Bioaugmentation with Thermoanaerobacterium thermosaccharolyticum W16 to enhance thermophilic hydrogen production using corn stover hydrolysate

In the present work, with corn stover hydrolysate as the substrate, an efficient hydrogen-producing thermophile, Thermoanaerobacterium thermosaccharolyticum W16, was added to three kinds of seed sludge (rotten corn stover (RCS), cow dung compost (CDC), and sludge from anaerobic digestion (SAD)) to investigate the effect of bioaugmentation on thermophilic hydrogen production. Batch test results indicate that the bioaugmentation with a small amount of the strain T. thermosaccharolyticum W16 (5% of total microbes) increased the hydrogen yield to varying degrees (RCS: from 8.78 to 9.90 mmol H₂/g utilized sugar; CDC: from 8.18 to 8.42 mmol H₂/g utilized sugar; SAD: from 8.55 to 9.17 mmol H₂/g utilized sugar). The bioaugmentation process also influenced the soluble metabolites composition towards more acetate and less butyrate production for RCS, and more acetate and less ethanol accumulation for SAD. Microbial community analysis indicates that Thermoanaerobacterium spp. and Clostridium spp. dominated microbial community in all situations and might be mainly responsible for thermophilic hydrogen generation. For RCS and SAD, the bioaugmentation obviously increased the relative abundance of the strain T. thermosaccharolyticum W16 in microbial community, which might be the main reason for the improvement of hydrogen production in these cases.     

Comparative study on bio-hydrogen production from corn stover: Photo-fermentation,dark-fermentation and dark-photo co-fermentation

In this study, three different fermentation methods, such as photo-fermentation (PF), dark-fermentation (DF) and dark-photo co-fermentation (DPCF) for bio-hydrogen production from corn stover were compared in terms of hydrogen production, substrate consumption, by-products formation and energy conversion efficiency. A modified Gompertz model was applied to perform the kinetic analysis of hydrogen production. The maximum cumulative hydrogen yield of 141.42 mL·(g TS)⁻¹ was achieved by PF, DF with the minimum cumulative hydrogen yield of 36.08 mL· (g TS)⁻¹ had the shortest lag time of 4.33 h, and DPCF had the maximum initial hydrogen production rate of 1.88 mL· (g TS)⁻¹·h⁻¹ and maximum initial hydrogen content of 44.40%. The results also indicated PF was an acid-consuming process with a low total VFAs concentration level of 2.90–4.19 g·L⁻¹, DF was a process of VFAs accumulation with a maximum total VFAs concentration of 12.66 g·L⁻¹, and DPCF was a synergistic process in which the total VFAs concentration was significantly reduced and the hydrogen production efficiency was effectively improved compared with DF. The energy conversion efficiency of PF, DF and DPCF were 10.12%, 2.58% and 6.45%, respectively.     

Optimization of alkaline pretreatment on corn stover for enhanced production of 1.3-propanediol and 2,3-butanediol by Klebsiella pneumoniae AJ4

Corn stover is one of the most promising lignocellulosic biomass that can be utilized for producing 1,3-propanediol and 2,3-butanediol. The pretreatment and enzymatic hydrolysis steps are essential for the bioconversion of lignocellulosic biomass to diols. For optimizing the pretreatment step, temperature, time, and NaOH concentration were evaluated based on total sugar recovery. Enzymatic hydrolysis for cellulose and hemicellulose were investigated at different solid-to-liquid ratios. The optimum conditions were found to be alkaline pretreatment with 0.25 mol dm⁻³ NaOH for 1 h at 60 °C followed by enzymatic hydrolysis at 50 °C for 48 h, with a solid slurry concentration of 100 g dm⁻³. Under these conditions, conversion rates of 92.55% and 78.82% were obtained from glucan and xylan, respectively. Diol production from fermentable sugars was 14.8 g dm⁻³, with a conversion yield and productivity of 0.46 g g⁻¹, and 0.98 g dm⁻³ h⁻¹, respectively. Our results are similar for diol production obtained using pure sugars under the same conditions. Therefore, mild alkaline pretreatment of corn stover facilitates delignification, significantly improving the rate of enzymatic saccharification and sugar recovery.     

Comparison of bio-hydrogen production from hydrolyzed wheat starch by mesophilic and thermophilic dark fermentation

Hydrogen gas production potentials of acid-hydrolyzed and boiled ground wheat were compared in batch dark fermentations under mesophilic (37 °C) and thermophilic (55 °C) conditions. Heat-treated anaerobic sludge was used as the inoculum and the hydrolyzed ground wheat was supplemented by other nutrients. The highest cumulative hydrogen gas production (752 ml) was obtained from the acid-hydrolyzed ground wheat starch at 55 °C and the lowest (112 ml) was with the boiled wheat starch within 10 days. The highest rate of hydrogen gas formation (7.42 ml H₂ h⁻¹) was obtained with the acid-hydrolyzed and the lowest (1.12 ml H₂ h⁻¹) with the boiled wheat at 55 °C. The highest hydrogen gas yield (333 ml H₂ g⁻¹ total sugar or 2.40 mol H₂ mol⁻¹ glucose) and final total volatile fatty acid (TVFA) concentration (10.08 g L⁻¹) were also obtained with the acid-hydrolyzed wheat under thermophilic conditions (55 °C). Dark fermentation of acid-hydrolyzed ground wheat under thermophilic conditions (55 °C) was proven to be more beneficial as compared to mesophilic or thermophilic fermentation of boiled (partially hydrolyzed) wheat starch.     

Hydrogen gas production from cheese whey powder (CWP) solution by thermophilic dark fermentation

Hydrogen gas production from cheese whey powder (CWP) solution by thermophilic dark fermentation was investigated at 55 °C. Experiments were performed at different initial total sugar concentrations varying between 5.2 and 28.5 g L⁻¹ with a constant initial bacteria concentration of 1 g L⁻¹. The highest cumulative hydrogen evolution (257 mL) was obtained with 20 g L⁻¹ total sugar (substrate) concentration within 360 h while the highest H₂ formation rate (2.55 mL h⁻¹) and yield (1.03 mol H₂ mol⁻¹ glucose) were obtained at 5.2 and 9.5 g L⁻¹ substrate concentrations, respectively. The specific H₂ production rate (SHPR = 4.5 mL h⁻¹ g⁻¹cells) reached the highest level at 20 g L⁻¹ total sugar concentration. Total volatile fatty acid (TVFA) concentration increased with increasing initial total sugar content and reached the highest level (14.15 g L⁻¹) at 28.5 g L⁻¹ initial substrate concentration. The experimental data was correlated with the Gompertz equation and the constants were determined. The optimum initial total sugar concentration was 20 g L⁻¹ above which substrate and product (VFA) inhibitions were observed.     

Hydrothermal pretreatment of switchgrass and corn stover for production of ethanol and carbon microspheres

Pretreatment of biomass is viewed as a critical step to make the cellulose accessible to enzymes and for an adequate yield of fermentable sugars in ethanol production. Recently, hydrothermal pretreatment methods have attracted a great deal of attention because it uses water which is a inherently present in green biomass, non-toxic, environmentally benign, and inexpensive medium. Hydrothermal pretreatment of switchgrass and corn stover was conducted in a flow through reactor to enhance and optimize the enzymatic digestibility. More than 80% of glucan digestibility was achieved by pretreatment at 190 °C. Addition of a small amount of K₂CO₃ (0.45-0.9 wt.%) can enhance the pretreatment and allow use of lower temperatures. Switchgrass pretreated at 190 °C only with water had higher internal surface area than that pretreated in the presence of K₂CO₃, but both the substrates showed similar glucan digestibility. In comparison to switchgrass, corn stover required milder pretreatment conditions. The liquid hydrolyzate generated during pretreatment was converted into carbon microspheres by hydrothermal carbonization, providing a value-added byproduct. The carbonization process was further examined by GC-MS analysis to understand the mechanism of microsphere formation.     

Analysis of shaking effect on photo-fermentative hydrogen production under different concentrations of corn stover powder

High solid phase and easily congeal affect the mass transfer in the photo-fermentative biohydrogen production when taken straw biomass as substrate. Hence, oscillator was adopted to provide the shaking condition to enhance the mass transfer situation in this paper. Diverse shaking velocity (0, 80, 120 and 160 rpm) and substrate concentration (0, 2, 4, 6, 8 and 10 g) were studied, to evaluate the influence on the hydrogen yield capacity. The results showed that shaking could help to accelerate of gas release, shorten the fermentation time, and improve hydrogen production rate. Hydrogen yield was significantly enhanced at high substrate concentration under shaking condition. Highest hydrogen yield of 57.08 ± 0.83, 57.62 ± 1.37, 62.28 ± 0.84 mL/g-volatile solids (VS) were observed at shaking velocities of 80, 120 and 160 rpm with 6, 8 and 10 g corn stover powder, respectively. On the contrary, shaking significantly reduced the potential of hydrogen yield at a low substrate concentration, and the lower hydrogen yield obtained at the higher shaking velocity. As the lowest hydrogen yields of 27.68 ± 1.02 and 41.93 ± 0.40 mL/g VS were obtained at shaking velocity of 160 rpm with 2 and 4 g corn stover powder, respectively.     

Enhance photocatalytic hydrogen evolution by using alkaline pretreated corn stover as a sacrificial agent

In this study, alkali pretreated corn stover was added as a sacrificial agent to the suspension of Pt/TiO₂ to significantly enhanced photocatalytic hydrogen evolution. The changes in structural characteristics of corn stover before and after alkali treatment and photocatalytic reactions were studied. According to the results, the removal of lignin and hemicellulose, as well as the changes in surface morphology, structural components, and functional groups of alkali pretreated corn stover may affect the photocatalytic hydrogen production process. Next, the effects of NaOH concentration, pretreatment time and temperature on hydrogen production were also investigated. Among them, within the scope of the experimental conditions, the optimal hydrogen production is 25.84 μmol h⁻¹. Moreover, the enhancement of photocatalytic hydrogen production is also achieved by using the waste liquid of alkali pretreated. The output of this study may provide a reference for the reuse of alkali-treated biomass residues and waste liquor in some industries, and make more comprehensive, efficient and green use of native lignocellulosic biomass.     

Biological hydrogen production from corn stover by moderately thermophile Thermoanaerobacterium thermosaccharolyticum W16

This study addressed the utilization of an agro-waste, corn stover, as a renewable lignocellulosic feedstock for the fermentative H₂ production by the moderate thermophile Thermoanaerobacterium thermosaccharolyticum W16. The corn stover was first hydrolyzed by cellulase with supplementation of xylanase after delignification with 2% NaOH. It produced reducing sugar at a yield of 11.2 g L⁻¹ glucose, 3.4 g L⁻¹ xylose and 0.5 g L⁻¹ arabinose under the optimum condition of cellulase dosage 25 U g⁻¹ substrate with supplement xylanase 30 U g⁻¹ substrate. The hydrolyzed corn stover was sequentially introduced to fermentation by strain W16, where, the cell density and the maximum H₂ production rate was comparable to that on simulated medium, which has the same concentration of reducing sugars with hydrolysate. The present results suggest a promising combined hydrogen production process from corn stover with enzymatic hydrolysis stage and fermentation stage using W16.     

Optimization of biohydrogen production of palm oil mill effluent by ozone pretreatment

The biohydrogen (H₂) production in batch experiments under varying concentrations of raw and ozonated palm oil mill effluent (POME) of 5000–30,000 mg COD.L⁻¹, at initial pH 6, under mesophilic (37 °C), thermophilic (55 °C) and extreme-thermophilic (70 °C) conditions. Effects of ozone pretreatment, substrate concentration and fermentation temperature on H₂ production using mesophilic seed sludge was undertaken. The results demonstrated that H₂ can be produced from both raw and ozonated POME, and the amounts of H₂ production were directly increased as the POME concentrations were increased. H₂ was successfully produced under the mesophilic fermentation of ozonated POME, with maximum H₂ yield, and specific H₂ production rate of 182 mL.g⁻¹ COD_removed (30,000 mg COD.L⁻¹) and 6.2 mL.h⁻¹.g⁻¹ TVS (25,000 mg COD.L⁻¹), respectively. Thus, indicating that the ozone pretreatment could elevate on the biodegradability of major constituents of the POME, which significantly enhanced yields and rates of the H₂ production. H₂ production was not achieved under the thermophilic and extreme-thermophilic fermentation. In both fermentation temperatures with ozonated POME, the maximum H₂ yield was 62 mL.g⁻¹ COD_removed (30,000 mg COD.L⁻¹) and 63 mL.g⁻¹ COD_removed (30,000 mg COD.L⁻¹), respectively. The highest efficiency of total and soluble COD removal was obtained at 44 and 37%, respectively following the mesophilic fermentation, of 24 and 25%, respectively under the thermophilic fermentation, of 32 and 20%, respectively under the extreme-thermophilic fermentation. The production of volatile fatty acids increased with an increased fermentation time and temperature in both raw and ozonated POME under all three fermentation temperatures. The accumulation of volatile fatty acids in the reactor content were mostly acetic and butyric acids. H₂ fermentation under the mesophilic condition of 37 °C was the better selection than that of the thermophilic and extreme-thermophilic fermentation.     

玉米秸秆预处理对厌氧发酵制氢影响的研究

为提高玉米秸秆的产氢能力,实验研究了蒸汽爆破预处理、硫酸预处理、氢氧化钠预处理、盐酸预处理和酸化(碱化)气爆预处理5种预处理方法对玉米秸秆发酵产氢能力的影响。结果表明,预处理可以将秸秆中相当一部分纤维素和半纤维素水解生成还原糖,其中质量分数为0.8%的H2SO4酸化汽爆预处理对秸秆的水解效果最好。在固-液比1∶10、H2SO4质量分数0.8%、保持微沸状态30min的处理条件下,秸秆的糖含量达到最大值24.57%,最大氢气产量为141mL/g。     

Co-digestion of swine manure and corn stover for bioenergy production in MixAlco™ consolidated bioprocessing

MixAlco™ consolidated bioprocessing (CBP) employs a mixed culture of terrestrial microorganisms to anaerobically ferment waste streams (e.g., animal manure, agriculture residues) into mixed carboxylate salts that can be further chemically converted to commodity chemicals (e.g., acetic acid, acetone) and liquid transportation fuels (e.g., ethanol, mixed alcohols, bio-gasoline). For countercurrent fermentations of 60% swine manure/40% lime-treated corn stover at 55 °C, the highest acid productivity [1.8 g/(L·d)] and highest conversion (73%) in this study occurred at an acid concentration of 25.1 and 17.0 g/L, respectively. The continuum particle distribution model (CPDM) predicted the experimental total acid concentrations and conversions around 11.1% and 17.2%, respectively. The CPDM prediction "map" for MixAlco™ CBP indicates that both high conversions (>79%) and high total acid concentrations (>35 g/L) are possible at industrial scale. The present study shows continuous co-digestion of corn stove and swine manure in the MixAlco™ process has the potential to produce annually 9.3 billion gallons of alcohol fuels (e.g., ethanol and mixed alcohols) in the United States.     

Optimization of biohydrogen production using acid pretreated corn stover hydrolysate followed by nickel nanoparticle addition

The dilute acid hydrolysis using corn stover (CS) to produce reducible sugars was optimized by the response surface methodology. The electron-equivalent balances of the main metabolites during the dark fermentation (DF) using acid hydrolysate were investigated to identify the evolutions of the electron sinks over the course of DF. The additions of nickel ion and Ni⁰ nanoparticles (NPs) were found to effectively enhance the hydrogen production at experimental conditions. The optimal condition (HCl 2.5 wt%, hydrolyzing duration 105 minutes, pH=5, S/B=3.5, Ni⁰ NPs=10 mg/L^-1) was achieved with Y_H2/S reaching 1.18 (mol.mol^-1-glucose). The Y_H2/S increased from 0.7 (mol.mol^-1-glucose) to 1.18 (mol.mol^-1-glucose) reaching 40% hydrogen yield increase when Ni⁰ NPs was added to the fermentation broth. Among the investigated significant soluble metabolites, the butyric acid was found to serve as the largest e-sink in the electron-equivalent balance. The additions of Ni⁰ NPs at low level (below 10 mg/L) were found to appreciably increase the hydrogen production. The increased pH and substrate to biomass ratio were found to skew the metabolic balance from hydrogen production to the biosynthesis (an increase of biomass). The proposed anaerobic digestion model with consideration of the inhibitory factors model presents a good agreement with the experimental data. The chemical addition such as nickel ions, Ni⁰ NPs was found to be a practical approach in enhancing biohydrogen production using CS acid hydrolysate as cultivation broth.     

Co-culture of Clostridium thermocellum and Clostridium thermosaccharolyticum for enhancing hydrogen production via thermophilic fermentation of cornstalk waste

A strategic method utilizing the co-culture of Clostridium thermocellum and Clostridium thermosaccharolyticum has been developed to improve hydrogen production via the thermophilic fermentation of cornstalk waste. The hydrogen yield in the co-culture fermentation process reached 68.2 mL/g-cornstalk which was 94.1% higher than that in the mono-culture. The hydrogen fermentation process was successfully scaled-up from 125 mL anaerobic bottles to an 8 L continuous stirred tank reactor, and the hydrogen production from cornstalk waste was significantly improved in the bioreactor system due to efficient mixing and mass transfer. The hydrogen yield in the bioreactor reached 74.9 mL/g-cornstalk which was 9.8% higher than that in the 125 mL anaerobic bottle. The present work indicates that the direct microbial conversion of lignocellulosic waste by co-culturing C. thermocellum and C. thermosaccharolyticum is a promising avenue for enhancing hydrogen production.     

Capnophilic lactic fermentation and hydrogen synthesis by Thermotoga neapolitana: An unexpected deviation from the dark fermentation model

The heterotrophic bacterium Thermotoga neapolitana produces hydrogen by fermentation of organic substrates. The process is referred to as dark fermentation and is typically complemented by production of acetic acid. Here we show that synthesis of products derived by reductive metabolism of pyruvate, mainly lactic acid, occurs to the detriment of acetic acid fermentation when the cultures of the thermophilic bacterium are flushed by saturating level of CO₂. Sodium bicarbonate in a very narrow range of concentrations (∼14 mM) also causes the same metabolic shift. The capnophilic (CO₂-requiring) re-orientation of the fermentative process toward lactic acid does not affect hydrogen productivity thus challenging the currently accepted dark fermentation model that predicts reduction of this gas when glucose is converted into organic products different from acetate.     

Co-producing hydrogen and methane from higher-concentration of corn stalk by combining hydrogen fermentation and anaerobic digestion

In this paper, the high concentration of corn stalk (60 g/L) was employed as feedstock to produce bio-hydrogen and methane by combining hydrogen fermentation and anaerobic digestion. In the first stage of hydrogen fermentation, the effects of several key parameters, such as strain enhancement technique, cetyl trimethyl ammonium bromide (CTAB), NH₄HCO₃ on hydrogen production from cornstalk were investigated and optimized. The maximum hydrogen yield of 79.8 ± 1.5 ml H₂/g-TS and hydrogen production rate of 3.78 ml/g-cornstalk h was observed at fixed acidizing cornstalk of 60 g/L, strains Bacillus sp. FS2011 dosage of 10%(v/v), CTAB of 30 mg/L, NH₄HCO₃ of 1.2 g/L and initial pH of 7.5 ± 0.5 at 36 ± 1 °C, respectively. In the second stage of anaerobic digestion, the effluent from hydrogen production bio-reactor was further employed as the feedstock to produce methane by methanogenic bacteria, the maximum methane yield of 227 ± 2.5 ml CH₄/g-COD and COD removal rate of 95  ± 1% was recorded. The interesting observations were that the total amount of the organic wastewater produced in a higher substrate concentration (60 g/l) by hydrogen fermentation was reduced by about two-thirds compared with that of traditional low substrate concentration (≤20 g/l).     

Thermophilic dark fermentation of acid hydrolyzed waste ground wheat for hydrogen gas production

Batch dark fermentation experiments were performed to investigate the effects of biomass and substrate concentration on bio-hydrogen production from acid hydrolyzed ground wheat at 55 °C. In the first set of experiments, the substrate concentration was constant at 20 g total sugar L⁻¹ and biomass concentration was varied between 0.52 and 2.58 g L⁻¹. Total sugar concentration was varied between 4.2 and 23.7 g L⁻¹ in the second set of experiments with a 1.5 g L⁻¹ constant biomass concentration. The highest cumulative hydrogen formation (582 mL, 30 °C, 1 atm), formation rate (5.43 mL h⁻¹) and final total volatile fatty acid (TVFA) concentration (6.54 g L⁻¹) were obtained with 1.32 g L⁻¹ biomass concentration. In variable substrate concentration experiments, the highest cumulative hydrogen (365 mL) and TVFA concentration (4.8 g L⁻¹) were obtained with 19.25 g L⁻¹ initial total sugar concentration while hydrogen gas formation rate (12.95 mL h⁻¹) and the yield (200 mL H₂ g⁻¹ total sugar) were the highest with 4.2 g L⁻¹ total sugar concentration.     

Effects of potential inhibitors present in dilute acid-pretreated corn stover on fermentative hydrogen production by Escherichia coli

This study reports the inhibitory compounds present in dilute sulfuric acid-pretreated corn stover for fermentative hydrogen production by Escherichia coli. The E. coli W3110-derived strain deficient in lactate and succinate production pathways was used. When dilute acid-treated corn stover liquor containing 175 mM acetate, 21 mM furfural, and 2.6 mM 5-hydroxymethylfurfural was added at fourfold dilution to the xylose-containing culture medium, the microbial growth and hydrogen production were almost completely inhibited. The addition of acetate, furfural, or 5-hydroxymethylfurfural, at up to 200, 80, or 40 mM, respectively, to the xylose- or glucose-containing medium led to dose-dependent inhibition of fermentative hydrogen production. No synergistic inhibition by furfural with acetate or 5-hydroxymethylfurfural at concentrations based on the corn stover hydrolysate was observed. Altogether, these results suggest that the inhibition of the E. coli-based hydrogen production by the corn stover hydrolysate is primarily attributed to acetate under the conditions used.     

Production costs of potential corn stover harvest and storage systems

Corn stover has potential as a bioenergy feedstock in North America. We simulated production costs for stover harvest (three-pass and two-pass with baling or chopping, and single-pass with baling or chopping) and on-farm storage (outdoor and indoor bales, outdoor wrapped bales, and chopped stover in bags, bunks, or piles). For three- and two-pass harvest, chopping was 33–45% more expensive than baling. For baling and chopping, two-pass harvest was 25% cheaper than three-pass. Single-pass chopping harvests were on average 42% cheaper than three-pass or two-pass chopping. Single-pass baling was cheaper (4–31%) than multi-pass baling at low rates of stover collection, but more expensive (1–39%) at high rates of collection. For bales, outdoor storage of wrapped bales was cheapest. Outdoor, unwrapped bale storage, even with 12% dry matter loss, was cheaper than indoor storage. For chopped stover, storage in bags was always cheapest, followed by piles, and then bunkers. With harvest and storage together, there were four least cost systems: single-pass, ear-snap baling with wrapped bale storage; single-pass chopping with silage bag storage; and two-pass baling with wrapped-bale storage. A second group of harvest/storage systems was 25% more expensive, including single-pass, whole-plant baling with wrapped-bale storage; two-pass chopping with silage-bag storage; and three-pass baling with wrapped-bale storage. The three-pass chop harvest with silage bag storage was most expensive. Our analysis suggests all harvest and farm storage systems have tradeoffs and several systems can be economically and logistically viable.     

Enhanced bio-hydrogen production from corn stalk by anaerobic fermentation using response surface methodology

The key process parameters of solid state enzymolysis for the generation of soluble sugar (SS) and bio-hydrogen production from corn stalk were optimized by the response surface methodology (RSM) based on a three factor-five level central composite design (CCD), respectively. The result showed that the optimal solid state enzymolysis condition from corn stalk was 47.7 °C, SCED of 0.054 g/g and 10.3 days for the maximum SS yield of 526 mg/g-TVS. Correspondingly, the optimal enzymolysis conditions from corn stalk appeared at 46.3 °C, SCED of 0.049 g/g and 7.5 days for the maximum hydrogen yield of 205.5 mL/g-TVS from the hydrolyzed substrate by the next dark fermentation. In addition, the bio-hydrogen production mechanism from corn stalk was preliminary investigated by XRD and SEM analyses. The results suggested that the solid state enzymolysis of substrate played a vital role in the effective conversion of corn stalk into bio-hydrogen by dark fermentation.