Study On 2-phenylethanol Biosynthesis Of Yeast

2-Phenethyl alcohol is an aromatic alcohol with a rose-honey scent,which is widely used in the cosmetics industry,the food industry and the pharmaceutical industry.At present,phenylethyl alcohol is synthesized mainly through natural extraction method,chemical synthesis method and microbial biotransformation synthesis method.In this study,Zygosaccharomyces rouxii and Saccharomyces cerevisiae are the research objects to explore their ability to synthesize phenylethanol.The main research work is as follows:gas chromatography tandem mass spectrometry identified this alcohol as2-phenylethanol(1)Z.rouxii M2013310 was fermented in YPD medium to the stable stage,and the volatile aromatic components of Z.rouxii M2013310 were dentified this alcohol as2-phenylethanol by gas chromatography-mass spectrometry(GC-MS),indicating that the strain had the ability to transform and synthesize 2-phenylethanol.The third-generation sequencing(Pac Bio)technology performs whole-genome sequencing,assembly and annotation of strains.The total length of the whole genome of yeast M2013310 is 7.3 Gb,and a total of 9043 genes have been annotated.The results show that this strain is a diploid strain with the shikimic acid pathway and the Ehrlich pathway for the synthesis of phenylethanol.(2)Z.rouxii M2013310 can tolerate up to 4 g/L phenethyl alcohol.After 72 h of fermentation of yeast M2013310 in M3(NH₄⁺),M3(Phe),and M3(Phe+NH₄⁺)media,the final yield of phenylethanol can reach 3.80 mg/L,3.58 g/L and 1.79 g/L,respectively.This suggested that Z.rouxii M2013310 can synthesize 2-phenylethanol via the shikimate pathway.Besides,the Ehrlich pathway is the main metabolic pathway for the synthesis of phenylethanol.The genes aromatic aminotransferases ARO8 and ARO9,phenylpyruvate decarboxylase ARO10 and PDC1,alcohol dehydrogenases ADH2,and control factor ARO80 are not nitrogen metabolism repressor(NCR)sensitive genes.The presence of ammonium sulfate in the culture medium mainly affects the transport of phenylalanine into the cell,resulting in a decrease in the production of phenylethanol.(3)S.cerevisiae FY20001 can tolerate 4 g/L phenethyl alcohol.S.cerevisiae FY20001 has the highest concentration of phenylethanol in the medium with phenylalanine as the sole nitrogen source,up to 3.90 g/L.The phenylalanine in the M3(Phe+NH₄⁺)medium is depleted and the yield of phenylethanol is as high as 3.20 g/L.The presence of the preferred nitrogen source ammonium sulfate will not affect the transport of phenylalanine into the cell.In addition,the real-time quantitative PCR data of S.cerevisiae FY20001 showed that the strains GAP1,aromatic aminotransferases ARO8 and ARO9,and phenylpyruvate decarboxylase ARO10 are sensitive genes for nitrogen metabolism repression,and the presence of ammonium sulfate inhibits the expression of these genes.