The current high demand of minimally processed or fresh-cut fruit and vegetables results from the consumer’s desire for healthy, convenient, fresh, and ready-to-eat plant food-derived commodities. Fresh-cut fruits and vegetables are usually packaged under active- or passive-modified atmosphere packaging, while its shelf life must be under refrigerated conditions. The most important goal to preserve quality and safety focuses on releasing the microbial spoilage flora, since every unit operation involved will influence the final load. Sanitation in the washing step is the only unit operation able to reduce microbial load throughout the production chain. Chlorine is widely used as an efficient sanitation agent, but some disadvantages force to find eco-friendly emerging alternatives. It is necessary to deal with aspects related to sustainability because it could positively contribute to the net carbon balance besides reducing its use. Several innovative techniques seem to reach that target. However, industrial changes for replacing conventional techniques request a fine knowledge of the benefits and restrictions as well as a practical outlook. This chapter reviews the principles of emerging eco-friendly techniques for preserving quality and safety of fresh-cut products in order to meet the expected market’s demand.
- minimally processed
- food safety
The benefits of fruit and vegetables consumption on human health are well known, being linked with prevention of a grand array of diseases such as degenerative disorders, cancer, and cardiovascular, among others . Consequently, their intake has been promoted among consumers by nutritionists, researchers, and even at a governmental level (i.e., campaigns like
This chapter summarizes the principles and development of eco-friendly techniques for preserving safety of FC products in order to meet the expected market’s demand.
2. Antimicrobial washing solutions
2.1. Peroxyacetic acid
Peroxyacetic acid (PAA), or peracetic acid, is a colorless organic peroxide that is a mixture of acetic acid and hydrogen peroxide. It is an eco-friendly sanitizer whose breakdown products are acetic acid, O2, CO2, and water. PAA is approved by the European Union as a disinfectant for drinking water and food areas and as an in-can preservative . PAA is also permitted by the U.S. Food and Drug Administration as an additive for food . The surface-cleaning concentrations range from 85 to 300 ppm, although 50 ppm has been reported to be enough . PAA is mainly used in fruit and vegetable processing due to tolerance to several factors such as temperature, pH (1–8), hardness, and soil contamination. A recommended combination of 11% hydrogen peroxide (H2O2) and 15% PAA, at 80 ppm, was proposed for the disinfection of plant surfaces .
2.2. Chlorine dioxide
Chlorine dioxide (ClO2) is a yellowish-green stable dissolved gas that has been used for the last decades for water treatment as a potential alternative to NaOCl. ClO2 has higher effectiveness over a broad range of pH, higher water solubility (10 times higher than NaOCl), higher oxidant capacity, lower reactivity with organic matter, and higher effectiveness at low concentrations than NaOCl . Nevertheless, ClO2 is a very unstable substance and is highly explosive as a concentrated gas when concentrations ≥10% are reached in air. Hence, ClO2 must be generated on-site by two different procedures: reacting an acid with sodium chlorite or the reaction of sodium chlorite with chlorine gas then being obtained in either aqueous or gaseous forms, respectively . The ClO2 is classified as a non-carcinogenic product since it does not ionize to produce weak acids (as occurred for chlorine and bromine) or to form carcinogenic by-products like trihalomethanes . Gaseous ClO2 treatment (100 ppm) of several fresh products (tomatoes, lettuce, cantaloupe, alfalfa sprouts, oranges, apples, and strawberries) did not leave any chemical residues on them . ClO2 is approved in the USA for usage in washing whole fresh fruits and vegetables and shelled beans and pears with intact cuticles at maximum levels of 5 ppm and 1 ppm for peeled potatoes .
ClO2 is considered as a strong microbicide at low levels such as 0.1 ppm, achieving also a rapid removal of biofilms which avoid bacterial re-growth . The bactericidal effect of ClO2 is explained by the interruption of several cellular processes (proteins production and changes in the cell structure) when organic substances in bacterial cells react with ClO2. On viruses, ClO2 reacts with peptone to prevent the protein formation being more effective than chlorine or ozone . Inoculated pathogens like
2.3. Hydrogen peroxide
Hydrogen peroxide (H2O2) is a strong oxidizer able to generate other cytotoxic-oxidizing species, like hydroxyl radicals, with strong bactericide (including spores) effect . H2O2 is an eco-friendly disinfectant since it is rapidly decomposed into water and oxygen in the presence of catalase. Likewise, it is colorless and non-corrosive. H2O2 is allowed for use in food processing and packaging but not as a sanitizing agent for fresh produce by the FDA . However, high H2O2 concentrations are needed to achieve good santising effects in FC products. However, such high concentrations may lead to browning being necessary thge use of anti-browning agents like sodium erythorbate . Accordingly, 2–3 × 104 ppm H2O2 were needed to reduce
2.4. Weak organic acids
Weak organic acids have been widely used as preservatives for the prevention of several quality degradation processes such as enzymatic and nonenzymatic browning, texture deterioration, and microbial spoilage. Contrary to NaOCl, weak organic acids do not produce toxic or carcinogenic compounds when they interact with organic molecules . Therefore, several weak organic acids are considered as GRAS (Generally Recognized as Safe) by the FDA and European Commission being well accepted by consumers. The antimicrobial effect of weak organic acids is related to the cytoplasm acidification, osmotic stress, disruption of proton motive force, and synthesis inhibition of macromolecules . Weak organic acids are more effective for bacteria than for yeasts and molds because of the low pH (2.1–2.7) of the applied solutions. Citric, acetic, lactic, and ascorbic acids are the most common acids applied in the food industry.
Citric acid, contrary to other acids, acts as a chelating agent of metallic ions of the medium, avoiding microbial growth [40, 41]. Citric acid treatment (0.52 mM) maintained microbial safety and visual quality of FC “Amarillo” melon during a shelf life of 10 days at 5℃ . A solution of 0.1 M citric and 0.5 M ascorbic acid achieved the same effectivity as 100 ppm NaOCl to control microbial growth and maintain quality of green celery crescents . Citric and lactic acid dippings of 0.5–1 × 104 ppm achieved comparable
2.5. Calcium, sodium, and potassium-derived salts
Several salts are recognized as GRAS being a low-cost material for the food industry with high acceptance by the consumers, since it is not toxic. Calcium is used to retain the firmness of plant commodities by interaction with pectin to form calcium pectate maintaining then the cell wall structure. FC lettuce treated with 15 × 103 ppm calcium lactate showed higher crispness than samples treated with 120 ppm NaOCl after 1 day at 4℃ . Latter authors hypothesized that such finding could be owed to the activation of texture-related enzymes, like PME, by the calcium absorption in the lettuce, or an increase in diffusive processes by temperature, including the calcium. Similar results were obtained by 15 × 103 ppm calcium lactate treatment at 50℃ of sliced FC carrots to maintain the cortex turgor of plant cells and reduce the lignification degree in cut surfaces . However, the calcium lactate antimicrobial properties have been scarcely studied. FC lettuce and carrots treated with 3 × 104 ppm calcium lactate showed similar microbial loads than 120 ppm NaOCl after 10 days at 4℃ . Similarly, 3 × 104 ppm calcium lactate treatment at 60℃ of FC melon induced 1–2 log CFU g−1 lower bacterial and yeasts and mold loads after 8 days at 5℃, while texture of such melon pieces was better maintained than samples washed with water at the same temperature . Latter authors also found that other calcium salt treatments (calcium chloride and calcium propionate) maintained lower microbial loads in the FC melon samples after 8 days at 5℃. Calcium pre-treatments may also help to prevent enzymatic browning reactions during high-pressure processing (HPP) of peaches if the penetration of Ca2+ reaches the target area, which is the tonoplast or vacuolar membrane as observed in peaches .
Sodium bicarbonate, sodium carbonate, sodium silicate, potassium bicarbonate, potassium carbonate, and potassium sorbate have been studied . Among them, sodium carbonate and sodium bicarbonate (at 3% w/v) reduced up to 100% disease (
2.6. Electrolyzed oxidizing water
Electrolyzed oxidizing water (EOW) is formed in an electrolysis chamber by the electrodialysis of a NaCl solution between an anode and a cathode . Acidic EOW (pH 2.5–3.5; oxidation-reduction potential (ORP) 1000–1200 mV) is produced in the anode and alkaline EOW (pH 10–11.5; ORP −800 to −900 mV) is produced in the cathode. HCl, HOCl, Cl2, OCl−, and O2 are formed in the anode, while the cathode produces hydroxyl ions, which can react with Na ions generating NaOH. EOW contains free chlorine as the main microbial inactivation agent showing higher microbicidal effect against pathogens and spoilage microorganisms than NaOCl [16, 54, 55]. EOW is considered as an eco-friendly technology that presents the following advantages over other sanitizing methods: easy-to-find and cheap materials (NaCl and water), simple and on-site production, and low operational expenses and trihalomethanes formation. Additionally, some cell electrodes, like boron-doped diamond electrodes, are able to oxidize organic matter, reducing then the environmental footprint of wastewater from fresh produce industry . Nevertheless, EOW has a short shelf life, in some cases, being necessary to be produced on-site and recommended in a ventilated area due to the Cl2 and H2 production . EOW has been approved at maximum concentration of 200 ppm by the FDA . Neutral EOW (pH 7; ORP 700 mV) can be produced by the mixture of acidic and alkaline EOW . The additional advantages from using neutral EOW are that it does not affect surface color, general appearance, or pH of FC vegetables .
EOW has been used in several works as an excellent disinfestation method of food equipment surfaces and tools reaching up to 9 log CFU cm−2 reductions for several pathogen biofilms like
The EOW (15–50 ppm free chlorine) was early studied on FC carrots, spinach, bell pepper, potato, and cucumber being considered as an effective disinfectant able to reduce microbial loads by 0.6–2.6 log units without product discoloration . Neutral EOW (100 ppm free Cl; pH 7) achieved 0.5, 1.3, and >2.1 log mesophilic, psychrophilic, and yeast and mold reductions, respectively, in FC kailan-hybrid broccoli, showing similar microbial loads and good sensory quality to NaOCl-treated (100 ppm) samples after 19 days at 5℃ . Neutral EOW also showed similar microbial effectiveness to NaOCl to reduce natural microflora of FC lettuce with no impact on its physical and sensory quality [63–65]. Acidic and neutral EOW treatments at 70 and 100 ppm free Cl were studied on two broccoli varieties showing neutral EOW (100 ppm) the best microbial reductions after shelf life comparing to NaOCl (100 ppm) . Furthermore, EOW-treated samples showed higher (up to 30%) total phenolic content and more stabilized myrosinase activity (the enzyme responsible for the formation of the bioactive isothiocyanates (ITC) in broccoli) than NaOCl-treated samples after shelf life . Microbial reductions of 1–2 log units were observed in FC mizuna baby leaves treated with acidic EOW (40–100 ppm free Cl) and neutral EOW (40–100 ppm free Cl), with similar microbial effectiveness to NaOCl 100 ppm, showing neutral EOW better bacteriostatic effects than acidic EOW in some cases . The sensory quality, physical structure, and health-promoting compounds of EOW-treated FC mizuna baby leaves were not significantly affected . Neutral EOW (100 ppm free Cl) treatment reduced counts of inoculated
The use of different organic and inorganic salts has been studied to increase electrolysis efficacy and avoid the corrosive effects of NaCl on equipment. Particularly, electrolyzed sodium bicarbonate allowed to control postharvest citrus rots as a result of direct inhibition and the induction of fruit resistance-response mechanisms showing normal electrolyzed water a less marked effect .
Ozone (O3) is a colorless gas with a pungent odor having an oxidizing potential (+2.07 V) 1.5 times higher than that of chlorine, which oxidizes the cell components of the microbial cell wall . Ozone half-life is very short, from seconds to hours depending on temperature and water quality . Thus, O3 is commercially generated on-site by submitting oxygen, or air, to ultraviolet radiation (285 nm) or through an electrical charge leading to the cleavage of oxygen molecules to form ozone [5, 72].
Ozone solubility is 12 times lower than that of NaOCl. Nevertheless, ozone concentrations as low as 1–5 ppm are enough to reach good antimicrobial reductions. Nevertheless, higher O3 concentrations are needed when it is applied as a gas treatment since its penetration into the cells, to achieve the disinfection effect, is affected by the humidity of the air [40, 73]. The O3 effectiveness may be increased at lower pH (more stable) and temperature (higher residual ozone concentrations), higher relative humidity of the storage room (increasing its solubility on the moisture present on produce surfaces), and purity of the water (ozone is consumed by the matrix components reducing its efficacy) . Ozone is spontaneously decomposed to the non-toxic O2 when applied. Accordingly, O3 has been approved as a GRAS product by the FDA to be used in the food industry . However, O3 can cause irritation to eyes and throat (at concentration higher than 0.2 ppm) of plant operators, is highly corrosive to the equipment, and the physicochemical properties of treated produce may be altered .
An ozonated water treatment at 0.4 ppm for 3 min has been recommended to maintain microbiological quality and firmness of tomato slices while it did not affect the physicochemical quality and organic acid contents . The levels of
Nevertheless, O3 seems to be not always highly effective. In that sense, low (<0.5 log units) inhibitory effect on mesophilic, psychrophilic, enterobacteria, molds, and LAB loads of FC “Galia” melon washed with ozonated water (0.4 ppm, up to 5 min) was observed after 10 days at 5℃, registering even higher yeast and molds compared to untreated samples . However, FC ‘Galia’ melon washed for just 1 min with 80 ppm PAA showed the lowest microbial loads after 10 days at 5℃. However, the latter PAA antimocrobial effect was reduced when it was combined with ozonated water treatment. On the other side, FC “Galia” melon samples treated with ozonated water reduced respiration rates, while sugar contents and vitamin C were better maintained . Shredded “Iceberg” lettuce treated with ozonated water (1 ppm, 120 s) showed lower sensory and microbiological quality than samples treated with NaOCl (200 ppm, 120 s) .
2.8. Essential oils
High antimicrobial properties have been studied with several plant essential oils (EOs) . Generally, EOs possessing the strongest antibacterial properties are those that contain phenolic compounds such as carvacrol, eugenol, and thymol [87, 88]. In general, the mechanism of action of EOs against microorganisms involves the interaction of phenolic compounds with the proteins (porins) in the cytoplasmic membrane that can precipitate and lead to ions leakage of and other cell contents causing cell lysis . Carvacrol solutions have shown good antimicrobial effects in different FC fruit and vegetables like lettuce, kiwifruit, apples, and melons treated with carvacrol-containing washing solutions [90–92]. Thymol (from thyme and oregano) and eugenol (from clove,
The EOs concentrations should be increased when tested
Isothiocyanates (ITC) are sulfur compounds that can be formed in the
3. Biological-based methods
Bacteriocins are toxins of protein nature that are synthetized by bacteria to inhibit the microbial growth of similar or closely related bacterial strains. The solubility of bacteriocins may increase at lower pH, facilitating diffusion of bacteriocin molecules . Nisin is a food-grade bacteriocin produced by
3.2. Biological control
The use of biological preservation (strains of
Bacteriophages are viruses that infect and replicate in a bacteria causing their lysis and death . Bacteriophages were earlier studied on FC fruit like apples and melons .
4. Physical-based treatments
4.1. Mild heat treatments
The use of mild heat treatments is a promising sanitizing technique for the FC industry, which may extend the FC product shelf life through microbial destruction and partial enzymatic inactivation. However, the treatment temperature and exposure time should be carefully selected for each product due to possible undesirable changes of sensory and nutritional quality. Fruit and vegetables treated by heat treatments within the range 40–60℃ for 1–5 min, depending of the commodity, are still considered as a fresh product as it is defined [17, 121].
Hot water and vapor treatments have been studied in several fresh-cut commodities like pomegranate arils [122, 123], kiwifruit , lemons , peaches , apples , sunchoke , lettuce [46, 129, 130], rocket , spinach [132, 133], celery , eggplants , and onions . PPO activity of fresh-cut pomegranate arils was 1.3-fold reduced by a hot water treatment at 55℃ for 30 s, while total anthocyanins contents were similar to those of untreated samples after 7 days at 5℃ .
Short vapor treatments, usually up to 15 s, usually steam jet-injection systems, have been also used as an alternative to hot water treatments due to less impact on sensory quality of the FC product. Accordingly, vapor treatment (95℃; 7–10 s) kept better quality of FC pomegranate arils  compared to the hot water (55℃; 30 s) treatment  during storage at 5℃, increasing the shelf life from 7 to 18 days. Furthermore, vapor heat treatments reduced up to 2-fold the total antioxidant capacity losses observed in fresh-cut pomegranate arils sanitized by conventional NaOCl treatment during shelf life . Steam jet-injection treatment of fresh-cut lettuce for 10 s reduced respiration rate, partially inactivated browning-related enzymes and kept the mesophilic load as low as with a conventional NaOCl treatment . Nevertheless, further research is needed to optimize the exposure conditions for FC commodities.
Heat treatments by microwave (750 W for 45–60 s) have been recently proposed to reduce natural microflora of FC carrots, which also prevented whitening and surface drying of samples during storage up to 7 days at 5℃ . The increased microbial growth due to plant cell disruption after the heat treatment may be controlled with the use of combined storage under modified atmosphere packaging.
4.2. UV radiation
Ultraviolet (UV) light is an electromagnetic radiation divided in four groups: UV-A, UV-B, UV-C, and vacuum UV . UV-C radiation (
The UV dose (
The UV-C effectiveness on natural microflora has also been studied in several FC fruit and vegetables. UV-C treatments (0.49, 4.9, and 9.8 kJ m−2) of FC zucchini slices reduced microbial activity and deterioration during subsequent storage at 5 or 10℃ . Mesophilic loads of FC pomegranate arils were reduced by approximately 1 log units after 4.5 kJ m−2, while yeasts were reduced >1.8 log units . Similarly, UV-C radiation doses (4.5–9 kJ m−2) of the kalian-hybrid broccoli reduced mesophilic loads by approximately 1.2 log units while enterobacteria and psychrophilic were unaffected [62, 154]. However, combination of UV-C with NEW in kalian-hybrid broccoli or with hot water (55℃ for 30 s) in FC pomegranate arils did not achieve further microbial inactivations [62, 139]. FC tomatoes treated with a UV-C dose of 4 kJ m−2 and stored for 21 days at 12℃ under MAP (5 kPa O2 +1 kPa CO2) retarded ripening and maintained better firmness and sensory attributes than UV-C treated samples stored under air conditions . The range of 0–2.5 kJ m−2 UV-C dose achieved the most important mesophilic reductions in treated date palm . The UV-C sanitizing effect has been studied in a wide FC products such as tomato , strawberry , watermelon , potatoes , and lettuce , among others.
Increases in bioactive compounds of several fruit and vegetables after UV treatments have been reported. Such enhancements of health-promoting compounds have been reported to be a consequence of the free radicals generated during irradiation that might act as stress signals and trigger stress responses leading to the observed bioactives increments . Broccoli exposed to several UV-C doses (1.5–15 kJ m−2) registered increases in its polyphenols content (up to 25%) after 19 days at 5℃ . FC tomatoes UV-C treated (0.97 kJ m−2) showed higher total phenolic content than other samples treated with hot water (40℃, 30 min), ultrasounds (45 kHz; power of 80%; 30 min), or its combination, after 30 days at 10℃ . The lycopene content of watermelon was preserved with a UV-C dose of 2.8 kJ m−2, although a lower dose of 1.6 kJ m−2 did not show the same benefit . However, immediate bioactive increments after UV-C have been observed in several FC fruit and vegetables, being probably attributed to an enhanced compound extraction due to plant cell disruption as a consequence of the UV radiation. Therefore, phenolic compounds and flavonoid contents of FC mangoes were increased after UV-C doses of 2.46 and 4.93 kJ m−2 . Hydroxycinnamoyl acid derivatives of FC broccoli were also increased by 4.5–4.8-fold after UV-C treatments (4.5–9.0 kJ m−2) . Total phenol and flavonoid contents of banana and guava were enhanced after UV-C treatment . FC carrot treated with UV-C (9 kJ m−2) showed higher chlorogenic content on total antioxidant capacity than untreated samples on processing day . Then, besides the interest of UV-C radiation as a microbial safety method, this non-ionizing radiation may also be used as a tool to enhance or better preserve the health-promoting compounds of plant products during shelf life .
Application of UV-B radiation (280–320 nm) has also been proposed as a friendly and cheap nonmolecular tool to enhance the phenolic compounds in carrots and other horticultural crops during postharvest life [166–168]. FC carrot shreds treated with a UV-B dose of 1.5 kJ m−2 showed 23% higher total phenolic content (mainly chlorogenic acid) than untreated samples after 72 h at 15℃ . UV-A has also been reported to induce biosynthesis of anthocyanins in cherries , although its effects has not been widely reported, and further research must be conducted.
Low-light conditions during storage have been recently proposed as an innovative and eco-friendly postharvest technique to highly prolong the shelf life of FC products. Accordingly, the shelf life of FC lettuce (butterhead and iceberg) was highly extended when it was stored under low-light conditions (≈5 μmol m−2 s−1 PAR; using either fluorescent or LED light) compared to samples stored under dark conditions . Thus, lighting delayed cut-edge browning, reduced ascorbic acid degradation while carbohydrates levels were highly increased, although light samples did not show net photosynthesis according to photosynthetic activity measurements. Latter authors hypothesized that the observed prolonged shelf life in lit samples could be due to the higher levels of sugar and ascorbate that may act as antioxidants, may maintain membrane integrity, and may supply enough respiratory substrate to prevent ATP depletion.
4.3. Pulsed light
Pulsed light (PL) is a preservation technology that involves the use of intense short duration (1 μs–0.1 s) pulses of polychromatic light from UV to near infrared (100–1100 nm) emitted by an inert gas (e.g., xenon) lamp . The microbicidal action of PL has been attributed to different mechanisms: photochemical, photothermal, and photophysical . Several studies have shown the effects of PL treatments on inoculated microorganisms, native microflora, and quality aspects of FC spinach, lettuce, cabbage, carrot, mushrooms, avocado, and watermelon [174–179]. Microbial reductions up to 2.2 log units have been reported in different products such as lettuce, celery, spinach, bean sprouts, white cabbage, and green bell pepper, with the different antimicrobial effectiveness in different produce dependent on the location of microorganisms as well as the presence of protective substances present in the product [179, 180]. Further investigation may cover the reported microbial photoreactivation after PL treatments [181–184] and the PL efficiency due to shadow effects that is one of the main industrial limitations of PL technology.
4.4. Pulsed electric fields
Pulsed electric fields (PEFs) are based on the application of DC voltages for very short periods of time, usually μs, to the food material which is placed between two electrodes. PEF equipment consists of a treatment chamber, pulse generator, control system, data acquisition, and material-handling equipment . PEF has been successfully applied for microbial inactivation in liquid food systems, although its application in plant tissues is limited due to the PEF-related plant cell disruption processes known as membrane breakdown, membrane permeabilization, or electroporation of the membrane . Accordingly, PEF treatment (wave bipolar pulses at 2 kV cm−1 electric field strength, 1 μs pulse width, and 100 pulses s−1) applied to blueberries (immersed in a saline fluid for PEF transmission) for just 2 min achieved 1 and 2 log unit reductions of inoculated
4.5. Cold plasma
Plasma is generated when an inert gas is in contact with electricity and is being considered as the fourth state of matter. Plasma is composed by charged particles, excited molecules, reactive species, and UV photons which induce microbial inactivation . Plasma is generally classified as cold (non-thermal) and thermal plasma. Thermal plasma generation requires temperature and high pressure with heavy electrons. Cold plasma is generated at temperatures of 30–60℃ under atmospheric or vacuum requiring low energy . Among the main advantages of cold plasma are lower cost operating temperature and water consumption, together with timely production of the acting agents and lack of residues during production when compared to thermal and chemical treatments [190–192]. Cold plasma can be generated using either of the following devices: resistive barrier discharge, dielectric barrier discharge, corona discharge, radio frequency discharge, glow discharge, and atmospheric pressure plasma jet . Plasma jet may be considered as the fastest plasma generation method to achieve microbial inactivation (4.3 ± 6.5 min) . The most widely used gas in the published research has been air, followed by pure Ar, mixtures of He/O2 and Ar/O2, and pure N2 . The plasma inactivation capacity depends on several factors like the type of technology used to generate the plasma, the voltage, the feed gas, the treatment time, the species, the direct or indirect exposure and the concentration of the tested microorganisms, and the structural characteristics of the produce . Cold plasma has a high potential to be used in the industry for fruit and vegetables according to studies of the last few years as recently reviewed . Generally, plasma treatments are able to achieve microbial reductions of 2.7 ± 1.4 log units, ranging from 1.5 ± 1.0 log units for bacilli and spores to 3.3 ± 1.6 Log for
Ultrasounds (US) are sound waves with amplitude higher than the upper audible limit of human threshold (above 20 kHz) that generate cavitation bubbles . The antimicrobial properties derived from US is based on the combination of mechanical (responsible for the disinfection action leading to detachment) and chemical energy (responsible for the free radicals formation leading to destruction), produced from the collapse of latter generated bubbles, which increase the cell membranes permeability . Consequently, DNA modifications of microbial cells are formed due to formed hot spots (due to collapse), with high temperatures and pressure, and released free radicals . The US treatment should always ensure that US pressure levels (70 dB at 20 kHz or 100 dB at ≥25 kHz) are not surpassed according to UK Health Protection Agency recommendations . The effectiveness of microbial inactivation achieved with US is influenced by microbial cell shape (high resistance of coccus), size (bigger cells are less resistant), Gram type (negative are less resistant), and cellular metabolism (anaerobes are less resistant) . Among treatment parameters, US effectiveness is influenced by fluid temperature (optimum at 60℃ which may be reduced to 20℃ to avoid losses of produce sensory quality without highly affecting microbial US inactivation), water hardness, and dissolved gases content . US treatment (40 kHz, 50 W) for 5 min of strawberries initially reduced natural mesophilic microflora by approximately 1 log unit . However, latter authors showed that such initial antimicrobial effect was lost after 5 days at 8℃ since similar reduction logs (regarding unwashed samples) to sterile water-washed (5 min) samples were achieved. On the other hand, such antimicrobial effects were maintained up to 9 days at 8℃ when US treatment was combined with PAA (40 ppm) even showing a synergistic effect on mesophiles and the highest initial reductions on inoculated
4.7. High-pressure processing
High-pressure processing (HPP) uses elevated pressures, with or without the addition of heat, also called high-hydrostatic pressure processing since water is the most used pressure-transmitting fluid . HPP is a promising eco-friendly sanitation treatment that may have a potential application in the FC industry. HPP may reach very high microbial inactivations as reviewed  while maintaining, or even enhancing, sensory properties of food products like aroma and taste. However, although the texture of tissues of firm FC products with low amounts of entrapped air remains unaffected, HPP may induce some alterations like water-soaked appearance of the product . Furthermore, HPP may either enhance (leading to enzymatic browning reactions due to loss of membrane permeability and sub-cellular compartmentalization) or inhibit the activity of enzymes related to cell wall degradation in FC products . HPP treatment (200–400 MPa, 3 min, 25℃) of FC persimmon induced changes in physicochemical quality (electrolyte leakage, texture, total soluble solids, pH, and color), which were a function of the amount of applied pressure compromising the consumer acceptance of the product . However, latter authors reported that HPP may improve carotenoid extractability and tannin polymerization of FC persimmon, which could enhance its functionality and eliminate astringency, respectively. Plant cell membranes of FC peaches have been tried to be stabilized prior to HPP treatment (200 MPa for 10 min; 23–28℃) through penetration of Ca2+ into the plasma membrane using calcium chloride or calcium lactate soaking treatments (1–2% w/v for 5 min). Nevertheless, latter authors reported that loss of cell integrity due to HPP was not avoided with the calcium soakings probably due to a low Ca2+ penetration into the tonoplast membrane being recommended for future research a higher calcium concentration and/or improved Ca2+ impregnation (e.g., using vacuum infusion). Higher pressure levels (585 MPa) were more effective to inactivate enzymes and to preserve color of FC peaches than longer times being optimized a HPP treatment of 585 MPa for 1 min . Nevertheless, due to the known baroresistance of some enzymes, like polyphenol oxidase (PPO), browning reactions may not be completely avoided during FC product shelf life, although MAP could limit such enzymatic activities due to low oxygen levels.
High-pressure carbon dioxide (HPCD) treatment has been proposed as another antimicrobial method being applied in FC carrots at 12 MPa (40℃) for 15 min and leading to complete inactivation of natural microflora being maintained after 28 days at 4℃ together with the enzymatic stability . HPCD treatment (12 MPa; 40℃; 20 min) of inoculated FC coconut also achieved
5. Packaging under non-conventional gas mixtures
Modified atmosphere packaging (MAP) is a postharvest preservation technique based on the packaging of a perishable product within an atmosphere that has been modified compared with air conditions. There are two types of MAP: active or passive. Active MAP consists in the replacement on the initial present gases by a desired mixture. Passive MAP is progressively generated as a result of respiration of the product and gas transfer through the film, which has a selected permeability to gases, until the desired gas equilibrium atmosphere is reached.
The use of superatmospheric O2 concentrations (>75 kPa O2) during modified atmosphere storage (HO—high oxygen conditions) reduces aerobic and anaerobic microbial growth, prevents anaerobic fermentation, avoids non-desirable flavor changes, and inhibits enzymatic browning. The microbial toxicity to HO may be explained due to the unfavorable effects on the oxidation-reduction potential of the system, the oxidation of enzymes having sulfhydryl groups or disulfide bridges, and the accumulation of toxic reactive O2 species . HO-controlled atmosphere (75 kPa O2 balanced with N2) inhibited the mesophilic count of FC lettuce during storage for 10 days at 7℃ [215, 216]. Chinese bayberries, strawberries, and blueberries stored under HO-controlled atmospheres (60–100 O2 kPa balanced with N2) inhibited decay during storage at 5℃ and subsequent 2 days at 20℃, while chemical parameters and surface color were only slightly affected compared to samples stored under air conditions .
High CO2 levels (maximum limits depending on the produce due to generation of related off-flavors) have also shown antimicrobial effects which are even stronger at low temperature because of enhanced CO2 solubility. Accordingly, controlled atmosphere with 15 kPa CO2 + 5 kPa O2 (balanced with N2) showed similar inhibitory effect to high O2 on mesophilic growth of FC lettuce than samples stored at 0 kPa CO2 + 75 kPa O2 (balanced with N2) . Interestingly, a combined high O2/CO2 effect may be obtained using active HO MAP. The latter beneficial gas conditions are a result of the produce respiratory activity that generates antimicrobial CO2 levels, while O2 is inevitably reduced due to a combined effect of respiration and film diffusivity processes. Hence, the addition of CO2 is unnecessary when high O2 atmospheres are injected during active HO MAP. Accordingly, kailan-hybrid broccoli stored under high O2/CO2 (initial O2/CO2 of 70/0.02 kPa changing to 50/30 after 19 days at 5℃) showed 2.8 log units lower natural microflora load safter 19 days at 5℃ compared to samples stored under passive MAP conditions (1.5–3.0 kPa O2 + 16–21 kPa CO2) . Similarly, active HO MAP of inoculated kailan-hybrid broccoli showed 1.4 and 2.3 lower
Enzymatic browning of FC produce has been shown to be reduced under HO atmospheres. Accordingly, active HO MAP (80 kPa O2 + 20 kPa CO2) delayed browning of FC lettuce during storage for 10 days at 5℃ . Active HO MAP (initial O2/CO2 of 80/0 kPa, balanced with N2) of FC pomegranate arils reduced enzymatic browning related to PPO [123, 137], while formed off-odors were highly controlled with active high CO2 MAP (20 kPa CO2 balanced with N2) . Active HO MAP of FC celeriac, mushrooms, and chicory endives (initial O2/CO2 of 95/0 kPa, balanced with N2, changing to 10–20/10–50 after 7 days at 4℃) were more effective to control enzymatic browning than low O2 atmospheres . It is hypothesized that high O2 may cause substrate inhibition of PPO or high contents of colorless quinones formed cause feedback PPO inhibition. High O2 levels kept the initial color and firmness of fresh-cut melon retarding anaerobic fermentation better than low O2 atmospheres . Pre-treatment of whole “Spartan” apple with 100 kPa O2 (up to 19 days at 1℃) before cutting decreased surface browning, flesh softening, and off-flavor in FC apple slices . Such inhibition of enzymatic browning was related to retention of cellular integrity while in low O2 pre-treatment (1 kPa O2, balanced with N2) would have another inhibitory browning mechanism on apple slices. Furthermore, the 100 kPa O2 pre-treatment before slicing apples can reduce the dependence on antioxidant additives to inhibit slices browning. Sensory quality of produce stored under high O2 atmospheres has been shown to be better than low O2 atmospheres due to fermentation processes .
High O2 levels are also considered as postharvest abiotic stresses able to increase PAL activity and consequently phenolic biosynthesis as observed in carrot shreds stored under high O2-controlled atmosphere (80 kPa O2 balanced with N2) . Furthermore, high O2 and CO2 levels may be tolerated by FC carrots maintaining their fresh characteristics and reducing microbial growth . Such abiotic stress may be used as a tool to increase the health-promoting compounds of plant material to subsequently obtain functional beverages after correspondent thermal or non-thermal treatments to ensure microbial quality and safety .
The use of MAP under mixtures of non-conventional gases such as Ar, He, Xe, or N2O has been proposed to maintain the quality of FC produce extending its shelf life. The latter gases may be chemically inert, but they have some physiological and/or antimicrobial properties, even though it does not seem to be through modification of enzyme activity . Ar, He, or N2 atmospheres mixed with low O2 showed different diffusive properties, since Ar and He are monoatomic and smaller in size than N2 . Treatment of asparagus spears for 24 h at 4℃ under an atmosphere of Ar and Xe at 2:9 (v:v) reduced RR and bract opening leading to a subsequent shelf life of 12 days at 4℃ showing better quality than those samples treated with an atmosphere of 5 kPa O2 + 5 kPa CO2 . Microbial quality and some bioactive compounds were highly preserved in FC red chard baby leaves stored under active He MAP (100%) during 8 days at 5℃ . N2O has a direct effect on cell metabolism achieving shelf life extension of FC produce. It may be explained since N2O has 77% solubility in fruit cell, while its absorption in tissues is completely reversible . FC spinach leaves stored under active N2O MAP (100%) showed low microbial growth after 8 days at 5℃, with chlorophylls and phenolics being well preserved . Different N2O and N2 combinations (including always 3% O2) were used as active MAP for FC lettuce and wild rocket during storage up to 12 days at 5℃, suggesting such results that N2O does not improve the produce quality compared to N2. Lower microbial loads have also been observed in FC watercress and arugula leaves at the end of cold storage when active MAP containing N2, Ar, He, Xe, or N2O were used [233, 234].
6. Future research needs and conclusions
FC plant produce is greatly vulnerable to microbial spoilage, and cultivar selection is probably the most important factor in FC overall quality and shelf life. With the intention of better inhibition of microbial spoilage, and subsequently decrease in decay and safety problems, genetic cultivar selection should turn to retard ripening and senescence, low ethylene production and/or sensitivity, and enhanced firmness, well adapted to minimal processing and increased antioxidant systems. While chlorine is widely used by the FC industry to ensure safety, new eco-friendly techniques/technologies are needed to replace the latter chemical treatment due to the production of carcinogenic compounds. Several eco-friendly strategies such as ozone, UV-C light, natural antimicrobial substances, GRAS chemical, and biological compounds can decrease microbial loads of FC products and extend their shelf life. Other advanced techniques such as pulsed electric fields, ultrasounds, and high-pressure processing, among others, are promising sanitizing strategies for the FC industry due to high microbicidal rates. However, the treatment parameters of latter technologies to be applied in FC products need further research to be optimized for each specific commodity. Furthermore, the potential and limits of these innovative eco-friendly techniques must be well defined and included in the regulations. In that way, modeling tools to predict microbial inactivation and product shelf life are very useful, principally to optimize production and distribution. Application of nanotechnology to FC products is also a promising future in order to produce products with extended shelf life and excellent quality meeting always the food safety. Nevertheless, regulations related to nanoparticles’ inclusion in food products need to be better defined by institutions. The combination of well-designed integrated production, handling, processing, and distribution chains for FC produces is crucial for achieving the high quality and safety demanded by consumers.
The authors are grateful to European Union, DG, for Science Research and Development (CRAFT Program Contracts SMT4-CT98-5530, QLK1-1999-707917, and OLK1-CT-2002-70791), 7th Framework Programme (FP7-BBBE.2013.1.2-02), Spanish Ministry for Education and Science and Spanish Ministry of Economy and Competitiveness (Projects ALI-95-0001, ALI-98-1006, AGL2005-08189-C02-01/ALI; AGL2007-63861/ALI; AGL2010-19201-C04-02-AGR; AGL2013-48830-C2-1-R), the Consejería de Educación y Cultura de la Comunidad Autónoma de la Región de Murcia (SUE-OA 06/02-0008), and Fundación Séneca de la Región de Murcia in Spain (Project AGR/92, PS96/CA/d3, AGR/3/FS/02 and 00553/PI/04) for financial support. We also acknowledge the following companies for financing research and development contract with us: Repsol Petróleo S.A. (Madrid), Plásticos de Alzira, S.A., Sakata Seed Iberica S.L.U. (Valencia), Primaflor SAT (Almería), Canarihorta (Las Palmas de Gran Canaria), Frutas Hermanos Mira S.L. (Alicante), Kernel Export S.L., Frutas Esparza S.L., Perichán SAT, Pozosur S.L. (Murcia), and Sakata Seed (Valencia). The good skills and work of all researches whenever involved in the Postharvest and Refrigeration Group are very much appreciated.