Isolation, Partial Purification And Some Properties Of Crude Gut Enzymes From The African Giant Snail (Achatina Achatina) Used For Yeast Protoplast Production:- Mbajiuka, Chinedu S

ABSTRACT

The crude gut enzyme ofthe African giant snail was partially purified 1 extraction with acetate buffer, fractional precipitation with ammonium sulpha and acetone followed by dialysis and heat treatment. The protein content of t , crude enzyme Extract was 1.05mg/ml, while that of ammonium sulphate ai •acetone fractions were 0.75mg/ml and 0.65mg/ml respectively. The partial purified enzyme fractions had enzyme activity of 4.70U/ml for ammoniu sulphate and 3.80U/ml for acetone while that ofthe crude extract was 1.60U/n The temperature range for the enzyme activity of both the crude and t partially purified fractions were 25°-60°C, while the pH range was 5.0- 6.0.T production of yeast protoplasts was achieved using both the crude and tl partially'purified fractions. There was 238% increase in protoplasts produced 1 the acetone fraction and 294% increase by the ammonium sulphate "fracti when compared with the number of protoplasts produced by the crude extra The statistical analysis showed significant differences (P<0.05) of protoplaj production from the various purification methods. Protoplasts products ’ increased with higher concentrations and higher incubation period. Optimu results of 3.95 x 106protoplasts/ml for Saccharomyces cerevisiae and 5.15 x 1 protoplasts/ml for Pichia pastoris were obtained when equal volume stabilized yeast cells and ammonium sulphate fraction of the partially purifu enzyme were mixed; incubated at 30°C for 120minutes with 1.0M sorbitol osmotic stabilizer and at a pH of 6.0. The results obtained from this study c; provide key factors for optimum yeast protoplasts production using the g enzymes oiAchatina achatina and offer a good means of commercial utilizatic of this otherwise wasted enzymes