Use of entrapped microorganisms as biological oxygen scavengers in food packaging applications

In this work a new method is proposed to produce oxygen-scavenger films using aerobic microorganisms as the “active compound”. The manufacturing cycle of the investigated oxygen-scavenger film was optimized both to prolong the microorganisms viability during storage and to improve the efficiency of the film to remove oxygen from the package headspace. It was found that it is possible to store the desiccated film over a period of 20 days without monitoring any appreciable decrease of microorganism viability. It was also pointed out that the highest respiratory efficiency of the proposed active film is obtained by entrapping the microorganisms into polyvinyl alcohol, and by using the active film as a coating for a high humidity food.     

The performance of several oxygen scavengers in varying oxygen environments at refrigerated temperatures: implications for low-oxygen modified atmosphere packaging of meat

The oxygen scavenging capacity of four commercially available iron-based oxygen scavengers was studied. Individual oxygen scavenger sachets were placed in pouches and filled with 1%, 2%, 6%, 12% or 22% oxygen, 40% carbon dioxide and balance nitrogen, and stored at 3 °C or 10 °C, with or without a drip pad infused with water and monitored over 24 h. The four scavengers all reduced oxygen from the packs at the oxygen concentrations and temperatures tested. However, for all of the conditions measured, the scavengers did not absorb their nominal capacity in the 24-h period. In anoxic modified atmosphere packaging of beef steaks, it is essential to reduce residual oxygen levels to below 0.05% as quickly as possible to minimise the formation of metmyoglobin. While the scavengers tested were effective in removing oxygen, the rate of removal would appear not to be fast enough to create the anoxic conditions required to prevent metmyoglobin formation in beef steaks, particularly in those cuts, which are highly susceptible to metmyoglobin formation. Reproducibility was also a critical issue for the scavengers, particularly at low oxygen concentrations. None of the scavengers had a coefficient of variation of less than 20% at the low oxygen concentrations. Therefore, to obtain consistent results, it is recommended that multiple scavengers be used.     

Extension of the display life of lamb with an antioxidant active packaging

Fresh lamb steaks were treated with three different preparations of natural antioxidants: one group was packaged with a rosemary active film, the second group was packaged with an oregano active film, and the third group was sprayed on the meat surface with a rosemary extract before packaging in a high-oxygen atmosphere. Samples were stored under illumination at 1 ± 1 °C for 13 days. Metmyoglobin formation, lipid oxidation (TBARS), instrumental colour (CIE a^*), psychrotrophic bacterial counts (PCA), sensory discolouration and off-odour were determined. The use of a rosemary extract, a rosemary active film or an oregano active film resulted in enhanced oxidative stability of lamb steaks. Active films with oregano were significantly more efficient than those with rosemary, exerting an effect similar to that of direct addition of the rosemary extract; in fact, they extended fresh odour and colour from 8 to 13 days compared to the control.     

The fate of Clostridium perfringens spores exposed to ozone and/or mild heat pretreatment on beef surfaces followed by modified atmosphere packaging

Clostridium perfringens is a natural contaminant of raw beef products that can proliferate to dangerous cell levels under conditions of temperature abuse. Spores of the bacterium were inoculated onto irradiated London broil beef at levels of 3 log₁₀ spores/g beef. Samples of beef (7.5×10.0×1.0 cm) were treated with aqueous ozone (5 ppm O₃ for 5 min), or heat (60°C for 30 min), or both and then vacuum-packaged to 2 kPa for up to 10 d storage at 37°C, 25°C, or 4°C. Storage at 37°C resulted in increases in viable counts after 1 d to over 7 log₁₀ cfu/g beef, whereas storage at 4°C prevented spore germination and growth for all treatments. At 25°C, heat-treated beef samples reached 6 log₁₀ cfu/g viable counts in 2 d and spores/vegetative cells on control or ozone-treated samples did not germinate or grow through the first day of vacuum-packaged storage. Modified atmospheres with increasing CO₂ concentration were also compared with regard to bacterial survival during beef storage at 25°C. C. perfringens spores remained dormant in control and ozone-treated beef during a 10-d storage at 25°C. Pretreatment with heat increased germination and outgrowth during storage of beef, whereas ozone treatment and no treatment controls were effective in inhibiting spore germination and outgrowth in combination with increasing CO₂ concentrations above 30% or refrigeration. These data support the avoidance of heat in the pretreatment of raw beef.     

Evaluating the migration of ingredients from active packaging and development of dedicated methods: a study of two iron-based oxygen absorbers

The behaviour of two commercial oxygen-scavenging products with respect to migration of active ingredients into foodstuffs was investigated. Migrants were identified, and by using appropriate analytical methods, migration was determined in a variety of liquid, solid or gelled food simulants and foods. Simulants were chosen to cover a range of water activities and viscosities. Foods and the gelled food simulant agar were packed with and without vacuum, and with the oxygen scavenger in various locations relative to the packed food. The main migrants, as identified by X-ray fluorescence spectrometry, infrared spectroscopy and scanning electron microscopy with energy-dispersive spectrometry were Na + and Cl - in non-acidic aqueous simulants, and Na + , Cl - and Fe 2+ in 3% acetic acid. Migration into aqueous simulants exceeded the current European Union limit for total migration from plastic materials (assumed to be currently applicable to these systems) and was probably excessive by any reasonable standard. However, neither oxygen scavenger appeared to release significant quantities of migrants into solid foods when the scavenger was properly located in the package and the packing process does not favour the contents becoming wet by water released from the food.     

Selection and evaluation of seafood-borne psychrotrophic lactic acid bacteria as inhibitors of pathogenic and spoilage bacteria

In this study, inhibitory psychrotrophic lactic acid bacteria were isolated and investigated for future use in biopreservation of seafood products. Screening of 5575 colonies isolated from various seafood products resulted in the selection of 132 colonies presenting inhibitory properties. Among them, 52 isolates had characteristics of LAB and showed growth at 15 °C but not at 30 °C. The inhibition spectrum of these 52 isolates against 14 target strains (Gram-positive and -negative) showed inhibition of typical seafood spoiling and pathogenic bacteria and enabled the formation of seven interesting clusters. Sequencing of the 16S rRNA gene of a representative isolate from each cluster identified three Leuconostoc gelidum, two Lactococcus piscium, one Lactobacillus fuchuensis and one Carnobacterium alterfunditum. Theses strains did not produce histamine nor tyramine, and showed no particular antibiotic resistance profile. Growth rate as a function of temperature was tested for one L. piscium and one L. gelidum isolate and confirmed their psychrotrophic behavior. One out of seven isolates showed bacteriocin-like activity. The inhibition mechanisms of the other isolates are still unknown but may be due to competition for substrate. Absence of a bacteriocin-like component could be a positive point to gain rapid authorization for food application in France. This collection of LAB is now ready for testing on products.     

Effects of modified atmosphere and vacuum packaging on the growth of spoilage and inoculated pathogenic bacteria on fresh poultry

Fresh chicken breast meats inoculated withYersinia enterocolitica andAeromonas hydrophila were packaged in glass jars either containing different compositions of modified atmospheres (MA) (100% CO₂; 80% CO₂/20% N₂), or in vacuo or containing air, and were stored at 3±1°C and 8±1°C. The changes in gas composition as well asY. enterocolitica, A. hydrophila, total aerobic bacterial, total psychrotrophic, Lactobacilli and Enterobacteriaceae counts were determined after 0,1,3,7,9,11 and 14 days of storage. The results show that while the growth ofY. enterocolitica andA. hydrophila were retarded following MA storage, the pathogens were capable of growth in MA and vacuum storage at both temperatures, for the inoculation levels studied. For total aerobic bacterial counts, there were no differences between the values for chicken breast meats kept in different atmospheres. Being packaged in CO₂ had the greatest inhibitory effect on the growth of psychrotrophic aerobic bacteria during the first 3 days. Lactic acid bacteria levels of samples stored in MA conditions and in vacuo increased rapidly when compared to those levels of samples stored in air. It was also found that the effect of MA storage increased at 3±1°C.     

Total RNA concentration as an index of microbial activity and oxygen supply in an oxidation ditch

Total RNA and chromosomal DNA concentrations at a municipal wastewater treatment plant with an oxidation ditch (OD) were monitored for 1.5 years using commercial extraction kits for DNA and RNA. No parameters correlated with the chromosomal DNA concentration. The total RNA concentration exhibited better correlation than the solids retention time and the mixed liquor suspended solids with the removal rate of total organic carbon, and can be regarded as an index of microbial activity. The total RNA concentration varied with a cycle of one year and increased at lower water temperatures in this OD. When diffusion theory was taken into account, it was found that the oxygen dissolution rate increased at lower temperature, and a small change in the oxygen dissolution rate caused a large variation in microbial activity and also affected nitrification and denitrification. The information was insufficient to clarify the various reaction relationships, but total RNA concentration will likely be useful as an index of microbial activity in actual wastewater treatment reactors.     

Evaluation of two packaging systems for regional cheese

Saloio cheese – a regional Portuguese cheese – is currently sold unpackaged or in a vacuum package. Neither of these packaging systems is acceptable: the first system yields a cheese too hard, because of excessive water loss, while the second yields a white cheese with poor textural properties. The use of a packaging system with a tailor-made moisture barrier, i.e., allowing for water loss, but at a lower rate, is a way of extending the cheese’s shelf-life.     

Effect of oxygen absorber, nitrogen flushing, packaging material oxygen transmission rate and storage conditions on quality retention of raw whole unpeeled almond kernels (Prunus dulcis)

The present study investigated the effect of active packaging, nitrogen flushing, container oxygen barrier and storage conditions on quality retention of raw whole unpeeled almonds. Almond kernels were packaged in: a) polyethylene terephthalate//low-density polyethylene (PET//LDPE), and b) low-density polyethylene/ethylene vinyl alcohol/low-density polyethylene (LDPE/EVOH/LDPE) pouches under N_2, with or without an oxygen absorber, heat-sealed and stored for a period of 12 months. Quality parameters monitored were: peroxide value (PV), hexanal content, color, fatty acid composition and volatile compounds. PV ranged between 0.17 for fresh almonds and 9.22 meq O₂/kg oil for almonds packaged in PET//LDPE pouches under N₂ exposed to light at 20 °C after 12 months of storage. Respective values for hexanal were <28.5 μg/kg and 4.88 mg/kg. Polyunsaturated fatty acids (PUFA) and saturated fatty acids (SFA) increased with a parallel decrease of monounsaturated fatty acids (MFA) after 12 months of storage in all treatments. Likewise, volatile compounds such as aldehydes, ketones, alcohols, alkanes and aromatic hydrocarbons increased indicating enhanced lipid oxidation. Color was the parameter least affected. Use of the oxygen absorber provided a shelf life of at least 12 months for all samples irrespective of container oxygen barrier, lighting conditions and storage temperature.     

Development of a novel bioactive packaging based on the incorporation of Lactobacillus sakei into sodium-caseinate films for controlling Listeria monocytogenes in foods

A novel packaging technology was developed based on the incorporation of Lactobacillus sakei cells into sodium-caseinate (SC) edible films. Incorporation was based either on direct addition of the cells in the film forming solution used for casting or by surface spraying of the culture on the preformed film, resulting in a population density of 10⁶ cfu/cm². Addition of sorbitol in the film matrix increased the viability of the cells, greater than 90%, upon storage under both refrigeration and ambient temperature conditions for 30 days. Incorporation of the viable protective culture did not affect the mechanical properties and the physico-chemical properties of the film. Application of the films to both laboratory medium (agar) and a food model system (fresh beef) inoculated with Listeria monocytogenes resulted in a rapid growth of L. sakei immobilized in the film following contact with the wet medium or the food surface and a significant inhibition of the pathogen growth compared to the control samples under both constant and dynamic storage temperature protocols. The present study indicated that biopolymer-based antimicrobial films containing cells of a protective culture can be used as an effective packaging technology for improving food safety.     

Describing and modeling the release of an antimicrobial agent from an active PP/EVOH/PP package for salmon

Natural antimicrobial active packaging is an emerging technology for fresh fish preservation in which a chemical compound of natural origin is purposely incorporated into a packaging material to be released into the food surface in order to protect it from spoilage by foodborne microorganisms. The maximum efficiency of an antimicrobial package can only be obtained when an adequate activity is achieved immediately after the packaging operation and is maintained constant throughout the product’s shelf life. This work develops an active package designed for the preservation of fresh farmed salmon in cubes or slices, made up of a rigid polypropylene (PP)/ethylene–vinyl alcohol copolymer (EVOH)/PP tray heat-sealed with an active PP/EVOH/PP film lid in which 6.5% carvacrol is incorporated in the EVOH kernel as an antimicrobial active agent. The work also includes the measurement of the carvacrol kinetics and equilibrium parameters in the preserved salmon fillets, and proposes a mathematical model based on the finite element method to describe and simulate the common performance of the developed package/food system, and to predict its behavior under different working conditions or system configurations with the objective of finding the optimum combination of variables that ensure the best packaging performance. The results obtained from the determination of parameters showed a rapid migration of the active compound through the fish muscle, and a low affinity of the agent molecules for the food matrix. The active package was successfully developed, and the proposed model was satisfactorily used to detect the key factors that govern the package performance, and also to improve the package design by modifying the thickness distribution of the multilayer active film.     

Effect of an active packaging with citrus extract on lipid oxidation and sensory quality of cooked turkey meat

An antioxidant active packaging was prepared by coating a citrus extract, consisting of a mixture of carboxylic acids and flavanones, on polyethylene terephthalate trays. The effect of the packaging in reducing lipid oxidation in cooked turkey meat and on meat pH, colour characteristics and sensorial parameters was investigated. An untrained sensory panel evaluated the odour, taste, tenderness, juiciness and overall acceptability of the meat, using triangle, paired preference and quantitative response scale tests. A comparison between the antioxidant effects of the different components of the extract was also carried out. The packaging led to a significant reduction in lipid oxidation. After 2 days of refrigerated storage the sensory panel detected differences in odour and, after 4 days, rated the meat stored in the active packaging higher for tenderness and overall acceptability. Citric acid appeared to be the most important component of the extract with regard to its antioxidant potency.     

Evaluation of polyethylene terephthalate as a packaging material for premium quality whole pasteurized milk in Greece

Chemical, microbiological and sensorial changes in premium quality whole pasteurized milk stored at 4 °C under fluorescent light was studied for a period of 13 days. Milk containers tested included 1 l bottles made of (a) clear PET + UV blocker, 350–400 μm in thickness bearing a transparent label, (b) clear PET + UV blocker, 350–400 μm in thickness bearing a white colored label, (c) clear PET 350–400 μm in thickness. Milk packaged in 1 l coated paperboard cartons and stored under the same experimental conditions served as the “commercial control” sample. Data were obtained for lipid oxidation, lipolysis, proteolysis, vitamin A, E and riboflavin content, microbial growth including mesophilic and psychrotrophic counts and sensorial attributes (odor and taste) of whole pasteurized milk. Results showed satisfactory protection of milk packaged in all containers with regard to microbiological and chemical parameters assessed over the 13-day period. Based on sensory analysis, the shelf life of premium quality whole pasteurized milk tested in the present study was 10–11 days for both samples packaged in clear PET + UV bottles and in paperboard cartons and 8–9 days for clear PET bottles. Vitamin E losses recorded after 10 days of storage were respectively 42.7, 53.6 and 43.9% for samples packaged in clear PET + UV protected bottles, clear PET and control samples. Respective losses for riboflavin were 38.7, 52.5 and 35.0%. Average losses for vitamin A were 20.6% for all packaging materials. Clear PET + UV provided equal or better protection to milk as compared to the paperboard carton. Clear PET was the least effective in retaining light-sensitive vitamins. Based on spectral transmission curves of packaging materials tested, it is suggested that the use of a UV blocking agent in combination with a dark color pigmentation (blue, green etc.) in fresh milk packaging will provide a better protection to light-sensitive vitamins in cases where the expected shelf life of milk exceeds 5–6 days.     

Evaluation of polyethylene terephthalate as a packaging material for premium quality whole pasteurized milk in Greece

Chemical, microbiological, and sensorial changes in premium quality whole pasteurized milk stored at 4 °C under fluorescent light for one day followed by storage in the dark for an additional 12 days was studied. Milk containers tested included 1 l bottles made of (a) clear PET + UV blocker, 350–400μm in thickness bearing a transparent label, (b) clear PET + UV blocker, 350–400 μm in thickness bearing a white colored label, (c) clear PET 350–400 μm in thickness. Milk packaged in 1 l coated paperboard cartons and stored under the same experimental conditions served as the “commercial control” sample. Data were obtained for lipid oxidation, lipolysis, proteolysis, vitamin A, E, and riboflavin content, microbial growth including mesophilic and psychrotrophic counts and sensorial attributes (odor and taste) of whole pasteurized milk. Results showed satisfactory protection of milk packaged in all containers with regard to microbiological and chemical parameters assessed over the 13 day period. Based on sensory analysis, the shelf life of premium quality whole pasteurized milk tested in the present study was 10–11 days for samples packaged in clear PET+UV bottles and 9–10 days for clear PET bottles and paperboard cartons. Vitamin A losses recorded after 10 days of storage were respectively 15.9, 20.6, and 14.3% for samples packaged in clear PET+UV protected bottles, clear PET, and control samples. Respective losses for Vitamin E were 26.4, 36.6, and 35.0% and for riboflavin 32.9, 38.3, and 32.5%. Clear PET + UV provided equal or better protection to milk as compared to the paperboard carton. Clear PET was the least effective in retaining light-sensitive vitamins. Based on spectral transmission curves of packaging materials tested, it is suggested that the use of a UV blocking agent in combination with a dark color pigmentation (blue, green etc.) in fresh milk packaging will provide a better protection to light-sensitive vitamins in cases where the expected shelf life of milk exceeds 5–6 days.     

Effect of lactate-enhancement, modified atmosphere packaging, and muscle source on the internal cooked colour of beef steaks

Earlier studies on lactate-mediated colour stability in beef did not address the possible influence on cooked colour. Our objective was to examine the effect of lactate-enhancement, muscle source, and modified atmosphere packaging (MAP) on the internal cooked colour of beef steaks. Longissimus lumborum (LL) and Psoas major (PM) muscles from 16 (n = 16) beef carcasses (USDA Select) were randomly assigned to 4 enhancement treatments (non-injected control, distilled water-enhanced control, 1.25% and 2.5% lactate), and fabricated into 2.54-cm steaks. Steaks were individually packaged in either vacuum (VP), high-oxygen MAP (HIOX; 80% O₂ + 20% CO₂), or carbon monoxide MAP (CO; 0.4% CO + 19.6% CO₂ + 80% N₂), and stored for 0, 5, or 9 days at 1 °C. At the end of storage, surface and internal colour (visual and instrumental) was measured on raw steaks. Steaks were cooked to an internal temperature of 71 °C, and internal cooked colour (visual and instrumental) was evaluated. Lactate-enhancement at 2.5% level resulted in darker (P < 0.05) cooked interiors than other treatments. Interior cooked redness decreased (P < 0.05) during storage for steaks in VP and HIOX, whereas it was stable for steaks in CO. Our findings indicated that the beef industry could utilise a combination of lactate-enhancement and CO MAP to minimise premature browning in whole-muscle beef steaks.     

Kinetic studies on high-pressure inactivation of Bacillus stearothermophilus spores suspended in food matrices

Bacillus stearothermophilus spores ATCC 7953 can effectively be inactivated by high-pressure treatment, but only if it is applied at elevated temperatures; however, these temperatures are much lower compared to the temperature level used in heat inactivation under atmospheric pressure. Temperature and pressure in a range between 60 and 120°C and 50–600 MPa were applied to inactivate spores suspended in mashed broccoli and in cocoa mass. Utilizing an empirical mathematical model, derived from nth order kinetics, the survival curves of the spore strain investigated could be described accurately. The model can predict the impact of combined action of pressure and temperature on spore reduction. It was demonstrated that the inactivation of B. stearothermophilus spores ATCC 7953 improved with increasing treatment intensity. Beside intrinsic microbial inactivation mechanisms, the role of the pressure-induced shift in crystallization temperature of fat on spore inactivation in cocoa mass is discussed.     

Influence of oxygen exclusion and temperature on pathogenic bacteria levels and sensory characteristics of packed ostrich steaks throughout refrigerated storage

Ostrich steaks (290) were obtained from Iliofibularis muscles. For microbiological and pH determinations, samples were inoculated with Listeria monocytogenes NCTC 11994 (80 steaks) or Escherichia coli ATCC 12806 (80), then air- or vacuum-packed and stored at either 4±1°C or 10±1°C. Analyses were carried out on days 0, 3, 6 and 9 of storage. For sensory evaluation, samples (130) were air- or vacuum-packed and stored at 4±1°C or at 10±1°C. Sensory attributes (odour, colour, drip loss, texture and general acceptability) were scored by six untrained judges using an unstructured nine-point hedonic scale on eleven sampling days (0, 3, 6, 9, 12, 15, 18, 21, 24, 27 and 30). Increases in microbial counts (log(10)cfu/g) were observed throughout storage in all groups of samples for both L. monocytogenes (from 6.39±0.43-6.62±0.32 at day 0 to 8.87±0.19-9.64±0.43 at day 9) and E. coli (from 5.57±0.15-5.68-0.40 to 7.79±0.96-9.64±0.17). Gas atmosphere influenced microbial counts from day 3 of storage with lower (P<0.05) values observed in vacuum- than in air-packed samples at 10°C (L. monocytogenes) or at 4 and 10°C (E. coli). Storage temperature significantly influenced bacterial counts throughout storage, especially in air-packed samples. Lower pH values in vacuum- than in air-packed samples were observed from day 6. Both effects (gas atmosphere and temperature) influenced the hedonic scores, with higher values assigned to vacuum-packed samples for most attributes (with the exception of drip loss) and sampling days. A marked influence of storage temperature on sensorial scores was obtained in air-packaged ostrich steaks. The shelf-life (time until the average general acceptability score fell below 5) was 6 (air-packed samples), 9 (vacuum-packed, 10°C), or 12 days (vacuum-packed, 4°C). The results being reported here suggest the importance of both oxygen exclusion and storage at low temperatures to reduce microbiological risks and improve the acceptability of ostrich meat. However, the short shelf-life of this product highlights the need to keep the time between slaughter and sale to a minimum.     

Quality evaluation of raw ground almond kernels (Prunus dulcis): Effect of active and modified atmosphere packaging, container oxygen barrier and storage conditions

The present study investigated the effect of active and modified atmosphere packaging, container oxygen barrier and storage conditions on quality retention of raw ground almonds. Ground almond kernels were packaged in: a) polyethylene terephthalate//low density polyethylene (PET//LDPE), and b) low density polyethylene/ethylene vinyl alcohol/low density polyethylene (LDPE/EVOH/LDPE), under N₂ or with an oxygen absorber and stored either under fluorescent light or in the dark at 4 or 20 °C for a period of 12 months. Quality parameters monitored were: peroxide value (PV), hexanal content, color, fatty acid composition and volatile compounds. Of the sensory attributes color, texture, odor and taste were evaluated. PV ranged between 0.26 for fresh almonds and 19.98 meq O₂/kg oil for almonds packaged in PET//LDPE pouches under N₂ exposed to light at 20 °C after 12 months of storage. Respective values for hexanal were < 28.5 µg/kg and 9.38 mg/kg. Polyunsaturated fatty acids (PUFA) and saturated fatty acids (SFA) increased with a parallel decrease of monounsaturated fatty acids (MFA) after 12 months of storage in samples stored with the oxygen absorber while in samples packaged in PET//LDPE under N₂, a decrease in PUFA and MUFA with a parallel increase in SFA was recorded. Likewise, volatile compounds such as aldehydes, ketones, alcohols, acids, alkanes and aromatic hydrocarbons increased during storage indicating enhanced lipid oxidation. Color parameters L, a and b remained unaffected in all treatments including the oxygen absorber while under a N₂ atmosphere L parameter showed a small but statistically significant (p < 0.05) decrease with a parallel increase (p < 0.05) of a and b values after 12 months of storage. The most pronounced color changes occurred for samples in PET//LDPE pouches irrespective of lighting conditions at 20 °C. Raw ground almonds retained acceptable quality for ca. 6–7 months packaged in PET//LDPE and ca. 8 months packaged in LDPE/EVOH/LDPE pouches under N₂ irrespective of lighting conditions at 20 °C while at 4 °C shelf life was extended by an additional month as compared to storage at 20 °C. Use of the oxygen absorber provided a shelf life of at least 12 months for all samples irrespective of container oxygen barrier, lighting conditions and storage temperature.

Industrial relevance

The use of oxygen absorbers is very effective in extending the shelf life of ground almonds commercially for at least 12 months irrespective of packaging material barrier to O₂, lighting conditions and storage temperature.