Mechanisms behind the impacts of CO₂ concentration on microbial methanogenesis in coal seams

Authors

• WANG Junyu, State Key Laboratory of Digital Intelligent Technology for Unmanned Coal Mining, Anhui University of Science & Technology, Huainan 232001, ChinaFollow
• LIU Bingjun, State Key Laboratory of Digital Intelligent Technology for Unmanned Coal Mining, Anhui University of Science & Technology, Huainan 232001, China; Joint National-Local Engineering Research Centre for Safe and Precise Coal Mining, Huainan 232001, ChinaFollow
• JIANG Xiangqing, Qinghai Energy Development (Group) Co. Ltd, Xining 810000, China; School of Safety Science and Engineering, Anhui University of Science & Technology, Huainan 232001, China
• CHEN Ruifeng, Xuangang Coal and Electricity Co., Ltd., Jinneng Holding Coal Industry Group, Xinzhou 034000, China
• XUE Sheng, Joint National-Local Engineering Research Centre for Safe and Precise Coal Mining, Huainan 232001, China
• ZHOU Wanlong, Xuangang Coal and Electricity Co., Ltd., Jinneng Holding Coal Industry Group, Xinzhou 034000, China
• YANG Yundu, State Key Laboratory of Digital Intelligent Technology for Unmanned Coal Mining, Anhui University of Science & Technology, Huainan 232001, China

Abstract

Background Coal seams exhibit the extensive distribution of a variety of indigenous functional microorganisms. Among these, some archaea enable methane metabolism using CO₂ as a substrate, providing the potential for the biological CO₂ storage and utilization in coal seams. However, conventional studies focus primarily on the degradation characteristics of coal matrix, leading to a lack of a systematic understanding of the mechanisms governing the adjustment of the methanation process in CO₂-bearing environments. Methods This study investigated the bituminous coals with a high volatile content in the Huainan mining area. Using laboratory-scale experiments on methanogenesis through anaerobic fermentation under varying CO₂ concentrations (volume fraction 0%‒100%), combined with untargeted metabolomics and microbiomics, this study systematically explored the mechanisms behind the impacts of CO₂ concentration on microbial methanogenesis. Furthermore, CO₂ consumption and methane production were calculated for the quantitative analysis of the conversion relationship between CO₂ and CH₄. Accordingly, the biochemical conversion pathways of CO₂ were established.Results The CO₂ concentration threshold was preliminarily observed under an atmosphere with a CO₂∶N₂ ratio of 4∶1 (C80 group). In this case, a peak methane production of 268.98 μmol/g coal was identified at day 45, suggesting a 42.11% increase compared to that of the atmosphere with a CO₂∶N₂ ratio of 0∶5 (189.27 μmol/g coal; CK group). In contrast, the methane production trended downward under atmospheres with CO₂∶N₂ ratios of 9∶1 (C90 group) and 1∶0 (C100 group). Meanwhile, the C80 group exhibited a 51.71% reduction in CO₂ within the headspace of the anaerobic bottle, with 1 mL CO₂ increasing about 0.38 mL CH₄. As the CO₂ concentration increased to 80%, the evolution patterns of the dominant genera of bacteria in the microbial community changed, with the dominant genera shifting from Paraclostridium and Enterococcus to Bacillus and Clostridium. The addition of exogenous CO₂ resulted in significant enrichment of methanogenic archaea, with mixotrophic Methanosarcina in the C80 group showing the most pronounced increase in relative abundance. Data from functional gene analysis also demonstrated that the intervention of exogenous CO₂ significantly increased both the Wood-Ljungdahl pathway of CO₂ fixation in Clostridium and the expression of methanogenesis-related functional genes. The analysis of liquid-phase differential metabolites before and after culture in various experimental groups shows that a high CO₂ concentration promoted the synthesis of metabolites associated with carbon fixation, such as cobamamide a,c-diamide and dihydroxyacetone phosphate (DAHP), while also accelerating the degradation of aromatic components in coals. In combination with results from microbial community composition, PICRUSt2 prediction, and untargeted metabolomics, this study determined the metabolic pathway that coupled the degradation of macromolecular organic matter in coals with CO₂-CH₄ conversion under the synergistic effects of bacteria and archaea.Conclusion The results of this study reveal that exogenous CO₂ enhance microbial methanogenesis through a triple mechanism, which (1) results in the enrichment of hydrogenotrophic/mixotrophic methanogenic archaea; (2) increases the abundance of functional genes involved in the Wood-Ljungdahl pathway of carbon fixation in Clostridium while also promoting CO₂ reduction; and (3) facilitates both the metabolism induced by the cleavage of aromatic rings and the synthesis of carbon fixation cofactors. The findings that the CO₂ concentration threshold is 80% and that the Wood-Ljungdahl pathway of carbon fixation can be activated by the directional enrichment of Clostridium-Sarcina synergetic microflora provide a theoretical basis and new control strategies for both coalbed methane bioengineering (CGB) and the conversion of geological to biological CO₂ storage.

Keywords

coal, methane metabolism, CO2 storage and utilization, untargeted metabolomics, anaerobic fermentation

DOI

10.12363/issn.1001-1986.25.07.0528

Recommended Citation

WANG Junyu, LIU Bingjun, JIANG Xiangqing, et al. (2026) "Mechanisms behind the impacts of CO₂ concentration on microbial methanogenesis in coal seams," Coal Geology & Exploration: Vol. 54: Iss. 4, Article 10.
DOI: 10.12363/issn.1001-1986.25.07.0528
Available at: https://cge.researchcommons.org/journal/vol54/iss4/10

Reference

[1] 孙腾民,刘世奇,汪涛. 中国二氧化碳地质封存潜力评价研究进展[J]. 煤炭科学技术,2021,49(11):10−20

SUN Tengmin,LIU Shiqi,WANG Tao. Research advances on evaluation of CO₂ geological storage potential in China[J]. Coal Science and Technology,2021,49(11):10−20

[2] 桑树勋,刘世奇,韩思杰,等. 中国煤炭甲烷管控与减排潜力[J]. 煤田地质与勘探,2023,51(1):159−175

SANG Shuxun,LIU Shiqi,HAN Sijie,et al. Coal methane control and its emission reduction potential in China[J]. Coal Geology & Exploration,2023,51(1):159−175

[3] 张泉,林进,折印楠,等. 我国二氧化碳地质封存潜力评价研究进展[J]. 中国石油和化工标准与质量,2022,42(11):144−146

[4] 唐相路,姜振学,邵泽宇,等. 第四系泥岩型生物气储层特征及动态成藏过程[J]. 现代地质,2022,36(2):682−694

TANG Xianglu,JIANG Zhenxue,SHAO Zeyu,et al. Reservoir Characteristics and Dynamic Accumulation Process of the Quaternary Mudstone Biogas[J]. Geoscience,2022,36(2):682−694

[5] 简阔,傅雪海,夏大平,等. 我国次生生物成因煤层气研究进展[J]. 煤矿安全,2023,54(4):11−21

JIAN Kuo,FU Xuehai,XIA Daping,et al. Research progress of secondary biogenic coalbed methane in China[J]. Safety in Coal Mines,2023,54(4):11−21

[6] 陈浩,李贵中,陈振宏,等. 微生物提高煤层气产量模拟实验研究[J]. 煤田地质与勘探,2016,44(4):64−68

CHEN Hao,LI Guizhong,CHEN Zhenhong,et al. Simulation experiment on enhancing coalbed methane production by microbes[J]. Coal Geology & Exploration,2016,44(4):64−68

[7] ZHANG Ji,LIANG Yanna. Evaluating approaches for sustaining methane production from coal through biogasification[J]. Fuel,2017,202:233−240.

[8] LIU Bingjun,WANG Yuewu,LI Yang,et al. Improved formation of biogenic methane by cultivable bacteria in highly volatile bituminous coals[J]. Journal of Cleaner Production,2022,366:132900.

[9] 苏现波,夏大平,赵伟仲,等. 煤层气生物工程研究进展[J]. 煤炭科学技术,2020,48(6):1−30

SU Xianbo,XIA Daping,ZHAO Weizhong,et al. Research advances of coalbed gas bioengineering[J]. Coal Science and Technology,2020,48(6):1−30

[10] AKIMBEKOV N S,DIGEL I,TASTAMBEK K T,et al. Hydrogenotrophic methanogenesis in coal–bearing environments:Methane production,carbon sequestration,and hydrogen availability[J]. International Journal of Hydrogen Energy,2024,52:1264−1277.

[11] CHENG H H,SYU J C,TIEN S Y,et al. Biological acetate production from carbon dioxide by Acetobacterium woodii and Clostridium ljungdahlii:The effect of cell immobilization[J]. Bioresource Technology,2018,262:229−234.

[12] GUO Hongyu,GAO Zhixiang,XIA Daping,et al. Biological methanation of coal in various atmospheres containing CO₂[J]. Fuel,2019,242:334−342.

[13] LIU Bingjun,CHEN Jian,LI Yang. Keystone microorganisms regulate the methanogenic potential in coals with different coal ranks[J]. ACS Omega,2022,7(34):29901−29908.

[14] LIU Bingjun,ZHOU Tianyao,XUE Sheng,et al. Improved formation of biomethane by enriched microorganisms from different rank coal seams[J]. ACS Omega,2024,9(10):11987−11997.

[15] 王增辉,赵彦军,姜淑珍,等. Gompertz曲线参数估计的一种加权方法[J]. 统计与决策,2018,34(20):26−29

WANG Zenghui,ZHAO Yanjun,JIANG Shuzhen,et al. A weighted method for parameter estimation of Gompertz curves[J]. Statistics and Decision,2018,34(20):26−29

[16] 杨霞,陈陆,王川庆. 16S rRNA基因序列分析技术在细菌分类中应用的研究进展[J]. 西北农林科技大学学报(自然科学版),2008,36(2):55−60

YANG Xia,CHEN Lu,WANG Chuanqing. Advance in application of 16S rRNA gene in bacteriology[J]. Journal of Northwest A & F University (Natural Science Edition),2008,36(2):55−60

[17] 朱大伟,武道吉,孙翠珍,等. 三维荧光光谱(3D–EEM)技术在溶解性有机质(DOM)分析中的应用[J]. 净水技术,2015,34(1):14−17

ZHU Dawei,WU Daoji,SUN Cuizhen,et al. Application of three–dimensional excitation–emission matrix (3 D–EEM) fluorescence spectrum technology in analysis of dissolved organic matter (DOM)[J]. Water Purification Technology,2015,34(1):14−17

[18] 周天尧,柳炳俊,薛生,等. 多手段表征生物降解煤基质分子结构变化特征及机制[J]. 煤炭学报,2025,50(增刊1):394−406

ZHOU Tianyao,LIU Bingjun,XUE Sheng,et al. Multi–method characterization of molecular structure changes and mechanisms of biodegradation of coal with different degrees of metamorphism[J]. Journal of China Coal Society,2025,50(Sup.1):394−406

[19] APARICI–CARRATALÁ D,ESCLAPEZ J,BAUTISTA V,et al. Archaea:Current and potential biotechnological applications[J]. Research in Microbiology,2023,174(7):104080.

[20] SCHULZ A,VOGT C,RICHNOW H H. Effects of high CO₂ concentrations on ecophysiologically different microorganisms[J]. Environmental Pollution,2012,169:27−34.

[21] GULLIVER D M,LOWRY G V,GREGORY K B. Comparative study of effects of CO₂ concentration and pH on microbial communities from a saline aquifer,a depleted oil reservoir,and a freshwater aquifer[J]. Environmental Engineering Science,2016,33(10):806−816.

[22] LI Yang,ZHANG Yuanyuan,XUE Sheng,et al. Actinobacteria may influence biological methane generation in coal seams[J]. Fuel,2023,339:126917.

[23] ZHOU Yixuan,SU Xianbo,ZHAO Weizhong,et al. Enhanced coal biomethanation by microbial electrolysis and graphene in the anaerobic digestion[J]. Renewable Energy,2023,219:119527.

[24] FAST A G,PAPOUTSAKIS E T. Stoichiometric and energetic analyses of non–photosynthetic CO₂–fixation pathways to support synthetic biology strategies for production of fuels and chemicals[J]. Current Opinion in Chemical Engineering,2012,1(4):380−395.

[25] AGLER M T,WRENN B A,ZINDER S H,et al. Waste to bioproduct conversion with undefined mixed cultures:The carboxylate platform[J]. Trends in Biotechnology,2011,29(2):70−78.

[26] JACK J,LO J,MANESS P C,et al. Directing Clostridium ljungdahlii fermentation products via hydrogen to carbon monoxide ratio in syngas[J]. Biomass and Bioenergy,2019,124:95−101.

[27] GUO Hongyu,GAO Zhixiang,XIA Daping,et al. Simulation study on the biological methanation of CO₂ sequestered in coal seams[J]. Journal of CO₂ Utilization,2019,34:171−179.

[28] SHI Jingxin,HAN Yuxing,XU Chunyan,et al. Anaerobic bioaugmentation hydrolysis of selected nitrogen heterocyclic compound in coal gasification wastewater[J]. Bioresource Technology,2019,278:223−230.

[29] 郭鑫,陈林勇,任恒星,等. 发酵罐中不同菌源降解甘肃煤和内蒙古煤产甲烷效果分析[J]. 煤炭转化,2022,45(4):46−54

GUO Xin,CHEN Linyong,REN Hengxing,et al. Analysis of methane production from Gansu coal and Inner Mongolia coal degraded by different bacteria in fermentor[J]. Coal Conversion,2022,45(4):46−54

[30] KORRE A,IMRIE C E,MAY F,et al. Quantification techniques for potential CO₂ leakage from geological storage sites[J]. Energy Procedia,2011,4:3413−3420.

[31] DE VRIEZE J,HENNEBEL T,BOON N,et al. Methanosarcina:The rediscovered methanogen for heavy duty biomethanation[J]. Bioresource Technology,2012,112:1−9.

[32] 李云嵩,郭红玉,符超勇,等. 煤层产甲烷菌对胍胶的生物降解实验[J]. 煤田地质与勘探,2019,47(3):56−60

LI Yunsong,GUO Hongyu,FU Chaoyong,et al. Study on the biodegradation of guanidine gum by methanogens in coal seam[J]. Coal Geology & Exploration,2019,47(3):56−60

[33] 周璇,李军英,伍刚,等. 维生素B₁₂在细菌与真核藻类相互作用中的功能研究进展[J]. 微生物学通报,2015,42(5):913−919

ZHOU Xuan,LI Junying,WU Gang,et al. The roles of vitamin B₁₂ in the interaction between bacteria and eukaryotic algae[J]. Microbiology China,2015,42(5):913−919

[34] KURADE M B,SAHA S,SALAMA E S,et al. Acetoclastic methanogenesis led by Methanosarcina in anaerobic co–digestion of fats,oil and grease for enhanced production of methane[J]. Bioresource Technology,2019,272:351−359.

[35] 严博,罗阳. 污泥厌氧发酵产酸的研究进展[J]. 工业微生物,2024,54(4):48−50

YAN Bo,LUO Yang. Research progress of VFA production by anaerobic fermentation of sludge[J]. Industrial Microbiology,2024,54(4):48−50

[36] 周艺璇,苏现波,赵伟仲,等. 煤微生物甲烷化的石墨烯强化机制[J]. 煤炭学报,2023,48(11):4145−4156

ZHOU Yixuan,SU Xianbo,ZHAO Weizhong,et al. Enhancement of bioconversion of coal to methane by graphene[J]. Journal of China Coal Society,2023,48(11):4145−4156

[37] SHI Chen,LIU Xiangrong,ZHAO Shunsheng,et al. Sequential degradations of Dananhu lignites by Nocardia mangyaensis and Bacillus licheniformis[J]. Fuel,2022,318:123623.

[38] ALI M,ELREEDY A,FUJII M,et al. Co–metabolism based adaptation of anaerobes to phenolic saline wastewater in a two–phase reactor enables efficient treatment and bioenergy recovery[J]. Energy Conversion and Management,2021,247:114722.

[39] ZHAO Weizhong,SU Xianbo,ZHANG Yifeng,et al. Microbial electrolysis enhanced bioconversion of coal to methane compared with anaerobic digestion:Insights into differences in metabolic pathways[J]. Energy Conversion and Management,2022,259:115553.

[40] LIU Yuchen,WHITMAN W B. Metabolic,phylogenetic,and ecological diversity of the methanogenic archaea[J]. Annals of the New York Academy of Sciences,2008,1125(1):171−189.