ABSTRACT

The crude gut enzyme of the African giant snail was partially purified by extraction with acetate buffer, fractional precipitation with ammonium sulphate and acetone followed by dialysis and heat treatment. The protein content of the crude. enzyme extract was 1 .05mg/mi, while that of ammonium suii5hate and acetone fractions were 0:75mg/mi and 0.65mg/ml respectively. The partially purified enzyme fractions had enzyme activity of 4.70U/mi for ammonium suiphite and 3 .80U/ml for acetone while that of the crude extract was 1 .60U/ml. The temperature range for the enzyme activity of both the crude and the jartially purified fractions were 25°-60°C, while the pH range was 5.0- 6.0.The production of yeast protoplasts was achieved using both the crude and the partially purified fractions. There was 238% increase in protoplasts produced by the acetone fraction and 294% increase by the ammonium sulphate fraction when compared with the number of protoplasts produced by the crude extract. The statistical analysis showed significant differences (P<0.05) of protoplasts production from the various purification methods. Protoplasts production increased with higher concentrations and higher incubation period. Optimum results of 3.95 x 106 protoplasts/mi for Saccharomyces cerevisiae and 115 x 1 6 protopiasts/tnl for Pichia pastoris were obtained when equal volume of • stabilized yeast cells and ammonium sulphate fraction of the partially purified enzyme were mixed; incubated at 30°C for 120minutes with 1.OM sorbitol as osmotic stabilizer and at a pH of 6.0. The results obtained from this study can 'provide key factors for optimum yeast protoplasts production using the gut enzymes of Achatina achatina and offer a good means of commercial utilization of this otherwise wasted enzymes.