Experiments And Optimization Of Biosynthesis Of Volatile Fatty Acids Into Microbial Lipids By Yarrowia Lipolytica

In recent years,volatile fatty acids(VFAs)are considered to be effective and promising alternate carbon sources for microbial lipid production by a few oleaginous yeasts,but the research is still in the early stage,lacking research results and theoretical findings.Moreover,the severe inhibitory effect of high-content(>10 g/L)VFAs on these yeasts has impeded the production of high lipid yields and their large-scale application.In this thesis,a promising yeast was selected through investigations combing the feature of large-scale production and the characteristic of VFA contained in organic-waste fermentate.Then,a systematic research on the lipid-producing performance of yeast cultivating in different conditions based on different VFA types and different VFA concentrations was conducted to obtain more understanding to optimize the process parameters.Finally,this thesis proposed an effective approach to solving the inhibition problem of high-concentration VFA and improving lipid production.The contributions to knowledge are summarized as follows.Under appropriate cultivation conditions,multiple oleaginous yeasts could take low concentrations of VFA,as the only carbon source,for lipid production.The yield of lipid,however,was low.Among the three yeasts used for the test,Yarrowia lipolytica(CICC-31596)yielded highest lipid.It also had a satisfactory tolerance for a specific hot environment at 38? and acid environment caused by non-adjusting pH which may occur in the practical application.The lipid-producing performance indexes of yeasts cultured with different types and concentrations of VFAs under different conditions were studied comprehensively and systematically,and the relevant conclusions and rules with guiding significance and reference value for follow-up research and actual production were summarized,mainly including:1)a series of common rules of yeast growth and lipid accumulation by using VFA as the only carbon source,such as the preference of yeasts for acetic acid;2)the characteristics of Yarrowia lipolytica in growth and lipid accumulation by using VFA,such as its step-wise utilization manner of different types of VFA in the mixed VFAs system;3)it was confirmed that in the low concentration system with VFA?10 g/L,the optimal lipid-production performance was obtained at the pH 6?7 due to the negligible inhibition of VFA and the yeast's adaptability towards environment.For Yarrowia lipolytica,the highest oil yield(1.17g/L)was obtained in the initial pH of 6,10 g/L acetic acid.The hypothesis that alkaline pH condition could effectively alleviate or even eliminate the inhibition of high concentration of VFA was proposed and verified,so as to realize the direct utilization of high concentration of VFA by yeast and greatly increase the oil yield.According to the results,the optimal pH condition suitable for the fermentation of high concentration of VFA by Yarrowia lipolytica was:the initial pH=8.Under this condition,the yeast could adapt to a concentration of up to 1 10g/L acetic acid and obtain the highest lipid production in the 70 g/L acetic acid system,which was 10.11g/L,achieving a significant increase in oil production.It was also proved in this study that the effects of alkaline environment to alleviate the toxicity of high concentration of VFA could be applied to food waste fermentate system(FW)and fruit and vegetable waste fermentate system(FVW),which contained high concentration of VFAs.When cultivated in undiluted FW fermentation broth(VFAs:35.35 g/L)and FVW fermentation broth(VFAs:22.18 g/L),cultures at the optimal alkaline condition(pH 8)had much better performance in cell growth and lipid accumulation than cultures at pH 6 and 7.The obtained biomass,lipid content and lipid concentration were FW:14.65 g/L,21.86%,3.20 g/L;FVW:11.84 g/L,26.02%,3.08 g/L.This study demonstrated the feasibility of utilizing synthetic and organic waste-derived VFA for lipid production by oleaginous yeasts.Through a series of systematic experiments,many meaningful research results and theoretical findings were provided.This study proposed and confirmed a method to overcome the inhibition of high concentrations of VFA,so as to effectively improve the cell growth and lipid synthesis capacity of the yeast in a high concentration VFA system.It could provide a new cultivation strategy to achieve simple utilizations of high-content VFAs and significantly increase the lipid yield.