Optimization for producing cell-bound lipase from Geotrichum sp. and synthesis of methyl oleate in microaqueous...

An integrated optimization strategy involving a combination of different designs was employed to optimize producing conditions of cell-bound lipase (CBL) from Geotrichum sp. Firstly, it was obtained by a single factorial design that the most suitable carbon source was a mixture of olive oil and citric acid and the most suitable nitrogen source was a mixture of corn steep liquor and NH4NO3. Then, the Plackett-Burman design was used to evaluate the effects of 13 variables related to CBL production, and three statistically significant variables namely, temperature, olive oil concentration, and NH4NO3 concentration, were selected. Subsequently, the levels of the three variables for maximum CBL production were determined by response surface analysis as follows: 1.64% (v/v) olive oil, 1.49% (w/v) NH4NO3, and temperature 33.00 degrees C. Such optimization resulted in a high yield of CBL at 23.15 U/ml, an enhanced 4.45-fold increase relative to the initial result (5.2 U/ml) in shake flasks. The dried CBL was used to synthesize methyl oleate in microaqueous hexane, resulting in 94% conversion after 24 h, and showed reusability with 70% residual activity and 69% conversion after eight cycles of batch operation, which indicating that CBL, as a novel and natural form of immobilized enzyme, can be effectively applied in repeated synthesis of methyl oleate in a microaqueous solvent.