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The effects of pore sizes on the in-situ utilization of synthesis gas (syngas, H 2 and CO) mixed culture fermentation (MCF) in the hollow-fiber membrane biofilm reactor (HfMBR) are not clear. Thus, the ultrafiltration (R1) and microfiltration (R2) HfMBRs were constructed. Syngas was totally consumed within the formed biofilm in R1; contrarily, it accumulated notably in R2. In the batch mode of R1 and R2, volatile fatty acids (VFAs) of acetate, butyrate and caproate were the main metabolites, but the production rate of total VFA in R1 (61.9 mmol-C/(L·d)) was higher than that of R2 (27.6 mmol-C/(L·d)). In the continuous mode, the R1 performance was much better than that of R2, and the biofilm in R2 was even washed out. Furthermore, Clostridium (30.0%) was the main genus in the enriched biofilm of R1, which converted syngas to VFAs. Thus, the ultrafiltration membrane shall be the suitable candidate for syngas MCF., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Alkaline mixed culture fermentation (MCF) is a promising technology for reducing organic waste and producing biochemicals. However, chemostat metabolite spectra that are necessary for constructing a model and analyzing factors are seldom reported. In the present study, the effects of pH on the metabolites distribution in mesophilic (35 °C), thermophilic (55 °C), and extreme-thermophilic (70 °C) alkaline MCF were demonstrated. A chemical oxygen demand balance above 80% was achieved, and the main metabolites included acetate, ethanol, propionate, lactate, and formate. The yields of ethanol and formate increased as pH was increased from 7.5 to higher pH under mesophilic and thermophilic conditions, while the yields of acetate, lactate, and/or propionate decreased. The yields of ethanol, acetate, and formate increased under extreme-thermophilic conditions as pH was increased from 7.5 to 9.0, whereas lactate and hydrogen yields decreased. Low biomass yield under thermophilic and extreme-thermophilic conditions benefited higher metabolite production and minimized the accumulation of sludge., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

There have been no reports of H 2 and CO 2 mixed-culture fermentation (MCF) at 25 °C in a hollow-fiber membrane biofilm reactor (HfMBR). In this study, H 2 and CO 2 MCF were conducted in an HfMBR at 25 °C producing metabolites including acetate, ethanol, butyrate, and caproate. Compared to pure culture fermentation (i.e., Clostridium carboxidivorans P7), the MCF in HfMBR at 25 °C produced a higher concentration of caproate in this study (3.4 g/L in batch 1 and 5.7 g/L in batch 2). The dominant genera were Clostridium_sensu_stricto_12 and Prevotella_7. The caproate was more likely formed from the pathway of acetate and ethanol rather than via butyrate and ethanol. Since caproate is more valuable than acetate and low temperature fermentation consumes less energy, this process of H 2 and CO 2 MCF at 25 °C is appropriate for industrial application., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

The mixed culture fermentation is an important environmental biotechnology that converts biodegradable organic wastes to valuable chemicals such as hydrogen, methane, acetate, ethanol, propionate, and so on. For the multistep process of hydrolysis, acidogenesis, acetogenesis/homoacetogensis, and methanogenesis, the typical metabolic reactions are firstly summarized. And then, since the final metabolites are always a mixture, the separation and purification processes are necessary to couple with anaerobic fermentation. Therefore, several typical coupling technologies including biogas upgrading, two-stage fermentation, gas stripping, membrane technology of pervaporation, membrane distillation, electrodialysis, bipolar membrane electrodialysis, and microbial fuel cells are summarized to separate the metabolites and recover energy. At last, the novel technologies such as the controlled metabolite production, medium chain carboxylic acid production, and high temperature ethanol recovery in thermophilic mixed culture fermentation are also reviewed. However, the novel concepts are still needed to meet the demands of better overall performances and lower total costs.

The control of metabolite production is difficult in mixed culture fermentation. This is particularly related to hydrogen inhibition. In this work, hydrogenotrophic methanogens were selectively enriched to reduce the hydrogen partial pressure and to realize efficient acetate production in extreme-thermophilic (70°C) mixed culture fermentation. The continuous stirred tank reactor (CSTR) was stable operated during 100 days, in which acetate accounted for more than 90% of metabolites in liquid solutions. The yields of acetate, methane and biomass in CSTR were 1.5 ± 0.06, 1.0 ± 0.13 and 0.4 ± 0.05 mol/mol glucose, respectively, close to the theoretical expected values. The CSTR effluent was stable and no further conversion occurred when incubated for 14 days in a batch reactor. In fed-batch experiments, acetate could be produced up to 34.4 g/L, significantly higher than observed in common hydrogen producing fermentations. Acetate also accounted for more than 90% of soluble products formed in these fed-batch fermentations. The microbial community analysis revealed hydrogenotrophic methanogens (mainly Methanothermobacter thermautotrophicus and Methanobacterium thermoaggregans) as 98% of Archaea, confirming that high temperature will select hydrogenotrophic methanogens over aceticlastic methanogens effectively. This work demonstrated a potential application to effectively produce acetate as a value chemical and methane as an energy gas together via mixed culture fermentation.

Thermophilic mixed culture fermentation (TMCF) has advantages such as high substrate degradation rate (e.g. 0.5 d−1 of 60 °C vs 0.2 d−1 of 35 °C), hydrogen and methane yields (e.g. 3–4 mol-H2/mol-glucose at 70 °C vs 1–2 mol-H2/mol-glucose at 35 °C) and low gas solubility (e.g. H2 decrease 10% and CH4 decrease 30%) for the treatment of hot wastewater. However, the low biomass concentration, complexity of metabolites and high-running cost still limit its application to some extent. Here, the typical metabolic reactions in this process are briefly summarized. Then, the recent advances in alkali TMCF, anaerobic membrane bioreactor, individual metabolite production, thermophilic microbial fuel cells and thermophilic syngas fermentation are reviewed. The following separation processes of biogas upgrading, gas stripping and electrodialysis are also reviewed to separate and purify the metabolites in TMCF. Lastly, to meet the demands of better overall performance and lower-running cost, the prospects in the fields of direct interspecies electron transfer, medium chain fatty acids production and energy cost reduction are highlighted, but new strategies in TMCF still need to be developed in the future. [ABSTRACT FROM AUTHOR]

Abstract Syngas (CO/H 2) fermentation provides an alternative method for organics utilization. The hollow-fiber membrane biofilm reactor (HfMBR) offers a potential way to use CO and H 2 effectively. However, studies on syngas utilization by mixed culture fermentation (MCF) in HfMBR were seldom reported. This study is the first to demonstrate and compare the in-situ production of volatile fatty acids via syngas by mesophilic and thermophilic MCF, in which CO and H 2 utilization exceeded 95% and even reached 100%. Acetate (4.22 g/L), butyrate (1.35 g/L), caproate (0.88 g/L), and caprylate (0.52 g/L) were detected at 35 °C, whereas acetate was the main metabolite at 55 °C. The proportion of acetate ranged from 47.5% to 62.7% at 35 °C and exceeded 90% at 55 °C in the batch mode. Notably, the acetate concentration and the production rate could reach 24.6 g/L and 16.4 g/(L.d), respectively, at pH 6.5 in the continuous mode at 55 °C. Thus, thermophilic conditions are preferable for high-purity and high-concentration acetate production from syngas. As revealed by Illumina high-throughput sequencing, Clostridium (41.6%) and Thermoanaerobacterium (92.8%) were the dominant bacteria under mesophilic and thermophilic conditions, respectively. This study demonstrates a potential technique for syngas (CO/H 2) utilization and biochemicals production by MCF in HfMBR. Graphical abstract Image 1 Highlights • Syngas (H 2 /CO) fermentation was conducted in hollow-fiber membrane biofilm reactors. • The main metabolites contain acetate, butyrate, caproate and caprylate at 35 °C. • Acetate and minimal butyrate (less than 1 g/L) were detected at 55 °C. • Clostridium (41.6%) was the dominant bacterium at 35 °C. • The dominant bacterium was Thermoanaerobacterium (92.8%) at 55 °C. [ABSTRACT FROM AUTHOR]

The inhibition of acetate under acidic pH is an ideal way to reduce methanogenesis in mesophilic mixed culture fermentation (MCF). However, the effects of acetate concentration and acidic pH on methanogenesis remain unclear. Besides, although hydrogenotrophic methanogens can be suitable targets in MCF, they are generally ignored. Therefore, we intentionally enriched hydrogenotrophic methanogens and found that free acetic acid (FAA, x) concentration and specific methanogenic activity (SMA, y) were correlated according to the equation: y = 0.86 × 0.31/(0.31 + x) (R 2  = 0.909). The SMA was decreased by 50% and 90% at the FAA concentrations of 0.31 and 2.36 g/L, respectively. The coenzyme M concentration and relative electron transport activity agreed well with the FAA concentration. Moreover, the methanogenic activity could not be recovered when the FAA concentration exceeded 0.81 g/L. These findings indicated that neither acetate nor acidic pH, but FAA was the key factor to inhibit methanogenesis in MCF. [ABSTRACT FROM AUTHOR]

Researches on environmentally friendly pathways in mixed culture fermentation (MCF) to convert synthesis gas (syngas), including carbon monoxide (CO) and hydrogen (H 2 ), to biofuels and chemicals are attracting worldwide attention. The hollow-fiber membrane biofilm reactor (HFMBR) offers a potential technology for in-situ utilization of syngas. In this work, the performance of syngas MCF in the HFMBR at acidic pH 4.5 was studied for the first time. Sole ethanol was produced using an HFMBR in batch mode and the maximum concentration reached 16.9 g/L. The results also showed that the partial pressure of H 2 (P H2 ) and CO (P CO ) could tune the acetate and ethanol production in HFMBR, in a manner whereby high P H2 and P CO favored ethanol production, while low P H2 and P CO benefited acetate production. Microbial analysis revealed that the dominant genus in the HFMBR biofilm was Clostridium (86.3%), which is consistent with the experimental results. Overall, the adjustable production of acetate and ethanol from syngas in an HFMBR would be useful for the utilization of syngas MCF in the future. [ABSTRACT FROM AUTHOR]

The microbial community compositions of a chemostat enriched in a thermophilic (55 °C) mixed culture fermentation (MCF) for hydrogen production under different operational conditions were revealed in this work by integrating denaturing gradient gel electrophoresis (DGGE), Illumina Miseq high-throughput sequencing, and 16S rRNA clone library sequencing. The results showed that the community structure of the enriched cultures was relatively simple. Clones close to the genera of Thermoanaerobacter and/or Bacillus mainly dominated the bacteria. And homoacetogens and archaea were washed out and not detected even by Illumina Miseq high-throughput sequencing which supported the benefit for hydrogen production. On the other hand, the results revealed that the metabolic shift was clearly associated with the change of dominated bacterial groups. The effects of hydrogen partial pressure (P) and pH from 4.0 to 5.5 on the microbial compositions were not notable and Thermoanaerobacter was dominant, thus, the metabolites were also not changed. While Bacillus, Thermoanaerobacter and Propionispora hippei dominated the bacteria communities at neutral pH, or Bacillus and Thermoanaerobacter dominated at high influent glucose concentrations, consequently the main metabolites shifted to acetate, ethanol, propionate, or lactate. Thereby, the effect of microbial composition on the metabolite distribution and shift shall be considered when modeling thermophilic MCF in the future. [ABSTRACT FROM AUTHOR]

[Display omitted] • Converting xylose to caproate (C6) by mixed culture fermentation was verified. • The caproate production from xylose (10 g/L) was 1.2 g/L in a CSTR reactor. • Dosing extra ethanol (5 g/L) slightly increased the caproate production by ∼ 30%. • Genus Caproiciproducens (>80%) was identified as the main caproate producer. • The fatty acid biosynthesis pathway was revealed via a metagenomic analysis. Caproate production from organic wastes is deemed as a novel strategy in mixed culture fermtation (MCF). However, producing caproate from natural sugar of xylose by MCF is seldom reported and the metabolic pathway is still unclear. Thus, the caproate production from xylose was investigated in this study by mesophilic MCF. The results showed that the caproate concentration from xylose (10 g/L) was 1.2 ± 0.17 g/L (equal to 2.7 gCOD/L) under pH 5.0. Dosing extra ethanol of 5 g/L could slightly increase the caproate production by ∼ 30% (i.e., 1.6 g/L). While dosing extra acetate of 5 g/L negatively affected the caproate production, which was just 0.2 g/L. The microbial analysis illustrated that genus Caproiciproducens was a main identified caproate producer, occupying over 80% of enriched mixed culture. The fatty acid biosynthesis pathway was identified via metagenomic analysis. These unexpected differences extended the understanding of caproate production from organic wastes. [ABSTRACT FROM AUTHOR]

The effects of pH, hydrogen partial pressure (P), and influent glucose concentration on the metabolic distribution in chemostat were investigated in this work in extreme-thermophilic mixed culture fermentation (MCF) process. The results showed that acetate, ethanol, and hydrogen were the main metabolites. A shift of ethanol to acetate and hydrogen was observed as pH increasing from 4.0 to 7.0 or P decreasing from 0.64 to 0.05 atm. The maximum hydrogen yield was 3.16 ± 0.16 mol/mol glucose at P 0.05 atm. Lactate was only accumulated at low pH or high influent glucose concentration, while others such as butyrate and formate were rather low. Thermodynamic analysis illustrated that a mixture of acetate, ethanol, and/or lactate was essential for hydrogen production in extreme-thermophilic MCF. The hydrogen-producing rate was also calculated, and the maximum value was 2.2 ± 0.1 L/(L-reactor/day) at P 0.05 atm. Except hydrogen, other metabolites, such as liquid fatty acids and biofuels, could also be the producing targets in extreme-thermophilic MCF. [ABSTRACT FROM AUTHOR]

Syngas fermentation is a promising route for resource recovery. Acetate is an important industrial chemical product and also an attractive precursor for liquid biofuels production. This study demonstrated high fraction acetate production from syngas (H 2 and CO 2) in a hollow-fiber membrane biofilm reactor, in which the hydrogen utilizing efficiency reached 100 % during the operational period. The maximum concentration of acetate in batch mode was 12.5 g/L, while the acetate concentration in continuous mode with a hydraulic retention time of 9 days was 3.6 ± 0.1 g/L. Since butyrate concentration was rather low and below 0.1 g/L, the acetate fraction was higher than 99 % in both batch and continuous modes. Microbial community analysis showed that the biofilm was dominated by Clostridium spp., such as Clostridium ljungdahlii and Clostridium drakei, the percentage of which was 70.5 %. This study demonstrates a potential technology for the in situ utilization of syngas and valuable chemical production. [ABSTRACT FROM AUTHOR]

[Display omitted] • Occurrences of MPs from WAS by THP were identified with dark color and a broad MW. • Dark-colored MPs were produced from glucose and tryptophan as the model components. • MPs were characterized by higher peak values in CV and typical peaks in 3D-EEM. • The methane production from MPs was reduced by 49.1 % (MPs160) and 89.8 % (MPs180). • Low methane was due to inhibit protease, acidogenesis, and acetoclastic methanogens. Thermal hydrolysis pretreatment (THP) is widely used to promote the hydrolysis of polysaccharides and proteins in waste activated sludge (WAS). However, it also generates unwanted refractory organics of melanoidins products (MPs), which should not be overlooked, especially concerning the occurrence and inhibition of hydrolases and anaerobes. In this study, the MPs occurrence by THP was investigated and inhibition mechanism on anaerobic fermentation was revealed for the first time. The results showed that SCOD in WAS significantly increased from 0.5±0.02 g/L in control to 4.1±0.02 g/L in THP at 180 °C. However, the color of MPs under a higher temperature (>120 °C) changed from colourless to dark brown. The components of MPs were mainly nitrogenous and humic acid-like refractory organics. Unwanted MPs generated above 160 °C reduced methane production by 49.1 %–89.8 %, which was due to the inhibition of protease, acidogenic bacteria, and acetoclastic methanogens. Inhibition of acetoclastic methanogens also resulted in the accumulation of VFAs. The microbial diversity clearly showed that the toxicity of MPs altered the microbial community structure, reducing the proportion of both acidogenic bacteria and acetoclastic methanogens. Therefore, the negative effect of MPs on anaerobic digestion demonstrated that THP should be re-evaluated in the production of biochemicals, which is important for achieving carbon neutrality of WWTPs in the future. [ABSTRACT FROM AUTHOR]

Thermophilic (55 °C) mixed culture fermentation (MCF) is a promising technology for converting organic wastes into bioproducts including hydrogen and chemicals. In this study, the effect of pH, hydrogen partial pressure ( P H 2 ) and influent glucose concentration on the metabolic distribution in thermophilic chemostat was investigated. The results showed that the metabolite composition was changed from acetate, butyrate and hydrogen to acetate, ethanol, propionate and format when pH increased from 4.0 to 7.0. The conversion of hydrogen to formate was found to be thermodynamically controlled. Reducing P H 2 did not significantly increase the hydrogen yield and consequently the metabolites did not change much. Lactate accumulation was observed only at high influent glucose concentration. The maximum hydrogen yield was 2.2 ± 0.16 mol/mol-glucose at P H 2 0.05 atm, while the maximum hydrogen production rate was 7.9 ± 0.1 L/(L-reactor·Day) at influent glucose concentration of 76.3 mmol/L. Thus, considering the operating cost, increasing influent glucose concentration to 76.3 mmol/L in this work could be a more practical approach to produce hydrogen in thermophilic MCF. Meanwhile, the impacts of precedent pH and influent glucose concentration on the metabolites distribution were negligible in thermophilic chemostat. The thermodynamic analysis illustrated that a mixture of acetate, butyrate, ethanol, propionate and/or lactate was essential for thermophilic hydrogen producing MCF. [ABSTRACT FROM AUTHOR]

• Converting xylose to caproate (C6) by mixed culture fermentation was verified. • The maximum concentration of caproate was 2.06 g/L in a batch reactor. • Caproate concentration were between 0.45 and 1.07 g/L at pH 5.0 in a CSTR reactor. • Genus Caproiciproducens was unexpectedly identified as the main caproate producer. • Fatty acid biosynthesis, not reverse β-oxidation, may be the pathway to produce C6. Caproate production by mixed culture fermentation (MCF) is economically attractive. Xylose is known as the second most abundant sugar in nature, however, producing caproate from xylose is never reported. In this study, caproate production from xylose by mesophilic MCF was firstly investigated. The results showed that as pH decreasing to 5.0, the caproate concentration was 2.06 g/L in a batch reactor and was between 0.45 and 1.07 g/L in a continuously stirred reactor. Microbial analysis illustrated that Caproiciproducens and Clostridium_sensu_stricto_12 , as two main identified caproate producers, occupied over 50% and around 10% of mixed culture, respectively. Thus, caproate production from xylose was proposed via the fatty acid biosynthesis pathway, not the well-known reverse β-oxidation pathway. These unexpected differences from literatures gains more understanding about caproate production from organic substrates via MCF. [ABSTRACT FROM AUTHOR]