Role of Polyphenol Oxidase and Peroxidase in the Generation of Black Tea Theaflavins

Abstract

It has been reported earlier that when macerated tea leaf is fermented at lower pH, the resultant black tea contains increased levels of theaflavin, an important quality marker in black tea. In an attempt to investigate the biochemistry and chemistry underlying this observation, in vitro oxidation experiments using polyphenol oxidase (PPO) from fresh tea leaves, horseradish peroxidase (POD), and tea catechins, precursors for theaflavins, were carried out. In vitro oxidation experiments using crude tea PPO resulted in higher content of theaflavins at pH 4.5 in comparison with pH 5.5, the normal pH of the macerated tea leaf. When purified PPO was used in the in vitro system, surprisingly a reversal of this trend was observed, with more theaflavins being formed at the higher pH. A combination of pure tea PPO and POD led to an observation similar to that with the crude enzyme preparation, suggesting a possible role for POD in the formation or degradation of theaflavin. POD was observed to oxidize theaflavins in the presence of H(2)O(2), leading to the formation of thearubigin, another black tea pigment. This paper demonstrates that tea PPO, while oxidizing catechins, generates H(2)O(2). The amount of H(2)O(2) produced is greater at pH 5.5, the optimum pH for PPO activity, than at pH 4.5. Hence, an observed increase of theaflavins in black teas fermented at pH 4.5 appears to be due to lower turnover of formed theaflavins into thearubigins.

Oxidation of Tea Catechins: Chemical Structures and Reaction Mechanism

“…There is another oxidation enzyme, peroxidase, which catalyzes the oxidation of polyphenols in the presence of hydrogen peroxide. The importance of peroxidase in tea fermentation is pointed out because hydrogen peroxide is generated during catechin oxidation (Subramanian et al, 1999;Nakayama et al, 2002). In our experiments, the initial oxidation was probably catalyzed by polyphenol oxidase, because the presence of excess amounts of catalase, which decompose hydrogen peroxide, did not affect the reaction in an experiment using Japanese pears.…”

Efeito do tipo de corte e de sanificantes no escurecimento de pequi minimamente processado

“…A peroxidase promove a oxidação de compostos fenólicos na presença de peróxido de hidrogênio (Dunford & Stillman, 1976). A liberação de peróxido de hidrogênio na oxidação de alguns compostos fenólicos, catalisada pela polifenoloxidase, poderia indicar uma possível ação sinergística entre essas duas enzimas, o que sugere a participação da peroxidase nos processos de escurecimento (Subramanian et al, 1999).…”

Alternative pathway implicated as an influencing factor in the synthesis of theaflavin

“…During the fermentation stage in the manufacturing of black tea, monomeric flavan-3-ols (catechins) undergo polyphenol oxidase (PPO)-dependent oxidative polymerization producing various quinones that go through condensation reactions, resulting in dimeric compounds known as theaflavins (TF). Peroxidase utilizes hydrogen peroxide generated by PPO during the oxidation of catechins, and catalyses the oxidation of the products of the PPO catalysed reactions, amounting to the conversion of formed theaflavins into thearubigins [33].…”

“…For this reason, various approaches have been employed for the synthesis of theaflavins from substrate materials, including fresh tea leaves [34], tea leaf juice, processed green tea leaves, green tea slurry [11], liquid green tea extract [16] and in vitro modelling of fermentation with purified catechins [27]. These approaches included the utilization of potassium ferricyanide as a catalyst in chemical oxidation along with enzymatic oxidation [33,39]. Enzymatic oxidation approaches entail the exploitation of endogenous enzymes in liquid plant extracts or crude tea leaves containing PPO [27,33], peroxidase-containing cultured plant cells such as cultured tea cells [34] and homogenates of various fruits [36].…”

“…These approaches included the utilization of potassium ferricyanide as a catalyst in chemical oxidation along with enzymatic oxidation [33,39]. Enzymatic oxidation approaches entail the exploitation of endogenous enzymes in liquid plant extracts or crude tea leaves containing PPO [27,33], peroxidase-containing cultured plant cells such as cultured tea cells [34] and homogenates of various fruits [36]. Many fruit homogenates are capable of synthesising theaflavins, and the mechanism was suggested to be similar to that present in tea fermentation.…”