产核黄素枯草芽孢杆菌代谢物组的初步研究-发酵工程专业论文.docxVIP

ABSTRACT According to comparison and optimization ，a scheme of metabolomics analysis byLCMS was developed. 叽le cold methanol (60% v/v) quenching proωcol has been used in 0町 labora阳ry for instant quenching of metabolic activity，however，cell leakage w制 found during the quenching procedure. Therefore，quick filtration method |‘ without quenching process was selected to separate cells 仕Om culture. In addition，we also compared t恤ee different extraction procedures，and fmally found that 4自 combination with the acetonitrile: me由anol: water and methanol: chlorofonn extraction method was more suitable for our research. The above metabolomics analysis method was applied in our research to distinguish various riboflavir←producing Bacillus subtilis by principal components analysis (PCA) and 16 metabolites were detennined by qualification analysis based on m.Iz ofmetabolites. The outcome showed that: Compared with the B.subtilis RH33，Ru5P，DRL，RIB w盯e significantly increased in B.subtilis RH33.SVZ ，B.subtilis RH33-SVG and B.subtilisRH33-VGZ ， whereas，PYR，CIT，SUC，MAL，OAA had great reduction. 叽le result ilIustrated that amplification of the key genes zl吧f and gnd can effectively increase the PP pathway flux，lower TCA cycle flux，change the flux distribution of glycolytic p削hwayand penωse phosphate pathway metabolic ，thus contribute to improve the yield of riboflavin. Meanwhile，due to the increase of PP fl岖， S-phosphoribose and NADPH for synthesis of the purine nucleotides and amino acids was adequately supplied，and L finally led ωthe increase of AIR， AMP，ADP，ATP，and Valine，Lysine，Phenylalanine ， Glutamate. f自‘ f KEY WORDS: metabolome Bacillus subtilis zl吃 gnd LιMS riboflavin 目录 龟 Ii. r-.， 第一章文献综述，...………………………………·…… 1 1.1 代谢组学简介…..………川..………………………… 1 1.2 代谢组学的提出 .11 …………..1 1.3代谢组学的发展……………………………………… ..2 1.4 代谢组学的研究方法………………………………………… 2 1.4.1 取样技术………………川…...3 1.4.2 灭活技术…………………………………