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Single-step bioconversion of lignocellulose to hydrogen using novel moderately thermophilic bacteria.

ABSTRACT: BACKGROUND:Consolidated bioprocessing (CBP) of lignocellulosic biomass to hydrogen offers great potential for lower cost and higher efficiency compared to processes featuring dedicated cellulase production. Current studies on CBP-based hydrogen production mainly focus on using the thermophilic cellulolytic bacterium Clostridium thermocellum and the extremely thermophilic cellulolytic bacterium Caldicellulosiruptor saccharolyticus. However, no studies have demonstrated that the strains in the genus Thermoanaerobacterium could be used as the sole microorganism to accomplish both cellulose degradation and H2 generation. RESULTS:We have specifically screened for moderately thermophilic cellulolytic bacteria enabling to produce hydrogen directly from conversion of lignocellulosic materials. Three new strains of thermophilic cellulolytic bacteria in the genus Thermoanaerobacterium growing at a temperature of 60°C were isolated. All of them grew well on various plant polymers including microcrystalline cellulose, filter paper, xylan, glucose, and xylose. In particular, the isolated bacterium, designated as Thermoanaerobacterium thermosaccharolyticum M18, showed high cellulolytic activity and a high yield of H2. When it was grown in 0.5% microcrystalline cellulose, approximately 82% cellulose was consumed, and the H2 yield and maximum production rate reached 10.86 mmol/g Avicel and 2.05 mmol/L/h, respectively. Natural lignocellulosic materials without any physicochemical or biological pretreatment also supported appreciable growth of strain M18, which resulted in 56.07% to 62.71% of insoluble cellulose and hemicellulose polymer degradation in corn cob, corn stalk, and wheat straw with a yield of 3.23 to 3.48 mmol H2/g substrate and an average production rate of 0.10 to 0.13 mmol H2/L/h. CONCLUSIONS:The newly isolated strain T. thermosaccharolyticum M18 displayed effective degradation of lignocellulose and produced large amounts of hydrogen. This is the first report of a Thermoanaerobacterium species presenting cellulolytic characteristics, and this species thus represents a novel cellulolytic bacterium distinguished from all other known cellulolytic bacteria. In comparison, the extraordinary yield and specific rate of hydrogen for strain M18 obtained from lignocellulose make it more attractive in monoculture fermentation. T. thermosaccharolyticum M18 is thus a potential candidate for rapid conversion of lignocellulose to biohydrogen in a single step.

SUBMITTER: Cao GL

PROVIDER: S-EPMC4052809 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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Single-step bioconversion of lignocellulose to hydrogen using novel moderately thermophilic bacteria.

Cao Guang-Li GL Zhao Lei L Wang Ai-Jie AJ Wang Zhen-Yu ZY Ren Nan-Qi NQ

Biotechnology for biofuels 20140603

<h4>Background</h4>Consolidated bioprocessing (CBP) of lignocellulosic biomass to hydrogen offers great potential for lower cost and higher efficiency compared to processes featuring dedicated cellulase production. Current studies on CBP-based hydrogen production mainly focus on using the thermophilic cellulolytic bacterium Clostridium thermocellum and the extremely thermophilic cellulolytic bacterium Caldicellulosiruptor saccharolyticus. However, no studies have demonstrated that the strains in ...[more]

PMID: 24920960

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Project description:BackgroundConsolidated bioprocessing (CBP) of lignocellulosic biomass to ethanol using thermophilic bacteria provides a promising solution for efficient lignocellulose conversion without the need for additional cellulolytic enzymes. Most studies on the thermophilic CBP concentrate on co-cultivation of the thermophilic cellulolytic bacterium Clostridium thermocellum with non-cellulolytic thermophilic anaerobes at temperatures of 55°C-60°C.ResultsWe have specifically screened for cellulolytic bacteria growing at temperatures >70°C to enable direct conversion of lignocellulosic materials into ethanol. Seven new strains of extremely thermophilic anaerobic cellulolytic bacteria of the genus Caldicellulosiruptor and eight new strains of extremely thermophilic xylanolytic/saccharolytic bacteria of the genus Thermoanaerobacter isolated from environmental samples exhibited fast growth at 72°C, extensive lignocellulose degradation and high yield ethanol production on cellulose and pretreated lignocellulosic biomass. Monocultures of Caldicellulosiruptor strains degraded up to 89-97% of the cellulose and hemicellulose polymers in pretreated biomass and produced up to 72 mM ethanol on cellulose without addition of exogenous enzymes. In dual co-cultures of Caldicellulosiruptor strains with Thermoanaerobacter strains the ethanol concentrations rose 2- to 8.2-fold compared to cellulolytic monocultures. A co-culture of Caldicellulosiruptor DIB 087C and Thermoanaerobacter DIB 097X was particularly effective in the conversion of cellulose to ethanol, ethanol comprising 34.8 mol% of the total organic products. In contrast, a co-culture of Caldicellulosiruptor saccharolyticus DSM 8903 and Thermoanaerobacter mathranii subsp. mathranii DSM 11426 produced only low amounts of ethanol.ConclusionsThe newly discovered Caldicellulosiruptor sp. strain DIB 004C was capable of producing unexpectedly large amounts of ethanol from lignocellulose in fermentors. The established co-cultures of new Caldicellulosiruptor strains with new Thermoanaerobacter strains underline the importance of using specific strain combinations for high ethanol yields. These co-cultures provide an efficient CBP pathway for ethanol production and represent an ideal starting point for development of a highly integrated commercial ethanol production process.

Project description:BACKGROUND:The selective lignin-degrading white-rot fungi are regarded to be the best lignin degraders and have been widely used for reducing the saccharification recalcitrance of lignocellulose. However, the biological delignification and conversion of lignocellulose in biorefinery is still limited. It is necessary to develop novel and more efficient bio-delignification systems. RESULTS:Physisporinus vitreus relies on a new versatile peroxidase (VP)-based delignification strategy to remove enzymatic recalcitrance of corn stover efficiently, so that saccharification of corn stover was significantly enhanced to 349.1 mg/g biomass (yield of glucose) and 91.5% (hydrolysis yield of cellulose) at 28 days, as high as levels reached by thermochemical treatment. Analysis of the lignin structure using pyrolysis-gas chromatography-mass spectrometry (Py-GC/MS) showed that the total abundance of lignin-derived compounds decreased by 54.0% and revealed a notable demethylation during lignin degradation by P. vitreus. Monomeric and dimeric lignin model compounds were used to confirm the ligninolytic capabilities of extracellular ligninases secreted by P. vitreus. The laccase (Lac) from P. vitreus could not oxidize nonphenolic lignin compounds and polymerized ?-O-4 and 5-5' dimers to precipitate which had a negative effect on the enzymatic hydrolysis of corn stover in vitro. However, the VP from P. vitreus could oxidize both phenolic and nonphenolic lignin model compounds as well as break the ?-O-4 and 5-5' dimers into monomeric compounds, which were measured by high-performance liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS). Moreover, we showed that addition of purified VP in vitro improved the enzymatic hydrolysis of corn stover by 14.1%. CONCLUSIONS:From the highly efficient system of enzymatic recalcitrance removal by new white-rot fungus, we identified a new delignification strategy based on VP which could oxidize both phenolic and nonphenolic lignin units and break different linkages in lignin. In addition, this is the first evidence that VP could break 5-5' linkage efficiently in vitro. Moreover, VP improved the enzymatic hydrolysis of corn stover in vitro. The remarkable lignin-degradative potential makes VP attractive for biotechnological applications.

Project description:Effective artificial microbial consortia containing microorganisms with desired biological functions have the potential to optimize the lignocellulose-based bioindustry. Thermobifida fusca was a dominant actinobacterium in high-temperature corn stalk composts, but it was unable to grow alone in corn stalk solid medium. Interestingly, T. fusca showed good growth and secreted enzymes when cocultured with Thermomyces lanuginosus. T. lanuginosus grew firstly during the initial stage, whereas T. fusca dominated the system subsequently during cocultivation. The secretome indicated that T. lanuginosus mainly degraded xylan by expressing a GH11 xylanase (g4601.t1, GenBank AAB94633.1; with relative secretion of 4.95 ± 0.65%). T. fusca was induced by xylan mainly to secrete a xylanase from GH11 family (W8GGR4, GenBank AHK22788.1; with relative secretion of 8.71 ± 3.83%) which could rapidly degrade xylan to xylo-oligosaccharide (XOS) and xylose within 2 min, while high concentrations (>0.5%, w/v) of XOS or xylose suppressed the growth of T. fusca; which may be the reason why T. fusca unable to grow alone in corn stalk solid medium. However, T. lanuginosus could utilize the XOS and xylose produced by xylanases secreted by T. fusca. During the synergistic degradation of lignocellulose by T. lanuginosus and T. fusca, xylan was rapidly consumed by T. lanuginosus, the residual cellulose could specifically induced T. fusca to express a GH10 xylanase with a CBM2 domain (Q47KR6, GenBank AAZ56956.1; with relative secretion of 5.03 ± 1.33%) and 6 cellulases (2 exocellulases and 4 endocellulases). Moreover, T. lanuginosus increased the secretion of cellulases from T. fusca by 19-25%. The order of T. lanuginosus and T. fusca was consistent with the multilayered structures of lignocellulose and could be regulated by different concentrations of XOS and xylose. The novel synergism of T. lanuginosus and T. fusca gave a new sight for revealing more synergetic relationships in natural environments and exploring efficient microbial inoculants and enzyme cocktails for lignocellulose degradation.

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Single-step bioconversion of lignocellulose to hydrogen using novel moderately thermophilic bacteria.

Publications

Single-step bioconversion of lignocellulose to hydrogen using novel moderately thermophilic bacteria.

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