Optimization Of Conditions Fermented By Candida Bombicola With Fish Oil And Characterization Of Sophorolipids

Sophorolipids were kinds of glycolipid biosurfactant with high surface activity,wide application and accessible degradation.In this study,Candida bombicola ATCC 22214 were used as yeast strain,and cheap crude fish oil and glucose were used as carbon sources to produce sophorolipids.Fermentation medium and parameters were optimized by shaking flask culture.On this basis,fermentation process was further optimized by 7.5 L fermentation cylinder,and the fermentation kinetic models were established.Crude fermentation product was extracted and their physicochemical and emulsifying properties were studied.The main results of this study were as follows:By single factor analysis,fish oil and glucose were used as carbon source and the ratio was 1:1.Yeast extract was used as nitrogen source.KH2PO4 and Na2HPO4·12H2O were used as phosphorus source and the ratio was 1:1.Plackett-Burman design was used to determine the concentration of carbon source,concentration of nitrogen source and inoculation volume as three significant factors affecting the yield of sophorolipids.The central combination of response surface design was determined by the steepest ascent design.Through response surface design,the optimized formulation(g/L)and fermentation parameters were obtained as follows:glucose 80.0,fish oil 80.0(V/V),yeast extract 1.9,KH2PO4 1.5,Na2HPO4·12H2O 1.5,MgSO4 0.3,the temperature during the early stage(0-48 h)was 28℃,the temperature during the late stage(48168 h)was 24℃,inoculation volume was 1.7%.Under these conditions,the yield of Sophorolipids was 105.856 g/L,which was 71.05%higher than that before optimization.On the basis of shaking flask fermentation,the yield of sophorolipids was 106.728 g/L by using 7.5 L fermentation cylinder under the control of 2 L/min ventilation rate and 300 r/min stirring speed.The Logistic equation and the Luedeking-Piret equation were used to establish kinetic models of the growth of cell,the synthesis of product and the consumption of substrate during fermentation process.These three equations could simulate the fermentation process well,which had a certain guiding significance for the further production of sophorolipids by a scale-up experiment.The product was extracted with ethanol,and then the brown and red viscous crude extract was obtained by vacuum distillation and drying.The results of HPLC-MS analysis showed that the crude extract of sophorolipids had the characteristics of longer carbon chains and more unsaturated bonds.The CMC of the crude extract was 45 mg/L,and the corresponding solution surface tension was 42.6 mN/m.The efficiency of the crude extract was higher than that of SDS.Above the CMC,temperature and salinity had little effect on the surface tension of sophorolipids.Except for extreme alkaline condition(pH=12)that the surface tension of sophorolipids solution were increased slightly,sophorolipid showed good stability under extreme conditions.Above the CMC,the emulsifying effect of sophorolipids on soya-bean oil or paraffin liquid was better than SDS.It also showed a better emulsifying stability than SDS.The above work indicated that the desired target product,sophorolipids,could be obtained through the fermentation by C.bombicola.The results obtained would be of great significance to further study on the fermentation mechanism of sophorolipids and expand the fermentation scale.More significantly,this study successfully utilized the crude fish oil recovered from Zhoushan fishery as one of the main carbon sources,providing a new path for the comprehensive utilization of seafood processing by-products.