Peroxidase activity falls steadily during fruit development (Gortner and Singleton, 1965), reaching a minimum of one-third the initial value during ripening. Acid peroxidase (pH optimum 5.0) does not appear to be related to chilling-injury symptom development (Teisson, 1977), though it does decline during storage (Teisson and Martin-Prevel, 1979). Some purification of the two chilling-related enzymes, peroxidase and polyphenol oxidase (PPO), has been accomplished (Teisson, 1977).
No ascorbic acid oxidase was detected in one report (van Lelyveld and de Bruyn, 1977); however, ammonium sulphate-precipitated protein has been shown to have ascorbic acid oxidase that has a high pH optimum (> 8) and no activity below pH 6.0 (Teisson, 1977). These different assay conditions may explain the published discrepancies. The PPO has optimum pH near 5.0 and a temperature optimum near 45°C (Teisson, 1977). Das et al. (1997) found three PPO isoforms, with the major isoform being a tetramer of identical subunits (c. 25 kDa) and having an optimum activity between pH 6 and 7. The difference in pH may be due to the different extraction methods used or different isoforms. The PPO is stable to heat when extracted, but loses over 50% of its activity following 20 min exposure to 60°C in vivo (Teisson, 1977), and thiols and bisulphite inhibit activity. Catalase and indole acetic acid oxidase have also been detected, but changes in activity during ripening or storage have not been reported. Indole acid acid oxidase has a pH optimum of 3.5, which is at a variance with that in other fruit (Teisson, 1977).
Acetone powders of pineapple stem extracts contain the proteinase bromelain and a family of polypeptide inhibitors of this enzyme (Reddy et al., 1975). Proteinase activity appears abruptly after flowering, remains high during fruit development (Gortner and Singleton, 1965) and declines during ripening (Gortner and Singleton, 1965; Lodh et al., 1972). The major cysteine proteinase is fruit bromelain (EC 22.214.171.124), a non-glycosylated proteinase that is immunologically distinct from the glycosylated stem bromelain (EC 126.96.36.199) (Rowan et al., 1990). Fruit brome-lain has an estimated molecular weight of between 23 and 31 kDa by different laboratories (Yamada et al., 1976; Ota et al., 1985;
Rowan et al., 1990). The fruit bromelain is an acidic protein and its isoelectric point (pI) is 4.6, different from the basic stem bromelain, the pI of which is 9.6 (Yamada et al., 1976). The amino-terminal end of the fruit brome-lain, has an additional alanine (Yamada et al., 1976). The sequence around the reactive cysteine is the same. Fruit bromelain has a 20% cross-reactivity with anti-stem bromelain (Sasaki et al., 1973).
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