Active packaging and colour control: the case of fruit and vegetables 421 50 ppm C2H4-65% initial red colour 50 ppm C2H4-85% initial red colour After 2 days at 180 Air-65% initial red colou Air-85%o initial red colour After 3 days at 7C Fig. 20.1 Changes in L' and a'* parameters of bell peppers with 65% and 85% of initial red colour degreened with 50ppm c2H4 during 2 days at 18%C followed by 3 days at 7C in air( Gomez et al., 2002) conversion of the chlorophyll to pheophytin can occur before a change of colour from bright green to olive brown could be observed (Lau et al., 2000) The rate of chlorophyll degradation can be lowered by several means However, a combination of them is more effective than one single method. The most important tool is chilling although there is a limit to temperature because some fruit and vegetables are susceptible to damage caused by low temperatures below their freezing point, suffering chilling injuries commonly accompanied by undesirable colour changes Ethylene greatly accelerates chlorophyll degradation. Usually, leafy vegetables do not produce much ethylene, but can be affected from ethylene coming from other sources. The inclusion of ethylene scavengers within packages containing these vegetables could provide protection against ethylene action(O Hare and Wong, 2000). Operations involved in fresh processing, like cutting, grating or peeling, stimulate ethylene biosynthesis that could cause physiological disorders, lowering the quality of the products. Some changes affecting colour include accumulation of phenolic compounds in carrots, red discoloration in chicory and endive, or russet spotting in lettuce(Artes, 2000b Van de velde and Hendrickx, 2001; Verlinden et al., 2001). It has been reported that ethylene produced during cutting of fresh processed spinach notably accelerates the loss of chlorophyll and damage is proportional to the ethylene level reached(Abe and Watada, 1991). Also celery sticks stored in atmospheres where ethylene is present showed a decrease in hue(Fig. 20.2)as colour anged from dark green to yellowish-green(Artes et al., 2002b). On the other
conversion of the chlorophyll to pheophytin can occur before a change of colour from bright green to olive brown could be observed (Lau et al., 2000). The rate of chlorophyll degradation can be lowered by several means. However, a combination of them is more effective than one single method. The most important tool is chilling although there is a limit to temperature because some fruit and vegetables are susceptible to damage caused by low temperatures below their freezing point, suffering chilling injuries commonly accompanied by undesirable colour changes. Ethylene greatly accelerates chlorophyll degradation. Usually, leafy vegetables do not produce much ethylene, but can be affected from ethylene coming from other sources. The inclusion of ethylene scavengers within packages containing these vegetables could provide protection against ethylene action (O’´Hare and Wong, 2000). Operations involved in fresh processing, like cutting, grating or peeling, stimulate ethylene biosynthesis that could cause physiological disorders, lowering the quality of the products. Some changes affecting colour include accumulation of phenolic compounds in carrots, red discoloration in chicory and endive, or russet spotting in lettuce (Arte´s, 2000b; Van de Velde and Hendrickx, 2001; Verlinden et al., 2001). It has been reported that ethylene produced during cutting of fresh processed spinach notably accelerates the loss of chlorophyll and damage is proportional to the ethylene level reached (Abe and Watada, 1991). Also celery sticks stored in atmospheres where ethylene is present showed a decrease in hue (Fig. 20.2) as colour changed from dark green to yellowish-green (Arte´s et al., 2002b). On the other Fig. 20.1 Changes in L* and a* parameters of bell peppers with 65% and 85% of initial red colour degreened with 50ppm c2H4 during 2 days at 18ºC followed by 3 days at 7ºC in air (Go´mez et al., 2002). Active packaging and colour control: the case of fruit and vegetables 421
422 Novel food packaging techniques 115 Air0°C AC5%C02+5%O2-0°C -AC5%CO2+5%02-5°C Initial 14 days Fig 20.2 Colou ges ( Hue)of celery sticks stored for 14 days in air and in ontrolled atmosphere (5%CO2+ 5%O2) free of ethy lene at 0 and 5C(Artes et al, hand, antioxidants are related to chlorophyll retention in leafy products. Two antioxidants commonly present in fruits and vegetables are ascorbic acid and B- carotene, which protect chlorophyll by inhibiting the reactions that degrade it, retarding yellowing(Schwartz and von Elbe, 1983) 20.4.1 Anthocyanin degradation Anthocyanins are very unstable pigments, particularly once removed from their natural environment and the protection provided by co-pigmentation, leading to unattractive yellowish and brownish pigments. This is particularly evident when minimal fresh processed products are prepared. When conditioning fruit and vegetables by techniques like peeling, cutting, slicing, etc, cell membranes are disrupted, allowing the mixing of phenolic substrates located in the vacuole and pecific polyphenol oxidases enzymes(PPO; EC 1. 14. 18. 1)associated to cell membranes, (mainly in the plastids) Washing the product immediately after cutting removes sugars and other substrates at the cut surfaces minimising It is well known that colour due to anthocyanins is particularly degraded by the nzymic hydrolysis in harvested products as recently reviewed (artes et al., 2002c) Anthocyanins are oxidised in the vacuole of the plant cells in the presence of molecular O2 and under appropriate conditions of pH, temperature and water activity, by the action of the enzyme tyrosinase(EC 1.10.3. 1)or polyphenol oxidase
hand, antioxidants are related to chlorophyll retention in leafy products. Two antioxidants commonly present in fruits and vegetables are ascorbic acid and - carotene, which protect chlorophyll by inhibiting the reactions that degrade it, retarding yellowing (Schwartz and von Elbe, 1983). 20.4.1 Anthocyanin degradation Anthocyanins are very unstable pigments, particularly once removed from their natural environment and the protection provided by co-pigmentation, leading to unattractive yellowish and brownish pigments. This is particularly evident when minimal fresh processed products are prepared. When conditioning fruit and vegetables by techniques like peeling, cutting, slicing, etc., cell membranes are disrupted, allowing the mixing of phenolic substrates located in the vacuole and specific polyphenol oxidases enzymes (PPO; EC 1.14.18.1) associated to cell membranes, (mainly in the plastids). Washing the product immediately after cutting removes sugars and other substrates at the cut surfaces minimising reactions responsible for changes in colour and nutritional quality. It is well known that colour due to anthocyanins is particularly degraded by the enzymic hydrolysis in harvested products as recently reviewed (Arte´s et al., 2002c). Anthocyanins are oxidised in the vacuole of the plant cells in the presence of molecular O2 and under appropriate conditions of pH, temperature and water activity, by the action of the enzyme tyrosinase (EC 1.10.3.1) or polyphenol oxidase Fig. 20.2 Colour changes (ºHue) of celery sticks stored for 14 days in air and in controlled atmosphere (5% CO2 + 5% O2) free of ethylene at 0 and 5ºC (Arte´s et al,. 2002b). 422 Novel food packaging techniques
Active packaging and colour control: the case of fruit and vegetables 423 (PPO). But anthocyanins are not direct substrates for PPO, which catalyses the hydroxylation of monophenols to o-diphenols(cresolase activity EC 1. 14. 18. 1)and the oxidation of o-diphenols to o-quinones(catecholase activity EC 1.10.3.1) Catecholases were considered as the main PPO enzymes responsible for browning in fruit and vegetables. These o-quinones are very reactive molecules that rapidly condense by combining with amino or sulfhidril groups of proteins and with reducing sugars, producing different brown, black or red polymers of high molecular weight and unknown structure known as melanins (artes et al, 1998 ) In contrast to the ethylene effect on chlorophyll, anthocyanin synthesis and ethylene production seem to be correlated. In fact, red cherries stored in air reached high ethylene levels and the highest anthocyanin content by the end of cold storage(Remon et al., 2000 The increase in pH and decrease in titratable acidity induced by high CO during CA storage of fruit and vegetables have a strong effect in anthocyanin expression and stability. The red flavylium cation(AH+) remains stable only in acidic conditions. Changes in anthocyanin stability can result from nucleophilic attacks by water molecules on the anthocyanin molecule to form a colourless pseudobase, hemiacetal, or carbinol. The flavylium form can be restored by acidification. The colourless carbinol can form chalcone(a yellow pigment) by the opening of the ring structure. As pH increases above 4, a blue quinonoidal base is formed. Increase in pH above 7 can result in the loss of a proton from the hydroxyl group to form a second quinonoidal base(holcroft and Kader, 1999b) In addition, these authors reported that phenylalanine ammonia lyase(PAL, EC 4.3.1.5)and flavonoid glucosyltransferase(GT, EC 2. 4.1.28), two key enzymes in the synthetic pathway of anthocyanins in strawberry, were adversely affected by high CO2 levels during cold storage On the other hand, it has been demonstrated that the degrading effect of vitamin C on anthocyanin stability leads to undesirable colour changes in model olution and in natural pomegranate juice systems(Marti et al., 2001). Exposure to light and heat also induced these degrading reactions. However, glucosylation provides protection against photodegradation and the formation of molecular copigmentation complexes and ion-pairs lowered the degradation thocyanins(Brouillard et al, 1997) 20.4.2 Browning Browning is the result of a chain of reactions that very often occurs in fruit and vegetables. The first step of that process takes place in the vacuole and it is the deamination of the amino acid phenylalanine by PAL. The product of that reaction is the cinnamic acid which is hydroxy lated into various phenolic compounds. When O2 is present, the PPO located in the cytoplasm(plastids) oxidises the compounds to o-quinones, which polymerise into brown compounds (Siriphanich and Kader, 1985). The relationship between PPO and browning was reported when it was found that CO2 competitively inhibited PPO activity in mushrooms retaining their colour, although at high concentrations increasing browning(Murr and Morris, 1974)
(PPO). But anthocyanins are not direct substrates for PPO, which catalyses the hydroxylation of monophenols to o-diphenols (cresolase activity EC 1.14.18.1) and the oxidation of o-diphenols to o-quinones (catecholase activity EC 1.10.3.1). Catecholases were considered asthe main PPO enzymes responsible for browning in fruit and vegetables. These o-quinones are very reactive molecules that rapidly condense by combining with amino or sulfhidril groups of proteins and with reducing sugars, producing different brown, black or red polymers of high molecular weight and unknown structure known as melanines (Arte´s et al., 1998). In contrast to the ethylene effect on chlorophyll, anthocyanin synthesis and ethylene production seem to be correlated. In fact, red cherries stored in air reached high ethylene levels and the highest anthocyanin content by the end of cold storage (Remo´n et al., 2000). The increase in pH and decrease in titratable acidity induced by high CO2 during CA storage of fruit and vegetables have a strong effect in anthocyanin expression and stability. The red flavylium cation (AH+) remains stable only in acidic conditions. Changes in anthocyanin stability can result from nucleophilic attacks by water molecules on the anthocyanin molecule to form a colourless pseudobase, hemiacetal, or carbinol. The flavylium form can be restored by acidification. The colourless carbinol can form chalcone (a yellow pigment) by the opening of the ring structure. As pH increases above 4, a blue quinonoidal base is formed. Increase in pH above 7 can result in the loss of a proton from the hydroxyl group to form a second quinonoidal base (Holcroft and Kader, 1999b). In addition, these authors reported that phenylalanine ammonia lyase (PAL, EC 4.3.1.5) and flavonoid glucosyltransferase (GT, EC 2.4.1.28), two key enzymes in the synthetic pathway of anthocyanins in strawberry, were adversely affected by high CO2 levels during cold storage. On the other hand, it has been demonstrated that the degrading effect of vitamin C on anthocyanin stability leads to undesirable colour changes in model solution and in natural pomegranate juice systems (Martı´ et al., 2001). Exposure to light and heat also induced these degrading reactions. However, glucosylation provides protection against photodegradation and the formation of intermolecular copigmentation complexes and ion-pairs lowered the degradation of anthocyanins (Brouillard et al., 1997). 20.4.2 Browning Browning is the result of a chain of reactions that very often occurs in fruit and vegetables. The first step of that process takes place in the vacuole and it is the deamination of the amino acid phenylalanine by PAL. The product of that reaction is the cinnamic acid which is hydroxylated into various phenolic compounds. When O2 is present, the PPO located in the cytoplasm (plastids) oxidises the compounds to o-quinones, which polymerise into brown compounds (Siriphanich and Kader, 1985). The relationship between PPO and browning was reported when it was found that CO2 competitively inhibited PPO activity in mushrooms retaining their colour, although at high concentrations increasing browning (Murr and Morris, 1974). Active packaging and colour control: the case of fruit and vegetables 423