultrasounds, pulsed electric fields, UV irradiation, microwaves, ozone and electrolyzed water, novel finishing agents, and biotechnological approaches resulted useful to reduce the contamination levels and the production of off-flavours. In addition, the application of the emerging technologies allowed the reduction of sulphur dioxide, considering its allergenic properties and the raise of Brettanomyces spp. resistance to this compound. The effect of these technologies on nutritional and sensory properties of wines is also discussed. Key findings and conclusions This review highlights some useful novel approaches to control Brettanomyces spp. during winemaking, allowing the producers to follow the “hurdle concept” and to minimize wine sensory changes.
Red chicory is a leafy vegetable, currently used for the preparation of ready-to-eat salads that ... more Red chicory is a leafy vegetable, currently used for the preparation of ready-to-eat salads that undergo heavy losses during cold storage mainly due to microbial soft rot development. In this work, chicory was stored at (1°C) in a passive refrigeration chamber with low doses of gaseous ozone (0.1-1 ppm) applied during 21 days in order to reduce postharvest losses. Conventional refrigeration in absence of gaseous ozone was used as the reference storage method. In addition, in order to evaluate the antimicrobial effect a challenge test was performed inoculating sound red chicory heads with a specific spoilage microbiota, accounting approximately 8 log cfu mL-1 bacteria and 7 log cfu mL-1 yeasts and moulds, responsible for soft rot development under cold storage. The results showed that soft rot developed from day 8, especially in the inoculated samples, even though the leaf appearance was unchanged. At the end of cold storage, non-inoculated red chicory heads stored under conventional conditions lost all characteristics related to vegetable fresh appearance; on the contrary, colour brightness and tissue turgidity were retained in samples stored under passive refrigeration. Oxidative discoloration of leaf margins was recorded in red chicory heads stored under 1 ppm ozone. At the day 21, microbial counts of spoiled vegetable regardless the different cold storage conditions did not show any significant difference. Conversely, as expected, evident difference in soft rot severity was recorded between inoculated and non-inoculated samples. Passive refrigeration determined a lower amount of food waste in comparison with conventional refrigeration. Control samples, inoculated or not, showed 100% of losses whereas those under passive refrigeration produced 40.09±4.24 and 90.69±12.03% of food waste for inoculated and non-inoculated samples, respectively. A further reduction of vegetable waste (ranging from 10 to 30%) was registered when passive refrigeration was associated with gaseous ozone treatment at 0.1 ppm. Higher ozone doses caused no significant additional improvements. Passive refrigeration alone was able to reduce significantly weight losses and preserve the quality of chicory up to 21 days. In addition, the combined application of gaseous ozone and passive refrigeration resulted a valid approach to counteract soft rot development on red chicory heads and to reduce the amount of food waste. However, ozone exposure needs to be carefully calibrated to preserve vegetable tissues from oxidative damages caused by this gas.
The present study was designed to evaluate possible sugar-based trophic interactions between acet... more The present study was designed to evaluate possible sugar-based trophic interactions between acetic acid bacteria (AAB) and non-Saccharomyces yeasts (NSY) involved in table grape sour rot, a disease in which berries spoilage is caused by the accumulation of several microbial metabolites. Acetobacter syzygii LMG 21419 (As) and Candida zemplinina CBS 9494 (Cz), a simplified AAB-NSY association responsible for table grape sour rot, grew differently in a minimal medium (YP) supplemented with glucose, ethanol, acetic and gluconic acid under monoculture conditions. In As-Cz co-culture media, after 24 h of incubation, As showed high relative abundance in YP-ethanol, whereas Cz was the dominant strain in YP-glucose medium. Co-culture in YP-glucose showed that glucose was converted into ethanol by Cz that, in turn, promoted the growth of As population. Gluconic acid was the main bacterial metabolite from glucose in monoculture, whereas acetic acid putatively derived from ethanol oxidation was found only in co-culture. However, gluconic acid showed inhibitory effect against As whereas acetic acid mainly inhibited Cz. Negative effects of both metabolites were mitigated in the glucose-supplemented medium. The results suggest a possible metabolic-based temporal succession between AAB and NSY during grape sour rot development. At the begin of sour rot, low glucose concentration promotes NSY producing ethanol, then, the AAB could take advantage from the oxidation of ethanol into acetic acid, becoming the dominant microbial sour rot population during the late stages of the process.
Ready-to-eat asparagus (Asparagus officinalis L.) is a tasty food with excellent nutraceutical pr... more Ready-to-eat asparagus (Asparagus officinalis L.) is a tasty food with excellent nutraceutical properties. In order to realize a new ready-to-eat product, in this study asparagus's spears were cooked by blanching or microwave, and then dehydrated until they reached a weight loss of 25%, and packaged in air or in modified atmosphere (30% CO 2 + 70% N 2). Sensorial, physico-chemical, biochemical, and microbiological parameters were evaluated during a 30 days storage period at 4 °C. The microwave cooking proved to be the most effective method to preserve green colour, improving the overall acceptability of the product. Moreover, the storage in the absence of O 2 and in the presence of high CO 2 percentage was the most effective method to preserve phytochemical composition, total antioxidant capacity, and hygienic quality. In conclusion, asparagus spears cooked by microwave , semi-dried, packaged in modified atmosphere and stored at 4 °C for 30 days retained their quality and sensorial properties.
Washing water used for processing fruits and vegetables can convey spoilage fungi and bacteria. T... more Washing water used for processing fruits and vegetables can convey spoilage fungi and bacteria. The common procedure to reduce microbial contamination involves the use of chlorine based compounds. Recently, electrolyzed water (EW) has been evaluated as an alternative measure in controlling microbial spoilage contamination occurring during washing steps. This work reviews results related to the application of EW for controlling microbial viability responsible for decay development during storage period. EW produced with sodium bicarbonate as electrolyte reduced Penicillium spp. population in water and, consequently, green mould decay in citrus fruits; the use of sodium chloride in EW production inactivated spores of Fusarium sp. in water and reduced pineapple decay during storage at 12°C for 20 days as well as controlled yeast and mould population in date fruit up to six months of cold storage. EW was also found effective in controlling spoilage bacteria on ready-to-eat produce. Pseudomonas fluorescens, Pantoea agglomerans, and Rhanella aquatilis were undetectable in electrolyzed process water amended with sodium chloride although similar treatment slightly reduced Erwinia carotovora load inoculated onto lettuce. EW at low free chlorine concentration reduced viability of Pseudomonas spp. and psychrotrophic bacteria in both simulated and industrial washing water. EW treatment of fresh cut lettuce dipped in microbial contaminated water reduced Pseudomonas spp. of about 1 log cfu g-1 delaying spoilage symptoms that occurred early in untreated vegetables. These results demonstrate that the use of EW can control spoilage microorganisms in washing water, reduce cross-contamination phenomena and delay fruit and vegetable decay.
In the present study, we evaluated the antimicrobial activity of neutral electrolyzed water (NEW)... more In the present study, we evaluated the antimicrobial activity of neutral electrolyzed water (NEW) against 14 strains of spoilage Pseudomonas of fresh cut vegetables under cold storage. The NEW, produced from solutions of potassium and sodium chloride, and sodium bicarbonate developed up to 4000 mg/L of free chlorine, depending on the salt and relative concentration used. The antimicrobial effect of the NEW was evaluated against different bacterial strains at 10 5 cells/ml, with different combinations of free chlorine concentration/contact time; all concentrations above 100 mg/L, regardless of the salt used, were found to be bactericidal already after 2 min. When catalogna chicory and lettuce leaves were dipped for 5 min in diluted NEW, microbial loads of mesophilic bacteria and Enterobacteriaceae were reduced on average of 1.7 log cfu/g. In addition, when lettuce leaves were dipped in a cellular suspension of the spoiler Pseu-domonas chicorii I3C strain, diluted NEW was able to reduce Pseudomonas population of about 1.0 log cfu/ g. Thanks to its high antimicrobial activity against spoilage microorganisms, and low cost of operation, the application of cycles of electrolysis to the washing water looks as an effective tool in controlling fresh cut vegetable microbial spoilage contamination occurring during washing steps.
This work aims at studying the efficacy of low doses of gaseous ozone in postharvest control of t... more This work aims at studying the efficacy of low doses of gaseous ozone in postharvest control of the table grape sour rot, a disease generally attributed to a consortium of non-Saccharomyces yeasts (NSY) and acetic acid bacteria (AAB). Sour rot incidence of wounded berries, inoculated with 8 NSYstrains, or 7 AAB, or 56 yeast-bacterium associations, was monitored at 25 °C up to six days. Sour rot incidence in wounded berries inoculated with yeast-bacterium associations resulted higher than in berries inoculated with one single NSY or AAB strain. Among all NSY-AAB associations, the yeast-bacterium association composed of Candida zemplinina CBS 9494 (Cz) and Acetobacter syzygii LMG 21419 (As) showed the highest prevalence of sour rot; thus, after preliminary in vitro assays, this simplified As-Cz microbial consortium was inoculated in wounded berries that were stored at 4 °C for ten days under ozone (2.14 mg m−3) or in air. At the end of cold storage, no berries showed sour-rot symptoms although ozonation mainly affected As viable cell count. After additional 12 days at 25 °C, the sour rot index of inoculated As-Cz berries previously cold-stored under ozone or in air accounted for 22.6 ± 3.7% and 66.7 ± 4.5%, respectively. Molecular analyses of dominant AAB and NSY populations of both sound and rotten berries during post-refrigeration period revealed the appearance of new strains mainly belonging to Gluconobacter albidus and Hanseniaspora uvarum species, respectively. Cold ozonation resulted an effective approach to extend the shelf-life of table grapes also after cold storage.
The microbial content of plant tissues has been reported to cause the spoilage of ca. 30% of chlo... more The microbial content of plant tissues has been reported to cause the spoilage of ca. 30% of chlorine-disinfected fresh vegetables during cold storage. The aim of this work was to evaluate the efficacy of antimicrobial peptides in controlling microbial vegetable spoilage under cold storage conditions. A total of 48 bacterial isolates were collected from ready-to-eat (RTE) vegetables and identified as belonging to Acinetobacter calcoaceticus, Aeromonas media, Pseudomonas cichorii, Pseudomonas fluorescens, Pseudomonas jessenii, Pseudomonas koreensis, Pseudomonas putida, Pseudomonas simiae and Pseudomonas viridiflava species. Reddish or brownish pigmentation was found when Pseudomonas strains were inoculated in wounds on leaves of Iceberg and Trocadero lettuce and escarole chicory throughout cold storage. Bovine lactoferrin (BLF) and its hydrolysates (LFHs) produced by pepsin, papain and rennin, were assayed in vitro against four Pseudomonas spp. strains selected for their heavy spoiling ability. As the pepsin-LFH showed the strongest antimicrobial effect, subsequent experiments were carried out using the peptide lactoferricin B (LfcinB), well known to be responsible for its antimicrobial activity. LfcinB significantly reduced (P ≤ 0.05) spoilage by a mean of 36% caused by three out of four inoculated spoiler pseudomonads on RTE lettuce leaves after six days of cold storage. The reduction in the extent of spoilage was unrelated to viable cell density in the inoculated wounds. This is the first paper providing direct evidence regarding the application of an antimicrobial peptide to control microbial spoilage affecting RTE leafy vegetables during cold storage.
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This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY)
This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY)