互營乙酸氧化菌研究進(jìn)展

吳 偉， 鄭珍珍， 麻婷婷， 承 磊， 張 輝

(1. 中國石化集團(tuán)勝利油田分公司油氣開發(fā)管理中心， 山東 東營 257097； 2. 農(nóng)業(yè)部沼氣科學(xué)研究所 農(nóng)業(yè)部農(nóng)村可再生能源開發(fā)利用重點(diǎn)實(shí)驗(yàn)室， 四川 成都 610041)

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互營乙酸氧化菌研究進(jìn)展

吳 偉1， 鄭珍珍2， 麻婷婷2， 承 磊2， 張 輝2

(1. 中國石化集團(tuán)勝利油田分公司油氣開發(fā)管理中心， 山東 東營 257097； 2. 農(nóng)業(yè)部沼氣科學(xué)研究所 農(nóng)業(yè)部農(nóng)村可再生能源開發(fā)利用重點(diǎn)實(shí)驗(yàn)室， 四川 成都 610041)

乙酸是沼氣發(fā)酵過程中的重要中間代謝產(chǎn)物，可以通過乙酸裂解途徑和互營乙酸氧化產(chǎn)甲烷途徑代謝產(chǎn)生甲烷。文章主要綜述了互營乙酸氧化菌的研究歷史和最新進(jìn)展，討論了影響互營乙酸氧化產(chǎn)甲烷代謝的環(huán)境因素，并展望了互營乙酸氧化菌的研究趨勢。

沼氣發(fā)酵； 互營乙酸氧化； 產(chǎn)甲烷途徑； 環(huán)境脅迫

沼氣發(fā)酵是在厭氧條件下，由多種不同類型的細(xì)菌和古菌微生物，通過互營代謝等協(xié)同作用，將復(fù)雜生物質(zhì)，如秸稈、畜禽糞便、石油烴等轉(zhuǎn)化為CH4和CO2的微生物學(xué)過程(見圖1)。沼氣發(fā)酵過程不僅發(fā)生在沼氣池等人工環(huán)境中，也普遍存在于油藏等地下缺氧環(huán)境中[1-2]。參與沼氣發(fā)酵的厭氧微生物是地下油藏的主要功能微生物，在油藏生物地球化學(xué)循環(huán)和提高石油采收率中起著重要的作用[3-4]。在沼氣發(fā)酵過程中，乙酸是重要的中間代謝產(chǎn)物。目前已知乙酸轉(zhuǎn)化為甲烷可通過兩種代謝途徑來實(shí)施，一是乙酸裂解型產(chǎn)甲烷途徑，乙酸營養(yǎng)型產(chǎn)甲烷烷古菌(AM)直接降解乙酸產(chǎn)生CH4和CO2，其中乙酸的甲基部分轉(zhuǎn)化為CH4，羧基端轉(zhuǎn)化為CO2。迄今已發(fā)現(xiàn)2個屬的產(chǎn)甲烷古菌(Methanothrix和Methanosarcina)可以通過該途徑降解乙酸產(chǎn)生甲烷[5]。另一種是互營乙酸氧化產(chǎn)甲烷途徑(SAO-HM)[6]，乙酸先分解為H2和CO2，再通過氫營養(yǎng)型產(chǎn)甲烷古菌(HM)轉(zhuǎn)化產(chǎn)生CH4，這需要互營乙酸氧化菌(SAOB)和氫營養(yǎng)型產(chǎn)甲烷古菌(HM)通過互營代謝作用來完成。本文主要介紹SAOB的研究歷史和國內(nèi)外研究進(jìn)展，以期讀者對沼氣發(fā)酵過程有一個全面的了解。

圖1 沼氣發(fā)酵過程的一般代謝途徑

1 國內(nèi)外研究現(xiàn)狀

1.1 互營乙酸氧化菌的研究歷史

1.2 互營乙酸氧化菌的熱力學(xué)特征

吉布斯自由能(△G)決定了一個反應(yīng)是否可以自發(fā)進(jìn)行，乙酸是氧化態(tài)最高的有機(jī)物之一，其進(jìn)一步發(fā)酵產(chǎn)生H2和CO2的反應(yīng)，在標(biāo)準(zhǔn)狀況下的吉普斯自由能(△G0’)為+104.6 kJ·mol-1(見表1反應(yīng)2)，難以自發(fā)進(jìn)行。HM可以利用H2和CO2產(chǎn)生CH4，其△G0’為 -135.6 kJ·mol-1。當(dāng)這兩個反應(yīng)耦聯(lián)，SAOB“發(fā)酵”乙酸產(chǎn)生的H2，可以快速的被HM轉(zhuǎn)化為CH4，整個反應(yīng)體系中保持著極低的氫分壓。SAOB-HM途徑的總反應(yīng)式是1 mol乙酸產(chǎn)生1 mol CH4和1 mol CO2，△G0’為-31.0 kJ·mol-1(見表1反應(yīng)3)，這樣SAOB產(chǎn)生H2和CO2的反應(yīng)就可以源源不斷地進(jìn)行。整個反應(yīng)與AM直接利用乙酸產(chǎn)CH4的△G0’一樣(見表1反應(yīng)1)，但是這些能量需要維持SAOB和HM這2種微生物的生命代謝活動，其能量劣勢導(dǎo)致了SAOB的代謝速率低、生長非常緩慢[29]。

表1 乙酸降解產(chǎn)甲烷的代謝反應(yīng)及其熱力學(xué)特征[6]

注：*代表乙酸的甲基碳的流向，標(biāo)準(zhǔn)吉布斯自由能數(shù)據(jù)參考文獻(xiàn)[30]。

1.3 互營乙酸氧化菌的微生物多樣性研究

目前已分離的SAOB都是嚴(yán)格厭氧菌，除AOR丟失外，共有5個被鑒定為新種，它們都具有與產(chǎn)甲烷古菌共培養(yǎng)，進(jìn)行互營乙酸氧化產(chǎn)甲烷代謝的功能(見表2)。根據(jù)文獻(xiàn)報道，其中有5個菌是嗜熱菌，最適生長溫度都在55℃ ～65℃，只有ClostridiumultunenseBST是中溫菌，最適生長溫度為37℃[10]。AOR,ThermacetogeniumphaeumPBT和ThermotogalettingaeTMOT不僅可以互營氧化乙酸，還具有進(jìn)行同型產(chǎn)乙酸功能[9，11-12](見表2)。多個SAOB可以利用甜菜堿、半胱氨酸、丙酮酸、葡萄糖等有機(jī)物生長，具有不同的代謝功能。SAOB生長緩慢，分離周期長，基于分離培養(yǎng)的傳統(tǒng)方法難以全面認(rèn)識SAOB。而基于未培養(yǎng)的微生物分子生態(tài)學(xué)方法和同位素示蹤技術(shù)的應(yīng)用，科學(xué)家發(fā)現(xiàn)SAOB廣泛分布在人工和自然環(huán)境中。Sun[21]等發(fā)現(xiàn)在13個連續(xù)流攪拌(CSTR)沼氣工程中，其中有10個反應(yīng)器中存在各種類型的SAOB。Lee[31]等還發(fā)現(xiàn)厭氧消化反應(yīng)器中SpirochaetesCluster II可能代表一類新的SAOB。Ito等通過RNA-SIP和MAR-FISH等證實(shí)Synergistes group 4中也含有SAOB[32]。在高氨[33-35]、高溫[19-36]、高濃度揮發(fā)性脂肪酸(VFA)[37]和低水力停留時間(HRT)[38]等環(huán)境條件下，互營乙酸氧化產(chǎn)甲烷代謝通常是有機(jī)質(zhì)降解產(chǎn)甲烷的主要途徑。在高溫水稻土中，Thermacetogenium和Thermoanaerobacteriaceae是乙酸互營氧化代謝產(chǎn)甲烷過程中的關(guān)鍵細(xì)菌類群[22-23]。在高溫油藏中也發(fā)現(xiàn)了參與互營乙酸氧化產(chǎn)甲烷的細(xì)菌類群。Nazina等發(fā)現(xiàn)中國大港油田，Thermoanaerobacteriales、Thermotogales、Nitrospirales和Planctomycetales是參與互營乙酸氧化產(chǎn)甲烷的主要細(xì)菌類群[28]，Mayumi發(fā)現(xiàn)Thermacetogenium和Methanothermobacter是日本Yabase 油藏互營乙酸氧化產(chǎn)甲烷的主要細(xì)菌和古菌類群[27]。Gieg[26]等推測SAO-HM途徑是原油降解過程中的主要產(chǎn)甲烷途徑。Conrad小組報道在以色列Kinneret湖底沉積物中(15℃ ～30℃)，互營乙酸氧化產(chǎn)甲烷也是乙酸代謝的主要途徑[24]，但是它們后來的同位素示蹤實(shí)驗(yàn)卻發(fā)現(xiàn)Methanothrix是代謝乙酸產(chǎn)甲烷的主要功能菌[39]。這些研究表明在很多獨(dú)特的生態(tài)系統(tǒng)中，還存在大量未分離培養(yǎng)的SAOB，它們在碳素生物地球化學(xué)循環(huán)中起著重要的生理生態(tài)學(xué)功能。

表2 互營乙酸氧化菌的純培養(yǎng)物

注：ND為未測出。

1.4 影響互營乙酸氧化產(chǎn)甲烷代謝的環(huán)境因子

1.5 互營乙酸氧化菌的生物強(qiáng)化作用

2 研究展望

綜上所述，通過培養(yǎng)和未培養(yǎng)方法，發(fā)現(xiàn)SAOB廣泛參與了沼氣發(fā)酵過程，并起著重要的生理生態(tài)學(xué)功能，但是迄今為止，還不清楚SAOB響應(yīng)和調(diào)控乙酸代謝的分子機(jī)理。氨,溫度,pH值,HRT和VFA等環(huán)境因子的改變，會影響SAOB的豐度和群落組成，但是，對SAOB應(yīng)答環(huán)境脅迫的分子機(jī)制研究還非常有限，這些有待更深入去探討。

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Recent Advances on Syntrophic Acetate Oxidation Bacteria /

WU Wei1, ZHENG Zhen-zhen2, MA Ting-ting2, CHENG Lei2, ZHANG Hui2/

(1. Center for oil and gas development of Shengli Oifield Company, China Petroleum & Chemical Corporation， Dongying 257091,China; 2. Biogas Institute of Ministry of Agriculture, Key Laboratory of Development and Application of Rural Renewable Energy of Ministry of Agriculture, Chengdu 610041, China )

Acetate is an important intermediate during biogas fermentation, which could be converted into methane through acetoclastic methanogenesis and syntrophic acetate oxidation coupled with hydrogenotrophic methanogenesis. This paper mainly reviewed the history and progress of research on syntrophic acetate oxidation bacteria, and discussed the environmental factors affecting the pathway of syntrophic acetate oxidation coupled hydrogenotrophic methanogenesis. And the further study was prospected.

Biogas fermentation； syntrophic acetate oxidation； methanogenic pathway； environmental stress

2015-12-20

項(xiàng)目來源： 國家高技術(shù)研究發(fā)展計(jì)劃(2013aa064401)； 中國農(nóng)業(yè)科學(xué)院基本科研業(yè)務(wù)費(fèi) (2013ZL001)； 微生物資源前期開發(fā)國家重點(diǎn)實(shí)驗(yàn)室項(xiàng)目(SKLMR-20150605)

吳 偉(1968- )，男，四川簡陽人，高級工程師，主要研究方向?yàn)樘岣呤筒墒章始夹g(shù)，E-mail: wuwei656.slyt@sinopec

張 輝，E-mail: zhanghuits@aliyun.com

S216.4;X172

A

1000-1166(2016)02-0003-06