The Effect of Oxidative Enzymes in Foods - Food Shelf Life Stability: Chemical, Biochemical, and Microbiological Changes - page 304

chemistry. Nevertheless, it is generally accepted that much of the browning in fresh
fruits is caused by the enzymic action of o -diphenol oxidoreductases now classified
as O 2 : o -diphenol oxidoreductase (EC.1.10.3.1). Apples are one of the most common
fruits worldwide and hence there is considerable interest in apple polyphenol oxi-
dases as a cause of excessive enzymic browning in juices, purees, and dried powder.
However, for the manufacture of cocoa, coffee, and tea, enzymic browning is not
only beneficial but is essential. In green coffee beans the main precursors of browning
identified by HPLC and UV-absorbtion spectra are the chlorogenic acids, 5- O -
caffeoylquinic acid (5-CQA) and 3-CQA and 4-CQA. 117 In tea catechin, epicatechin
and their gallate esters are oxidized to quinones that then undergo condensation and
polymerization to form a range of colored compounds including theaflavins and
thearubigens. Undesirable enzymic browning caused by PPO on the surface of foods
is of great concern, as the products are less acceptable to consumers. It is also thought
that enzymic browning contributes significantly to undesirable color formation in
sugar cane juice and that PPO is the main enzyme involved. 118 The enzymes are
often referred to as PPO, catechol oxidases, phenolase, tyrosinase, and cresolase. A
survey of recent reports on the enzyme in fruits and vegetables and their products
adds several new species to an already long list of those previously reported. 119,120
The recent literature abounds with more studies on catechol oxidases in deciduous
fruits, grapes and wine, potatoes, avocados, olives, bananas, mangoes, tea, coffee,
cocoa, and mushrooms. The enzymes are abundant in tubers, storage roots, and
fruits. High levels of enzyme are usually found in tissues that are also rich in phenolic
compounds. The levels of PPO and its substrates often change markedly during the
life cycle of the plant particularly in fruits and vegetables. Multiple forms of polyphe-
nol oxidase are invariably found in extracts of fresh fruits and vegetables, juices,
and concentrates. Although post-translational modifications might be responsible for
the formation of multiple forms, the observations described suggest that at least part
of the multiplicity encountered may be artifactual. Kiwi fruit 121 and DeChaunac
hybrid grapes 122 have been reported to possess from 3 to 14 isoenzymes of various
sizes. The multiple forms are easily observed on PAGE and IEF electrophoretograms.
Although multiplicity in a hybrid could result from genetic differences between the
parent species, it is probably not the only reason for the multiple forms observed.
The isolation of PPOs is fraught with problems especially as the quinones react with
the enzyme and thus cause inactivation to varying degrees. The membranous location
of the enzyme in many cases necessitates the use of harsh extraction procedures.
Tanning often seems to affect the multiple forms of the enzyme as well as the
apparent molecular weight, specificity for substrates (especially the hydroxylation
reaction), enzyme kinetics, and sensitivity to inhibitors. PPOs are often “tanned”
proteins to which phenols and their oxidation products are attached. Antibodies
prepared against such preparations are likely to give erroneous results if related to
the phenolic epitope. The removal of phenolics and prevention of browning has been
shown to markedly reduce the number of “multiple forms” of PPO. 123
Although catechol oxidase is apparently not ubiquitous in plants, 119 there are
few fruits and vegetables in which it has not been found. Histochemical methods
using 3,4-dihydroxyphenylalanine (DOPA) may be used to locate the cellular dis-
tribution of PPO and distinguish them from laccases detected by their oxidation of