Simultaneous thermophilic hydrogen production and phenol removal from palm oil mill effluent by Thermoanaerobacterium-rich sludge

Thermoanaerobacterium-rich sludge was used for hydrogen production and phenol removal from palm oil mill effluent (POME) in the presence of phenol concentration of 100–1000 mg/L. Thermoanaerobacterium-rich sludge yielded the most hydrogen of 4.2 L H₂/L-POME with 65% phenol removal efficiency at 400 mg/L phenol. Butyric acid and acetic acid were the main metabolites. The effects of oil palm ash, NH₄NO₃ and iron concentration (Fe²⁺) on hydrogen production and phenol removal efficiency from POME by Thermoanaerobacterium-rich sludge was investigated using response surface methodology (RSM). The RSM results indicated that the presence of 0.2 g Fe²⁺/L, 0.3 g/L NH₄NO₃ and 20 g/L oil palm ash in POME could improved phenol removal efficiency, with predicted hydrogen production and phenol removal efficiency of 3.45 L H₂/L-POME and 93%, respectively. In a confirmation experiment under optimized conditions highly reproducible results were obtained, with hydrogen production and phenol removal efficiency of 3.43 ± 0.12 L H₂/L-POME and 92 ± 1.5%, respectively. Simultaneous hydrogen production and phenol removal efficiency in continuous stirred tank reactor at hydraulic retention time (HRT) of 1 and 2 days were 4.0 L H₂/L-POME with 85% and 4.2 L H₂/L-POME with 92%, respectively. Phenol degrading Thermoanaerobacterium-rich sludge comprised of Thermoanaerobacterium thermosaccharolyticum, Thermoanaerobacterium aciditolerans, Desulfotomaculum sp., Bacillus coagulans and Clostridium uzonii. Phenol degrading Thermoanaerobacterium-rich sludge has great potential to harvest hydrogen from phenol-containing wastewater.     

Magnetite nanoparticles enhanced glucose anaerobic fermentation for bio-hydrogen production using an expanded granular sludge bed (EGSB) reactor

The feasibility and efficiency of magnetite nanoparticles (Fe₃O₄NPs) enhanced bio-hydrogen production from glucose anaerobic fermentation were evaluated in this study. The results demonstrated that the maximum hydrogen yield (HY) of 12.97 mL H₂/g-VSS was obtained with 50 mg/L and 40–60 nm of Fe₃O₄NPs in batch experiments. Moreover, the optimum dosage of Fe₃O₄NPs produced hydrogen production (HP) of 4.95 L H₂/d in an expanded granular sludge bed (EGSB) reactor. Fe₃O₄NPs involved could promote ethanol and acetic acid accumulation. Fe²⁺ as by-product of iron corrosion could effectively promote the activity of key coenzymes and soluble microbial products (SMPs). Importantly, Fe₃O₄NPs addition resulted in the formation of electronic conductor chains to enhance the electron transport efficiency in the granular sludge. Microbial community analysis revealed that the relative abundance of butyrate-hydrogen-producing bacteria (Clostridium) decreased from 40.55% to 11.45%, while the relative abundance of ethanol-hydrogen-producing bacteria (Acetanaerobacterium and Ethanoligenens) increased from 19.62% to 35.35% with Fe₃O₄NPs involved, confirming that the fermentation type was transformed from butyrate-type to ethanol-type, which finally facilitated more hydrogen production.     

Hydrogen production in reactors: The influence of organic loading rate,inoculum and support material

Hydrogen production was evaluated in two thermophilic structured bed (USBR) reactors. USBR1was inoculated with auto-fermented sugarcane vinasse and low-density polyethylene cubes were used as support material. USBR2 was inoculated with anaerobic sludge from an up-flow anaerobic sludge blanket (UASB) reactor treating sugarcane vinasse, and polyurethane foam matrices was used as support material. The reactors were operated in parallel with sugar cane molasses at organic loading rate (OLR) from 30 to 120 g COD L⁻¹d⁻¹ during 45 days. Hydrogen production was detected during the whole operational period, with maximum values of 1123 mL H₂ d⁻¹L⁻¹ and 2041 mL H₂ d⁻¹L⁻¹ for USBR1 and USBR2, respectively. The number of gene copies encoding for Fe-hydrogenase was higher in USBR2 for all OLR applied. DNA sequences related to Thermoanaerobacterium and Clostridium sensu stricto were predominant in USBR1. In USBR2, in addition to these microorganisms, Lactobacillus, Pseudomonas and Thermotuga, and sequences with low frequency of abundance (<5%) involved directly and indirectly in hydrogen production were also present. The taxonomical and functional more diverse inoculum of USBR2 was associated with a higher hydrogen production. Besides fermentation, an unknown metabolism was relevant in USBR2, revealing the importance of physiological characterization of the microbial community present.     

Bio-hydrogen production from thin stillage using conventional and acclimatized anaerobic digester sludge

To assess the viability of biohydrogen production from thin stillage, a comparative evaluation of anaerobic digester sludge (ADS) and acclimatized anaerobic digester sludge (AADS) for biohydrogen production over a wide range of S⁰/X⁰ ratio (0.5-8 gCOD/gVSS) was performed. A maximum hydrogen yield of 19.5 L H₂/L thin stillage was achieved for the AADS while tests with ADS achieved a maximum yield of only 7.5 L H₂/L thin stillage. The optimum range of S⁰/X⁰ ratio for hydrogen production was found to be 1 to 2 gCOD/gVSS using conventional ADS and 3 to 6 gCOD/gVSS using AADS. The biomass specific hydrogen production rate for the AADS was 3.5 times higher than rate for the ADS throughout the range of S⁰/X⁰ ratio examined in this study. The DGGE profiles of the 16S rDNA gene fragments confirmed the superior performance of the AADS over the ADS, showing that the widely known hydrogen producers Clostridium acetobutyricum, Klebsiella pneumonia, Clostridium butyricum and Clostridium pasteurianum were the predominant species.     

Microbial culture selection for bio-hydrogen production from waste ground wheat by dark fermentation

Hydrogen formation performances of different anaerobic bacteria were investigated in batch dark fermentation of waste wheat powder solution (WPS). Serum bottles containing wheat powder were inoculated with pure cultures of Clostridium acetobutylicum (CAB), Clostridium butyricum (CB), Enterobacter aerogenes (EA), heat-treated anaerobic sludge (ANS) and a mixture of those cultures (MIX). Cumulative hydrogen formation (CHF), hydrogen yield (HY) and specific hydrogen production rate (SHPR) were determined for every culture. The heat-treated anaerobic sludge was found to be the most effective culture with a cumulative hydrogen formation of 560 ml, hydrogen yield of 223 ml H₂ g⁻¹ starch and a specific hydrogen production rate of 32.1 ml H₂ g⁻¹ h⁻¹.     

Thermophilic dark fermentation of untreated rice straw using mixed cultures for hydrogen production

Biohydrogen production from untreated rice straw using different heat-treated sludge, initial cultivation pH, substrate concentration and particle size was evaluated at 55 °C. The peak hydrogen production yield of 24.8 mL/g TS was obtained with rice straw concentration 90 g TS/L, particle size <0.297 mm and heat-treated sludge S1 at pH 6.5 and 55 °C in batch test. Hydrogen production using sludge S1 resulted from acetate-type fermentation and was pH dependent. The maximum hydrogen production (P), production rate (R_m) and lag (λ) were 733 mL, 18 mL/h and 45 h respectively. Repeated-batch operation showed decreasing trend in hydrogen production probably due to overloading of substrate and its non-utilization. PCR-DGGE showed both hydrolytic and fermentative bacteria (Clostridium pasteurianum, Clostridium stercorarium and Thermoanaerobacterium saccharolyticum) in the repeated-batch reactor, which perhaps in association led to the microbial hydrolysis and fermentation of raw rice straw avoiding the pretreatment step.     

Enhancement of hydrogen production by the filamentous non-heterocystous cyanobacterium Arthrospira sp. PCC 8005

The efficiency of hydrogen production by different cyanobacterial species depends on several external factors. We report here the factors enhancing hydrogen production by filamentous non-heterocystous cyanobacterium Arthrospira sp. PCC 8005. Cells adapted to dark-anaerobic conditions produced hydrogen consistent with increased hydrogenase activity when supplemented with Fe²⁺. Stimulation of hydrogen production could be achieved by addition of reductants, either dithiothreitol or β-mercaptoethanol with higher production observed with the latter. Additionally, Fe²⁺ and β-mercaptoethanol added to nitrogen- and sulphur-deprived cells significantly stimulated H₂ production with maximal value of 5.91 ± 0.14 μmol H₂ mg Chla⁻¹ h⁻¹. Glucose and a small increase of osmolality imposed by either NaCl or sorbitol enhanced hydrogen production. High rates of hydrogen production were obtained in cells adapted in nitrogen-deprived medium with neutral and alkaline external pH, significant decrease of hydrogen production occurred under acidic external pH.     

Bioaugmentation on hydrogen production from food waste

The aim of this work was to evaluate the effect of two hydrolytic (Paenibacillus polymyxa and Bacillus subtilis) and two fermentative (Clostridium saccharobutylicum and Clostridium beijerinckii) strains on hydrogen (H₂) production in dark fermentation by batch testing. Food waste was used as a substrate, pretreated anaerobic sludge was used as the inoculum, and different concentrations of the evaluated microorganisms were used. Bioaugmentation with 3.5 × 10⁹ CFU/mL/L_reactor B. subtilis showed the best performance, obtaining a production of 84.5 mL H₂/g SV and a reduction in the lag phase (from 7.9 h in control to 3.5 h). Bioaugmentation with B. subtilis in an anaerobic sequencing batch reactor exhibited a significant effect on volumetric productivity, reaching a maximal increase of 344% of H₂ production in comparison with that obtained without the addition of the strain. The increase in H₂ was observed in a short period of time (4 cycles), after which H₂ production returned to the original H₂ production baseline. During all reactor operations, the main volatile fatty acids produced were acetic acid and butyric acid. Microbial community analysis when bioaugmentation was applied showed an importance of lactic acid bacteria abundance, such as that of Bifidobacterium and Lactobacillus, whose metabolic activity was crucial in reactor performance. The added concentration of microorganisms is a critical parameter for the bioaugmentation process.     

Biohydrogen production from waste glycerol and sludge by anaerobic mixed cultures

Key factors affecting biohydrogen production from waste glycerol and sludge by anaerobic mixed cultures were optimized using response surface methodology (RSM) with central composite design (CCD). Investigated parameters were waste glycerol concentration, sludge concentration, and the amount of Endo–nutrient addition. Concentrations of waste glycerol and sludge had a significant individual effect on hydrogen production rate (HPR) (p ≤ 0.05). The interactive effect on HPR (p ≤ 0.05) was found between waste glycerol concentration and sludge concentration. The optimal conditions for the maximum HPR were: waste glycerol concentration 22.19 g/L, sludge concentration 7.16 g-total solid (TS/L), and the amount of Endo–nutrient addition 2.89 mL/L in which the maximum HPR of 1.37 mmol H₂/L h was achieved. Using the optimal conditions, HPR from a co-digestion of waste glycerol and sludge (1.37 mmol H₂/L h) was two times greater than the control (waste glycerol without addition of sludge) (0.76 mmol H₂/L h), indicating a significant enhancement of HPR by sludge. Major metabolites of the fermentation process were ethanol, 1,3-propanediol (1,3-PD), lactate, and formate.     

Effect of initial pH,nutrients and temperature on hydrogen production from palm oil mill effluent using thermotolerant consortia and corresponding microbial communities

Thermotolerant consortia were obtained by heat-shock treatment on seed sludge from palm oil mill. Effect of the initial pH (4.5–6.5) on fermentative hydrogen production palm oil mill effluent (POME) showed the optimum pH at 6.0, with the maximum hydrogen production potential of 702.52 mL/L-POME, production rate of 74.54 mL/L/h. Nutrients optimization was investigated by response surface methodology with central composite design (CCD). The optimum nutrients contained 0.25 g urea/L, 0.02 g Na₂HPO₄/L and 0.36 g FeSO₄·7H₂O/L, giving the predicted value of hydrogen production of 1075 mL/L-POME. Validation experiment revealed the actual hydrogen production of 968 mL/L-POME. Studies on the effect of temperature (25–55 °C) revealed that the maximum hydrogen production potential (985.3 mL/L-POME), hydrogen production rate (75.99 mL/L/h) and hydrogen yield (27.09 mL/g COD) were achieved at 55, 45 and 37 °C, respectively. Corresponding microbial community determined by the DGGE profile demonstrated that Clostridium spp. was the dominant species. Clostridium paraputrificum was the only dominant bacterium presented in all temperatures tested, indicating that the strain was thermotolerant.     

Hydrogen production from sludge of poultry slaughterhouse wastewater treatment plant pretreated with microwave

This study aims to produce hydrogen from sludge of poultry slaughterhouse wastewater treatment plant (5% total solid) by anaerobic batch fermentation with Enterobactor aerogenes or mixed cultures from hot spring sediment as the inoculums. Sludge was heated in microwave at 850 W for 3 min. Results indicated that a soluble chemical oxygen demand (sCOD) of pretreated sludge was higher than that of raw sludge. Pretreated sludge inoculated with E. aerogenes and supplemented with the Endo nutrient had a higher hydrogen yield (12.77 mL H₂/g tCOD) than the raw sludge (0.18 mL H₂/g tCOD). When considered the hydrogen yield, the optimum initial pH for hydrogen production from microwave pretreated sludge was 5.5 giving the maximum value of 12.77 mL H₂/g tCOD. However, when considered the hydrogen production rate (R_m), the optimum pH for hydrogen production would be 9.0 with the maximum R_m of 22.80 mL H₂/L sludge·h.     

The effects of seed sludge and hydraulic retention time on the production of hydrogen from a cassava processing wastewater and glucose mixture in an anaerobic fluidized bed reactor

The potential for co-fermentation of a cassava processing wastewater and glucose mixture was studied in anaerobic fluidized bed reactors. The effects of different hydraulic retention times (HRTs) (10–2 h) and varying sources of inoculum are reported. The sludge from a UASB reactor that had been used to treat poultry slaughterhouse wastewater (SP) resulted in the highest yields of hydrogen (HY) and ethanol (EtOHY) of 1.0 mmol H₂ g⁻¹ COD (10 h) and 3.0 mmol EtOH g⁻¹ COD (6 h). The sludge from a UASB reactor used for the treatment of swine wastewater (SW) resulted in a maximum HY of 0.65 mmol H₂ g⁻¹ COD (6 h) and EtOHY of 2.1 mmol g⁻¹ COD (10 and 8 h). Methane was produced with a maximum production of 9.68 L CH₄ d⁻¹ L⁻¹. Based on phylogenetic analysis of 16S rRNA, bacteria and methanogenic archaea similar to Lactobacillus and Methanobacterium, respectively, were identified.     

Effects of Fe0 and Ni0 nanoparticles on hydrogen production from cotton stalk hydrolysate using Klebsiella sp. WL1316: Evaluation of size and concentration of the nanoparticles

Fe⁰ and Ni⁰ nanoparticles (NPs) of certain size were synthesized and added to the hydrogen production system from cotton stalk hydrolysate using Klebsiella sp. WL1316. Fe⁰ and Ni⁰ NPs with a size of 50 nm at all concentrations effectively improve hydrogen production during mid to late fermentation stages; particularly, the highest daily hydrogen production obtained following treatment with 50 nm Fe⁰ NPs at 30 mg/L fermented for 96 h significantly increased by 61% comparing to the control treatment. The reducing sugar consumption in cotton stalk hydrolysate and ΔOD₆₀₀ could be improved to some extent by Fe⁰ and Ni⁰ NPs supplementation. Addition of Fe⁰ or Ni⁰ NPs of 50 nm at a concentration of 30 mg/L resulted in enhanced cumulative hydrogen production with improvement of hydrogen yield reached higher than 20%, and the values of Y(H₂/S) were all higher than 90 mL/g _substrate, reflecting good hydrogen production and substrate consumption. The analysis of the main soluble metabolites profile revealed that supplementation with Fe⁰ and Ni⁰ NPs of suitable size and concentration may decrease the metabolic flux in the competitive branch of hydrogen production and increase the metabolic flux of the key node that leads to hydrogen generation, thus promoting biohydrogen synthesis.     

Effects of rice husk particle size on biohydrogen production under solid state fermentation

As a renewable energy source bio-hydrogen production from lignocellulosic wastes is a promising approach which can produce clean fuel with no CO₂ emissions. Utilization of agro-industrial residues in solid state fermentation (SSF) is offering a solution to solid wastes disposal and providing an economical process of value-added products such as hydrogen.In this study three different particle size of rice husk (<2000 μm, <300 μm, <74 μm) was subjected to batch SSF with a Clostridium termitidis: Clostridium intestinale ratio of 5:1. C. termitidis is a cellulolytic microorganism that has the ability to hydrolyze cellulosic substances and C. intestinale is able to grow on glucose having a potential of enhancing hydrogen production when used in the co-culture. 5 g dw rice husk with 75% humidity was used as substrate in SSF under mesophilic conditions. The highest HF Volume (29.26 mL) and the highest yield (5.9 mL H₂ g⁻¹ substrate) were obtained with the smallest particle size (<74 μm). The main metabolites obtained from the fermentation media were acetic, butyric, propionic and lactic acids. The second best production yield (3.99 mL H₂ g⁻¹ substrate) was obtained with the middle particle size (<300 μm) rice husk with a HF of 19.71 mL.     

Enhanced hydrogen production from sewage sludge by cofermentation with wine vinasse

The cofermentation of sewage sludge and wine vinasse at different mixing ratios to enhance hydrogen production was investigated. Batch experiments were carried out under thermophilic conditions with thermophilic sludge inoculum obtained from an acidogenic anaerobic reactor. The results showed that the addition of wine vinasse enhances the hydrogen production of sewage sludge fermentation. The highest hydrogen yields, 41.16 ± 3.57 and 43.25 ± 1.52 mL H₂/g VS_added, were obtained at sludge:vinasse ratios of 50:50 and 25:75, respectively. These yields were 13 and 14 times higher than that obtained in the monofermentation of sludge (3.17 ± 1.28 mL H₂/g VS_added). The highest VS removal (37%) was obtained at a mixing ratio of 25:75. Cofermentation had a synergistic effect the hydrogen yield obtained at a sludge:vinasse ratio of 50:50 was 40% higher, comparing to the sum of each waste. Furthermore, kinetic analysis showed that Cone and first-order kinetic models fitted hydrogen production better than the modified Gompertz model.     

Co-hydrothermal gasification of microbial sludge and algae Kappaphycus alvarezii for bio-hydrogen production: Study on aqueous phase reforming

In this study, wastewater obtained from a sewage treatment plant was treated successively by using microbial consortium and macroalgae Kappaphycus alvarezii to generate microbial sludge and algal biomass. The production of green fuel was carried out via co-gasification of microbial sludge and macroalgae Kappaphycus alvarezii for a duration of 60 min, feedstock to solvent ratio (5 to 20 g of feedstock in 200 mL), sludge to algae ratio (ranging from 1:1 to 3:1) and temperature (300–400 °C) respectively. Maximum bio-hydrogen yield was 36.1% and methane yield was 38.4% at a temperature of 360 °C at a feedstock to solvent ratio of 15:200 g/mL and sludge to algae ratio of 2:1 individually. The liquid by product of co-gasification process was later subjected to photocatalytic reforming, resulted in an enhanced hydrogen composition of 61.25%.     

Biohydrogen production from Enterobacter cloacae IIT-BT 08 using distillery effluent

Distillery effluent poses severe environmental pollution problem mainly due to its high organic content. During alcohol fermentation, most of the essential macro- and micro-nutrients get utilized. Therefore, supplementation of these nutrients becomes imperative for the improvement of biohydrogen production. In the present study, starch based distillery effluent was used for dark fermentative hydrogen production using Enterobacter cloacae IIT-BT 08. Hence, this study was undertaken to evaluate the effect of supplementation of yeast extract, malt extract, Fe⁺⁺, Cu⁺⁺ and Mg⁺⁺ on biohydrogen production. The interaction among supplements and their mutual effect on the hydrogen production was studied using five factor–five level central composite design). Optimum hydrogen yield of 7.4 mol H₂/kg COD_reduced was predicted by the model, which showed an excellent correlation with experimental hydrogen yield of 7.38 ± 0.24 mol H₂/kg COD_reduced. An average hydrogen production rate of 80 mL/L h was achieved after supplementation, having 2.2 times higher hydrogen yield as compared to non-supplemented distillery effluent.     

Effects of pretreatment of natural bacterial source and raw material on fermentative biohydrogen production

Effects of pretreatment of natural bacterial source and raw material on biohydrogen production in fermentative biohydrogen production process were investigated systematically. Biohydrogen production from stale corn slurry was found to be feasible and effective by A1 (water soak) pretreated compost and B1 (gelatinization) pretreated stale corn. The results were further confirmed by the verification test with working volume of 8 L, in which the maximum hydrogen yield of 262 mL H₂/g-substrate, hydrogen production rate of 39 mL/g h⁻¹ and the corresponding hydrogen content of 50% was observed at fixed substrate concentration of 10 g/L, working pH 5.0-5.5 and 36 ± 1 °C. The effluent was mostly composed of acetate and butyrate. Subsequently, two new hydrogen producing strains were isolated from the effluent sludge in the running bioreactor, and they were preliminarily identified as Clostridium and Enterobacter, respectively, according to the routine screening examinations.