一株木质素降解细菌的筛选及其降解途径

doi:10.16085/j.issn.1000-6613.2020-1541

摘要/Abstract

摘要：

经高温碱性盐法预处理的玉米秸秆预处理液中木质素和碱性盐含量高，为了反复利用含盐预处理液，需脱除其中的可溶性木质素。本实验从腐木中筛选分离到一株木质素降解菌株CCZU-WJ6，通过形态学观察和16S rDNA序列分析，鉴定该菌株属于无色杆菌属（Achromobacter sp.）。测定了CCZU-WJ6木质素降解相关酶的活性，及其在玉米秸秆预处理液中的生长趋势、预处理液的COD去除率和木质素降解率，通过GC-MS和FTIR检测预处理液中组分及其化学键的变化。通过单因素实验确定了菌株CCZU-WJ6处理预处理液的最佳条件为pH 7.0、温度30℃、预处理液稀释倍数10倍、接菌量0.5g/mL。在处理6天后，预处理液COD脱除率为40.9%，木质素的降解率为32.1%，CCZU-WJ6分泌木质素过氧化物酶和锰过氧化物酶的酶活分别达到215.5U/L、200.1U/L。CCZU-WJ6会破坏木质素结构中的苯环结构、醚键以及C＝O键，降解产物中含有愈创木酚和对香豆酸，推断其降解途径为β-芳基醚代谢途径和阿魏酸代谢途径。CCZU-WJ6可用来处理含木质素的工业废水，在工业方面具有广阔的应用前景。

关键词: 木质素, 降解途径, 木质素过氧化物酶, 锰过氧化物酶

Abstract:

The content of lignin and alkaline salt in the pretreatment solution of corn straw obtained by high temperature alkaline salt treatment was high. In order to reuse the pretreatment solution containing salt, soluble lignin should be removed. In this experiment, a lignin-degrading strain CCZU-WJ6 was screened and isolated from rotten wood and was identified as Achromobacter sp. by morphological observation and 16S rDNA sequence analysis. The activities of enzymes related to lignin degradation in CCZU-WJ6, the growth trend of CCZU-WJ6 in corn straw pretreatment solution, COD removal rate and lignin degradation rate of the pretreatment solution were measured. GC-MS and FTIR were used to detect the changes of components and chemical bonds in the pretreatment solution. Through single-factor experiment, the optimal conditions for degradation of the pretreatment solution by CCZU-WJ6 were determined as pH 7.0, temperature 30℃, dilution factor 10 times, and inoculation amount 0.5g/mL. After 6 days of treatment, the removal rate of COD and the degradation rate of lignin were 40.9% and 32.1%, respectively. The activities of lignin peroxidase and manganese peroxidase secreted by CCZU-WJ6 were 215.5U/L and 200.1U/L, respectively. CCZU-WJ6 can destroy the benzene ring structure, ether bond and C＝O bond in the lignin structure, and the degradation products contained guaiacol and p-coumaric acid, which inferred that the degradation pathways are β-aryl ether metabolic pathway and ferulic acid metabolic pathway. CCZU-WJ6 can be used to degrade industrial wastewater containing lignin and has broad prospects in industrial applications.

Key words: lignin, degradation pathway, lignin peroxidase, manganese peroxidase

中图分类号:

X703

王晶, 倪金荧, 王利群, 卿青, 严生虎, 张跃. 一株木质素降解细菌的筛选及其降解途径[J]. 化工进展, 2021, 40(7): 4021-4026.

WANG Jing, NI Jinying, WANG Liqun, QING Qing, YAN Shenghu, ZHANG Yue. Screening of a lignin degrading bacterium and its degradation pathway[J]. Chemical Industry and Engineering Progress, 2021, 40(7): 4021-4026.

图/表 8

图1 菌株CCZU-WJ6的系统进化树

图2 不同条件对菌株CCZU-WJ6降解性能的影响

图3 CCZU-WJ6在木质素培养液中的酶活

图4 CCZU-WJ6在预处理液中的生长曲线和COD降解曲线

图5 预处理液全波段扫描图

表1 GC-MS产物分析

编号	保留时间/min	名称	结构式	降解时间/d
编号	保留时间/min	名称	结构式	0	3	6
A1	11.589	三苯基磷	C₁₈H₁₆P	+	-	-
A2	13.684	三苯基磷胺	C₁₈H₁₆NP	+	-	-
A3	14.527	三苯基氧化磷	C₁₈H₁₅OP	+	-	-
B1	9.008	愈创木酚	C₇H₈O₂	-	+	+
B2	10.082	对香豆酸	C₉H₈O₃	-	+	+

图6 预处理液红外图谱

图7 预处理液中木质素的降解途径推断B1—愈创木酚；B2—对香豆酸

参考文献 13

1	LOPES A M C, GOAO K G, RUBIK D F, et al. Pre-treatment of lignocellulosic biomass using ionic liquids: wheat straw fractionation [J]. Bioresource Technology, 2013, 142: 198-208.
2	GREGOROVA A, CIBULKOVA Z, KOSIKOVA B, et al. Stabilization effect of lignin in polypropylene and recycled polypropylene [J]. Polymer Degradation and Stability, 2005, 89(3): 553-558.
3	CARDOSO M, OLIVEIRA E D D, PASSOS M L. Chemical composition and physical properties of black liquors and their effects on liquor recovery operation in Brazilian pulp mills [J]. Fuel, 2009, 88 (4): 756-763.
4	FONSECA M I, FARINA J I, CASTRILLO M L, et al. Biopulping of wood chips with Phlebia brevispora BAFC 633 reduces lignin content and improves pulp quality [J]. International Biodeterioration & Biodegradation, 2014, 90: 29-35.
5	CHEN Yuehui, CHAI Liyuan, TANG Chongjian, et al. Kraft lignin biodegradation by Novosphingobium sp. B-7 and analysis of the degradation process [J]. Bioresource Technology, 2012, 123: 682-685.
6	JOY S, RAHMAN P K S M, SHARMA S. Biosurfactant production and concomitant hydrocarbon degration potentials of bacteria isolated from extreme and hydrocarbon contaminated environments [J]. Chemical Engineering Journal, 2017, 317: 232-241.
7	TABASSUM S, JI Qinhong, HENA S, et al. Treatment of coal gasification wastewater by anaerobic SBR-aerobic SBR process for elimination of toxic organic matters—A lab scale study [J]. RSC Advances, 2015, 5(72): 58334-58344.
8	LARA M A, RODRI GUEZ-MALAVER A J, ROJAS O J, et al. Black liquor lignin biodegradation by Trametes elegans [J]. International Biodeterioration & Biodegradation, 2003, 52(3): 167-173.
9	苏玉龙. 高效降解烟杆菌株筛选及其特性研究[D]. 北京: 中国农业科学院, 2016.
	SU Yulong. Screening and characteristics of high-efficiency degrading tobacco stem strains [D]. Beijing: Chinese Academy of Agricultural Sciences, 2016.
10	CHEN Y H, CHAI L Y, ZHU Y H, et al. Biodegradation of kraft lignin by a bacterial strain Comamonas sp. B-9 isolated from eroded bamboo slips [J]. Journal of Applied Microbiology, 2012, 112 (5): 900-906.
11	WANG W, YANG S, HUNSINGER G B, et al. Connecting lignin-degradation pathway with pre-treatment inhibitor sensitivity of Cupriavidus necator [J]. Front Microbiol., 2014, 5: 247.
12	CHEN Yuehui, CHAI Liyuan, TANG Chongjian, et al. Depolymerization and decolorization of kraft lignin by bacterium Comamonas sp. B-9 [J]. Applied Microbiology and Biotechnology, 2014, 98(4): 1907-1912.
13	ZHU Daochen, ZHANG Peipei, XIE Changxiao, et al. Biodegradation of alkaline lignin by Bacillus ligniniphilus L1 [J]. Biotechnology for Biofuels, 2017, 10(1): 44.

[1]	戴欢涛, 曹苓玉, 游新秀, 徐浩亮, 汪涛, 项玮, 张学杨. 木质素浸渍柚子皮生物炭吸附CO₂特性[J]. 化工进展, 2023, 42(S1): 356-363.
[2]	关红玲, 杨辉, 井红权, 刘玉琼, 谷守玉, 王好斌, 侯翠红. 木质素基控释材料及其在药物输送和肥料控释中的应用[J]. 化工进展, 2023, 42(7): 3695-3707.
[3]	于丁一, 李圆圆, 王晨钰, 纪永升. pH响应性木质素水凝胶的制备及药物控释[J]. 化工进展, 2023, 42(6): 3138-3146.
[4]	任建鹏, 吴彩文, 刘慧君, 吴文娟. 木质素-聚苯胺复合材料的制备及对刚果红的吸附[J]. 化工进展, 2023, 42(6): 3087-3096.
[5]	杨程瑞雪, 黄琪媛, 冉建速, 崔耘通, 王健健. 磷酸修饰二氧化硅负载钯催化剂用于木质素衍生物高效水相低温加氢脱氧[J]. 化工进展, 2023, 42(10): 5179-5190.
[6]	张鹏, 王绍庆, 李志合, 张安东, 高亮, 万震, 宋宁. 赤泥/木质素共热解制备复合吸附材料及其性能[J]. 化工进展, 2022, 41(S1): 407-414.
[7]	蒲福龙, 伍尚炜, 郑映玲, 郑玉意, 侯雪丹. 基于乳酸的深度共熔溶剂提取秸秆木质素对纤维素酶水解效率的影响[J]. 化工进展, 2022, 41(9): 4937-4945.
[8]	龙垠荧, 杨健, 管敏, 杨怡洛, 程正柏, 曹海兵, 刘洪斌, 安兴业. 木质素基材料在混合型超级电容器电极材料中的研究进展[J]. 化工进展, 2022, 41(9): 4855-4865.
[9]	汪兴, 赵子龙, 张小山, 王宏杰, 董文艺, 陈慧慧. 制备条件对生物炭载铁催化剂催化破络Ni-EDTA性能及活性组分浸出的影响[J]. 化工进展, 2022, 41(9): 4831-4839.
[10]	张伟, 安兴业, 刘利琴, 龙垠荧, 张昊, 程正柏, 曹海兵, 刘洪斌. 木质素纳米颗粒/天然纤维基活性碳纤维材料的制备及其电化学性能[J]. 化工进展, 2022, 41(7): 3770-3783.
[11]	张丽珠, 王欢, 李琼, 杨东杰. 木质素衍生吸附材料及其在废水处理中的应用研究进展[J]. 化工进展, 2022, 41(7): 3731-3744.
[12]	娄瑞, 刘钰, 田杰, 张亚男. 纳米木质素基多孔炭的制备及其电化学性能[J]. 化工进展, 2022, 41(6): 3170-3177.
[13]	申琪, 薛雨源, 杨涛伟, 张妍, 李胜任. 木质素荧光研究进展[J]. 化工进展, 2022, 41(5): 2672-2685.
[14]	王鲁元, 金春江, 陈惠敏, 程星星, 安东海, 张兴宇, 孙荣峰, 耿文广. 一步热解活化法制备纳米木质素基多孔炭材料[J]. 化工进展, 2022, 41(5): 2582-2592.
[15]	张雷, 王海英, 韩洪晶, 陈彦广, 王程昊. 木质素催化热解用催化剂的研究进展[J]. 化工进展, 2022, 41(5): 2429-2440.