生物聯(lián)合加工菊芋生產乙醇的釀酒酵母菌株構建
發(fā)布時間：2018-08-25 18:33
【摘要】：利用生物質資源通過生物煉制生產燃料乙醇是近年來研究的熱點。生物聯(lián)合加工策略(Consolidated bioprocessing,CBP)集成產酶、酶解和發(fā)酵,是具有良好發(fā)展前景的生物燃料生產技術。菊芋可在干旱和鹽堿地等邊際土地生長且生物質產量高,是可選擇的能源作物,但利用菊芋發(fā)酵生產乙醇的研究報道很少。菊芋的主要生物量來自塊莖和秸稈,塊莖主要成分是菊粉,是提取附加值較高菊粉產品的主要原料,而秸稈可用于生產燃料乙醇。本研究的目的是基于原料組成特點,選育具有高發(fā)酵活性的釀酒酵母CBP菌株并優(yōu)化工藝過程的操作單元,提高乙醇發(fā)酵水平。首先研究了菊芋秸稈乙醇發(fā)酵。結果表明:2%(w/v)氫氧化鈉預處理可以有效去除木質素,纖維素含量由39.4%提高到58.2%,且保留16.4%的菊粉。20%秸稈補料批次糖化總還原糖為115.8 g/L。通過細胞表面展示在耐高溫工業(yè)釀酒酵母安琪酵母中整合菊粉酶基因,構建CBP菌株Saccharomyces cerevisiae MK01,其水解秸稈中菊粉性能良好,發(fā)酵液中乙醇濃度為38.3 g/L,得率為0.361 g/g,為理論得率的71%。其次,針對菊芋大規(guī)模種植塊莖產量可能過剩的問題,采用S.cerevisiaeMK01以塊莖為底物進行CBP乙醇發(fā)酵的研究。結果表明:溫度控制在38-42℃,c,S.cerevisiae MK01生長沒有明顯差異,但在42 ℃時菊粉酶活力提高,乙醇發(fā)酵過程加快,72 h內消耗198.2 g/L菊粉水解糖,產生89.3 g/L乙醇;以185.7 g/L菊芋塊莖水解總還原糖為底物,72 h內產生85.2 g/L乙醇,乙醇得率為0.459 g/g,達理論得率的90%。最后,探討了有利于工業(yè)生產環(huán)境的耐高溫CBP菌株選育,并對其耐高溫分子機理進行了研究。通過導入人工轉錄因子,獲得了一株表面展示纖維素酶且耐高溫的CBP菌株。與親本相比工程菌株可以在43℃條件下生長,與對照菌株相比,二者在胞內海藻糖含量、活性氧自由基積累和超氧化物歧化酶活力方面存在明顯差異。工程菌株磷酸膨化纖維素的降解率提高了 45.8%,此外在43 ℃時以菊芋秸桿為底物的乙醇發(fā)酵周期縮短了 24 h。比較轉錄組學分析表明,工程菌株細胞壁完整性、PKC1-MPK1代謝通路及γ-氨基丁酸代謝旁路(GABA shunt)關鍵酶基因表達上調,表明人工轉錄因子通過對代謝的整體調控改善了細胞膜組成,進而改善了高溫耐性。這些研究進展為菊芋秸稈和塊莖綜合利用生產燃料乙醇,奠定了良好技術基礎。
[Abstract]:Using biomass resources to produce fuel ethanol through biological refining is a hot topic in recent years. Consolidated bioprocessing,CBP integrated enzyme production, enzymatic hydrolysis and fermentation is a promising biofuel production technology. Jerusalem artichoke (Jerusalem artichoke) is an alternative energy crop which can grow in marginal land such as arid and saline-alkali land and has high biomass yield. However, there are few reports on the production of ethanol by using Jerusalem artichoke fermentation. The main biomass of Jerusalem artichoke comes from tuber and straw. The main component of tuber is chrysanthemum powder, which is the main raw material to extract higher added value inulin product, and straw can be used to produce fuel ethanol. The aim of this study was to select the yeast CBP strain with high fermentative activity and optimize the operation unit of the process based on the characteristics of raw material composition to improve the ethanol fermentation level. Firstly, ethanol fermentation of Jerusalem artichoke straw was studied. The results showed that sodium hydroxide pretreatment (w / v) could effectively remove lignin, increase cellulose content from 39.4% to 58.2%, and retain 16.4% inulin. 20% straw batch saccharification total reducing sugar was 115.8 g / L. The inulinase gene was integrated on the cell surface of yeast An Qi, a thermotolerant industrial Saccharomyces cerevisiae, and CBP strain Saccharomyces cerevisiae MK01, was constructed. The hydrolyzed straw showed good inulinase performance. The ethanol concentration in fermentation broth was 38.3 g / L, and the yield was 0.361 g / g, which was 71g / g of the theoretical yield. Secondly, aiming at the possible excess of tuber yield in large scale planting of Jerusalem artichoke, S.cerevisiaeMK01 was used as substrate for CBP ethanol fermentation. The results showed that there was no significant difference in the growth of S.cerevisiae MK01 at 38-42 鈩，
本文編號：2203746
[Abstract]:Using biomass resources to produce fuel ethanol through biological refining is a hot topic in recent years. Consolidated bioprocessing,CBP integrated enzyme production, enzymatic hydrolysis and fermentation is a promising biofuel production technology. Jerusalem artichoke (Jerusalem artichoke) is an alternative energy crop which can grow in marginal land such as arid and saline-alkali land and has high biomass yield. However, there are few reports on the production of ethanol by using Jerusalem artichoke fermentation. The main biomass of Jerusalem artichoke comes from tuber and straw. The main component of tuber is chrysanthemum powder, which is the main raw material to extract higher added value inulin product, and straw can be used to produce fuel ethanol. The aim of this study was to select the yeast CBP strain with high fermentative activity and optimize the operation unit of the process based on the characteristics of raw material composition to improve the ethanol fermentation level. Firstly, ethanol fermentation of Jerusalem artichoke straw was studied. The results showed that sodium hydroxide pretreatment (w / v) could effectively remove lignin, increase cellulose content from 39.4% to 58.2%, and retain 16.4% inulin. 20% straw batch saccharification total reducing sugar was 115.8 g / L. The inulinase gene was integrated on the cell surface of yeast An Qi, a thermotolerant industrial Saccharomyces cerevisiae, and CBP strain Saccharomyces cerevisiae MK01, was constructed. The hydrolyzed straw showed good inulinase performance. The ethanol concentration in fermentation broth was 38.3 g / L, and the yield was 0.361 g / g, which was 71g / g of the theoretical yield. Secondly, aiming at the possible excess of tuber yield in large scale planting of Jerusalem artichoke, S.cerevisiaeMK01 was used as substrate for CBP ethanol fermentation. The results showed that there was no significant difference in the growth of S.cerevisiae MK01 at 38-42 鈩，
本文編號：2203746
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