Purification and partial characterization of milk-clotting enzyme extracted from glutinous rice wine mash liquor

摘要

Glutinous rice wine mash liquor is a traditional food of south of China and its ability to coagulate the milk has been proved. The aim of this work was to extract milk-clotting enzyme from glutinous rice wine mash liquor. A partial purified extract of enzyme was obtained by fractional precipitation with (NF_4)_2SO_4 The fractions obtained by precipitation, 40-90% possessed the milk-clotting activity (MCA) (145.72 U/mg). The 40-90% (NH_4)_2SO_4, fraction was further purified by sephadex G-100 and DEAE-sephadex A-50 with MCA (4,360±50 U/mg), which was confirmed by SDS-PAGE that showed only one band with a molecular mass of 36.0 kDa. Highest MCA was attained at 36℃. The enzyme was completely inactivated by heating for 20 min at 60℃. The MCA increased with the decreasing of milk pH from 8.0 to 5.5, and it was active at the wide range of pH 1 to 7. The metal ions Mg~(2+), Ca~(2+), Ba~(3+), MN~(2+), Al~(3+), Fe~(2+) had a very clear function to accelerate milk coagulation whereas Na~+ and K~+ decelerated the activity slightly. The curd effect of the milk-clotting enzyme has primarily been studied.