Study on factors influencing stability of whole cell during biodiesel production in solvent-free and tert-butanol system

Utilizing whole cell biocatalyst for biodiesel production has some advantages since it can avoid the complex procedures of isolation, purification and immobilization of extracellular lipase. However, during repeated use of Rhizopus oryzae (R. oryzae) IFO4697 whole cell for biodiesel production in solvent-free system, the whole cell exhibited very poor stability; while the whole cell stability has been found to be significantly improved in tert-butanol system compared to that in solvent-free system. The difference in whole cell stability was found to be due to the difference of product accumulation between solvent-free and tert-butanol system. After 144 h reaction, glycerol and methyl ester accumulated in the cell in solvent-free system came up to about 1000 mg/g and 350 mg/g dry biomass, respectively, while in tert-butanol system, glycerol and methyl ester accumulation was kept at a relatively low level, approximately 100 mg/g and 2 mg/g dry biomass, respectively. The accumulated glycerol influenced whole cell stability through mass transfer limitation only, while the accumulated methyl ester influenced whole cell stability through both mass transfer limitation and product inhibition. Further study showed that a slight loss in enzymatic activity in tert-butanol system was caused by protein leakage.