Study On Fermentation Technology Of Microbial Sterols With Fatty Acid And Its Mechanism For High-yield Sterols Production

Sterols is a kind of steroid widely found in nature.And sterols is a kind of monohydric secondary alcohol of full cyclopentane hydrogen skeleton,which given four-ring steroid nucleus.Sterol can resist oxidation,decrease blood lipids,diminish inflammation and resist cancer,and has the ability of promoting animal growth and protecting skin.Sterol also can be widely used in medicine,food,cosmetics and feed industries.In this study,oleic acid was fermented with C.tropicalis 1253.The fermentation conditions of sterol-rich cells and the extraction conditions were researched.The main results are as follows:1)The optimization of culture conditions of sterol-rich yeast cells.The suiatable amount of oleic acid was 40 g/L.The best nitrogen source was the compound of urea,beef extract and yeast extract.Ammonium dihydrogen phosphate significantly increased the yield of sterol.The optimal growth factor was a combination of glycine and alanine.The highest sterol yield was obtained when emulsifier OP10 was added.The optimal medium combination was: oleic acid 40 g/L,1 g/L urea + 1g/L beef extract + 0.5 g/L NH4NO3,0.1 g/L MgSO4,0.2 g/L ZnSO4,0.01% glycine,0.01% alanine,0.5 g/L Na2HPO4,1% SDBS + 1% PEG400.The bed shaking condition was 30℃,200 r/min.The total content of sterol was 5.93%(wet weight).2)Study on the extraction of sterol by acid-heating method.The cells of C.tropicalis 1253 was extracted with acid-heating method,which was fermented by oleic acid.The study found that the extract obviously divided into three layers,whichi were named the organic phase,the upper of water phase and lower of water phase.Sterols mainly existed in the organic phase and the upper of water phase.The appropriate extraction conditions of sterols in the organic phase: concentration of hydrochloric acid was 5 mol/L,water bath temperature was 90℃,water bath time was 2h.-20℃ refrigerator was used to cool to room temperature.The appropriate extraction conditions of sterols in the upper water phase: material liquid ratio was 15:1,extraction time was 5 min,extraction temperature was 35℃,the origanic solvent was n-hexane.The upper of water phase wds dried firstly.The total extraction yield of sterols by acid-heating method was 80.16%.3)The preliminary study on the high yielding mechanism of stero in C.tropicalis 1253.When the carbon source was fatty acids,the total sterols content in the yeast cells was much higher than the carbon source was glucose.It suggested that the reason was the transformation of carbon source.Sodium hexametaphosphate,acetyl CoA,pyruvate had great role in promoting the synthesis of sterols.The preliminary inference on the high yielding mechanism of sterols in C.tropicalis 1253: the oxidation of fatty acid provided a number of key precursor such as acetyl CoA and a lot of energy substances such as ATP for sterol synthesis.