THE FORMATION OF THEARUBIGIN-LIKE SUBSTANCES BY IN-VITRO POLYPHENOL OXIDASE-MEDIATED FERMENTATION OF INDIVIDUAL FLAVAN-3-OLS

An enzymic in-vitro model tea fermentation system has been used in combination with gradient elution reversed-phase high-performance liquid chromatography to study the formation from individual flavan-3-ols of resolvable and unresolvable thearubigin-like substances. It has been found that in 30 min at pH 5.6 tea polyphenol oxidase (EC 1.14.18.1) produces a distinctive range of resolvable thearubigin-like products from each flavan-3-ol substrate examined ((+)-catechin, (-)-epicatechin, (-)-epigallocatechin, (-)-epicatechin gallate and (-)-epigallocatechin gallate). However, (-)-epicatechin, and to a much lesser extent (+)-catechin, were the only flavan-3-ols converted to an unresolvable thearubigin-like hump under these conditions. In addition, it was demonstrated that varying the pH of incubation in the range 5.0-6.0 had little effect on the profile of products obtained from the simple catechins, but a pronounced effect in the case of the gallocatechins. It is concluded that the combination of (-)-epicatechin and polyphenol oxidase is the simplest model of tea fermentation that can be used to generate in quantity thearubigin-like materials for structure elucidation.