Phytochemicals are a heterogeneous group of substances found in all plant products, therefore constituting an important component of human diets. It is estimated that there are 250 000–500 000 plant species on Earth (Borris, 1996) and approximately 1–10% of these are used as food by human or other animals. Besides food, plants have also been used for centuries as remedies for human diseases because they contain components (phytochemicals) with therapeutic values. Phytochemicals have been isolated from several herbs and plants, and they have shown potent biological activities (Cowan, 1999; Negi et al., 1999; Beuchat, 2001; Burt, 2004; Negi and Jayaprakasha, 2004; Jayaprakasha et al., 2007; Tiwari et al., 2009; Raybaudi-Massilia et al., 2009; Negi et al., 2010; Negi, 2012). Food can be used as a vehicle for the delivery of phytochemicals that provide health benefits for increased well-being. Consumption of foods rich in phytochemicals has been reported to protect against various degenerative diseases, but their beneficial properties may alter as food products undergo processing and subsequent storage prior to consumption, which can affect stability of phytochemicals. Given the recent trend of health promotion through diet, understanding processing and storage effects is critical for conserving active phytochemicals. Simultaneously, there should not be any compromise on maintaining quality and enhancing shelf life of the food after addition of phytochemicals.
Phytochemical content in plants can vary greatly with variety, maturity, growing conditions, and agro climatic factors; therefore it is difficult to pinpoint whether the differences in composition of commercial samples are due to agronomic reasons or degradation during storage and processing methods followed prior to storage. In general, carotenoids are very susceptible to degradation, and the major mechanism of degradation is oxidation. The rate of carotenoid oxidation depends on carotenoid structure, oxygen, temperature, light, water activity, pH, enzymes, presence of metals and unsaturated lipids, type and physical state of the carotenoids present, severity and duration of processing, packaging material, storage conditions, and the presence of pro- and anti-oxidants (Namitha and Negi, 2010). Anthocyanins are destabilized by heat, high pH, light exposure, dissolved oxygen, and enzymes such as PPO, whereas copigmentation with acids or other flavonoids and metals enhances their color during storage (Fennema and Tannenbaum, 1996). Anthocyanin stability during storage follows first order kinetics (Garzon and Wrolstad, 2002; Turker et al., 2004; Brenes et al., 2005). Anthocyanins are found in their monomeric form in fresh fruit and juices, and the monomeric anthocyanins may undergo a condensation reaction to form polymeric pigments during storage (Bishop and Nagel, 1984; Dallas et al., 1996; Es-Safi et al., 2000; Hillebrand et al., 2004), which results in greater color stability of the matrix during further processing and storage (Gutierrez et al., 2004). Light exposure reduces betalain stability and metal cations are capable of accelerating betalain degradation (Herbach et al., 2006). Both betanidin and betanin were reported to be unstable in the presence of oxygen (Pasch and von Elbe, 1978; Schwartz et al., 2008).
The phytochemical stability of freeze-dried apple products during storage (up to 45 days) at 30 °C was most affected by aw and highest losses were observed at highest moisture activity (Corey et al., 2011). Phytochemical degradation for added green tea extracts also occurred more rapidly at higher moisture contents, except for caffeine, which was stable throughout the storage period irrespective of moisture content. Similarly, absorption of moisture by the acerola concentrate affected the stability of phytochemicals and almost half of the phenolics were lost within the first hour of storage at 65% RH (Ikonte et al., 2003). Superior stability of Amaranthus pigment powders as compared to the respective aqueous solutions may be attributed to lower aw values. Spray drying was found to increase stability of betacyanins, probably by increasing dry matter. Therefore, it was recommended that moisture content of pigment should be kept below 5% for enhancing their stability (Cai et al., 2005). Some matrix compounds like pectin, guar gum, and locust bean gum were shown to enhance storage stability of red beet solutions, probably by lowering the aw value (Herbach et al., 2006).
The stability of a phytochemical depends on storage environment and length of storage. It has been stated that temperature is the most important factor to affect the overall stability of various phytochemicals during storage (Rodrigues et al., 1991; Su et al., 2003). In Hawthorn fruit, Procyanidin B2 (PC-B2), (-)Epicatechin (EC), Chlorogenic acid (ChA), Hysperoside (HP), and Isoquercitrin (IQ) were stable for six months at a storage temperature of 4 °C. At room temperature their stability varied from relatively stable (HP and IQ, 8% loss), to intermediate stable (ChA, 30% loss), to quite unstable (EC and PC-B2, 50% loss), however, these compounds were unstable at the higher temperature of 40 °C (Chang et al., 2006). In Hawthorn drink also these compounds were stable for six months at lower temperatures (4 °C) and relatively unstable at higher temperatures (23 and/or 40 °C). Wang and Stretch (2001) observed that in cranberries storage temperature has profound effect on anthocyanins content as storage at 15 °C promoted anthocyanins biosynthesis as compared to storage at higher or lower temperature. Fresh cactus fruit of Yellow spineless also showed an increase in the concentration of active substances during three to four weeks storage at 5–8 °C (Nazareno et al., 2009). In milled rice also there was a consistent decrease in phenolic acid content during storage and the decline was greater at 37 °C than at 25 °C (Thanajiruschaya et al., 2010).
Uddin et al. (2002) studied the effects of degradation of ascorbic acid in dried guava during storage and observed that as the storage time and temperature increased, there was a progressive decrease in ascorbic acid content. Similarly, total vitamin C was found to decrease with increase in storage temperature and duration in green leaves (Negi and Roy, 2001a, 2001b) and carrots (Negi and Roy, 2000). Total vitamin C in grapefruit juice was retained higher at the lowest temperature (10 °C) at the end of 12 weeks of storage (Smoot and Nagy, 1990). However, Rodriguez et al. (1991) observed that ascorbic acid degradation of an alcoholic orange juice beverage was not influenced by temperature while other quality parameters such as degree of browning, accumulation of furfural, and limonene content were highly correlated with temperature of storage.
Fruit juices should be kept refrigerated to increase the stability of phytochemicals as the ascorbic acid present in most of the fruit combines with anthocyanins, which may be mutually destructive, more so at higher temperatures (Brenes et al., 2005; Choi et al., 2001). This destruction has been linked to the formation of dehydroascorbic acid breakdown products, but the exact mechanism for their adverse interaction has not been completely elucidated. Ascorbic acid stability is important in anthocyanin containing juice blends since the degradation products of ascorbic acid can degrade anthocyanins (Es-Safi et al., 1999, 2002; Brenes et al., 2005). Brenes et al. (2005) reported a 12% decrease in total anthocyanins in a grape juice (Vitis vinifera) model system with and without added ascorbic acid. Pozo-Insfran et al. (2007) reported that the added ascorbic acid decreased the anthocyanin stability, as opposed to the wine where polyphenolics helps to stabilize anthocyanins (Gutierrezz et al., 2004).
Juice containing cyanidin-3-glucoside retained 90% of its color for 60 days at 10 °C (Fossen et al., 1998), whereas at 25 °C storage color loss was 10–17%, which accelerated further to 35–49% at 38 °C. In thermally processed blueberry (Vaccinium myrtillus) juices degradation of anthocyanins was also significantly accelerated with increasing storage temperatures. Combined pressure temperature treatment (100–700 MPa, 40–121 °C) of pasteurized juice led to a slightly faster degradation of total anthocyanins during storage compared to heat treatments at ambient pressure (Buckow et al., 2010). Phytochemicals were stable after high hydrostatic pressure processing (400 and 550 MPa for 15 min) in ascorbic acid-fortified muscadine grape juice at 25 °C for 21 days, and addition of rosemary and thyme polyphenolic extracts increased muscadine grape juice color, antioxidant activity, and also reduced phytochemical losses during storage (Pozo Insfran et al., 2007). Addition of rosemary extract readily forms copigment complexes with anthocyanins in concentration-dependent manner and increases its antioxidant activity (Talcott et al., 2003a). Condensation of the other polyphenolic compounds with the anthocyanins may cause higher retention of polyphenolics, as observed in blood orange juice (Hillebrand et al., 2004). Guava juice showed a protecting effect on several of the juice blends, which is attributed to the guava polyphenolics forming more stable polymeric compounds with the anthocyanins (Bishop and Nagel, 1984; Dallas et al., 1996).
Storage of commercial tea leaves at 20 °C for six months resulted in a progressive decrease in the total phytochemical content, most of which were attributed to losses in the epigallocatechin 3-gallate and epicatechin 3-gallate (Friedman et al., 2009). Epigallocatechins were shown to be isomerized into (-)-catechin during storage at 40 °C after a few days (Komatsu et al., 1993; Wang and Helliwell, 2000). Changes in the antioxidant capacity of a green tea infusion were directly related to the changes in catechins that showed considerably higher stability at lower pH. Green tea catechins used in oil/water emulsions were found to decrease to 70% of the initial content at room temperature and almost negligible amount remained at 40 °C after six months (Frauen et al., 2000). Spanos et al. (1990) also observed a complete degradation of procyanidins, including catechin, epicatechin, and procyanidins B1, B2, B3, and B4, after the storage of concentrated apple juice at 25 °C for nine months. Similarly, isoquercetin and kaempferol 3-glucoside present in red raspberry jam decreased slightly after six months of storage (Zafrilla et al., 2001). In a model system, catechin was more stable in aqueous solutions stored in the dark as compared to illuminated storage, and its stability was further enhanced when stored at refrigeration temperature (4 °C) over a storage period of 80 days (Callemien and Collin, 2007). Degradation of catechins in fruit juice was greater at high storage temperature (23 °C) than low storage temperature (4 °C), and the degradation pathway was related to oxidative processes (Chang et al., 2006). Although, oxidation can occur under a variety of temperature conditions, reaction rates are generally faster at higher temperatures and depend on the dissolved oxygen in aqueous solution (Devlin and Harris, 1984; Alnaizy and Akgerman, 2000).
Individually quick frozen black raspberries retained anthocyanins during long-term storage at −20 °C, but heating followed by storage for six months resulted in dramatic losses in total anthocyanins ranging from 49–75% (Hager et al., 2008). Blueberries stored at −25 °C for six months after initial heat processing showed 62–85% losses in total anthocyanins (Brownmiller et al., 2008). Heat processing for different durations at 95 °C did not have an effect on the initial concentration of tea catechins, but it significantly influenced the stability of these compounds during storage. The heat treatment decreased the storage stability of all tea catechins, and the duration of heating was not a factor in polyphenolic stability. In green tea, mild heat pasteurization (85 °C) retains characteristic color and flavor better for longer storage duration than higher temperatures treatments (Kim et al., 2007). High temperature also induces a negative effect by lowering polyphenolic stability during storage regardless of time duration, which may be attributed to the loss of ascorbic acid by heat treatment.
A reduction in the carotene content of fresh vegetables irrespective of storage conditions has been reported. Unfavorable relative humidity and temperature has been shown to hasten the loss of carotenes during storage of fresh produce, wherein spinach lost almost 63.5% of the original carotenoids after wilting (Akpapunam, 1984). Negi and Roy (2003) reported up to 85% losses in β-carotene in fresh green leaves depending on duration and storage conditions with packaging helping in retaining higher β-carotene. During storage of fresh carrots, a steady decrease (Negi and Roy, 2000) and a slight increase followed by decrease (Lee, 1986) in β-carotene content have been reported. Total carotenoids in hand peeled carrot disks were significantly higher than fine or coarse carborundum plates abrasion peeled carrot disks throughout eight days of storage at 4 °C (Kenny and O’Beirne, 2010). During storage of tomatoes and their processed products all-trans-lycopene was more stable at 20 °C compared to −10, 2, and 37 °C, as re-isomerization from cis- to trans- is favored at this temperature (Lovric et al., 1970).
The Phytochemical composition of five varieties of black soybeans (Glycine max) and their stability at room temperature, 4 and −80 °C over 14 months were determined by Correa et al. (2010). No significant decrease was found in total phenols of black soybeans during storage for 14 months. On the other hand, lutein and γ-tocopherol degraded significantly within a month of storage at room temperature, whereas they remained stable up to six months at 4 °C and up to 14 months at −80 °C. Storage at low temperature can reduce the loss of fat-soluble phytochemicals in black soybeans over an extended period of time; however no significant decrease occurs in total phenols even at room temperature for 14 months.
Koski et al. (2002) found that the content of α-tocopherol in cold pressed rape seed oil declined to nil from the initial value of about 200 mg/kg in fresh oil within 7–11 days of storage at a temperature of 60 °C, while γ-tocopherol was retained to 5–10% of the initial value (600 mg/kg) after two weeks of storage. Morello et al. (2004) also found that α-tocopherol was totally absent in olive oil after 12 months of storage at room temperature, but at lower temperatures a slower rate of reduction of α-tocopherol (60% loss after 12 months) in virgin olive oil was observed (Okogeri and Tasioula-Margari, 2002).
Kopelman and Augsburger (2002) determined the influence of capsule shell composition and sealing on the stability of the phytochemical in fresh and formulated Hypericum perforatum extract capsules stored at 25 °C/60% RH for 60 days. Phytochemicals had varying stability towards capsule shell composition and sealing. Except with gelatin capsules of neat Hypericum perforatum extract, sealing of the capsules did not offer much protection. Neat Hypericum perforatum extract was typically more sensitive to the effects of shell composition and sealing relative to formulated Hypericum perforatum extract.
Long-term storage at −24 °C of raw carrot cubes reduced the falcarinol content by almost 35%. Blanching before storage reduces almost one-third of the falcarinol content of carrot, although no further reduction in the falcarinol content was reported after steam blanching during long-term storage (Hansen et al., 2003). Studies testing the stability of Policosanol (PC) supplement under environments that favor acid hydrolysis, basic hydrolysis, oxidation, photolytic degradation, and thermolysis indicated a shelf life of five years for the original PC supplement (Mas, 2000; Cabrera et al., 2002; Castano et al., 2002; Cabrera et al., 2003). Storage temperature has no effect on the degradation of bixin during initial storage period, but at later stages the degradation accelerated with temperature, and it followed second-order rate kinetics. The reaction rates increased by increasing the interaction between oxygen molecules and bixin, and temperature had a positive effect on reaction rate causing faster degradation (PrabhakaraRao et al., 2005).
Besides water activity and temperature, light exposure is also an important factor to influence the stability of phytochemicals during storage (Schwartz et al., 2008). It is known that light induced oxidation of carotenoids, proteins, lipids, and vitamins are common in many food systems (Wishner, 1964; Pesek and Warthesen, 1987; Solomon et al., 1995). Carotenoids are very susceptible to degradation, particularly once they have been extracted from biological tissues. The major cause of carotenoid destruction during the storage of food is oxidation (Zanoni et al., 1998), and they are susceptible to oxidation when exposed to light (Saguy et al., 1985) and enzymes (Gregory, 1996), but reduced water activity of the medium has a protective role (Minguez and Galan, 1995). Oxidation of carotenoids occurs as a result of either auto-oxidation in the presence of oxygen, or by photo-oxidation in the presence of light (MacDougall, 2002). Overall, the rate of carotenoid oxidation depends on carotenoid structure, oxygen, temperature, light, water activity, pH, metals, enzymes, presence of unsaturated lipids, type and physical state of the carotenoids present, severity and duration of processing, packaging material, storage conditions, and the presence of pro- and anti-oxidants (Rodriguez-Amaya, 2003; Namitha and Negi, 2010). Oxidation of carotenoids results in the formation of colorless end products such as compounds with epoxy, hydroxyl, and carbonyl groups (MacDougall, 2002). Therefore, while designing the delivery system for their use as functional food ingredients, appropriate measures should be taken to protect them. Addition of carotenoids to functional foods should be done by incorporating into edible oil as it makes them more bioavailable than carotenoids in a plant cellular matrix (Lakshminarayana et al., 2007).
Lycopene stability in products such as guava nectar is also a function of light, water activity, oxygen, pH, temperature, and the presence of pro-oxidants or antioxidants (Chou and Breene, 1972). Oxygen-independent reactions affect Yellow passion fruit juice (Passiflora edulis) color and antioxidant activity, and ascorbic acid and sucrose fortification increases stability of carotenoids (Talcott et al., 2003b). Dehydrated vegetables lose color due to the oxidation of highly unsaturated molecules upon exposure to air during storage and β-carotene degradation is associated with the development of an off flavor in dehydrated carrots (Ayer et al., 1964).
The three most predominant phenolic compounds in tea (ECG, EGCG, and EGC) showed higher stability at lower temperature in the dark, indicating that the two storage conditions (temperature and light) were a significant factor to influence phenolic stability during green tea storage (Callemien and Collin, 2007). During storage for six months of virgin olive oil under diffused light in the temperature range of 6–18 °C, an almost 60% decrease in the total phenols occured, whereas storage in darkness resulted in a decrease of 50% of total phenols after 12 months (Okogeri and Tasioula-Margari, 2002). Tsimidou et al. (1992) also found significant losses of phenolic compounds in virgin olive oil stored in the dark at 20 °C in closed bottles. A significant decrease of phenol content in virgin olive oil after 12 months of storage in darkness at room temperature with subsequent loss of oxidative stability was also established in the study by Morello et al. (2004). The total anthocyanins contents in colored rice were retained under low O2 concentrations (0, 5, and 10%). Polyphenol contents significantly declined during four months of storage with free and soluble conjugated phenolic contents showing minimum losses at 0% O2 storage, whereas minimum loss of insoluble bound phenolics was detected in samples stored at 5% O2 (Htwe et al., 2010).
Ascorbic acid was not affected by light exposure in juice stored in air-tight containers for 52 days at 8 °C (Solomon et al., 1995); however, commercial juice in foil-covered bottles retained higher ascorbic acid than clear bottles during 18 days of storage at 3 °C (Andrews and Driscoll, 1977). Similarly, green tea stored in lightproof packaging retained higher ascorbic acid (Yaminish, 1996). Light exposure reduced betalain stability (von Elbe et al., 1974; Bilyk et al., 1981; Cai et al., 2005, Herbach et al., 2007) and detrimental effects of light were observed at temperatures below 25 °C, but no effect of light was observed at storage temperatures above 40 °C (Attoe and von Elbe, 1981; Huang and von Elbe, 1986). High ascorbic acid concentrations were capable of reducing betalain degradation (Pasch and von Elbe, 1978) and supplementation with ascorbic and isoascorbic acids was reported to enhance betalain stability by oxygen removal (Attoe and von Elbe, 1982).
Additive effects of light and oxygen were observed as light alone caused 15.6% betanin degradation, and oxygen alone caused 14.6% betanin degradation, whereas their simultaneous presence was responsible for 28.6% betanin decomposition (von Elbe et al., 1974). Attoe and von Elbe (1981) reported that light-induced degradation was oxygen dependent as the detrimental effects of light were found to be negligible under anaerobic conditions (Huang and von Elbe, 1986). Supplementation of red beet and purple pitaya juices with acids has been shown to inhibit light-induced betacyanin degradation during juice storage (Bilyk et al., 1981; Herbach et al., 2007). The exposure to light during storage showed a more pronounced decrease (15% at 38 °C) in color than those kept in the dark in rose extracts (PrabhakaraRao et al., 2005). Similarly, the effect of light on degradation of bixin was seen from the initial days of the storage period in both oleoresin and dye (Balaswamy et al., 2006). Betanin stability decreases linearly with increasing oxygen concentration (Czapski, 1985), and storage in a nitrogen atmosphere significantly increased its stability (Attoe and von Elbe, 1982; von Elbe and Attoe, 1985). In addition to oxygen, hydrogen peroxide was also reported to accelerate betanin degradation (Wasserman et al., 1984).
The stability of phenolic compounds is highly pH dependent and varies depending on the structural conformation. Flavan-3-ols show high storage stability under acidic conditions but are unstable in neutral pH (Komatsu et al., 1991; Suematsu et al., 1992; Zhu et al., 1997; Chen et al., 1998; Xu et al., 2003). Addition of acids confer stability to tea beverages since a lower pH is more effective for stabilizing tea catechins during storage (Chen et al., 1998), while neutral pH degraded tea catechins faster. Adding ascorbic acid or organic acids (citric and malic acid) to mimic citrus flavor improves storage stability and flavor of green tea (Aoshima and Ayabe, 2007), probably by lowering the pH. Moreover, ascorbic acid is more stable at lower pH, indicating that the protective effect on polyphenolics is higher at lower pH (Gallarate et al., 1999). Lowering pH is effective in slowing down the reduction of predominant compounds (chlorogenic acid and its isomers) formed during oxidative degradation (Schmalko and Alzamora, 2001). In green tea, changing pH affects the rate of hydrogen peroxide production, and when the pH of green tea infusion was lowered, the production rate of hydrogen peroxide and superoxide was significantly reduced (Akagawa et al., 2003). Presence of 3-deoxyanthocyanins, which lacks the hydroxyl group at 3 position of C ring in sorghum, increases the anthocyanin stability at high pH making it a good colorant for food use (Awika et al., 2004).
Polyphenols are readily oxidized during storage, which results in the production of H2O2 (Akagawa et al., 2003; Chai et al., 2003; Aoshima and Ayabe, 2007). The H2O2 produced during storage can degrade a polyphenol-rich product (Long et al., 1999), and ascorbic acid may be effective in reducing the rate of oxidative degradation during storage by quenching of H2O2. Ascorbic acid fortification may reduce free radical production in polyphenol-rich beverages by lowering pH, while no protective effect was observed when pH of the tea beverage was neutral (Aoshima and Ayabe, 2007). Ascorbic acid present in guava juice is known to stabilize lycopene (Mortensen et al., 2001) by a radical scavenging mechanism, although this protecting effect was not observed during heating probably due to degradation of ascorbic acid.
In general, it has been found that a higher concentration of bioactive compounds is required to achieve similar efficacy in foods as demonstrated in in vitro experiments (Shelef, 1983; Tassou et al., 1995; Smid and Gorris, 1999; Burt, 2004; Holley and Patel, 2005; Negi, 2012), and experiments have proved that the concentration of essential oils to achieve the desired antibacterial effect should be approximately two-fold in semi-skimmed milk (Karatzas et al., 2001), ten-fold in pork liver sausage (Pandit and Shelef, 1994), 50-fold in soup (Ultee and Smid, 2001), and up to100-fold in soft cheese (Mendoza-Yepes et al., 1997). Most studies on food application of phytochamicals are limited to examining their bioactive efficacy rather than their stability during storage (Burt, 2004; Fisher and Phillips, 2008; Negi, 2012). Stability of phytochemicals have been discussed in detail elsewhere in this book (Chapters 14 and 15).
The use of edible coatings to extend the shelf life and improve the quality of fruits and vegetables has been studied extensively due to their ecofriendly and biodegradable nature. Edible coatings can provide a supplementary and sometimes essential means of controlling physiological, morphological, and physicochemical changes in fruit and preserves the phytochemicals present in them.
The functionality of edible coatings can be improved by incorporating natural or synthetic antimicrobial agents, antioxidants, and functional ingredients such as minerals and vitamins. The addition of preservatives is of special interest for minimally processed fruit and vegetables, which have an extremely short shelf life because of microbiological concerns as well as sensory and nutritional losses that occur during their distribution and storage. Antioxidants are added to edible coatings to protect fruit against oxidative rancidity and discoloration (Baldwin et al., 1995). The antioxidants were also added in edible coatings to control oxygen permeability and reduce vitamin C losses in apricots during storage (Ayranci and Tunc, 2004). Anti-browning agents (McHugh and Senesi, 2000; Baldwin et al., 1996; Lee et al., 2003; Perez-Gago et al., 2006) and texture enhancers like CaCl2 (Wong et al., 1994) and milk proteins (Le Tien et al., 2001) have also been used in edible coatings for preservation purpose. Eswaranandam et al. (2006) extended the shelf life of fresh-cut cantaloupe melon by incorporating malic and lactic acid into soy protein coatings. Antimicrobial and antioxidant coatings have advantages over direct incorporation of the antimicrobial or antioxidant agents because they can be designed for slow release of the active compounds from the surface of the coated commodity. By slowing their diffusion into coated foods, the preservative activity at the surface of the food is maintained for a longer storage period, and a smaller amount of antimicrobials/antioxidants would come into contact with the food compared to dipping, dusting, or spraying the preservatives onto the surface of the food to achieve a target shelf life (Min and Krochta, 2005).
Use of natural antimicrobials in the development of coatings, which use inherently antimicrobial polymers as a support matrix has been studied in detail using chitosan, which is mainly obtained from the deacetylation of crustacean chitin and is one of the most effective antimicrobial film forming biopolymers (it is out of purview of this topic but readers can refer to Vargas et al., 2006; El Gaouth et al., 1991; Zhang and Quantick, 1997, 1998; Romanazzi et al., 2003; Devlieghere et al., 2004; Park et al., 2005). Chitosan-based edible coatings can be also used to carry other antimicrobials compounds such as organic acids (Outtara et al., 2000), essential oils (Zivanovich et al., 2005), spice extracts (Pranoto et al., 2005), lysozyme (Park et al., 2004), and nisin (Pranoto et al., 2005; Cha et al., 2003). Natural antimicrobial compounds have been incorporated into protein or polysaccharide-based matrices, thereby obtaining a great variety of multi-component antimicrobial coatings by adding oregano, rosemary, and garlic essential oils (Seydim and Sarykus, 2006). Rojas-Grau et al. (2006) used apple puree and high methoxyl pectin combined with oregano, lemon grass, or cinnamon oil at different concentrations as coatings for enhancing phytochemical stability. Greater details about effect of phytochemicals on minimally processed fruit and vegetables can be found in Chapter 10 of this book.
Although several coatings have shown their efficacy in in vitro tests against a range of microorganisms, they were not tested in food systems and therefore information about their possible impact on the aroma and flavor of the coated products is not available. The influence of the incorporation of antimicrobial phytochemicals into edible films and coatings on sensory properties of coated commodities needs much deeper investigation.
The use of elevated CO2 as the packaging gas reduced the overall antioxidative capacity of cranberries during the initial storage period. The antioxidant status of air packaged fruit decreased initially but increased on further storage. Berries stored under elevated O2 exhibited good antioxidative capacity over the first four days of storage but this declined with prolonged storage, possibly due to O2 promoted oxidation of the constitutive anthocyanins and phenolics. However, during the first four days of storage the effect of elevated O2 on antioxidative status was minimal. High levels of oxygen in controlled atmosphere storage had little effect on post-harvest anthocyanins development and total phenolics in cranberries (Gunes et al., 2002).
Modified atmospheres with controlled concentrations of CO2 and O2 have been used to maintain the quality of fresh-cut spinach. The total flavonoid content remained constant during storage in both air and MAP atmospheres, while vitamin C was better preserved in MAP stored spinach. Ascorbic acid was transformed to dehydroascorbic acid during storage, and its concentration was higher in MAP-stored tissues. A decrease in the total antioxidant activity was observed during storage in MAP-stored spinach, which may be due to higher content of dehydroascorbic acid and lower content of both ascorbic acid and antioxidant flavonoids in the MAP-stored samples (McGill et al., 1966; Izumi et al., 1997). Neither controlled atmospheric nor cold storage had any adverse effect on antioxidant activity in apples. After 25 weeks of cold storage there was no decrease in chlorogenic acid, but catechin content decreased slightly. Storage at 0 °C for nine months had little effect on phenolic content of apple peel (Goulding et al., 2001). Lattanzio et al. (2001) also found that after 60 days of cold storage the concentration of total phenolics in the skin of Golden Delicious apples increased. Quercetin glycosides, phloridzin, and anthocyanin content of various apple cultivars were not affected by 52 weeks of storage in controlled atmospheric conditions, although chlorogenic acid and total catechins decreased slightly in Jonagold apples; total catechin concentration decreased slightly in Golden Delicious; and chlorogenic acid concentrations remained stable during storage period (van der Sluis et al., 2001) indicating stability of phytochemicals under modified atmosphere is a function of crop, variety, and phytochemical in question.
Fiber content in asparagus increases significantly during storage of up to 13 days irrespective of storage temperatures (10 and 15 °C), but a slow increase in fiber content was observed during MAP-stored asparagus (at 4 °C) up to 30 days of storage (Sothornvit and Kiatchanapaibul, 2009). Asparagus stored in different packaging conditions at 10 °C for four days showed an increase in lignin content also (Huyskens-Keil and Kadau, 2003).
Total phenolic content decreased during storage of fresh-cut jackfruit bulbs during 35 days of storage at 6 °C. Bulbs dipped in a solution containing CaCl2, ascorbic acid, citric acid, and sodium benzoate coupled with MAP resulted in significantly lower loss in phenolics (Saxena et al., 2009). Mateos et al. (1993) also found inhibition of enzyme mediated phenolic metabolism in fresh-cut lettuce stored under low O2 and high CO2 atmosphere. Alasalvar et al. (2001) reported that storage under low O2 conditions reduces the accumulation of total phenols in shredded oranges and purple carrots as compared to air or high O2 storage.
Bioactive packaging is a process in which a food package or coating plays the unique role of enhancing impact of food over the consumer’s health. The bioactive packaging material should be capable of withholding desired bioactive principle in optimum conditions until their eventual release into the food product either during storage or just before consumption. Bioactive packaging can be achieved by integration and controlled release of bioactive components or nanocomponents from a biodegradable packaging system, micro or nano encapsulation of active substances in the packaging, and packaging with active enzymes exerting a health-promoting benefit through transformation of specific food components (Lagaron, 2005). Method of fabrication of the films, the optimal time temperature conditions for mixing the biomaterial with phytochemical, and the suitable mechanism to attain the desired release rate just upon packaged food opening and before consumption are important for phytochemical based bioactive packaging systems. An antimicrobial agent releasing plastic film for cheese packaging has been developed (Han, 2002) that has the potential to incorporate other phytochemicals using a similar system. The edible films can be modified using polysaccharide (starch, alginates, etc.), protein (gelatin, soy protein, wheat gluten, etc.), and lipids (waxes, triglycerides, fatty acids, etc.) to contain phytochemicals for food use.
Microencapsulation is defined as “the technology of packaging solid, liquid and gaseous materials in small capsules that release their contents at controlled rates at specific conditions over prolonged periods of time” (Champagne and Fustier, 2007). Release can be solvent activated or signaled by changes in pH, temperature, irradiation, or osmotic shock. As the encapsulated materials are protected from moisture, heat, or other extreme conditions, their stability is enhanced and they maintain viability for longer durations. Lopez-Rubio (2006) observed that microencapsulation is suitable for incorporating functional ingredients that are very susceptible to lipid oxidation or to mask off odors or tastes expected in foods after addition of phytochemicals. Microencapsulation promotes the delivery of active ingredients without their interaction with food components. As it is used to provide barriers between the sensitive bioactive materials and the environment (food or oxygen), it can also be used to mask unpleasant flavors and odors, or to modify texture or preservation properties (Fang and Bhandari, 2010). Omega-3 and omega-6 fatty acids are used for food fortification, but the taste and smell of these oils and their tendency to oxidize rapidly is a problem in their food application (Augustin and Sanguansri, 2003). It was demonstrated that the consumption of food enriched with microencapsulated fish oil obtained by emulsion spray-drying was as effective as the daily intake of fish oil gelatine capsules in meeting the dietary requirements of omega-3 fatty acid (Wallace et al., 2000).
Several technologies have been used for microencapsulation of bioactive ingredients, which basically include three steps; formation of a wall around the material, prevention of undesirable leakage, and leaving undesirables out of encapsulated material (Gibbs et al., 1999; Mozafari et al., 2008). The current encapsulation techniques include spray-drying, spray-chilling, fluidized-bed coating, extrusion, liposome entrapment, coacervation, and nanoemulsions (Arneado, 1996; Gibbs et al., 1999; Tan and Nakajima, 2005; Garti et al., 2005; Weiss et al., 2006; Flanagan and Singh, 2006; Augustin and Hemar, 2009).
Spray-drying has been traditionally used for the encapsulation of oil-based vitamins and fatty acids. For many emulsions, spray-chilling and liposome techniques have shown potential for the controlled release of bioactive compounds. Spray-chilling and fluidized-bed coatings are the most popular methods for encapsulating water-soluble vitamins, whereas spray-drying of emulsions is generally recommended for the encapsulation of lipid-soluble vitamins (Kirby et al., 1991; Arnaud, 1995; Reineccius, 1995; Augustin et al., 2001; Guimberteau et al., 2001; McClements, 2005; Goula and Adamopoulos, 2012; Wang et al., 2012). In spray-chilling and spray-cooling, the core and wall mixtures are atomized into the cooled or chilled air, which causes the wall to solidify around the core. The coating materials used are vegetable oils or their derivatives, fats and stearin, and mono- and di-acylglycerols (Cho et al., 2000; Taylor, 1993). Atomization causes quick and intimate mixing of droplets with the cooling medium and evaporation does not occur due to low temperatures, therefore it yields droplets of almost perfect spheres to give free-flowing powders. Microcapsules are insoluble in water as oils are used as a coating material, therefore this technique can be utilized for encapsulating water-soluble core materials such as minerals, water-soluble vitamins, enzymes, acidulants, and flavors (Lamb, 1987). Fluidized-bed coating involves fluidization of the solid particles in a temperature- and humidity-controlled chamber of high velocity air where the coating material is atomized (Balassa and Fanger, 1971; Zhao et al., 2004). Wall materials used in this technique include cellulose derivatives, dextrins, emulsifiers, lipids, protein derivatives, and starch derivatives, which may be used in a molten state or dissolved in an evaporable solvent either by top-spray, bottom-spray, or tangential spray (Jackson and Lee, 1991). Top spray method was used to obtain microencapsulated ascorbic acid after fluidization with hydrophobic coating materials (Knezevic et al., 1998). Microfluidization involves high pressure homogenization to produce fine emulsion, which can be further evaporated to obtain nano particles (O’Donnell and McGinity, 1997; Couvreur et al., 1997), and Salvia-Trujilo et al. (2013) showed that the microfluidization has the potential for obtaining nano-emulsions of essential oils.
The Liposome entrapment technique utilizes liposomes, which consist of an aqueous phase that is completely surrounded by a phospholipid-based membrane and both the aqueous and lipid-soluble materials can be enclosed in the liposome. Permeability, stability, surface activity, and affinity of liposomes can be varied through size and lipid composition variations (Gregoriadis, 1984; Kirby and Gregoriadis, 1984). Encapsulation of ascorbic acid in a liposome together with vitamin E produces a synergistic antioxidant effect (Reineccius, 1995).
Encapsulation by extrusion involves forcing a core material in a molten carbohydrate mass through a series of dyes into a bath of dehydrating liquid. In this encapsulation method, the pressure is kept around 100 psi and temperature rarely goes beyond 115 °C (Reineccius, 1989). The coating material hardens on contacting the liquids, forming an encapsulating matrix to entrap the core material. The extruded filaments are then separated from the liquid bath, dried, and sized (Shahidi and Han, 1993). The carrier used may be composed of more than one ingredient, such as sucrose, maltodextrin, glucose syrup, glycerine, and glucose (Arshady, 1993). Several polyphenolic antioxidants from medicinal plants were encapsulated using extrusion procedure by Belscak-Cvitanovic et al. (2011).
Centrifugal suspension separation involves mixing the core and wall materials and then adding to a rotating disk. The core material then leaves the disk with a coating of residual liquid, which is then dried or chilled (Sparks, 1989), whereas centrifugal extrusion is a liquid co-extrusion process that uses nozzles consisting of concentric orifice located on the outer circumference of a rotating cylinder through which coating and core materials are pumped separately on the outer surface of the device. While the core material passes through the center tube, coating material flows through the outer tube. As the cylinder rotates, the core and coating materials are co-extruded and the coating material envelops the core material. The wall materials used include gelatin, sodium alginate, carrageenan, starches, cellulose derivatives, gum acacia, fatty acids, waxes, and polyethylene glycol (Schlameus, 1995). Using alginate or alginate- hydroxy propyl methyl cellulose combinations, stability of avocado oil encapsulated by co-extrusion process was maintained for 90 days at 37 degree C (Sun-Waterhouse et al., 2011).
Many emulsion and coating technologies offer significant opportunities for the co-encapsulation of various hydrophobic and hydrophilic bioactives (Champagne and Fustier, 2007). Co-crystallization utilizes sucrose syrup as a wall material, which is concentrated to the supersaturated state and maintained at a temperature high enough to prevent crystallization. A predetermined amount of core material is then added to the concentrated syrup with vigorous mechanical agitation until the agglomerates are discharged from the vessel. The encapsulated products are then dried to the desired moisture and screened to a uniform size (Rizzuto et al., 1984). Yerba mate (Ilex paraguariensis) extract was encapsulated by co-crystallization in a super saturated sucrose solution (Lorena et al., 2007). Coacervation involves the separation of a liquid phase of coating material from a polymeric solution followed by the coating of that liquid phase around suspended core particles followed by solidification of the coating. The coacervation process consists of three steps, which involves formation of a three-immiscible chemical phase consisting of a liquid vehicle phase, a core material phase, and a coating material phase; deposition of the coating by controlled physical mixing; and solidification of the coating by thermal, cross-linking, or desolventization techniques to form a self-sustaining microcapsule. The coating materials used for coacervation microencapsulation include gelatin-gum acacia, gliadin, heparin-gelatin, carrageenan, chitosan, soy protein, polyvinyl alcohol, gelatin-carboxymethylcellulose, β-lactoglobulin-gum acacia, and guar gum-dextran (Gouin, 2004). Using coacervation of gelatin A with sodium carboxy methyl cellulose, neem seed oil was encapsulated (Devi and Maji, 2011), whereas gelatin and gum arabic were used for obtaining microcapsules of peppermint oil by complex coacervation process (Dong et al., 2011). Molecular inclusion (Inclusion Complexation) achieves encapsulation at a molecular level, which typically employs β-cyclodextrin as an encapsulating medium. The external part of the cyclodextrin molecule is hydrophilic, whereas the internal part is hydrophobic, therefore the apolar flavor compounds can be entrapped into the apolar internal cavity through a hydrophobic interaction and are entrapped inside the hollow center of a β-cyclodextrin molecule (Pagington, 1986). Nunes and Mercadante (2007) encapsulated lycopene using beta-cyclodextrin as an encapsulating medium by molecular inclusion process, but reported a slight decrease in its purity after encapsulation.
The solubility of functional ingredients in food formulations is a major consideration as the bioavailability of water insoluble or low-water-soluble ingredients gets reduced in many foods. Nano sized particles have shown substantial increase in solubility in water, which improves bioavailability (Grau et al., 2000; Muller et al., 1999; Trotta et al., 2001). Tan and Nakajima (2005) evaluated stability of β-carotene nanodispersions prepared by emulsification evaporation technique. They observed significant effect of homogenization pressure and homogenization cycle on the size of particles, which in-turn was responsible for β-carotene stability during storage. High homogenization pressure ensured a good emulsification and led to the formation of smaller sized particles, but had an adverse influence on the stability of β-carotene. The smaller particles showed higher losses during 12 weeks of storage, probably on account of increase in surface area in comparison to higher diameter particles.
Phytochemicals are effective in promoting health and reducing the disease risk to human beings. Phytochemicals are often lost during many of the commonly practiced processes and subsequent storage and food preparation. Modified atmospheres with controlled concentrations of CO2 and O2 have been used to maintain the quality of several fruits and vegetables to extend shelf life and preserve the phytochemicals present in them. Edible coating technology is a promising method for preserving the quality of fresh and minimally processed fruit, and research efforts have resulted in an improvement of the functional characteristics of the coatings. Microencapsulation and nanoencapsulation are promising techniques that can be potentially used to incorporate phytochemicals into edible coatings. Investigations related to the additional benefits of microencapsulation on the stability of bioactive ingredients in the gastric environment and on the release of bioactive ingredients into the GI tract needs attention.
Processing is a critical aspect of phytochemical production, especially due to the low yield of extracts. Processing methods are usually based on traditional methods such as water or solvent extraction. New innovative methods such as microwave and ultrasound assisted techniques or supercritical fluid extraction to obtain phytochemicals need to be explored for production of a higher yield of phytochemicals at lower operating costs and faster production times. There are a number of stability issues that must be overcome before phytochemicals can be successfully used as functional food ingredients since extracted phytochemicals can be less stable than naturally occurring phytochemicals in tissues.
Packaging can affect quality of phytochemicals and phytochemical-containing foods by influencing browning, flavor, and nutrient losses during storage, but studies reporting the phytochemical stability as affected by various packaging materials are lacking. Bioactive packaging, a novel technology for enhanced delivery of phytochemicals, is being investigated, but the issues related to feasibility, stability, and bioactivity of phytochemicals for food industry are yet to be studied.
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