【作者】 王亮；

【导师】 孙付保；

【作者基本信息】 江南大学 ， 发酵工程， 2015， 硕士

【摘要】 目前纤维素乙醇成本偏高的根本原因在于没有达到淀粉质乙醇发酵水平的“三高”(高浓度、高转化率和高效率)指标,提高预处理后基质的纤维素纯度、水解糖液浓度和避免发酵抑制物来实现浓醪发酵,是解决问题的关键。本论文对木质纤维素原料转化为生物乙醇过程中的预处理及浓醪发酵等问题进行了研究。甘蔗渣经过甘油蒸煮溶液蒸煮后,其可酶解性显著提高,本文通过对蒸煮过程中关键参数蒸煮温度以及蒸煮时间进行了优化,得出最佳工艺参数:甘蔗渣10 g、70%甘油水溶液200 g、蒸煮温度220℃和蒸煮时间2 h。此时预处理的纤维素保留率为95%,半纤维素和木质素脱除率分别为70%和65%;预处理物料在酶载量16 FPU·g-1干基质时24h酶解率可达90%以上。通过一系列现代新型分析表征手段(热重分析、扫描电镜、原子力显微镜、激光共聚焦显微镜、傅里叶红外、X射线衍射以及固体核磁共振)对甘油处理前后的麦草及甘蔗渣物料进行了分析表征。结果表明常压甘油自催化这种新型预处理可以从三个方面有效地打破木质纤维素基质复杂的抗逆结构:1)组成上可以有选择性地脱除半纤维素、木质素和乙酰基团;2)物理结构上朝着有利于酶解的方向改变物料的物理特性;3)化学结构上打破了物料分子间及分子内的主要化学键和功能性基团导致了向同质异晶纤维素的转化。上述特征充分解释了预处理后物料酶解性提高的原因。以常压甘油自催化预处理麦草作为基质,进行了纤维素乙醇浓醪发酵的探索。在对发酵培养基和发酵条件等优化选择基础上,通过采用同步糖化发酵、半同步糖化发酵和分批补料式半同步糖化发酵等一系列强化策略,逐步实现了该基质的浓醪发酵。在同步糖化发酵条件(15%基质浓度、加酶量30 FPU·g-1干基质、硫酸铵10 g·L-1;接种量10%,温度37℃,初始p H4.8,全程震荡发酵)下发酵72 h时乙醇产量为31.2 g·L-1,发酵强度0.43 g·(L·h)-1;改用半同步糖化发酵方式,预酶解时间24 h时乙醇产量提高到33.7 g·L-1,纤维素乙醇转化率将近80%,发酵强度0.47 g·(L·h)-1,其中(半)同步糖化发酵中90%以上纤维素已被糖化水解用于发酵;进一步采用分批补料式半同步糖化发酵,使基质浓度相当于30%,发酵96 h时乙醇产量达到52.8 g·L-1,纤维素乙醇转化率为62%,发酵强度为0.55 g·(L·h)-1;最后通过一系列强化措施(增加预处理后基质的纤维素纯度、增加发酵基质浓度、添加表面活性剂、延长补料时间、换用新型酶制剂)使得基质浓度相当于35%,发酵72 h后乙醇产量达到了73.1 g·L-1,发酵强度为1.02 g·(L·h)-1,乙醇浓度和发酵强度两个指标都达到了乙醇经济蒸馏的要求(乙醇浓度>40 g·L-1,发酵强度>1 g·(L·h)-1),纤维素乙醇转化率为67%。发酵96 h后乙醇产量达到了75.4 g·L-1,发酵强度为0.79 g·(L·h)-1。该发酵抑制物很少,无糠醛和5-羟甲基糠醛类发酵抑制物,乙酸含量不足4 g·L-1。因此,常压甘油自催化预处理木质纤维素原料适用作基质,进行纤维素乙醇浓醪发酵。更多还原

【Abstract】 The expensive production of bioethanol is due to that it has not yet reached the‘THREE-HIGH’(High-titer, high-conversion and high-productivity) technical level as the starchy ethanol production. To cope with it, it is necessary to manage a high-gravity mash bioethanol production(HMBP), in which the sugar hydrolysate is thick, the purity of cellulose is high and fermentation-inhibitive compounds are negligible. In this thesis, a new pretreatment and bioethanol fermentation of lignocellulosic materials was studied.The hydrolyzability of sugarcane bagasse was improved by an atmospheric aqueous glycerol autocatalytic organosolv pretreatment(AAGAOP). With a preliminary optimization on some key factors such as cooking temperature and cooking time, the pretreatment of sugarcane bagasse was processed as below: 70% glycerol solution at 220 ℃ for 2 h. At the optimized condition, the pretreatment had a good cellulose recovery(95%), and high hemicellulose(70%) and lignin(65%) removals. Interestingly, the cellulose conversion of the pretreated substrate was up to 90% with an enzyme loading of 16 FPU·g-1 dry substrate.Several modern analytic techniques(TGA, SEM, AFM, CLSM, FT-IR, XRD and CP/MAS 13C-NMR) were used to characterize changes of wheat straw and sugarcane bagasse before and after the AAGAOP. The good hydrolyzability of substrates subjected to AAGAOP is mainly due to that the AAGOAP can effectively disrupt the complex, recalcitrant architecture of lignocellulosic substrates, which divides into three hierarchies:compositionally to selectively remove some component barriers(i.e., lignin, hemicellulose and acetyl group), structurally to dissect the native physical structure into some features helpful for hydrolyzability, and structure-chemically to dissociate key chemical bonds and functional groups(i.e., β-ether bond, β-ester band and hydrogen bond) of inter- and intramolecules, resulting in allomorphous transformations(from crystalline to amorphous or para-crystalline).Finally, HMBP from wheat straw after the AAGAOP was carried out with different fermentation strategies. Under an optimized condition(15% substrate concentration,(NH4)2SO4 10 g·L-1, 30 FPU·g-1 dry matter, inital p H4.8, fermentation temperature 37 ℃,10%(v/v) inoculum ratio), the HMBP was at 31.2 g·L-1with a shaking simultaneous saccharification and fermentation(SSF) for 72 h, which reached a conversion of 73% and a productivity of 0.43 g·(L·h)-1. Further by a semi-SFF with a pre-hydrolysis time of 24 h, the HMBP reached 33.7 g·L-1, with 79% of the conversion and 0.47 g·(L·h)-1 of the productivity.During the SSF and semi-SSF, more than 90% of the cellulose in both substrates was found to hydrolyze into fermentable sugars. Again, a fed-batch semi-SFF was developed with an initial substrate concentration of 15%. The HMBP achieved 52.8 g·L-1for 96 h with a productivityof 0.55 g·(L·h)-1 and a cellulose conversion of 62%. Finally, such several strategies as purifying the substrate cellulose, increasing substrate concentration, extending the fed-batch time, adding the surfactant and using the new cellulase preparation Cellic CTec2, were used.With an equivalent substrate concentration of 35%, the HMBP reached 73.1 g·L-1for 72 h with a high productivity of 1.02 g·(L·h)-1 and a cellulose conversion of 67%. Notably, the fermentation inhibitive compound was mainly the acetic acid at less than 4.0 g·L-1, and there were no other inhibitors detected, commonly as such furfural and hydroxymethylfurfural existing in the slurry, during the three types of fermentation. The data indicate that the lignocellulosic substrate subjected to the AAGAOP is very applicable for the HMBP.更多还原