Substrate Specificity of β-Primeverosidase, A Key Enzyme in Aroma Formation during Oolong Tea and Black Tea Manufacturing

Abstract

We synthesized nine kinds of diglycosides and a monoglycoside of 2-phenylethanol to investigate the substrate specificity of the purified β-primeverosidase from fresh leaves of a tea cultivar (Camellia sinensis var. sinensis cv. Yabukita) in comparison with the apparent substrate specificity of the crude enzyme extract from tea leaves. The crude enzyme extract mainly showed β-primeverosidase activity, although monoglycosidases activity was present to some extent. The purified β-primeverosidase showed very narrow substrate specificity with respect to the glycon moiety, and especially prominent specificity for the β-primeverosyl (6-O-β-D-xylopyranosyl-β-D-glucopyranosyl) moiety. The enzymes hydrolyzed naturally occurring diglycosides such as β-primeveroside, β-vicianoside, β-acuminoside, β-gentiobioside and 6-O-α-L-arabinofuranosyl-β-D-glucopyranoside, but were unable to hydrolyze synthetic unnatural diglycosides. The purified enzyme was inactive toward 2-phenylethyl β-D-glucopyranoside. The enzyme hydrolyzed each of the diglycosides into the corresponding disaccharide and 2-phenylethanol. These results indicate the β-primeverosidase, a diglycosidase, to be a key enzyme involved in aroma formation during the tea manufacturing process.

• substrate specificity
• β-primeverosidase
• diglycosidase
• β-primeveroside
• Camellia sinensis