Isolation and study of the degradation mechanism of hydroxyl-terminated polybutadiene-degrading strain

doi:10.11949/0438-1157.20250053

Abstract

Abstract:

Hydroxyl-terminated polybutadiene (HTPB) is an important component of butyl hydroxy propellant. The disposal of waste butyl hydroxy propellant, resulting from the retirement and upgrading of weapons, poses notable environmental challenges. In this study, a Pseudomonas sp. HM6 was isolated from refinery sludge, which used HTPB as the sole carbon source. The molecular mass distribution of HTPB before and after degradation was determined by using gel permeation chromatography. The degradation products were characterized by using Fourier transform infrared spectroscopy, proton nuclear magnetic resonance spectroscopy, and gas chromatography-mass spectrometry. After degradation, the degradation products exhibit two distinct molecular mass,which decreased from an initial value of 9523 g/mol to 6809 g/mol and 366 g/mol. The degradation products contained aldehydes, alcohols, and carboxylic acid compounds. Through genomic and transcriptional analysis, genes encoding copper oxidase, aldehyde dehydrogenase, hydrolase and other genes were found, and their expression levels were upregulated. Based on chemical characterization and genomic analysis, a degradation pathway of HTPB by strain HM6 was proposed. This study provides a theoretical basis for the use of microorganisms to degrade waste butyl hydroxyl propellant.

Key words: Pseudomonas, hydroxyl-terminated polybutadiene, degradation, biotechnology, polymers

摘要：

端羟基聚丁二烯（HTPB）是丁羟推进剂的重要组分，随着武器弹药的退役升级，废弃丁羟推进剂的处理对环境带来了重大的挑战。本研究从炼厂污泥中分离出一株以HTPB为唯一碳源的假单胞菌HM6。采用凝胶渗透色谱法测定了降解前后HTPB的分子量分布，采用傅里叶变换红外光谱、核磁共振氢谱和气相色谱-质谱联用法对降解产物进行了表征。HTPB降解后产生两种低分子量化合物，数均分子量由降解前的9523 g/mol分别降低至6809 g/mol和366 g/mol，中间代谢产物包括醛类、醇类和羧酸类等化合物。通过基因组和转录分析，发现编码铜氧化酶、醛脱氢酶、水解酶等基因且表达量均上调。基于化学表征和基因组分析，提出了菌株HM6对HTPB降解的途径。本研究为利用微生物降解废弃丁羟推进剂提供了理论依据。

关键词: 假单胞菌, 端羟基聚丁二烯, 降解, 生物技术, 聚合物

CLC Number:

TQ 319

Xiaochen ZHANG, Zhongshan LU, Teng GUO, Heng GUI, Hongbing SONG, Meng XIAO. Isolation and study of the degradation mechanism of hydroxyl-terminated polybutadiene-degrading strain[J]. CIESC Journal, 2025, 76(8): 4205-4216.

张晓晨, 鲁中山, 郭腾, 桂恒, 宋红兵, 肖盟. 一株端羟基聚丁二烯降解菌的筛选及降解机理研究[J]. 化工学报, 2025, 76(8): 4205-4216.

Figures/Tables 14

References 35

[1]	Kohga M. From cross-linking to plasticization–characterization of glycerin/HTPB blends[J]. Propellants, Explosives, Pyrotechnics, 2009, 34(5): 436-443.
[2]	Mohan Y M, Raju K M. Synthesis and characterization of HTPB-GAP cross-linked co-polymers[J]. Designed Monomers and Polymers, 2005, 8(2): 159-175.
[3]	Luo Y L, Miao Y, Xu F. Synthesis, phase behavior, and simulated in vitro degradation of novel HTPB-b-PEG polyurethane copolymers[J]. Macromolecular Research, 2011, 19(12): 1233-1241.
[4]	Medchill C, Perezselsky A A, Cortopassi A, et al. Transient plasma enhanced combustion of solid rocket propellants[J]. Aerospace Science and Technology, 2024, 148: 109107.
[5]	李立远, 张丽华, 王鹏, 等. 废弃丁羟推进剂的处理与再利用研究进展[J]. 河南化工, 2010, 27(24): 3-7.
	Li L Y, Zhang L H, Wang P, et al. Research progress on treatment and reuse of waste HTPB propellants[J]. Henan Chemical Industry, 2010, 27(24): 3-7.
[6]	鲁彦玲, 张力, 施冬梅, 等. 废弃发射药的再利用研究[J]. 环境科学与技术, 2006, 29(S1): 147-150.
	Lu Y L, Zhang L, Shi D M, et al. Study on the reusing obsolete propellant[J]. Environmental Science & Technology, 2006, 29(S1): 147-150.
[7]	Ganesh K, Sundarrajan S, Kishore K, et al. Primary pyrolysis products of hydroxy-terminated polybutadiene[J]. Macromolecules, 2000, 33(2): 326-330.
[8]	Williams P T. Pyrolysis of waste tyres: a review[J]. Waste Management, 2013, 33(8): 1714-1728.
[9]	仪建华, 赵凤起, 李上文, 等. 美俄废弃火箭发动机装药绿色销毁与回收技术研究进展[J]. 化学推进剂与高分子材料, 2006, 4(6): 25-27, 37.
	Yi J H, Zhao F Q, Li S W, et al. Research progress of green destruction and reclamation technology of obsolete rocket motor charge in America and Russia[J]. Chemical Propellants & Polymeric Materials, 2006, 4(6): 25-27, 37.
[10]	Zhang Y, Zou M, Lodhi A F, et al. Microbial degradation and community structure analysis of hydroxyl-terminated polybutadiene (HTPB)[J]. AMB Express, 2021, 11(1): 180.
[11]	Wu K, Liu Y C, Ma Y, et al. Pretreatment of hydroxy-terminated polybutadiene (HTPB)/toluene diisocyanate (TDI) binder system for biodegradation[J]. Advanced Composites and Hybrid Materials, 2021, 4(1): 96-103.
[12]	吴凯. 废弃丁羟推进剂黏合体系生物降解方法及机理[D]. 太原: 中北大学, 2021.
	Wu K. Biodegradation method and mechanism of binder system waste from HTPB propellant[D]. Taiyuan: North University of China, 2021.
[13]	Imai S, Ichikawa K, Muramatsu Y, et al. Isolation and characterization of Streptomyces, Actinoplanes, and Methylibium strains that are involved in degradation of natural rubber and synthetic poly(cis-1,4-isoprene)[J]. Enzyme and Microbial Technology, 2011, 49(6/7): 526-531.
[14]	Bröker D, Dietz D, Arenskötter M, et al. The genomes of the non-clearing-zone-forming and natural-rubber-degrading species Gordonia polyisoprenivorans and Gordonia westfalica harbor genes expressing Lcp activity in Streptomyces strains[J]. Applied and Environmental Microbiology, 2008, 74(8): 2288-2297.
[15]	Ibrahim E M A, Arenskötter M, Luftmann H, et al. Identification of poly(cis-1, 4-isoprene) degradation intermediates during growth of moderately thermophilic actinomycetes on rubber and cloning of a functional lcp homologue from Nocardia farcinica strain E1[J]. Applied and Environmental Microbiology, 2006, 72(5): 3375-3382.
[16]	Roy R V, Das M, Banerjee R, et al. Comparative studies on crosslinked and uncrosslinked natural rubber biodegradation by Pseudomonas sp.[J]. Bioresource Technology, 2006, 97(18): 2485-2488.
[17]	Yikmis M, Arenskötter M, Rose K, et al. Secretion and transcriptional regulation of the latex-clearing protein, Lcp, by the rubber-degrading bacterium Streptomyces sp. strain K30[J]. Applied and Environmental Microbiology, 2008, 74(17): 5373-5382.
[18]	Rose K, Tenberge K B, Steinbüchel A. Identification and characterization of genes from Streptomyces sp. strain K30 responsible for clear zone formation on natural rubber latex and poly(cis-1,4-isoprene) rubber degradation [J]. Biomacromolecules, 2005, 6(1): 180-188.
[19]	Nayanashree G, Thippeswamy B. Biodegradation of natural rubber by laccase and manganese peroxidase enzyme of Bacillus subtilis [J]. Environmental Processes, 2015, 2(4): 761-772.
[20]	Vivod R, Oetermann S, Hiessl S, et al. Oligo(cis-1,4-isoprene) aldehyde-oxidizing dehydrogenases of the rubber-degrading bacterium Gordonia polyisoprenivorans VH2[J]. Applied Microbiology and Biotechnology, 2017, 101(21): 7945-7960.
[21]	Song F, Li C, Zhang N, et al. A novel endophytic bacterial strain improves potato storage characteristics by degrading glycoalkaloids and regulating microbiota[J]. Postharvest Biology and Technology, 2023, 196: 112176.
[22]	Zhang N, Ding M Z, Yuan Y J. Current advances in biodegradation of polyolefins [J]. Microorganisms, 2022, 10(8): 1537.
[23]	Iiyoshi Y, Tsutsumi Y, Nishida T. Polyethylene degradation by lignin-degrading fungi and manganese peroxidase[J]. Journal of Wood Science, 1998, 44(3): 222-229.
[24]	Zhang Y, Pedersen J N, Eser B E, et al. Biodegradation of polyethylene and polystyrene: from microbial deterioration to enzyme discovery[J]. Biotechnology Advances, 2022, 60: 107991.
[25]	Cheng Y J, Wei Y C, Wu H L, et al. Biodegradation of vulcanized natural rubber by enriched bacterial consortia[J]. Chemical Engineering Journal, 2024, 481: 148685.
[26]	Ma Y X, Yan F, An L L, et al. Transcriptome analysis of changes in M. aeruginosa growth and microcystin production under low concentrations of ethinyl estradiol[J]. Science of the Total Environment, 2023, 859: 160226.
[27]	Wang F, Du W, Huang W X, et al. Linkages of volatile fatty acids and polyhexamethylene guanidine stress during sludge fermentation: metagenomic insights of microbial metabolic traits and adaptation[J]. Chinese Chemical Letters, 2023, 34(6): 107890.
[28]	Rozen R, Vockley J, Zhou L B, et al. Isolation and expression of a cDNA encoding the precursor for a novel member (ACADSB) of the acyl-CoA dehydrogenase gene family[J]. Genomics, 1994, 24(2): 280-287.
[29]	Manow R, Wang C, Garza E, et al. Expression of acetaldehyde dehydrogenase (aldB) improved ethanol production from xylose by the ethanologenic Escherichia coli RM10[J]. World Journal of Microbiology & Biotechnology, 2020, 36(4): 59.
[30]	Zhou S F, Ainala S K, Seol E, et al. Inducible gene expression system by 3-hydroxypropionic acid[J]. Biotechnology for Biofuels, 2015, 8: 169.
[31]	Harris D J, Assink R A, Celina M. NMR analysis of oxidatively aged HTPB/IPDI polyurethane rubber: degradation products, dynamics, and heterogeneity[J]. Macromolecules, 2001, 34(19): 6695-6700.
[32]	康莹, 陈智群, 陈曼, 等. 复合固体推进剂长贮中HTPB胶微结构损伤研究[J]. 固体火箭技术, 2015, 38(4): 519-522, 527.
	Kang Y, Chen Z Q, Chen M, et al. Microstructure damage of HTPB glue in the long-time storage of composite solid propellants[J]. Journal of Solid Rocket Technology, 2015, 38(4): 519-522, 527.
[33]	Lv G Z, Cen L, Tan X W, et al. Preparation of epoxidized acrylonitrile butadiene rubber and its compatibilization effect on water-swellable rubber composites[J]. Journal of Applied Polymer Science, 2019, 136(26): 47694.
[34]	Huerta Lwanga E, Thapa B, Yang X M, et al. Decay of low-density polyethylene by bacteria extracted from earthworm's guts: a potential for soil restoration[J]. Science of the Total Environment, 2018, 624: 753-757.
[35]	Cheng X T, Xia M L, Yang Y. Biodegradation of vulcanized rubber by a gut bacterium from plastic-eating mealworms[J]. Journal of Hazardous Materials, 2023, 448: 130940.

KEGG	基因编号	名称	基因描述	显著性	上/下调情况
K01692	gene2244	paaF	crotonase/enoyl-CoA hydratase family protein	yes	up
K00059	gene1731	fabG	MULTISPECIES: 3-oxoacyl-ACP reductase FabG	yes	up
K00059	gene2659	fabG	MULTISPECIES: 3-oxoacyl-ACP reductase FabG	yes	up
K00249	gene0772	acd	acyl-CoA dehydrogenase family protein	yes	up
	gene2751	bcd	acyl-CoA dehydrogenase	yes	down
K22552	gene1654		copper oxidase	yes	up
K00257	gene2983	acd	acyl-CoA/acyl-ACP dehydrogenase	yes	up
K00128	gene0641	aldB	MULTISPECIES: aldehyde dehydrogenase	yes	down
	gene0924		MULTISPECIES: aldehyde dehydrogenase family protein	yes	up
	gene2891	aldB	MULTISPECIES: aldehyde dehydrogenase	yes	down
	gene4094	mmsA	MULTISPECIES: CoA-acylating methylmalonate-semialdehyde dehydrogenase	yes	up

KEGG	基因编号	名称	基因描述	显著性	上/下调情况
K01692	gene2244	paaF	crotonase/enoyl-CoA hydratase family protein	yes	up
K00059	gene1731	fabG	MULTISPECIES: 3-oxoacyl-ACP reductase FabG	yes	up
K00059	gene2659	fabG	MULTISPECIES: 3-oxoacyl-ACP reductase FabG	yes	up
K00249	gene0772	acd	acyl-CoA dehydrogenase family protein	yes	up
	gene2751	bcd	acyl-CoA dehydrogenase	yes	down
K22552	gene1654		copper oxidase	yes	up
K00257	gene2983	acd	acyl-CoA/acyl-ACP dehydrogenase	yes	up
K00128	gene0641	aldB	MULTISPECIES: aldehyde dehydrogenase	yes	down
	gene0924		MULTISPECIES: aldehyde dehydrogenase family protein	yes	up
	gene2891	aldB	MULTISPECIES: aldehyde dehydrogenase	yes	down
	gene4094	mmsA	MULTISPECIES: CoA-acylating methylmalonate-semialdehyde dehydrogenase	yes	up

降解时间	组分1			组分2
降解时间	M_n/ (g/mol)	M_w/ (g/mol)	多分散性	M_n/ (g/mol)	M_w/ (g/mol)	多分散性
原样	9523	22364	2.350	—	—	—
对照组（28d）	9426	20882	2.220	—	—	—
5 d	9179	19808	2.160	682	784	1.150
10 d	8767	19406	2.210	716	806	1.330
14 d	6932	16141	2.328	383	652	1.701
21 d	6857	15968	2.329	394	648	1.644
28 d	6809	15795	2.320	366	617	1.685

降解时间	组分1			组分2
降解时间	M_n/ (g/mol)	M_w/ (g/mol)	多分散性	M_n/ (g/mol)	M_w/ (g/mol)	多分散性
原样	9523	22364	2.350	—	—	—
对照组（28d）	9426	20882	2.220	—	—	—
5 d	9179	19808	2.160	682	784	1.150
10 d	8767	19406	2.210	716	806	1.330
14 d	6932	16141	2.328	383	652	1.701
21 d	6857	15968	2.329	394	648	1.644
28 d	6809	15795	2.320	366	617	1.685

序号	保留时间/min	化学式	鉴定产物
1	2.518	CH₄O	甲醇
2	18.079	C₁₇H₃₄O₂	棕榈酸甲酯
3	19.665	C₁₉H₄₀O	十九烷醇
4	20.019	C₁₉H₃₈O₂	硬酯酸甲酯C18
5	20.758	C₂₂H₄₄O₂	正二十醋酸
6	21.575	C₂₀H₄₂	正二十烷
7	22.069	C₂₁H₃₈O₄	一亚油酸甘油酯
8	22.420	C₂₃H₃₂O₂	抗氧剂2246
9	23.990	C₃₆H₇₄	正三十六烷
10	25.465	C₃₄H₇₀	正三十四烷
11	26.917	C₄₄H₉₀	四十四烷