The Effect of Oxidative Enzymes in Foods - Food Shelf Life Stability: Chemical, Biochemical, and Microbiological Changes - page 305

syringaldiazine or phenylenediamine. Care must be taken to test for peroxidase
activity as this enzyme, arising from its ability to generate free radicals with aromatic
compounds, may also oxidize these reagents. PPOs are frequently found in thyla-
koids in higher plants, whereas in fungi the enzyme seems to be present in the
cytosol and not membrane-bound organelles. In higher plants, e.g., potato tubers,
sugar beet roots, and bananas, 119 the enzyme is mainly believed to be mem-
brane-bound in non-senescing tissues. However, lack of enzyme activity should not
be taken as proof of the absence of catechol oxidase which may be due to the
presence of endogenous inhibitors. 119,120,124 Zawistowski et al. 125 consider that
cytochemical investigations show that PPO is solely a plastid enzyme localized in
a diverse series of organs and tissues. It is generally accepted that the enzyme is not
active until released from the organelle when it comes in contact with the polyphenol
substrates located in the vacuole. Clearly, although it is universally accepted that
the enzyme has a deleterious effect on the quality of fresh fruits and vegetables, it
is nevertheless regarded by botanists as part of a natural plant defense system against
insects and pathogens. Defense mechanisms in higher plants, attack mechanisms in
fungi, pigment formation, or free radical scavenging are all possible candidates for
functions of PPOs. Activation of PPO occurs during cellular disruption and therefore
is caused by infection and mechanical injury. The enzymic products are primarily
quinones which then react with other constituents including amino acids and possibly
membrane bound peptides to form higher polymers to provide a barrier against
further infection as well as with the infective agents themselves.
Monophenolase and Diphenolase Activity
Enzymic browning is caused by the oxidation of o -diphenols by the O 2 : o -diphe-
noloxidoreductase activity resulting in the formation of o -quinones which may
polymerize and also react further with amino acids and peptides to form melanins.
Spread through the literature are claims for very high levels of enzyme, but these
must be treated with considerable caution as this may be due more to greater amounts
of polyphenols or in other cases lower activity might be due to the presence of
inhibitors, such as ascorbic acid and other reducing agents. A wide range of phenolic
compounds is found in many fruits, all of which are potential substrates for polyphe-
nol oxidases. Recently, Murata et al. 126 have reported that the Japanese apple cultivar
“Tsugaru” contained smaller amounts of polyphenols than the browning variety
“Matsu” and was due to the oxidation of catechins rather than chlorogenic acids.
However, the most common natural substrates are chlorogenic acid, catechin, epi-
catechin, and 3,4-dihydroxyphenylalanine (DOPA) and in bananas 3,4-dihydroxy-
phenylethylamine (dopamine). p -Coumaroyl and caffeoyl derivatives of tartaric acids
have been stated to be oxidized by grape diphenolases, while dates contain a range
of caffeoyl-shikimic acids. 127 The flavonoid pigments have been claimed to be poor
substrates, but can possibly undergo co-oxidation in the presence of chlorogenic
acid, 127 although a mechanism for this reaction, which may involve the release of
free radicals as with lipoxygenases catalyzed oxidation of carotenoids, has not been
elucidated. All o -diphenol oxidases require the o -diphenol group. Suitable test sub-
strates are catechol, 4-methyl catechol, and 1,4-dihydroxytoluene. As referred to