In contrast to previous knowledge of chlorophyllase activity in higher plants, significant enzyme activity was isolated from tea leaves in a soluble state. Soluble chlorophyllase was partially purified by procedures including ammonium sulfate fractionation (Preparation I). The insoluble fraction was extracted, by solubilizing it with SDC, from the methanol-acetone powder of sediments of the leaf homogenate, from which the water-soluble enzyme had been completely removed by repeated extraction. This initially insoluble enzyme was also partially purified (Preparation II). Specific activities (mg chlorophyll a hydrolyzed per hr per mg protein, 7.2 for Preparation I, and 12.4 for Preparation II), were much higher than those reported for other plant material. The soluble enzyme was more resistant to PCMB, lipase and heat treatment. The two enzymes differed in optimum temperature and optimum acetone concentration needed for the reaction, but showed the same optimum pH, and same Km value. The Km value was the same (7 μM) for reactions with 30% and 50% acetone. These results suggest that, in spite of differences in location and extractability, activities of the soluble and insoluble (solubilized) chlorophyllase in tea leaves are attributable to the same enzyme.
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Nagao Ogura (1972) studied this question.