Preliminary Exploration Of The Biorefinery Process For The Production Of Tyrosol From Cellulose

Lignocellulose is one of the best sources of renewable energy and biofuels.It is abundant in nature,and is of great importance in biorefinery technology.Lignocellulose can be used as a source to produce a variety of biofuels and chemicals to alleviate environmental pollutions and energy fuel deficiencies.This study was based on the construction of a biorefinery process by using lignocellulosic biomass to produce high value product tyrosol.The improvement of cellulose saccharification efficiency and the optimization of yeast fermentation process in tyrosol production were studied.A new process of simultaneous saccharification and fermentation to produce tyrosol and whole cell transformation of tyrosol to hydroxytyrosol was preliminary explored.The CBM of Cel7A from Trichoderma reesei was studied in order to improve the saccharification efficiency of cellulose.The amino acid residues at different positions were mutated,and five variants with improved saccharification efficiency were obtained,named Cel7A-CBMT1D,Cel7A-CBMI11D,Cel7A-CBMG22D,Cel7A-CBMS14D/S21D/T24D/P30D,Cel7A-CBMS14D/V18D/S21D/T24D/P30D,The variant with the highest increase in saccharification efficiency was Cel7A-CBMS14D/V18D/S21D/T24D/P30D,and the saccharification efficiency was increased by 59.0%when Avicel was used as substrate.Then,the effects of different carbon sources on the metabolic engineering of yeast-produced tyrosol were studied.The fermentation process was explored and optimized by shake flask and fermenter,and the yield of tyrosol under fermenter conditions was finally improved by 3.85 times to 3.35 g/L.In addition,it was found that ethanol as a feed carbon source has a higher carbon conversion rate than glucose.Finally,an attempt was made to use filter paper as the substrate to produce tyrosol by the simultaneous saccharification and fermentation process,and the tyrosol yield of 0.3g/L was obtained eventually,which initially achieved a biorefinery process for producing tyrosol from cellulose.Simultaneously,the Escherichia coli expressing 4-hydroxyphenylacetate-3 monooxygenase(EC 1.14.14.9)and flavin reductase(EC 1.5.1.36)was constructed,and whole cell transformation was carried out using tyrosol as a substrate,which was further converted into hydroxytyrosol.