CARBOXYLESTERASES OF RAPANA VENOSA DIGESTIVE GLANDS HOMOGENATES

Abstract

Aim. Investigation of biochemical and physico-chemical properties of carboxylesterases in homogenates gland and hepatopancreas of Rapana venosa; study of the features of enantioselective enzymatic hydrolysis of 3-hydroxy-1,4- benzodiasepine-2-one ester, potential anxiolytic and hypnotic drug. Methods. Digestive glands of Rapana venosa were homogenized for subsequent investigations, the protein content in them was determined by the Lowry-Hartree method and esterase activity – according to 1-, 2-naphthyl acetates. Enzymatic hydrolysis of 3-hydroxy-5-phenyl-7-bromo-1,2-dihydro-3H-1,4-benzodiasepine-2-one was conducted for 2,5 h in the solution: dimethyl sulfoxide: Na-phosphate buffer 0,0167 M, pH 7,0 in volume ratio 2:3, at 37 °C. Determination of enantiomeric excess was conducted with help of high performance liquid chromatography in SHIMATSU system, equipped with ChіraDex HR 5μm(4mm×250mm) column. Results. For the first time it was shown, that esterase activity of hepatopancreas and esophageal gland homogenates for 2-naphtyl acetate as substrate, was 3,6-fold and 6,7- fold greater, than for 1-naphtyl acetate, respectively. The belonging of esterases in homogenates to the family of carboxylesterase was confirmed by the total suppression of their activity by the selective carboxylesterase inhibitor, di-(p-nitrophenyl)phosphate (2.0 mmol/dm3). It was shown, that pH-optima of esterase activity in homogenates of esophageal gland and hepatopancreas equals 7,5 and 5,5, respectively. The features of 3-hydroxy-1,4-benzodiazepine-2-one ester hydrolysis, catalyzed by carboxylesterase of Rapana venosa digestive glands homogenates were studied. The preferential formation of the R-enantiomer of substrate was shown (enantiomeric excess of R- enantiomer was 42%). Conclusion. The regioselectivity of carboxylesterase to 1-, 2-naphtyl acetates and enantioselectivity of 3-hydroxy-1,4-benzodiazepine-2-one ester were found.

Keywords

• карбоксилестераза
• травні залози rapana venosa
• регіоселективність
• енантіоселективність