木质素结构单元对纤维素酶吸附的影响

doi:10.11707/j.1001-7488.LYKX20210766

摘要/Abstract

摘要：

目的: 以稻草、芦草和竹子中不同结构单元木质素为研究对象，借助耗散型石英晶体微天平（QCM-D）技术，原位、实时探究不同结构单元分离木质素与酶吸附的动态行为，为木质素对生物质酶水解的影响提供理论基础、对木质纤维素的高效利用进行定向调控。方法: 原料化学成分参照标准方法测定，分离木质素结构经红外、凝胶渗透色谱和碱性硝基苯氧化表征，采用原子力显微镜观察木质素薄膜表面形貌，借助QCM-D技术分析纤维素酶在木质素薄膜上的吸附行为。结果: 分离木质素均为GSH型木质素，纯度高，含糖量约10%，分子质量接近，约5 000；硝基苯氧化结果显示，稻草、芦草和竹子中木质素结构单元存在差异，竹子中木质素未缩合单元得率最高，为444.4 g·kg^?1，S/G比为1∶0.6，芦草、稻草中木质素S/G比均为1∶1.1；经QCM-D分析，竹子木质素薄膜对纤维素酶吸附速率最快、吸附量最大，芦草和稻草木质素薄膜对纤维素酶的吸附速率和最大吸附量相近；停止通酶后，用缓冲液冲洗，纤维素酶附着在木质素上难以被洗脱。酶吸附过程中3种木质素表面均具有较好的黏弹性，且黏弹性差异不大。结论: 稻草、芦草和竹子中木质素对纤维素酶均有明显吸附作用，来源不同的木质素但结构单元相同，对纤维素酶吸附能力相同；木质素结构单元不同，对纤维素酶吸附能力不同。纤维素酶在木质素薄膜表面上结合紧密，难以脱附，S/G比较高的木质素，其纤维素酶吸附能力较强，在纤维素酶水解中产生更多无效吸附，导致葡聚糖转化效率降低。

关键词: 木质素, 结构单元, 酶吸附, 纤维素酶水解, 石英晶体微天平

Abstract:

Objective: Using lignin with different structural units isolated from straw, reed and bamboo as the research material, the dynamic behavior between isolated lignin and enzyme adsorption was studied based on quartz crystal microbalance with dissipation (QCM-D) in situ and in real-time. The results obtained could provide targeted regulation for the efficient utilization of lignocellulose. Method: The chemical composition of raw materials was determined according to the standard method, the structural units of isolated lignin were characterized by infrared, alkaline nitrobenzene oxidation and gel permeability chromatography. The surface morphology of lignin film was detected by using an atomic force microscope, and the adsorption behavior of cellulase on lignin film was studied by QCM-D. Result: The isolated lignin was GSH type with high purity, the sugar content was about 10%, and their molecular weight was close, about 5 000. The nitrobenzene oxidation results indicated that structural units of isolated lignins were different, uncondensed units yield of bamboo lignin was highest, 444.4 g·kg^?1, and its S/G mole ratio was 1∶0.6. The lignin S/G ratio in reed was 1∶1.1, as well as rice straw. Bamboo lignin film showed the fastest adsorption rate of cellulase, and its adsorption amount was also the maximum via QCM-D analysis. The adsorption rate and maximum adsorption amount of cellulase in reed and straw lignin films were similar. After stopped the cellulase injection, the lignin film was rinsed with buffer solution while the cellulase attached to lignin was difficult to be eluted. All three lignin surfaces during cellulase adsorption had good viscoelasticity with little difference. Conclusion: Lignin present an obvious adsorption effect on cellulase, lignin with the same structural units, even from different sources, showed the same adsorption capacity of lignin to cellulase. Different structural units of lignin have different adsorption capacities for cellulase. Cellulase was hard to be removed because of its binding closely on the surface of lignin film. The lignin with higher S/G ratio showed more strong adsorption capacity to cellulase, which produced more ineffective adsorption in cellulase hydrolysis, and led to reduction of glucan conversion efficiency in enzymatic hydrolysis.

Key words: lignin, structural unit, enzyme adsorption, cellulose enzyme hydrolysis, QCM-D (quartz crystal microbalance with dissipation)

中图分类号:

S781.4

邹春阳,吴文娟. 木质素结构单元对纤维素酶吸附的影响[J]. 林业科学, 2023, 59(6): 141-148.

Chunyang Zou,Wenjuan Wu. Effect of Lignin Structural Unit on Cellulase Adsorption[J]. Scientia Silvae Sinicae, 2023, 59(6): 141-148.

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木质素 Lignin	葡聚糖 Glucan/ (g·kg^?1)	木聚糖 Xylan/ (g·kg^?1)	其他 Others/ (g·kg^?1)	总糖 Sum/ (g·kg^?1)
芦草 Phragmites australis	25.1±0.1	41.6±0.0	29.3±0.2	96.0±0.3
稻草 Oryza sativa	27.8±0.1	48.6±0.1	26.7±0.1	103.1±0.3
竹子 Bambusoideae	36.8±0.0	66.3±0.1	7.0±0.1	110.1±0.2

木质素 Lignin	数均分子质量 Number-average molecular weight (M_n)	重均分子质量 Mass average molar mass (M_w)	PDI (M_w / M_n)
芦草 Phragmites australis	5 456	10 418	1.91
稻草 Oryza sativa	6 118	12 265	2.00
竹子 Bambusoideae	5 043	10 831	2.15

木质素 Lignin	结构单元比例Molar ratio of structural units			结构单元得率 Yield of structural unit / (g·kg^?1)			总得率 Total yield/ (g·kg^?1)
木质素 Lignin	S	G	H	S	G	H	总得率 Total yield/ (g·kg^?1)
芦草 Phragmites australis	1	1.1	0.7	180.9±1.1	158.2±1.2	81.1±1.1	420.2±1.1
稻草 Oryza sativa	1	1.1	0.6	138.8±1.2	124.3±1.2	53.1±1.3	316.2±1.1
竹子 Bambusoideae	1	0.6	0.4	248.2±1.2	129.9±1.3	67.3±1.3	444.4±1.2

木质素 Lignin	葡聚糖 Glucan(%)	木聚糖 Xylan (%)	总糖 Sum (%)
芦草 Phragmites australis	86.6	62.1	75.1
稻草 Oryza sativa	81.0	69.3	66.1
竹子 Bambusoideae	71.5	66.2	68.9

木质素 Lignin	粗糙度 Roughness/nm
芦草 Phragmites australis	1.67±0.09
稻草 Oryza sativa	1.76±0.07
竹子 Bambusoideae	2.16±0.06