When fruit tissue is cut or injured, physiological changes occur. These changes are exemplified by the production of ethylene, by a change in color due to oxidative reactions, by moisture migration, and by a diversity of enhanced enzymatic activity, all of which lead to deterioration in the quality of the fruit tissue and a decline in sensory attributes, such as the texture and flavor typically associated with fresh fruit. The organoleptic characteristics of fruit are greatly modified by the accumulation of brown pigments that replace the natural color of the fruit tissue. Although some browning may be nonenzymatic, most of the browning in fruit tissue is due to enzymatic oxidation of phenolic compounds when they mix with polyphenol oxidases upon cutting or wounding of the tissue. Such changes are considered to be signs of deterioration that lower the fruit quality both visually and with regard to other sensory attributes, such as flavor and texture. Deterioration can subsequently lead to increased microbial growth.
The retention of fresh flavor and texture are also very important to the eating quality of freshly-cut fruit, and it is the delicate and complex flavors of fruit that are extremely susceptible to change when fruit is freshly cut. Driven by consumer interest and demand for convenience, freshly cut vegetables have received the most emphasis by technologists. As new cut vegetable products with acceptable shelf life have been successfully introduced into the market, there has been an increasing interest in the development of freshly cut fruit as the next generation of ready-to-eat cut produce.
Discoloration of freshly cut fruit is the most noticeable loss of quality and is of considerable economic importance. As the food becomes unsightly, it signifies that the food is no longer edible. The appearance of cut fruit is generally considered significant from a safety point of view. Cut fruit is considered safe for consumption only as long as it looks good. In fact, it is important that it look bad before it actually becomes unsafe to eat due to the presence of undesirable microorganisms.
Discoloration of freshly cut fruit has received the most attention by the technologists because of the visibility and economic impact of discoloration. The browning and blackening of cut tissue is due to the mixing of enzyme and substrate. This discoloration response to cutting is often attributed to the oxidation of the substrates, i.e. o-hydroxyphenols, such as chlorogenic acid, which is catalyzed by polyphenol oxidase (PPO) enzyme.
Oxidative browning of cut fruits has been the subject of several reviews (Vamos-Vigyazo, L. Crit. Rev. Food Sci. Nutr. 1981, Vol. 15, pp. 49-127 1981; Whitaker, 1985; Mayer, A. M. Phytochemistry 1987 Vol. 26, pp. 11-20; Marques, L. In Enzymatic Browning and its Prevention, 1995, pp. 90-102). The majority of polyphenoloxidases that are involved in the browning of fruits are catecholoxidases (Mayer, A. M.; Harel, E. In Food Enzymology; Fox, P. F., Eds; Elsevier Applied Sciences London, 1991, pp. 373-398), also designated o-diphenoloxidases. Overall studies indicate high heterogeneity in expression of PPO in the fruit that may be due to differential genomic expression in species, the physiological age of the fruit, or the biochemical and physiological nature of the tissue studied. The activity and mode of action of PPO has been reviewed by Vamos-Vigyazo, supra; Whitaker, supra; Mayer, 1987 supra. It is a copper-requiring enzyme that in the presence of oxygen catalyzes the oxidation of phenolic substrates, such as chlorogenic acid, to quinones, which through subsequent reactions are polymerized to brown, pink or black pigments.
There are numerous reports of polyphenol oxidase (PPO) isozymes causing discoloration in apples, potatoes, grapes, avocados, parsnips, peaches and pears. PPO activity has been detected in all fruit tissue including the peel, flesh, and cortex (Macheix, J. J.; Fleuriet, A.; Billot, J. J., Fruit Phenolics; CRC Press; Boca Raton, Fla., 1990, pp. 295-312). Activity is mainly concentrated in the peel and the cortex with chlorogenic acid oxidase localized mainly at the core of the apple fruit and near the skin (Munata, P.,1981, Am. Pot. J. Vol. 58, pp. 85). Munata supra reports that enzymatic browning in potatoes can be mitigated by inhibiting PPO activity by adjusting the pH, adding bisulfite or sulfhydryl compounds, chelating the copper from PPO, using reducing compounds that reduce the whole- quinones to the o-hydroxyphenol state, or applying chemicals which react with o-quinone to yield colorless additive products.
According to Mayer and Harel (Mayer, A. M.; Harel, E., 1979, Phytochemistry Vol. 18, pp. 193-215), inhibitors of PPO can be grouped into two classes: compounds that interact with the copper moiety and those that affect the active site for the phenolic substrate, such as chlorogenic acid. Grom (U.S. Pat. No. 3,895,119 issued July, 1975) teaches the application of calcium chloride in combination with sulfite or ascorbic acid to the cut surface of plant tissue, wherein the calcium helps to maintain texture and the sulfite or ascorbic acid inhibits discoloration. Steiner (U.S. Pat. No. 4,818,549 issued April, 1989 and U.S. Pat. No. 4,911,940 issued March, 1990) teaches the combination of citric acid, calcium chloride, and sodium chloride for use in maintaining the color of the cortex of the fruit tissue, and Warren (U.S. Pat. No. 5,055,313 issued August, 1991) presents a combination of ascorbic acid, calcium chloride, or other sources of halides, citric acid, and sodium acid pyrophosphate to maintain the integrity of cut plant tissue. However, the methods of Steiner supra and Warren supra provide shelf life of less than four days.
Pizzocaro, et al. (J. Food Proc. and Pres. 1993, Vol. 17, pp. 21) has reported inhibition of PPO activity in apples using a mixture of ascorbic acid, citric acid and sodium chloride. Sapers (Sapers, G. M., Ziolkowski, M. A.,; J. Food Sci. 1987, Vol. 52, pp. 1732-1733) reported the use of ascorbic acid and erythorbic acid to inhibit PPO activity in apple tissue and in apple cider. Sapers et al. (Sapers, G. M. et al. J. Food Sci. 1989, Vol. 54, pp. 997) evaluated ascorbic acid derivatives for their capacity to inhibit browning in apple juice and apple tissue. Ascorbic acid -2-phosphate and -triphosphate showed promise for cut apple pieces but were not effective in juice. Ascorbic acid -6-fatty acid esters showed anti-browning activity in juice.
Monsalve-Gonzalez et al. (Monsalve-Gonzalez, A. et al., J. Food Sci. 1993, Vol. 58, pp. 797) demonstrated efficacy of 4-hexylresorcinol in retarding browning in apple pieces and further demonstrated that in combination with vacuum packaging browning inhibition could be achieved for several weeks. McEvily (U.S. Pat. No. 5,059,438 issued October 1991) discloses the use of resorcinol derivatives as inhibitors of enzymatic browning in foods and beverages. Cysteine, as a reducing agent, has been demonstrated to mitigate browning of tissue by reacting with o-quinones to form colorless addition products or by reducing the o-quinones to their phenol precursors (Richard-Forget, F. C., et al., J. Agric. Food Chem. 1992, Vol. 40, pp. 2108) (Richard, 1991). The authors report that cysteine has no direct effect on apple PPO activity. Janovitz-Klapp (Janovitz-Klapp, A. H. et al., J. Agric. Food Chem. 1990, Vol. 38, pp. 926) carried out studies on the inhibition of PPO. When they added ascorbic acid, cysteine, or bisulfite color formation was retarded. Molnar-Perl and Freidman (Molnar-Perl, I.; Friedman, M.; 1990 J. Agric. Food Chem. Vol. 38, pp. 1652) revealed that SH-containing N-acetyl-L-cysteine and reduced glutathione were almost as effective as sodium sulfite in preventing enzymatic and nonenzymatic browning of apple juice.
Calcium chloride has been reported to maintain the texture of fruit tissue, but calcium has not been implicated in the inhibition of browning.
In 1994, apple production in the United States was reported to be 10,634.8 million pounds, with per capita consumption at about 17 lbs. With continued planting of apple trees and continued increase in yields and total fruit harvested, the search is on for the development of diverse apple markets to maintain the demand for apples and to sustain price. A considerable volume of apples are sliced, treated with an anti-browning inhibitor and refrigerated or frozen to provide apple pieces for the baking industry. More recently there has evolved considerable interest in supplying fresh-like quality apple pieces to retail and food service markets where consumer expectations are for a fresh-tasting apple slice.
The problems with the apple pieces that are currently sold to the baking industry is that they still discolor to some degree even though they are treated with sulfites and preservatives (and are thus no longer "fresh" by FDA definition), and they are subjected to handling and freezing operations that result in the loss of texture associated with freshly-cut quality. Sulfites also tend to cause translucency of the apple tissue. Furthermore, most mixtures of anti-browning treatments now in use alter the flavor of the apple pieces, or fail to maintain a flavor profile similar to a freshly cut apple slice. To date no treatment other than sulfite-based treatments has been able to maintain the color of freshly cut apple pieces. Thus, to date, technology does not exist that will maintain the quality of freshly cut apple pieces over an acceptable shelf life without the use of prior art sulfites and preservatives.
While no single agent has been found to replace sulfites, numerous combinations of agents have been tried, but all have failed to date to provide shelf life stability to freshly cut apple pieces and to maintain the sensory characteristics associated with freshly-cut apple pieces.