Isolation and Purification of Lipase from the Midgut of Fifth Instar Larvae of Antheraea mylitta drury

Abstract

Lipases obtained from healthy and pebrinised larvae were purified by 45-85% (NH4) 2SO 4 fractionation followed by Sephadex S-100 gel filtration and CM-Sepharose. In both the samples final enzyme purification reported was 19.02 and 17.7 folds(magnitude) and the recovery of final purified enzyme was 19.32% and 17.14% with a specific activity of 7.87 and 7.52 µmol/min/mg. Results also show that activity of purified lipase was highest at pH 8 in both the samples. Highest lipase activity was recorded between 37°C to 40°C tem perature and lipase activity was maximum at 37°C temperature in healthy sample and 38°C tempe rature in pebrinised sample. The enzyme activity reduced with addition of NaCl, Urea and MgCl 2 whereas EDTA and CaCl 2 increased the activity. The molecular weight of the purified enzyme was 30 kDa as determined by SDSpolyacrylamide gel electrophoresis. Aim: The present study was conducted to isolate, purify and characterize lipases from the midgut of both healthy and pebrinised fifth instar larvae of Antheraea mylitta drury . Study Design: Study involves dissection of midgut from the fifth instar larvae of fourth day of both healthy and pebrinised larvae, Lipases obtained from both the samples were purified by 45-85%