【作者】 王堃；

【导师】 孙润仓；

【作者基本信息】 北京林业大学 ， 林产化学加工工程， 2011， 博士

【摘要】 本论文针对木质纤维原料生产燃料乙醇过程中的预处理技术进行了系列研究。重点探讨了蒸汽爆破和纤维素溶解预处理技术与机理,并对木质纤维原料和造纸黑液木质素的主要组分进行分离及结构研究。本文研究了蒸汽爆破预处理过程中不同蒸汽压力和维压时间对胡枝子杆物理化学性质、微观结构和酶解转化效率的影响；提出基于蒸汽爆破预处理和碱性溶剂后处理的清洁分离方法,对其主要组分的物理性质及化学结构进行分析；采用不同纤维素溶剂体系预处理木质纤维原料,阐明溶解机理,建立纤维素微晶结构与酶解转化效率的构效关系；采用有机溶剂分级分离方法纯化造纸黑液的木质素,并对各组分进行结构表征。所得主要结论如下：1、蒸汽爆破预处理过程明显地破坏了细胞壁的致密结构,降解并溶出大量的化学成分。胡枝子杆主要组分纤维素、半纤维素和酸不溶木质素的含量分别为44.6%、29.3%和17.0%。在所采用的蒸汽爆破预处理条件下(1.5—2.5 MPa,2—10 mmin),处理后样品纤维素含量为43.6%—47.1%,相对结晶度由处理前的10.55%增加到16.30%。同时,半纤维素组分明显降解,其含量最低可达3.7%,而木质素含量相应提高到17.7%—23.3%。2、蒸汽爆破预处理显著地提高纤维素酶对纤维素的可及度,从而提高了酶解糖化效率。蒸汽压力2.25 MPa时,蒸汽爆破预处理后底物的平均酶解糖化率分别为78%、93%和97%,较未预处理胡枝子提高约2.8倍。并且延长维压时间能进一步提高平均酶解糖化率。维压时间4 min时,蒸汽爆破压力的增加能够显著提高原料的平均酶解糖化率：1.5 MPa时,平均酶解糖化率仅提高约20%；而当压力升高到1.75 MPa时,平均酶解糖化率提高到原来的2倍。3、基于蒸汽爆破预处理技术和温和条件的后处理工艺,对胡枝子原料的主要组分进行了分离,提出了木质生物质原料清洁分离的新技术。分离后所得固体残渣主要为纤维素,其葡萄糖含量最高达94.3%。由于蒸汽爆破预处理纤维束化作用以及其它成分在后处理过程中的有效提取,分离得到的纤维素具有较高的可接触面积和较低的聚合度有利于随后的生物转化。在大于蒸汽压力2.0 MPa和维压时间4 min条件下聚合度可明显降低,最小值为207。分离得到的三种半纤维素组分在蒸汽爆破过程中明显降解,其分子量分别由47960 g/mol、63300 g/mol和102400 g/mol降至6380g/mol、8100 g/mol和16200 g/mol。大量的脱支链反应破坏了阿拉伯糖与木聚糖主链的连接,使阿拉伯糖与木糖比例的明显降低。此外,剧烈条件下部分纤维素发生降解。在2.5MPa,4 mmin条件下,经1%NaOH碱性乙醇水(60%,V/V)溶液分离得到的半纤维素中葡萄糖的含量达86.6%。木质素组分经蒸汽爆破预处理后较未预处理原料分离效率显著提高,1M NaOH碱性水溶液和1%NaOH碱性乙醇水(60%,V/V)溶液后处理最高可分别得到73.4%和93.7%的木质素组分,且其纯度有明显的提高,糖组分的含量最低均可达到0.8%。经1H NMR定量分析,木质素结构单元间β-O-4醚键,在2.25MPa蒸汽压力条件下,其含量由维压时间2 min时的11%降低至10 mmin时的7%。在剧烈条件下,大量缩合反应的发生形成了更不易降解、热稳定性较高的碳碳键,使木质素分子量显著增加,而由C5位的甲氧基取代基所造成立体位阻效应使愈创木基木质素较紫丁香基木质素更多的参与到缩合反应中。实验结果表明,中低强度的蒸汽爆破预处理与碱性溶剂后处理相结合的清洁分离新技术能显著提高各组分的提取效率,同时提取的半纤维素和木质素具有较高的纯度,有利于其进一步高值化利用。4、氢氧化锂/脲素、浓磷酸、氮甲基氧化吗啉和离子液体([BMIM]Cl)四种纤维素溶剂处理将原有纤维素Ⅰ结构转化为了纤维素Ⅱ结构或无定型结构,相对结晶度和聚合度最低可分别降至8%和60。相比于未处理样品(平均酶解转化率33%),经该四种溶剂处理后样品72小时的平均酶解转化率分别达到81%、92%、83%和80%,而且转化速率有了显著提高。同时,半纤维素的回收率也由未处理前的18.5%提高到了39.4%—49.6%。5、有机溶剂可将蒽醌硫酸盐法制浆黑液中回收的木质素依据溶解度的大小分级分离。随着溶剂希尔德布兰德系数从7.3 (cal/cm3)1/2增加到14.5 (cal/cm3)1/2,得到的木质素组分分子量由493 g/mol增加到2468 g/mol,其分散度也相应地由1.0增加到1.5。经1H和13CNMR分析,木质素结构单元间主要的β-O-4醚键在大分子组分中被包裹而得到保护,相对于小分子组分其含量较高。基于以上内容,本文首次对胡枝子蒸汽爆破预处理过程进行了系统的研究,为胡枝子用于第二代燃料乙醇的生产进行了探索性的研究工作。本研究提出了蒸汽爆破预处理及温和碱性溶剂后处理相结合的连续分离技术,实现了木质生物质主要组分的清洁分离,为木质纤维原料全组分、高质化利用提供了基础理论依据；利用纤维素溶解方法破坏纤维素结晶结构,突破了酶解转化过程中关键障碍;采用有机溶剂对造纸黑液回收的粗木质素进行了精制,为其高值化利用提供了合理的技术途径。更多还原

【Abstract】 In this study, the pretreatment process for second-generation bioethanol production from lignocellulosic materials was mainly discussed. The research focused on the optimal conditions of steam explosion pretreatment, main components and high-value utilization of lignocellulosic materials, mechanism and effect of cellulose-dissolving pretreatment, and recycle and fractionation of crude lignin for further application. Lespedeza crytobotrya, as an excellent source for bioethanol production, was employed for bioconversion to produce sugars. As an environmentally friendly pretreatment method, the efficiency of steam explosion, mainly steam pressure and incubation time, was successfully reflected by the physicochemical characteristics, microscopic structure and following enzymatic hydrolysis. A new fractionation process was proposed, based on the synergistic effect of steam-explosion pretreatment and alkaline solution post-treatment. The structural characteristics of the main fractions were completely investigated, aiming to obtain some evidence for the further high-value application. The structure-activity relationship among cellulose solvent, crystal structure and enzymatic conversion was established. Besides, organic solvents were used to fractionate the crude Kraft-AQ lignin. We expected to get more results and data for the utilization of lignocellulosic materials, and provide some theoretic basis for its production in industrial scale. The results of this study were summarized as follows:1. The main components of Lespedeza crytobotrya stalks are cellulose 44.6%, hemicelluloses 29.3% and lignin 17.0%. Steam explosion significantly decreased the size of the materials, loosed the compact structure of lignocellulosic material, and degraded the components in the cell wall. Under the given conditions (1.5-2.5 MPa steam pressure, 2～10 min incubation time), the content of cellulose kept the same level (43.6%-47.1%) regardless of the severity of steam explosion, and the relative crystallinity of the pretreated samples were slightly increased from 10.55% to 16.30%, probably due to the removal of hemicelluloses, degradation of the amorphous cellulose and rearrangement of amorphous and para-crystalline cellulose to crystalline form. Hemicelluloses were significantly degraded during the pretreatment process and reached lowest content of 3.7%; however, the content of lignin was correspondingly increased to 17.7%-23.3%.2. Steam explosion is an efficient pretreatment method, which significantly increased the accessibility of cellulose and further increased the enzymatic hydrolysis efficiency. Based on the optimal enzymatic condition derived from the single-factor and orthogonal experiment, the average enzymatic conversion rates of the steam-pretreated sample at 2.25 MPa were 78%,93% and 97%, respectively, corresponding to the commercial cellulase products cellulaseⅠ,ⅡandⅢ,2.8 times higher than the raw materials. For the pretreated samples incubated for 4 min, elevating steam pressure obviously increased the enzymatic hydrolysis. At 1.5 MPa, the enzymatic conversion was slightly increased by about 20%. In comparison, reducing sugars analysis demonstrated a factor of two improvements in the bioconversion rate after steam exploded at 1.75 MPa for 4 min.3. A two-step process based on steam explosion pretreatment followed by alkaline ethanol solution post-treatment was used to fractionate Lespedeza cyrtobotrya stalks. The residues were mainl consisted of cellulosic components and the highest content of glucose was up to 94.3%. The significant effect of defibrillation of microfibers and isolation of non-cellulosic components led to the increasing reactive area. The degree of polymerization was obviously decreased as the steam pressure was higher than 2.0 MPa and incubation time was longer than 4 min, and reached the lowest level of 207. Both were benefit for the following enzymatic bioconversion process. Hemicelluloses, as the most thermally instable component, were extensively degraded during steam explosion process, evidenced by the average weight molecular weights decreased from 47960,63300 and 102400 g/mol to 6380,8100 and 16200 g/mol, respectively. The comprehensive debranching reaction cleaved the linkages between the mainchain of xylan and arabinose, lead to the obvious decrease of Ara/Xyl ratio from 0.08 to 0. At 2.5 MPa for 4 min,86.6% glucose was determined in the isolated hemicellulosic fraction, indicating the partial degradation of cellulose and co-precipitation with non-cellulosic polysaccharides. Due to the remarkable selectivity with respect to lignin, up to 73.4% and 93.7% lignin component was extracted from 1M NaOH solution and 60% ethanol solution containing 1% NaOH post-treatment, respectively. The significant cleave of the linkages between carbohydrate and lignin induced the extremely lower content of sugar (0.80%) in the lignin fraction. Based on the quantitative analysis of 1H NMR spectrum, the content of/β-O-4 ether bonds were decreased from 11% to 7% as prolonging the incubation time from 2 min to 10 min at 2.25 MPa. The depolymerization reactions were accompanied with the comprehensive repolymerization reactions during the steam explosion process, leading to the formation of new carbon-carbon bonds, which were chemically and thermally stable. Due to the steric hindrance of methoxyl group at C5 position in syringyl units, more guaiacyl units were participated in the condensation reactions, and S/G ratio was correspondingly increased. The results indicated the existence of the restricting interrelation between efficient fractionation of cell wall components and maintainance of structural characteristics of hemicelluloses and lignin. Taking Lespedeza stalks as the starting material, steam explosion pretreatment at middle or lower severity and alkaline solution post-treatment was proposed as a new environmentally friendly fractionation process, which is benefit for the further high-value utilization of the main components.4. Cellulose dissolving and following enzymatic hydrolysis aims to establish an efficient pretreatment process using cellulose-dissolution solvents to enhance the enzymatic saccharifi cation. LiOH/Urea, concentrated phosphoric acid, N-methyl-morpholine-N-oxide (NMMO) and ionic liquid (1-butyl-3-methylimidazolium chloride [BMIM]C1) were selected as the efficient agents, and the regenerated samples exhibited the significant crystal transformation from cellulose I to cellulose II or amorphous region. Consequently, the relative crystallinity and degree of polymerization (DP) were reduced to the lowest level of 8% and 60, respectively. Comparing to the untreated sample (33%), obvious enhancement on the bioconversion efficiency from dissolving process was observed not only on glucose yield, also hydrolysis rate. After 72 h enzymatic hydrolysis, the conversion of cellulose reached to 81%,92%,83% and 80%, respectively. Meanwhile, the recovery of xylose was also increased from 18.5% to 39.4%-49.6%.5. Kraft-AQ pulping lignin was sequentially fractionated by organic solvent extractions (hexane, diethylether, methylene chloride, methanol and dioxane) and the molecular properties of each fraction were characterized in detail. The average molecular weight and polydispersity of each lignin fraction increased with its hydrogen-bonding capacity (Hildebrand solubility parameter), from 493 g/mol to 2468 g/mol, and to 13651 g/mol. Based on 1H and 13C NMR spectra,β-0-4 ether bonds, the main subunit in lignin macromolecule, were relatively packed and protected, and rich in the fractions with higher molecular weight.更多还原