Properties of tea-polyphenol-complexed bromelain

Abstract

Tea polyphenols extracted from Chinese green tea were used to complex and separate bromelain from pineapple juice. The antioxidant activity of tea polyphenols and their effects on the properties of bromelain were studied. The thermal stability of bromelain was enhanced after complexing with tea polyphenols. By complexing, the activity half-life of bromelain at 60°C was lengthened from 60 to 105 min. The improved thermal stability of the tea-polyphenol-complexed (TPC) bromelain is attributed to the antioxidant activity of tea polyphenols. Iso-electric focusing showed that six iso-enzymes fractions of bromelain could complex tea polyphenols with no change in their iso-electric points. Complexing with tea polyphenols did not change the optimum temperature (60°C) for the enzyme activity but the optimum pH value range was narrowed down from pH 6.8–9.0 to around 7.8. The Michaelis constant (K) of the TPC bromelain for casein substrate was determined to be 1.42×10⁻⁵ mol/l, higher than that of free bromelain (8.06×10⁻⁶ mol/l). Tea polyphenols were found to act as a competitive inhibitor on bromelain in the hydrolysis of casein.

Introduction

Polyphenolic compounds such as tannic acid, catechin, epicatechin and proanthocyanidin can react with proteins and enzymes to form light-scattering hazes. Sediments are formed when the hazes further complex each other and grow to colloidal size. This phenome non must be prevented in the production of beer, tea beverages, fruit juices and wine because it would impair consumer appeal. The mechanism of interaction between polyphenols and proteins, and the characteristics of haze-active proteins and haze-active polyphenols have been described in details in the literature (Asano, Ohtsu, Shinagawa and Hashimoto, 1984, Siebert, Carrasco and Lynn, 1996, Siebert, Troukhanova and Lynn, 1996, Van Buren and Robinson, 1969). Polyphenolic substances are present in green tea in large amount. The major polyphenolic compounds found in tea are the flavanols, which include catechin and gallocatechin, their esters with gallic acid, catechin gallate, and gallocatechin gallate, and their epimers. It has been reported that tea, as a natural beverage, has many beneficial health effects. These include antimutagenic activity, antioxidation, scavenger of free radicals, cancer prevention, and heart and blood vessel disease prevention (Akaike, T., Ijiri, S., Sato, K., Katsuki, T. and Maeda, 1995, Jovanovic, Hara, Steenken and Simic, 1997, Lin, Juan, Chen, Liang and Lin, 1996, Yen, Chen and Peng, 1997, Yu, Jiao, Duh, Krishnamurthy, Tan and Ant, 1997). The health benefits of tea are largely attributed to the antioxidant activity of tea polyphenols and their interaction with large bio-molecules (including proteins and enzymes). Some studies showed that enzymes and proteins such as lipoxygenase, α-amylase and xanthine oxidase were denatured by tea polyphenols (Hara and Honda, 1990, Sekiya, Kajiwara, Monma and Hatanaka, 1984, Yeo, Park, Kim, Kim and Park, 1995). However, the strong antioxidant activity and protein-complexing property of tea polyphenols may help to stabilize some sulfhydryl-group containing proteases such as bromelain, papain and actinidin, which tend to lose activity at room temperature because of oxidation. In this present investigation, tea polyphenols extracted from Chinese green tea were used to complex and separate bromelain from pineapple juice. The properties of the free and the tea-polyphenol-complexed (TPC) bromelain were determined.