【作者】 毛健贞；

【导师】 许凤；

【作者基本信息】 北京林业大学 ， 林产化学加工工程， 2014， 博士

【摘要】 本文选择在我国北方地区广泛分布的多年生草本植物乌拉草为研究对象,经过连续的碱抽提出碱木质素、半纤维素、酶酸解木质素,结合湿法化学和现代仪器分析手段,系统表征了乌拉草木质素、半纤维素、酶酸解木质素的化学结构特征,并比较了白腐菌C7784预处理和稀酸预处理后对乌拉草木质素结构的影响。对纤维素酶解残渣进行了0.1M盐酸的90%二氧六环-水溶液抽提和2%氢氧化钠抽提,提取液经三倍乙醇沉淀得到了两个半纤维素组分H1和H2,进一步浓缩在酸水中沉淀得到两个木质素组分L1和L2。分离得到的木质素达到原木木质素29.5%和13.5%,木质素样品中的总糖浓度分别为糖的总浓度达到11.20mg L-1, L2中残留糖的总浓度仅有0.36mg L-1。碱法提取的溶液经乙醇沉淀除去的组分占原半纤维素12.2%,而酸性二氧六环提取的溶液提取的溶液除去的组分仅占原半纤维素4.2%。总的来说,通过乙醇沉淀半纤维素以达到纯化木质素产品的目的只对碱法提取的过程行之有效。酶酸解木质素与碱木质素都是典型的GSH型木质素,主要的连接方式为β-O-4’连接,其次是β-5’连接,再次是β-β’的连接。酶酸解L]中阿魏酸和对香豆酸均以酯键与其相连,碱木质素木质素L2中只有对香豆酸以酯键与其相连,并且黄酮的信号消失。采用了七步连续抽提对乌拉草原料中的木质素进行分离,条件依次为0.25%NaOH-70%乙醇溶液、0.5%、1%、1.5%和2%NaOH溶液以固液比1：25(g mL-1)在80℃水浴加热3h,得到的木质素质量占原本克拉森木质素质量的4.4%,6.9%,10.9%,13.2%,7.9%和4.9%,得到的木质素样品总量占原本克拉森木质素质量的48.2%。随着使用的碱浓度的升高,得到的木质素样品的分子量降低(从4590降到2120g mol-1),纯度升高(中性糖和糖醛酸总量从8.52%降到3.46%)。从红外谱图可以看出,6个木质素样品具有相似的化学结构,,以木质素样品L2和L3的核磁共振谱图解析推断木质素结构得出：乌拉草碱溶性木质素样品为典型的GSH型木质素,主要的连接键类型为β-O-4芳基醚键连接,此外还含有少量的β-β,β-5’的连接键类型。采用了七步连续抽提对乌拉草原料中的半纤维素进行分离,条件依次为0.25%NaOH-70%乙醇溶液、0.5%、1%、1.5%、2%和3%NaOH溶液以固液比1：25(gmL-1)在80℃水浴加热3h,得到的半纤维素质量分别占原料的初始质量的1.8%,5.2%,5.9%,4.1%,4.9%,4.0%和4.4%,占原本半纤维素质量的5.8%,16.7%,18.9%,13.1%,15.7%,12.8%和14.1%,得到的半纤维素样品总量占原木半纤维素质量的97.1%。水溶性的半纤维素平均分子量低(16690g mol-1)分支度高(木糖相对含量5.6%),碱溶性的半纤维素平均分子量高分支度低,并且随着碱浓度的升高,所得半纤维素样品的平均分子量升高(34620-106210g mol-1),分支度降低(木糖相对含量48.6-71.2%)。糖分析结果看出,碱溶性半纤维素是以木糖为骨架,支链为阿拉伯糖(11.9-18.1%)和糖醛酸(8.1-10.5%),而水溶性半纤维素则主要以葡萄糖为骨架(相对含量43.5%),半乳糖(相对含量23.9%)和阿拉伯糖(相对含量10.3%)为分支。从红外分析结果可以看出,七个半纤维素样品具有相似的结构,对半纤维素样品的H3的核磁共振谱图分析可以得到半纤维素样品结构的示意图：(1→4)[4-O-甲基-α-葡萄糖醛酸-(1→2)][α-L-阿拉伯呋喃-(1→3)]-β-D-木聚糖白腐菌T. hirsute C7784不仅降解乌拉草中的木质素,也降解其中的纤维素和半纤维素,而且菌处理三周时,半纤维素降解最多其次是纤维素,木质素未发生降解,降解六周后,木质素才发生降解。从分子量的数据可以看出,菌处理三周时,木质素样品中低分子量的部分得到了降解,六周时大分子量的木质素得到了降解。从核磁结果看来,乌拉草木质素中主要的连接键类型是β-O-4醚键,其次是苯基香豆满和少量的麦黄酮在菌处理前后未发生减少。菌处理6周后,一部分碳-碳键和少量的甲氧基,像β-β’和β-5’明显地发生了降解。对于木质素的结构单元来说,愈创木单元比紫丁香单元更容易被白腐菌T. hirsute C7784降解。更多还原

【Abstract】 In this thesis, lignin, hemicelluloses, and enzymatic mild acidolysis lignin (EMAL) were isolated from Carex meyeriana Kunth by successive treatments with incremental concentration alkaline solvents. The structural characteristics of the isolated hemicelluloses, lignin, and EMAL were investigated., and the lignin structure changes were comparatively studied after white-rot fungus treatment and dilute acid treatment.The enzymatic hydrolysis residue were successively extracted with0.1M HCI and2%NaOH, the filtration was adjusted pH5.5, concentration and the hemicellulosic fractions Hiand H2were precipitated in3-time volue ethanol. The filtration was further adjusted pH to2to obtain lignin fractions L1and L2, accounting to the29.5%and13.5%of Klason lignin. The sugar and uronic acid concentration in lignin fractions were11.20and1.14mg L-1. The precipitated hemicellulosic fractions were12.2and4.2%of the origin hemicelluloses with alkali and acidified dioxane extraction. The EMAL and alkali lignin were both typical GSH linin, the linkage between lignin units were β-O-4’, β-5,β-β’. The ester bond signals of lignin with ferulic acid and p-courmaric acid signals presented in HSQC of L1, while the ester bond signal between lignin and ferulic acid was disappeared in HSQC of L2.Successive extractions with distilled water,0.25%NaOH-95%ethanol solution,0.5%,1%,1.5%and2%NaOH aqueous solutions at a solid-liquid ratio of1:25(g mL"1) at80℃for3h from Carex meyeriana Kunth released4.4%,6.9%,10.9%,13.2%,7.9%and4.9%of the original lignin, respectively. The physico-chemical properties and structural features of these lignin fractions were comprehensively characterized by HPLC, GPC, NMR spectroscopy, and TGA. The alkali concentration increment had a positive influence on the purity of the lignin fractions, but a negative influence on the average molecular weight and thermal stability. The FT-IR spectra showed that the lignin fractions had similar structural features with those of GSH type lignin. The 1H and13C and2D NMR spectra illustrated that typical lignin fractions had predominantly β-O-4aryl ether linkages followed by β-β and β-5’ones.Successive extractions of de-waxed Carex meyeriana Kunth with distilled water,0.25%NaOH-70%ethanol solution,0.5%,1%,1.5%,2%and3%NaOH aqueous solutions at a solid-liquid ratio of1:25(g mL"1at80℃for3h released5.8%,16.7%,18.9%,13.1%,15.7%,12.8%and14.1%of the original hemicelluloses, respectively. The GPC analysis showed that the weight-average molecular weight (Mw) of alkali-soluble hemicelluloses was obviously higher than that of water-soluble hemicelluloses (16690g mol-1) and with the increasing of alkali concentration the Mw increased from34620to106210g mol-1. Comparison of these hemicellulosic indicated that more branched and smaller molecular size of hemicelluloses fractions (xylose5.6%) were more easily extracted by the hot water treatment. While the alkali-soluble hemicellulosic fractions contained less branched and larger molecular size of the polymers, in which xylose (48.6-71.2%) was found to be the major sugar component together with small amounts of arabinose (11.9-18.1%), uronic acids (8.1-10.5%), galactose (4.5-13.1%), glucose (2.7-9.2%), and a trace amount of rhamnose (0.7-1.5%). The results obtained by the analysis of FT-IR, sugar composition,1H and13C NMR indicated that there were no considerable differences in the structural features of the water-and alkali-soluble hemicellulosic fractions, which were mainly comprised of (1→4)[4-O-Me-β-D-GlcpA-(1→2)][a-L-Araf-(1→3)]--β-D-Xylp units.Carex meyeriana Kunth was subjected to biological pretreatment with white-rot fungus F. palustris C7784, and the structural changes of the lignin were investigated. Results showed that there was a slight decrease in carbohydrate content after pretreatment for3weeks, but a noticeable decrease in lignin and carbohydrate contents for6weeks. Detail structural analysis of the isolated lignin from the samples revealed that Carex meyeriana Kunth lignin consisted mainly of p-hydroxyphenyl (H), guaiacyl (G), and syringyl (S) units with minor p-coumarate and ferulate units. The predominate lignin interunits were β-O-4’ ether linkages, followed by phenylcoumaran, together with a lower proportion of resinol and tricin substructures. After pretreatment for6weeks, the contents of β-β’and β-5’ linkages notably decreased, suggesting that the fungus preferentially attacked these linkages. The pretreatment led to an increase of S/G ratio from0.64in the control to0.83in the sample pretreated for6weeks. The comprehensive understanding of the structural changes of lignin offers new insights into the biological degradation of Carex meyeriana Kunth by white-rot fungus.更多还原