Innovative Systems for the Delivery of Naturally Occurring Antimicrobial Volatiles in Active Food-Packaging Technologies for Fresh and Minimally Processed Produce: Stimuli-Responsive Materials.

Certain naturally occurring volatile organic compounds are able to mitigate food spoilage caused by microbial growth. Their considerable vapor pressure enables them to create an antimicrobial atmosphere within a package, and this property can be used for the development of active food-packaging technologies. The volatility of these molecules, however, makes their stabilization difficult and limits their effectiveness. Whilst much research is being undertaken on the use of natural antimicrobial volatiles for inhibiting microbial growth in food, less attention has been paid to the design of controlled-release mechanisms that permit the efficient application of these compounds. Most studies to date either spray the volatile directly onto the fresh product, immerse it in a solution containing the volatile, or embed the volatile in a paper disc to create a vapor in the headspace of a package. More sophisticated alternatives would be delivery systems for the sustained release of volatiles into the package headspace. Such systems are based on the encapsulation of a volatile in organic or inorganic matrices (cyclodextrins, electrospun non-wovens, polymer films, micelles, molecular frameworks, etc.). However, most of these devices lack an efficient triggering mechanism for the release of the volatile; most are activated by humidity. All of these techniques are revised in the present work, and the most recent and innovative methods for entrapping and releasing volatiles based on reversible covalent bonds are also discussed.

Agro-Industrial Protein Waste and Co-Products Valorization for the Development of Bioplastics: Thermoprocessing and Characterization of Feather Keratin/Gliadin Blends.

Biopolymers based on plant and animal proteins are interesting alternatives in the development of films with future prospects as food packaging. Considering that in recent years there has been an increasing interest in the valorization of agro-industrial residues and by-products and that the blending of polymers can lead to materials with improved properties, in this work, keratin-rich feather fibers and gliadins were blended at different ratios in order to develop sustainable and biodegradable films. Control gliadin G100, feather F100 films, and their blends at 3:1 (G75F25), 2:2 (G50F50), and 1:3 (G25F75) ratios were successfully developed through thermoprocessing. The physical properties were differentiated as a function of the concentration of both polymeric matrices. Although gliadins showed higher hydrophilicity as confirmed by their highest swelling degree, films with high gliadin ratios exhibited lower water vapor permeability values at low and medium relative humidities. On the other hand, the feather fiber-based films displayed the highest Young's modulus values and provided an oxygen barrier to the blends, principally at the highest relative humidity. In conclusion, the blend of these protein-based polymers at different ratio resulted in interesting composites whose physical properties could be adjusted.

ZIF-8 encapsulation improves the antifungal activity of benzaldehyde and methyl anthranilate in films.

In this work, two ZIF-8-based biocomposites were obtained by entrapping the biomolecules benzaldehyde and methyl anthranilate via direct impregnation with fast encapsulation kinetics and high molecule payloads were achieved. The obtained biocomposites exhibit an enhanced antifungal activity against Penicilium expansum after integration in biopolymeric zein films in comparison with the action of free molecules, making these biomaterials promising candidates for food preservation and packaging applications.

Designing an Oxygen Scavenger Multilayer System Including Volatile Organic Compound (VOC) Adsorbents for Potential Use in Food Packaging.

Oxygen scavengers are valuable active packaging systems because several types of food deterioration processes are initiated by oxygen. Although the incorporation of oxygen scavenger agents into the polymeric matrices has been the trend in recent years, the release of volatile organic compounds (VOC) as a result of the reaction between oxygen and oxygen scavenger substances is an issue to take into account. This is the case of an oxygen scavenger based on a trans-polyoctenamer rubber (TOR). In this work, the design of an oxygen scavenger multilayer system was carried out considering the selection of appropriate adsorbents of VOCs to the proposed layer structure. Firstly, the retention of some representative organic compounds by several adsorbent substances, such as zeolites, silicas, cyclodextrins and polymers, was studied in order to select those with the best performances. A hydrophilic silica and an odor-adsorbing agent based on zinc ricinoleate were the selected adsorbing agents. The principal VOCs released from TOR-containing films were carefully identified, and their retention first by the pure adsorbents, and then by polyethylene incorporated with the selected compounds was quantified. Detected concentrations decreased by 10- to 100-fold, depending on the VOC.

Plastic ingestion by two cetacean groups: Ziphiidae and Delphinidae.

The presence of plastic in our environment is having a massive impact on today's marine biota. Whales and dolphins are becoming sentinels of litter pollution as plastic entanglement and ingestion affect them with unknown consequences. Although information exists about this anthropogenic interaction, the compilation of this data on metastudies is difficult due to the use of varied methodologies. A combination of our own data as well as a review of historical data was used to complete an extensive study of how cetaceans are interacting with macro and micro-litter at a global level. Here, we identify the plastic uptake by two cetacean families: Ziphiidae and Delphinidae, thus allowing for a better understanding in order to offer a global overview of their current status. Additionally, analysis was run on the plastic found in the digestive contents of stranded specimens of two Cuvier's beaked whales and fourteen striped dolphins in the Alboran Sea, in the Western Mediterranean, a hotspot for marine megafauna. Out of 623 stranded cetaceans from datasets, beaked whales displayed the highest concentration of macro, meso and microplastic in the Western Pacific Ocean. Regarding striped dolphins, Eastern Spain was the location with the highest plastic ingestion. Moreover, deep divers such as beaked whales ingested more plastic than striped dolphins which could be as a consequence of their feeding behavior or habitat. Thus, this overview provides useful information concerning conservation issues on how cetacean hotspots are highly affected by marine plastic ingestion.

pH modulates antibacterial activity of hydroxybenzaldehyde derivatives immobilized in chitosan films via reversible Schiff bases and its application to preserve freshly-squeezed juice.

Antimicrobial food grade hydroxybenzaldehyde derivatives were immobilized on the surface of chitosan films by means of reversible Schiff bases. Spectroscopy and elemental analysis evidenced the different ability of the aldehydes to form Schiff bases with chitosan. Chitosan films modified with Schiff bases of aldehydes exerted antimicrobial properties against E. coli under mild acidic environments. The efficacy of the films lied on the reversibility of synthetized imine bonds and release of the aldehydes which was promoted in mildly acid aqueous solutions. Besides acidity, imine bond reversibility depended on the chemical structure of the aldehyde covalently bonded. Films carrying salicylaldehyde presented the highest in vitro antimicrobial performance and thus, they were chosen to evaluate their effectivity in inhibiting E. coli proliferation in freshly-squeezed carrot-orange juice. Films were successfully activated by the acid environment of the juice and reduced the population of the inoculated pathogen. Salicylaldehyde migrated to the juice did not exert toxic effects on Caco-2 cells.

Pilot-Scale Processing and Functional Properties of Antifungal EVOH-Based Films Containing Methyl Anthranilate Intended for Food Packaging Applications.

Antimicrobial packaging has emerged as an efficient technology to improve the stability of food products. In this study, new formulations based on ethylene vinyl alcohol (EVOH) copolymer were developed by incorporating the volatile methyl anthranilate (MA) at different concentrations as antifungal compound to obtain active films for food packaging. To this end, a twin-screw extruder with a specifically designed screw configuration was employed to produce films at pilot scale. The quantification analyses of MA in the films showed a high retention capacity. Then, the morphological, optical, thermal, mechanical and water vapour barrier performance, as well as the antifungal activity in vitro of the active films, were evaluated. The presence of MA did not affect the transparency or the thermal stability of EVOH-based films, but decreased the glass transition temperature of the copolymer, indicating a plasticizing effect, which was confirmed by an increase in the elongation at break values of the films. Because of the additive-induced plasticization over EVOH, the water vapour permeability slightly increased at 33% and 75% relative humidity values. Finally, the evaluation of the antifungal activity in vitro of the active films containing methyl anthranilate showed a great effectiveness against P. expansum and B. cinerea, demonstrating the potential applicability of the developed films for active food packaging.

Presence and implications of plastics in wild commercial fishes in the Alboran Sea (Mediterranean Sea).

The presence of plastic in the environment has become a major problem for marine ecosystems. The identification of the global micro and mesoplastic uptake by commercial fish populations may allow for a better understanding of their impact. This study aims to determine the presence and composition of plastic in two pelagic fish (Engraulis encrasicolus and Scomber scombrus) and two demersal species (Scyliorinus canicula and Mullus barbatus) from the Alboran Sea (western Mediterranean) to quantify the relationship between plastic prevalence and the environment and feeding behavior in the selected fish species. Samples of these four fish species from sites in the Alboran Sea were studied for ingested plastics. These localized samples were also compared to published values which covered a broader geographical range. Samples from the Alboran Sea study sites showed that the predominant fiber color was black and the predominant plastic polymers were polyethylene and cellulose. At the Alboran Sea study site the highest plastic occurrence was found in S. scombrus, whereas in the published literature the highest occurrence of plastics in digestive tracts was found in E. encrasicolus. The general prevalence of marine plastic pollution and levels of macro- and micro-plastic ingested by commercial fish species in this study support the idea that quantifying plastic presence and composition may be essential to understanding potential impacts on marine ecosystems.

Exploiting the Redox Activity of MIL-100(Fe) Carrier Enables Prolonged Carvacrol Antimicrobial Activity.

The design of efficient food contact materials that maintain optimal levels of food safety is of paramount relevance to reduce the increasing number of foodborne illnesses. In this work, we develop a smart composite metal-organic framework (MOF)-based material that fosters a unique prolonged antibacterial activity. The composite is obtained by entrapping a natural food preserving molecule, carvacrol, into a mesoporous MIL-100(Fe) material following a direct and biocompatible impregnation method, and obtaining particularly high payloads. By exploiting the intrinsic redox nature of the MIL-100(Fe) material, it is possible to achieve a prolonged activity against Escherichia coli and Listeria innocua due to a triggered two-step carvacrol release from films containing the carvacrol@MOF composite. Essentially, it was discovered that based on the underlying chemical interaction between MIL-100(Fe) and carvacrol, it is possible to undergo a reversible charge-transfer process between the metallic MOF counterpart and carvacrol upon certain chemical stimuli. During this process, the preferred carvacrol binding site was monitored by infrared, Mössbauer, and electron paramagnetic resonance spectroscopies, and the results are supported by theoretical calculations.

Trans-2-Hexenal-Based Antifungal Packaging to Extend the Shelf Life of Strawberries.

Strawberries are valuable because of their nutritional value, but they are also highly perishable fruits. Fungal decay is the overriding factor that alters the overall quality of fresh strawberries. Because no hygienic treatments to reduce the initial microbial load are feasible, molds develop during postharvest when using conventional packaging. In this study, an antifungal packaging system for strawberries was developed to improve safety and quality. Trans-2-hexenal (HXAL), a natural compound in strawberries, was incorporated into the modified atmosphere packaging (MAP) systems. Zero, 100, and 250 µL of HXAL were included in cellulosic pads and were covered with a polyamide coating to control its release. The pads were placed on the bottom of plastic trays; an amount of250 g of strawberries was added, flow packed in micro-perforated PP bags, and stored at 4 °C for 14 days. Fungal infection was monitored during the storage period, and the optical and textural properties of the strawberries were measured at days 0 and 14. Analysis of the package headspace was conducted to check for the HXAL concentration. HXAL was partially retained in the fruits and was converted into hexyl acetate and 2-hexen-1-ol acetate, but this was only measurably present in the headspace of the active systems. Mold growth was fully inhibited in active packaging although the strawberries were softer and darker than those in the control packages. The active package was not as efficient if the fruits were stored under thermal-abuse conditions (15 and 22 °C).

Hot-Melt-Extruded Active Films Prepared from EVOH/Trans-Cinnamaldehyde Blends Intended for Food Packaging Applications.

In this work, novel active films based on ethylene vinyl alcohol copolymer (EVOH) and cinnamaldehyde (CIN) were successfully obtained employing a hybrid technique consisting of a two-step protocol involving the preparation of a polymeric EVOH-CIN masterbatch by solvent-casting for its further utilization in the preparation of bioactive EVOH-based films by melt extrusion processing. The influence of CIN over the EVOH matrix was studied in terms of optical, morphological, thermal, and mechanical properties. Optically transparent films were obtained and the incorporation of cinnamaldehyde resulted in yellow-colored films, producing a blocking effect in the UV region. A decrease in the glass transition temperature was observed in the formulations containing cinnamaldehyde, indicating a plasticizing effect. This phenomenon was confirmed by an increase in the elongation at break values of the extruded films. Results from thermogravimetric analysis determined a slight decrease in the thermal stability of EVOH provoked by the vaporization of the bioactive compound. Bioactive properties of the films were also studied; the presence of residual cinnamaldehyde in EVOH after being subjected to an extrusion process conferred some radical scavenging activity determined by the DPPH assay whereas films were able to exert antifungal activity in vapor phase against Penicillium expansum. Therefore, the present work shows the potential of the hybrid technique employed in this study for the preparation of bioactive films by a ready industrial process technology for food packaging applications.

Development of antifungal biopolymers based on dynamic imines as responsive release systems for the postharvest preservation of blackberry fruit.

This study describes the synthesis and reversibility of Schiff bases from chitosan and bioactive compounds, and their application in the antifungal packaging of fruit. Imine bonds between primary amine groups of chitosan and carbonyl groups of antifungal aldehydes were synthesised and their reversibility was assayed in an aqueous medium under different acidic conditions. The mechanism of action of the dynamers is based on the hydrolysis of imine bond and the release of the active agent. The new films were effective at inhibiting the growth of Penicillium expansum and Botrytis cinerea, and their effectivity depended on the degree of hydrolysis achieved which was greater when the bonds were hydrolysed in a mild acidic medium. A double bottom cylindrical tray was used for the responsive antimicrobial packaging of blackberries. The package extended shelf-life of berries from 3 to 12 days without causing phytotoxic effects on the fruit being safe for human consumption.

Designing Biodegradable and Active Multilayer System by Assembling an Electrospun Polycaprolactone Mat Containing Quercetin and Nanocellulose between Polylactic Acid Films.

The design of multilayer systems is an innovative strategy to improve physical properties of biodegradable polymers and introduce functionality to the materials through the incorporation of an active compound into some of these layers. In this work, a trilayer film based on a sandwich of electrospun polycaprolactone (PCL) fibers (PCLé) containing quercetin (Q) and cellulose nanocrystals (CNC) between extruded polylactic acid (PLA) films was designed with the purpose of improving thermal and barrier properties and affording antioxidant activity to packaged foods. PCLé was successfully electrospun onto 70 µm-thick extruded PLA film followed by the assembling of a third 25 µm-thick commercial PLA film through hot pressing. Optical, morphological, thermal, and barrier properties were evaluated in order to study the effect of PCL layer and the addition of Q and CNC. Bilayer systems obtained after the electrospinning process of PCL onto PLA film were also evaluated. The release of quercetin from bi- and trilayer films to food simulants was also analyzed. Results evidenced that thermal treatment during thermo-compression melted PCL polymer and resulted in trilayer systems with barrier properties similar to single PLA film. Quercetin release from bi- and trilayer films followed a similar profile, but achieved highest value through the addition of CNC.

Development of Biodegradable Films Loaded with Phages with Antilisterial Properties.

The inhibitory and bactericidal capacity of Listex P100 bacteriophage has been studied against different concentrations of Listeria monocytogenes in stationary and exponential phases. Three different matrices were employed to developed films incorporating Listex P100: (1) sodium caseinate, (2) sodium alginate mixed with gelatin, and (3) polyvinyl alcohol (PVOH). All the films were successfully developed by casting at room temperature. These active biodegradable films were optical, structural, and thermally characterized, and their antimicrobial capacities against L. monocytogenes were studied. The incorporation of phages did not affect the morphology, colour, opacity, and thermal stability of polymers. The antimicrobial analysis revealed the bacteriophage presented a high antimicrobial capacity against L. monocytogenes in the stationary phase (4.40 and 6.19 log reduction values or bactericide effect depending on the initial inoculum of the pathogen). Developed films showed antimicrobial capacity close to 1 log after 24 h of incubation at 30 °C. The effectiveness of PVOH films was greater under refrigeration conditions, reaching 2 log reduction after eight days of incubation. The use of these films as a coating in a food or as part of a packaging could improve food safety against the growth of pathogenic microorganisms such as Listeria monocytogenes.

Broadening the antimicrobial spectrum of nisin-producing Lactococcus lactis subsp. Lactis to Gram-negative bacteria by means of active packaging.

Cast films obtained from polyvinyl alcohol (PVOH) blended with casein hydrolysates (HCas) in a weight ratio of 1:1 were employed to carry nisin-producing L. lactis and phytic acid in order to broaden the antimicrobial spectrum of L. lactis to Gram-positive and Gram-negative spoilage and pathogen bacteria. For this purpose, the effect of the antimicrobial activity of various film formulations and combinations of films on the growth of E. coli at 37 °C for 24 h was studied. The film system that showed antimicrobial activity against Gram-negative bacteria consisted of phytic acid and L. lactis incorporated in separate films. When the active agents were in the same film the viability of L. lactis decreased considerably and it did not exert antimicrobial activity against the bacterium. Therefore, the combination of L. lactis and phytic acid in separate films was chosen as the reliable system, and the effect of its activity on the growth of Gram-negative bacteria (E. coli, Salmonella enterica, and Pseudomonas fluorescens) and Gram-positive bacteria (Listeria monocytogenes) in liquid culture medium was tested at refrigeration temperature (4 °C), and with simulated breaks in the cold chain (14 °C and 24 °C). The survival of L. lactis in coexistence with these bacteria was also studied. The film system exerted an antimicrobial effect against the Gram-negative bacteria tested, and the activity depended on the bacteria and the temperature assayed. With regard to the antimicrobial activity against L. monocytogenes, phytic acid improved the antimicrobial capacity of L.lactis. The survival of L. lactis was maintained at 7-8 log (CFU/mL) culture in liquid medium throughout the storage period. The films developed were intended to be used as coatings in the design of a double-sided active bag for a non-fermented dairy product. The bags were filled with homemade preservative-free pastry cream, and the microbiological shelf life and evolution of pH of the packaged ready-to-eat food stored at 4 °C was studied for 20 days. The results showed a reduction in the growth of spoilage bacteria and therefore an increase in the shelf life of the packaged product. The films developed could be applied in the design of packages for perishable dairy foods in order to increase their microbiological shelf life.

Machine learning approach for predicting Fusarium culmorum and F. proliferatum growth and mycotoxin production in treatments with ethylene-vinyl alcohol copolymer films containing pure components of essential oils.

Fusarium culmorum and F. proliferatum can grow and produce, respectively, zearalenone (ZEA) and fumonisins (FUM) in different points of the food chain. Application of antifungal chemicals to control these fungi and mycotoxins increases the risk of toxic residues in foods and feeds, and induces fungal resistances. In this study, a new and multidisciplinary approach based on the use of bioactive ethylene-vinyl alcohol copolymer (EVOH) films containing pure components of essential oils (EOCs) and machine learning (ML) methods is evaluated. Bioactive EVOH-EOC films were made incorporating cinnamaldehyde (CINHO), citral (CIT), isoeugenol (IEG) or linalool (LIN). Several ML methods (neural networks, random forests and extreme gradient boosted trees) and multiple linear regression (MLR) were applied and compared for modeling fungal growth and toxin production under different water activity (aw) (0.96 and 0.99) and temperature (20 and 28 °C) regimes. The effective doses to reduce fungal growth rate (GR) by 50, 90 and 100% (ED50, ED90, and ED100) of EOCs in EVOH films were in the ranges 200 to >3330, 450 to >3330, and 660 to >3330 µg/fungal culture (25 g of partly milled maize kernels in Petri dish), respectively, depending on the EOC, aw and temperature. The type of EVOH-EOC film and EOC doses significantly affected GR in both species and ZEA and FUM production. Temperature also affected GR and aw only affected GR and FUM production of F. proliferatum. EVOH-CIT was the most effective film against both species and ZEA and FUM production. Usually, when the EOC levels increased, GR and mycotoxin levels in the medium decreased although some treatments in combination with certain aw and temperature values induced ZEA production. Random forest models predicted the GR of F. culmorum and F. proliferatum and ZEA and FUM production better than neural networks or extreme gradient boosted trees. The MLR mode provided the worst performance. This is the first approach on the ML potential in the study of the impact that bioactive EVOH films containing EOCs and environmental conditions have on F. culmorum and F. proliferatum growth and on ZEA and FUM production. The results suggest that these innovative packaging systems in combination with ML methods can be promising tools in the prediction and control of the risks associated with these toxigenic fungi and mycotoxins in food.

Melt-Processed Bioactive EVOH Films Incorporated with Ferulic Acid.

In this work, antimicrobial and antioxidant films based on ethylene vinyl alcohol (EVOH) copolymer containing low amounts of ferulic acid (FA) were successfully developed by melt extrusion. Optically transparent films were obtained, and the presence of FA provided some UV blocking effect. The characterization of the thermal and barrier properties of the developed films showed that the addition of FA improved the thermal stability, decreased the glass transition temperature (Tg) and increased the water vapor and oxygen transmission rates when ferulic acid was loaded above 0.5 wt.%, associated with its plasticizing effect. Mechanical characterization confirmed the plasticizing effect by an increase in the elongation at break values while no significant differences were observed in Young's modulus and tensile strength. Significant antioxidant activity of all active films exposed to two food simulants, 10% ethanol and 95% ethanol, was also confirmed using the 2,2-diphenyl-1-pricylhydrazyl (DPPH) free radical scavenging method, indicating that FA conserved its well-known antioxidant properties after melt-processing. Finally, EVOH-FA samples presented antibacterial activity in vitro against Escherichia coli and Staphylococcus aureus, thus showing the potential of ferulic acid as bioactive compound to be used in extrusion processing for active packaging applications.

PVOH/protein blend films embedded with lactic acid bacteria and their antilisterial activity in pasteurized milk.

PVOH-based polymer matrices in the form of films were evaluated as carriers of living Lactococcus lactis subsp. Lactis. These lactic acid bacteria are capable of producing nisin, which is an effective antilisterial peptide. A low percentage (1:0.125 w/w) of yeast extract, gelatin, sodium caseinate, gelatin, or casein hydrolysates was incorporated in PVOH matrices with the aim of increasing the viability of bacteria in the film. The films were obtained by casting after incorporating L. lactis. Then they were evaluated for antilisterial activity in liquid medium at 37 °C for 24 h, and also at 4 °C for 21 days in order to simulate the storage of liquid foods in refrigeration conditions. The survival of the lactic acid bacteria was also evaluated at both temperatures during the experiment. L. lactis remained viable in all the films tested at 37 and 4 °C. The antimicrobial activity of the films was greater at 4 °C than at 37 °C. With regard to the effect of the film composition, the activity of the films was higher when protein hydrolysates and sodium caseinate were incorporated in the formulation. Films supplemented with protein hydrolysates or sodium caseinate inhibited growth of the pathogen during the 21 days of storage at 4 °C. At 37 °C, after 24 h the films had slowed the growth of the inoculated pathogen by between 2 and 4 log CFU/mL. Finally, as the films developed are intended to be used in the design of active packaging of foods, they were tested in pasteurized milk inoculated with 4 log CFU/mL of Listeria monocytogenes and stored at 4 °C for 21 days. The pathogen began to grow after the second day of storage with or without film, but when the films were added to the medium the growth of the pathogen was slowed down, without reaching >6 log CFU, whereas the control reached a maximum growth of 8.5 log CFU. The pH of the milk was monitored throughout the experiment, and it decreased with time. This was due to the generation of organic acids by the lactic bacteria. Buffering the food stabilized the pH without modifying the activity of the films. Thus, the current study shows that PVOH films supplemented with nutrients can act as carriers of L. lactis, and they can help to increase the safety of refrigerated dairy beverages and sauces.

Chromatic Sensor to Determine Oxygen Presence for Applications in Intelligent Packaging.

A chromatic sensor has been designed for the detection of oxygen in package headspace. The sensor is based on the redox change of methylene blue (MB) to its leuco form. Its formulation includes the pigment, glycerol, as a sacrificial electron donor, TiO2, as a photocatalyst and ethylene-vinyl alcohol copolymer (EVOH), as a structural polymer matrix. The final sensor design that allows its manufacture by conventional printing and laminating technologies consists of the sensing polymer matrix (MB-EVOH) sandwiched in a suitable transparent multilayer structure. The outer layers protect the sensor from the external atmosphere and allow visualization of the colour. The inner layer is sufficiently opaque to facilitate sensor reading from the outside, is thick enough to avoid direct contact with food (functional barrier), and is oxygen-permeable to expose the sensing material to the internal package atmosphere. In the absence of oxygen, the sensor becomes white by irradiation with halogen lamps in less than 60 s. All components are substances permitted for food contact except the pigment, but specific migration analysis showed no trace of migration thanks to the functional barrier included in the design.