Hydrogen production from alcohol wastewater by an anaerobic sequencing batch reactor under thermophilic operation: Nitrogen and phosphorous uptakes and transformation

The objective of this study was to investigate hydrogen production from alcohol wastewater using an anaerobic sequencing batch reactor (ASBR) under thermophilic operation and at a constant pH of 5.5. Under the optimum COD loading rate of 68 kg/m³d, the produced gas contained 43% H₂ without methane and the system provided a hydrogen yield and specific hydrogen production rate of 130 ml H₂/g COD removed and 2100 ml H₂/l d, respectively, which were much higher than those obtained under the mesophilic operation. Under thermophilic operation, both nitrogen and phosphate uptakes were minimal at the optimum COD loading rate for hydrogen production and most nitrogen uptake was derived from organic nitrogen. Under the thermophilic operation for hydrogen production, the nutrient requirement in terms of COD:N:P was found to be 100:6:0.5, which was much higher than that for the methenogenic step for methane production under both thermophilic and mesophilic operations and for the acidogenic step for hydrogen production under mesophilic operation.     

Enzymatic hydrolysis and anaerobic biological treatment of fish industry effluent: Evaluation of the mesophilic and thermophilic conditions

Enzymatic hydrolysis and anaerobic treatment of effluent similar to that generated in the fish processing industry were evaluated at 30 °C and 50 °C. Hydrolysis used lipase produced by fungus Penicillium simplicissimum in solid state fermentation with babassu cake as substrate, which has optimal activity at 50 °C. Hydrolysis kinetics was conducted with mixtures of effluent (containing 1500 mg oils and greases/L) and different lipase activities (0–0.67 U/ml of effluent), verifying that with 0.16 U/ml of effluent, 9.69 μmol/ml of free acids were produced after 4 h at 50 °C. Anaerobic biodegradation assays were conducted with effluent submitted to three different treatments: thermophilic (hydrolysis and anaerobic treatment at 50 °C), mesophilic (hydrolysis and anaerobic treatment at 30 °C) and hybrid (hydrolysis at 50 °C and anaerobic treatment at 30 °C). The best results (97.5% of chemical oxygen demand [COD] removal and 105.4 ml CH₄/g COD_removed) were obtained with the hybrid treatment in only 68 h. The thermophilic hydrolysis not only reduced the amount of enzyme and the hydrolysis time but also reduced the time and the cost of mesophilic anaerobic treatment, favoring the application of this treatment on an industrial scale.     

Simultaneous saccharification and fermentation of cellulose for bio-hydrogen production by anaerobic mixed cultures in elephant dung

The objective of this study was to optimize the culture conditions for simultaneous saccharification and fermentation (SSF) of cellulose for bio-hydrogen production by anaerobic mixed cultures in elephant dung under thermophilic temperature. Carboxymethyl cellulose (CMC) was used as the model substrate. The investigated parameters included initial pH, temperature and substrate concentration. The experimental results showed that maximum hydrogen yield (HY) and hydrogen production rate (HPR) of 7.22 ± 0.62 mmol H₂/g CMC_added and 73.4 ± 3.8 mL H₂/L h, respectively, were achieved at an initial pH of 7.0, temperature of 55 °C and CMC concentration of 0.25 g/L. The optimum conditions were then used to produce hydrogen from the cellulose fraction of sugarcane bagasse (SCB) at a concentration of 0.40 g/L (equivalent to 0.25 g/L cellulose) in which an HY of 7.10 ± 3.22 mmol H₂/g cellulose_added. The pre-dominant hydrogen producers analyzed by polymerase chain reaction-denaturing gel gradient electrophoresis (PCR-DGGE) were Thermoanaerobacterium thermosaccharolyticum and Clostridium sp. The lower HY obtained when the cellulose fraction of SCB was used as the substrate might be due to the presence of lignin in the SCB as well as the presence of Lactobacillus parabuchneri and Lactobacillus rhamnosus in the hydrogen fermentation broth.     

Continuous thermophilic hydrogen production and microbial community analysis from anaerobic digestion of diluted sugar cane stillage

The aim of this study was to promote biohydrogen production in an thermophilic anaerobic fluidized bed reactor (AFBR) at 55 °C using a mixture of sugar cane stillage and glucose at approximately 5000–5300 mg COD L⁻¹. During a reduction in the hydraulic retention time (HRT) from 8, 6, 4, 2 and 1 h, H₂ yields of 5.73 mmol g COD_added⁻¹ were achieved (at HRT of 4 h, with organic loading rate of 52.7 kg COD m⁻³ d⁻¹). The maximum volumetric H₂ production of 0.78 L H₂ h⁻¹ L⁻¹ was achieved using stillage as carbon source. In all operational phases, the H₂ average content in the biogas was between 31.4 and 52.0%. Butyric fermentation was the predominant metabolic pathway. The microbial community in accordance with the DGGE bands profile was found similarity coefficient between 91 and 95% without significant changes in bacterial populations after co-substrate removal. Bacteria like Thermoanaerobacterium sp. and Clostridium sp. were identified.     

Flux balance analysis of mixed anaerobic microbial communities: Effects of linoleic acid (LA) and pH on biohydrogen production

The internal fluxes of mixed anaerobic cultures fed 2000 mg l⁻¹ linoleic acid (LA) plus glucose at 6 initial pH conditions and maintained at 37 °C were estimated using a flux balanced analysis (FBA). In cultures fed LA at pH 7, less than 8% of the flux was diverted to CH₄. At an initial pH ≥ 5.5, the quantity of glucose removed was greater than 95%; however, at pH 4.5 and 5.0 the quantity consumed were 38% and 75%, respectively. The FBA output showed that the acetogenic H₂-consumers were responsible for more than 20% of the H₂ consumed. Adding LA and decreasing the pH was ineffective in reducing the activity of acetogenic H₂-consumers. As the initial pH decreased, the acetogenic H₂-consuming flux decreased in the presence of 2000 mg l⁻¹ LA. A maximum H₂ yield of 1.55 mol mol⁻¹ glucose consumed (peak hydrogenase flux (R12)) was attained when the acetogenic H₂-consuming flux reached 0.42 mol at a pH of 5.5.     

Performance and composition of bacterial communities in anaerobic fluidized bed reactors for hydrogen production: Effects of organic loading rate and alkalinity

This study evaluated the effects of the organic loading rate (OLR) and pH buffer addition on hydrogen production in two anaerobic fluidized bed reactors (AFBRs) operated simultaneously. The AFBRs were fed with glucose, and expanded clay was used as support material. The reactors were operated at a temperature of 30 °C, without the addition of a buffer (AFBR1) and with the addition of a pH buffer (AFBR2, sodium bicarbonate) for OLRs ranging from 19.0 to 140.6 kg COD m⁻³ d⁻¹ (COD: chemical oxygen demand). The maximum hydrogen yields for AFBR1 and AFBR2 were 2.45 and 1.90 mol H₂ mol⁻¹ glucose (OLR of 84.3 kg COD m⁻³ d⁻¹), respectively. The highest hydrogen production rates were 0.95 and 0.76 L h⁻¹ L⁻¹ for AFBR1 and AFBR2 (OLR of 140.6 kg COD m⁻³ d⁻¹), respectively. The operating conditions in AFBR1 favored the presence of such bacteria as Clostridium, while the bacteria in AFBR2 included Clostridium, Enterobacter, Klebsiella, Veillonellaceae, Chryseobacterium, Sporolactobacillus, and Burkholderiaceae.     

Fabrication of microcrystalline silicon solar cells on a SnO2 coated substrate using seed layer insertion

We fabricated hydrogenated microcrystalline silicon (μc-Si:H) solar cells on SnO₂ coated glass using a seed layer insertion technique. Since rich hydrogen atoms from the μc-Si:H deposition process degrade the SnO₂ layer, we applied p-type hydrogenated amorphous silicon (p-a-Si:H) as a window layer. To grow the μc-Si:H layer on the p-a-Si:H window layer, we developed a seed layer insertion method. We inserted the seed layer between the p-a-Si:H layer and intrinsic bulk μc-Si:H. This seed layer consists of a thin hydrogen diluted silicon buffer layer and a naturally hydrogen profiled layer. We compared the characteristics of solar cells with and without the seed layer. When the seed layer was not applied, the fabricated cell showed the characteristics of a-Si:H solar cell whose spectral response was in a range of 400-800 nm. Using the seed layer, we achieved a μc-Si:H solar cell with performance of V_oc=0.535 V, J_sc=16.0 mA/cm², FF=0.667, and conversion efficiency=5.7% without any back reflector. The spectral response was in the range of 400-1100 nm. Also, the fabricated device has little substrate dependence, because a-Si:H has weaker substrate selectivity than μc-Si:H.     

Effects of pH value and substrate concentration on hydrogen production from the anaerobic fermentation of glucose

A series of batch experiments were conducted to investigate the effects of pH and glucose concentrations on biological hydrogen production by using the natural sludge obtained from the bed of a local river as inoculant. Batch experiments numbered series I and II were designed at an initial and constant pH of 5.0–7.0 with 1.0 increment and four different glucose concentrations (5.0, 7.5, 10 and 20 g glucose/L). The results showed that the optimal condition for anaerobic fermentative hydrogen production is 7.5 g glucose/L and constant pH 6.0 with a maximum H₂ production rate of 0.22 mol H₂ mol⁻¹ glucose h⁻¹, a cumulative H₂ yield of 1.83 mol H₂ mol⁻¹ glucose and a H₂ percentage of 63 in biogas.     

Effect of inoculum pretreatment on the microbial community structure and its performance during dark fermentation using anaerobic fluidized-bed reactors

The effect of two different inoculum pretreatments, thermal and cell wash-out (A1 and A2, respectively) on the performance of anaerobic fluidized bed reactors for hydrogen production was determined. The reactors were operated for 112 days under the same operational conditions using glucose as substrate at increasing organic loading rates and decreasing hydraulic retention times. Both treatments were effective avoiding methanogenesis. Reactor A2 showed better performance and stability than reactor A1 in each one of the different operational conditions. Cell wash-out treatment produced higher hydrogen volumetric production rates and yields than thermal treatment (7 L H₂/L-d, 3.5 mol H₂/mol hexose, respectively). DGGE analysis revealed that the microbial communities developed were affected by the inoculum treatment. Organisms from the genera Clostridium and Lactobacillus predominated in both reactors, with their relative abundances linked to hydrogen production. Resilience was observed in both reactors after a period of starvation.     

Effect of pH on continuous biohydrogen production from liquid swine manure with glucose supplement using an anaerobic sequencing batch reactor

pH is considered as one of the most important factors governing the hydrogen fermentation process. In this project, five pH levels, ranging from 4.4 to 5.6 at 0.3 increments, were tested to evaluate the pH effect on hydrogen production from swine manure supplemented with glucose in an anaerobic sequencing batch reactor system with 16 h of hydraulic retention time (HRT). The optimal hydrogen yield (1.50 mol H₂/mol glucose) was achieved at pH 5.0 when the maximum production rate of 2.25 L/d/L was obtained. Continuous hydrogen production was achieved for over 3 weeks for pH 5.0, 4.7, and 4.4, with no significant methane produced. However, as pH increased to 5.3 and 5.6, methane production was observed in the biogas with concurrent reductions in hydrogen production, indicating that methanogens could become increasingly activated for pH 5.3 or higher. Acetate, propionate, butyrate, valerate, and ethanol were the main aqueous products whose distribution was significantly affected by pH as well.     

Hydrogen production from cassava wastewater using an anaerobic sequencing batch reactor: Effects of operational parameters,COD:N ratio,and organic acid composition

In this work, hydrogen production from cassava wastewater using anaerobic sequencing batch reactors (ASBR) was investigated to determine the optimum number of cycles per day, chemical oxygen demand (COD) loading rate, and COD:N ratio. The system operated at a COD loading rate of 30 kg/m³d and 6 cycles per day provided maximum hydrogen production performance in terms of specific hydrogen production rate (SHPR) (388 ml H₂/g VSS d or 3800 ml H₂/l d) and hydrogen yield (186 ml H₂/g COD removed). The effect of nitrogen supplementation was also studied by adding NH₄HCO₃ into the system at the COD:N ratios of 100:2.2, 100:3.3, and 100:4.4 under the COD loading rate of 30 kg/m³d and 6 cycles per day. The maximum SHPR and hydrogen yield of 524 ml H₂/g VSS d (5680 ml H₂/l d) and 438 ml H₂/g COD removed, respectively, were obtained at the stoichiometric COD:N ratio of 100:2.2. An excess nitrogen was found to promote the productions of higher organic acids and ethanol, resulting in lowering hydrogen production efficiency.     

Impact of furfural on biological hydrogen production kinetics from synthetic lignocellulosic hydrolysate using mesophilic and thermophilic mixed cultures

The impact of furfural on hydrogen production and microbial growth kinetics was assessed using mixed anaerobic cultures at mesophilic and thermophilic conditions. Mesophilic experiments showed a hydrogen yield of 1.6 mol H₂/mol initial sugars at 1 g/L furfural which is a 45% enhancement from the control (0 g/L furfural) at a substrate-to-biomass ratio (S°/X°) of 4 gCOD/gVSS. On the other hand, thermophilic experiments showed no enhancement at 1 g/L furfural but rather a 53% decrease in hydrogen yield from its control. Furfural inhibition threshold limit was observed to be greater than 1 g/L for mesophilic experiments and less than 1 g/L for thermophilic experiments. In both cases, 4 g/L was the most recalcitrant furfural concentration, with propionate and lactate the most predominant soluble metabolites in the mesophilic and thermophilic experiments respectively. It was also noted that in the presence of furfural, hydrogen-producers in both mesophilic and thermophilic mixed cultures were inactivated as no hydrogen was produced until furfural was completely degraded irrespective of sugars degradation. This study also presents the kinetics of microbial growth and substrate degradation obtained using the Monod model on MATLAB^®, ignoring an inhibition term. IC₅₀ of the mesophilic and thermophilic experiments were 1.03 g/L and 0.5 g/L respectively indicating that the thermophilic hydrogen producers were more strongly affected by furfural than the mesophilic cultures.     

Effect of inhibitors on hydrogen consumption and microbial population dynamics in mixed anaerobic cultures

The impact of different chemical microbial stressors (2-bromoethanesulfonate (BES), furfural, fish oil, lauric acid (LUA) and linoleic acid (LA)) on the inhibition of mesophilic hydrogen (H₂) consumption was examined in this study. Hydrogen consumption half-life values were used to compare the extent of inhibition by the different microbial stressing agents. A statistical analysis of the percent H₂ consumed using Tukey's analysis revealed the following trend: Control > fish oil = linoleic acid (LA (C18:2)) = furfural > BES > lauric acid (LUA (C12:0). The terminal restriction fragment length polymorphism (T-RLFP) results indicated that aceticlastic methanogens (Methanosaeta sp., Methanosarcina sp.) and hydrogenotrophic methanogens (Methanococcus sp.) were inhibited by the different chemical stressing agents. Cultures fed LUA and LA had a high abundance of Clostridium sp., Clostridium propionicum and Propionibacterium acnes. In comparison, BES and furfural fed cultures contained large fractions of Clostridium sp., Eubacteria sp. and Bacteroides sp. while in the fish oil fed cultures, the dominant organism detected was Eubacteria sp. This study indicated that H₂ consumption was affected by the chemical stressing agent concentration.     

Effect of pH and sulfate concentration on hydrogen production using anaerobic mixed microflora

The effects of varying sulfate concentrations with pH on continuous fermentative hydrogen production were studied using anaerobic mixed cultures growing on a glucose substrate in a chemostat reactor. The maximum hydrogen production rate was 2.8 L/day at pH 5.5 and sulfate concentration of 3000 mg/L. Hydrogen production and residual sulfate level decreased with increasing the pH from 5.5 to 6.2. The volatile fatty acids (VFAs) and ethanol fractions in the effluent were in the order of butyric acid (HBu) > acetic acid (HAc) > ethanol > propionic acid (HPr). Fluorescence In Situ Hybridization (FISH) analysis revealed the presence of hydrogen producing bacteria (HPB) under all pH ranges while sulfate reducing bacteria (SRB) were present at pH 5.8 and 6.2. The inhibition in hydrogen production by SRB at pH 6.2 diminished entirely by lowering to pH 5.5, at which activity of SRB is substantially suppressed.     

Effects of linoleic acid and its degradation by-products on mesophilic hydrogen production using flocculated and granular mixed anaerobic cultures

The effects of linoleic acid (LA (C18:2)) and its degradation by-products on hydrogen (H₂) production were examined at 37 °C and an initial pH value of 5.0 using granular and flocculated mixed anaerobic cultures from the same source. In the flocculated cultures, the H₂ consumers were inhibited to a greater extent when compared to the granular cultures. The maximum H₂ yields were 2.52 ± 0.2 and 1.9 ± 0.2 mol mol⁻¹ glucose in the flocculated and granular cultures, respectively. The major long chain fatty acids (LCFAs) detected at which H₂ attained a maximum value were LA (750 mg L⁻¹) and myristic acid (MA) (500 mg L⁻¹).     

Effects of various pretreatment methods of anaerobic mixed microflora on biohydrogen production and the fermentation pathway of glucose

The influence of different pretreatment methods on anaerobic mixed inoculum was evaluated for selectively enriching the hydrogen (H₂) producing mixed culture using glucose as the substrate. The efficiency of H₂ yield and the glucose fermentation pathway were found to be dependent on the type of pretreatment procedure adopted on the parent inoculum. The H₂ yield could be increased by appropriate pretreatment methods including the use of heat, alkaline or acidic conditions. Heat pretreatment of the inoculum for 30 min at 80 °C increased the H₂ yield to 53.20% more than the control.When the inoculum was heat-pretreated at 80 °C and 90 °C, the glucose degraded via ethanol (HEt) and butric acid (HBu) fermentation pathways. The degradation pathways shifted to HEt and propionate (HPr) types as the heat pretreatment temperature increased to 100 °C. When the inoculum was alkali- or acid-pretreated, the fermentation pathway shifted from glucose to a combination of the HPr and HBu types. This trend became obvious as the acidity increased. As the fermentation pathway shift from the HEt type to the HPr and HBu types, the H₂ yield decreased.     

Focusing on the process diagnosis of anaerobic fermentation by a novel sensor system combining microbial fuel cell,gas flow meter and pH meter

Process diagnosis is essential to ensure anaerobic fermentation stable and efficient. Here, a novel sensor system combining microbial fuel cell (MFC), gas flow meter and pH meter was developed to evaluate its feasibility for probing the anaerobic process established on a model high-rate bioreactor. Repeated transient responses of electrical signal, proton concentration, and gas flow rate, were observed subject to external disturbances. The transient response lasted from <1 h to 6 h. In addition, MFC obtained compatible signal variations with other sensors, and biofilm MFC (MFC_Biofilm) resulted in better agreements than control MFC (MFC_Control). These results revealed that 1) the composite sensor system was capable to probe anaerobic process, suggesting a novel approach for process analysis and diagnosis of biogas or biohydrogen production; 2) the variations of sensor signals might provide more valuable information for process diagnosis than sensor signals themselves.     

Influence of linoleic acid,pH and HRT on anaerobic microbial populations and metabolic shifts in ASBRs during dark hydrogen fermentation of lignocellulosic sugars

In this study, controlling an anaerobic microbial community to increase the hydrogen (H₂) yield during the degradation of lignocelluosic sugars was accomplished by adding linoleic acid (LA) at low pH and reducing the hydraulic retention time (HRT) of an anaerobic sequencing batch reactor (ASBR). At pH 5.5 and a 1.7 d HRT, the maximum H₂ yield for LA treated cultures fed glucose or xylose reached 2.89 ± 0.18 mol mol⁻¹ and 1.94 ± 0.17 mol mol⁻¹, respectively. The major soluble metabolites at pH 5.5 with a 1.7 day HRT differed between the control and LA treated cultures. A metabolic shift toward H₂ production resulted in increased hydrogenase activity in both the xylose (13%) and glucose (34%) fed LA treated cultures relative to the controls. In addition, the Clostridia population and the H₂ yield were elevated in cultures treated with LA. A flux balance analysis for the LA treated cultures showed a reduction in homoacetogenic activity which was associated with reducing the Bacteriodes levels from 12% to 5% in the glucose fed cultures and 16% to 10% in the xylose fed cultures. Strategies for controlling the homoacetogens and optimal hydrogen production from glucose and xylose are proposed.     

Effect of hydraulic retention time on hydrogen production and chemical oxygen demand removal from tapioca wastewater using anaerobic mixed cultures in anaerobic baffled reactor (ABR)

This study evaluated hydrogen production and chemical oxygen demand removal (COD removal) from tapioca wastewater using anaerobic mixed cultures in anaerobic baffled reactor (ABR). The ABR was conducted based on the optimum condition obtained from the batch experiment, i.e. 2.25 g/L of FeSO₄ and initial pH of 9.0. The effects of the varying hydraulic retention times (HRT: 24, 18, 12, 6 and 3 h) on hydrogen production and COD removal in a continuous ABR were operated at room temperature (32.3 ± 1.5 °C). Hydrogen production rate (HPR) increased with a reduction in HRT i.e. from 164.45 ± 4.14 mL H₂/L.d (24 h HRT) to 883.19 ± 7.89 mL H₂/L.d (6 h HRT) then decreased to 748.54 ± 13.84 mL H₂/L.d (3 h HRT). COD removal increased with reduction in HRT i.e. from 14.02 ± 0.58% (24 h HRT) to 29.30 ± 0.84% (6 h HRT) then decreased to 21.97 ± 0.94% (3 h HRT). HRT of 6 h was the optimum condition for ABR operation as indicated.