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A coupled system of dark fermentation and microalgal cultivation was applied to investigate the hydrogen and lipid production potentials from three main substrates of food waste. Compared with waste proteins and waste fats, waste carbohydrates were more appropriate for energy production, which exhibited the highest cumulative hydrogen volume of 668.3 ± 36.5 mL L −1 and hydrogen yield of 133.66 ± 7.31 mL g −1 substrate. Most by-products of dark fermentation were consumed in algal culture process, and the maximum lipid content and lipid productivity reached 33.2 ± 1.8% and 52.6 ± 1.1 mg L −1  d −1 , respectively. The energy conversion efficiency significantly enhanced from 10.14% (dark fermentation) to 24.06% (coupled system). In continuous operation, energy production was steady and efficient, and the average hydrogen production rate of 741.4 mL L −1  d −1 and average lipid concentration of 0.4 g L −1 were obtained. This study provides a promising and sustainable route for efficient energy recovery from waste substrates by using coupled hydrogen and lipid production system in long-term operation. [ABSTRACT FROM AUTHOR]

Poor flocculation of photo fermentative bacteria resulting in continuous biomass washout from photobioreactor is a critical challenge to achieve rapid and stable hydrogen production. In this work, the aggregation of Rhodopseudomonas faecalis RLD-53 was successfully developed in a photobioreactor and the effects of different carbon sources on hydrogen production and aggregation ability were investigated. Extracellular polymeric substances (EPS) production by R. faecalis RLD-53 cultivated using different carbon sources were stimulated by addition of L-cysteine. The absolute ζ potentials of R. faecalis RLD-53 were considerably decreased with addition of L-cysteine, and aggregation barriers based on DLVO dropped to 15-43 % of that in control groups. Thus, R. faecalis RLD-53 flocculated effectively, and aggregation abilities of strain RLD-53 cultivated with acetate, propionate, lactate and malate reached 29.35, 32.34, 26.07 and 24.86 %, respectively. In the continuous test, hydrogen-producing activity was also promoted and reached 2.45 mol H/mol lactate, 3.87 mol H/mol propionate and 5.10 mol H/mol malate, respectively. Therefore, the aggregation of R. faecalis RLD-53 induced by L-cysteine is independent on the substrate types, which ensures the wide application of this technology to enhance hydrogen recovery from wastewater dominated by different organic substrates. [ABSTRACT FROM AUTHOR]

Waste activated sludge is a valuable resource containing multiple nutrients, but is currently treated and disposed of as an important source of pollution. In this work, waste activated sludge after ultrasound pretreatment was reused as multiple nutrients for biofuel production. The nutrients trapped in sludge floc were transferred into liquid medium by ultrasonic disintegration during first 30 min, while further increase of pretreatment time only resulted in slight increase of nutrients release. Hydrogen production by Ethanoligenens harbinense B49 from glucose significantly increased with the concentration of ultrasonic sludge, and reached maximum yield of 1.97 mol H 2 /mol glucose at sludge concentration of 7.75 g volatile suspended solids/l. Without addition of any other chemicals, waste molasses rich in carbohydrate was efficiently turned into hydrogen with yield of 189.34 ml H 2 /g total sugar by E. harbinense B49 using ultrasonic sludge as nutrients. The results also showed that hydrogen production using pretreated sludge as multiple nutrients was higher than those using standard nutrients. Acetic acid produced by E. harbinense B49 together with the residual nutrients in the liquid medium were further converted into hydrogen (271.36 ml H 2 /g total sugar) by Rhodopseudomonas faecalis RLD-53 through photo fermentation, while ethanol was the sole end product with yield of 220.26 mg/g total sugar. Thus, pretreated sludge was an efficient nutrients source for biofuel production, which could replace the standard nutrients. This research provided a novel strategy to achieve environmental friendly sludge disposal and simultaneous efficient biofuel recovery from organic waste. [ABSTRACT FROM AUTHOR]

Anaerobic sludge (AS) and microalgae were co-cultured to enhance the energy conversion and nutrients removal from starch wastewater. Mixed ratio, starch concentration and initial pH played critical roles on the hydrogen and lipid production of the co-culture system. The maximum hydrogen production of 1508.3 mL L −1 and total lipid concentration of 0.36 g L −1 were obtained under the optimized mixed ratio (algae:AS) of 30:1, starch concentration of 6 g L −1 and initial pH of 8. The main soluble metabolites in dark fermentation were acetate and butyrate, most of which can be consumed in co-cultivation. When sweet potato starch wastewater was used as the substrate, the highest COD, TN and TP removal and energy conversion efficiencies reached 80.5%, 88.7%, 80.1% and 34.2%, which were 176%, 178%, 200% and 119% higher than that of the control group (dark fermentation), respectively. This research provided a novel approach and achieved efficient simultaneous energy recovery and nutrients removal from starch wastewater by the co-culture system. [ABSTRACT FROM AUTHOR]

A photo-fermentative bacterium for hydrogen production, Rhodopseudomonas sp. nov. strain A7, was isolated from sludge in a running bioreactor. The strain A7 has a flagellum and capsule. The hydrogen production performance exhibited by the strain A7 under several key parameters was investigated in batch culture. The maximum hydrogen yield achieved was160 ml-H 2 /60 ml-vessel using 60 mmol/l acetate, 15 mmol/l glutamate at a pH 7.0 and an inoculum time of 14 h. This study determined that the strain A7 can utilize glucose and, especially, fructose and sucrose for cell growth and hydrogen production. The hydrogen yield obtained with these substrates reached 33, 30 and 43 ml-H 2 /60 ml-vessel, respectively. These results imply that the strain A7 possesses a wide range of substrate utilization capabilities and is a potential candidate for photo-hydrogen production. [ABSTRACT FROM AUTHOR]

Poor hydrogen production performance and low biomass limit the practical application of photo-fermentation. To improve the immobilization capability of bacteria and hydrogen production performance, activated carbon fibers ( ACFs) were modified by acidic, alkaline, and neutral solutions. The modified ACFs were further used in the anaerobic fluidized bed photo-reactor ( AFBPR) to explore its continuous operation characteristics. Results showed that among the three reagents, nitric acid was the most efficient for ACF modification, and the maximum yield and production rate of hydrogen increased between about 33.6% and 65.8% compared to the control. Furthermore, with the optimal influent glutamate concentration (10 mmol L−1) and light intensity (4000 lux), the AFBPR gave efficient and stable performance with hydrogen yield of 2.26 mol H2 mol−1 acetate and hydrogen production rate of 25.8 mL L−1 h−1. The results showed the potential of using the AFBPR with HNO3-modified ACF carriers for the large-scale production of bio-hydrogen. [ABSTRACT FROM AUTHOR]

The practical application of photofermentative hydrogen production process has been generally limited by the low feedstock conversion efficiency. In this work, material flow analysis ( MFA) was applied into continuous photohydrogen to investigate the material flow of feedstock. Maximum feedstock utilization for hydrogen production (31.87%) was obtained at an hydraulic retention time of 60 h and influent acetate concentration of 40 mmol l−1, corresponding to maximum hydrogen yield of 1.89 mol H2 mol−1 acetate. MFA showed that due to the poor flocculation, photofermentative bacteria ( PFB) cannot be efficiently separated from supernatant and rushed out with effluent continuously. To replenish the biomass washout, most of feedstock was continuously utilized for cell growth rather than hydrogen production, which caused low feedstock conversion efficiency. The results showed that poor flocculation of PFB resulting in low biomass retention capacity of photobioreactor was the origin of low feedstock conversion efficiency. Therefore, enhancing flocculation of PFB and using high biomass retaining operation modes are essential to further develop photofermentative hydrogen production process. [ABSTRACT FROM AUTHOR]

Highlights: [•] Dark fermentation and algal cultivation were combined to produce H2 and lipid. [•] The sequential system successfully converted wastewater into bioenergy. [•] The performance of the two-stage process was evaluated for the first time. [•] Energy conversion efficiency raised from 18.6% (single stage) to 37.4% (two-stage). [ABSTRACT FROM AUTHOR]

Abstract: The applications of photo-fermentative bacteria (PFB) for continuous hydrogen production are generally subjected to a serious biomass washout from photobioreactor, resulting from poor flocculation of PFB. In this study, through reducing the absolute ζ-potentials of PFB, Ca2+ greatly decreased total interaction energy barrier of PFB based on DLVO theory, thus promoted the bioflocculation of Rhodopseudomonas faecalis RLD-53. Average floc size of PFB increased with the Ca2+ concentration, and reached maximum 30.07 μm at 4 mmol/l. Consequently, biomass retention capacity of photobioreactor significantly enhanced after 30 min settling with half working volume discharge. In the continuous photo-fermentative sequencing batch reactor, compared with the free cell culture, bioflocculation reached a higher steady-state hydrogen production rate of 879 ml H2/l/d and hydrogen yield of 2.64 mol H2/mol acetate, respectively. Therefore, bioflocculation promoted by calcium ion was an effective strategy for retaining PFB in photobioreactor to produce hydrogen continuously. [Copyright &y& Elsevier]

Abstract: In this work, the kinetic characterization of hydrogen production by the photofermentative bacteria Rhodopseudomonas faecalis RLD-53 was investigated at different growth phase. During entire fermentation, 89.30% of total biomass was accumulated in exponential growth phase, while hydrogen yield was only 1.82 mol H2/mol acetate at the expense of 51.25% substrate. In the stationary phase, biomass synthesis was minimal (7.51%), and 38.17% of the substrate was directly converted into hydrogen. As a result, hydrogen (59.19%) was mainly produced in stationary phase with highest hydrogen yield of 3.67 mol H2/mol acetate. Consequently, bacteria in stationary phase were most effective for hydrogen production. Based on these findings, a novel membrane photobioreactor was developed to retain bacteria during stationary phase in reactor through membrane separation. Maximum rate (32.82 ml/l/h) and yield (3.27 mol H2/mol acetate) of hydrogen production were achieved using membrane photobioreactor under the continuous operation. Therefore, using bacteria in stationary phase as hydrogen producer can offer considerable benefits for enhancing photo-hydrogen production. [Copyright &y& Elsevier]

Abstract: A novel photo-fermentative sequencing batch reactor (PFSBR) process assisted by activated carbon fibers (ACFs) was used to continuously produce hydrogen gas by Rhodopseudomonas faecalis RLD-53. Feasibility of continuous hydrogen production in PFSBR operation at different hydraulic retention times (HRTs) (48, 96, 144 and 192 h) and influent acetate concentrations (20, 40, 60 and 80 mmol/l) was investigated. The rate and yield of hydrogen production increased with HRTs from 48 to 144 h, and then decreased with the HRTs from 144 to 192 h. Regulation of the proper influent acetate concentration (60 mmol/l) not only increased hydrogen production by PFSBR, but also maintained quality of the effluent with high substrate removal efficiency (97.70%). Free R. faecalis RLD-53 was adsorbed on the surface of ACFs, initially isolated cells, then monolayer, and finally mature biofilm with three dimensional multilayers structures. The PFSBR reached a maximum hydrogen yield (3.12 mol H2/mol acetate), and achieved a steady state when mature biofilm developed on ACFs. Therefore, photo-fermentative sequencing batch reactor was a promising process for continuous photo-fermentative hydrogen production. [Copyright &y& Elsevier]

Abstract: Activated carbon fibers (ACFs) were firstly applied as fluidized solid carrier to immobilize photo-fermentative bacteria (PFB) for H2 production in batch culture. The observations by scanning electronic microscopy (SEM) demonstrated the close interaction between ACFs and PFB. The amount of immobilized bacteria and the performance of H2 production were strongly affected by specific surface area, length and amount of ACFs, respectively. Large specific surface area provided more surface attachment sites and more PFB were immobilized. ACFs with proper length avoided intertwining with each other and better fluidized during reactor operation. Excessive amount of ACFs not only limited the light conversion efficiency, but also increased biofilm detachment, resulting in low H2 yield. The maximum yield (3.08 mol H2 mol−1 acetate) and rate (32.85 ml l−1 h−1) of H2 production were obtained, using specific surface area (1500 m2 g−1), length (1 mm) and amount (0.8 g l−1) of ACFs. Compared with the conventional solid carriers, ACFs were effective solid carriers to immobilize PFB for improving H2 production, due to bacteria immobilized on the external surface of fluidized ACFs and formed a layer of dense biofilm. [Copyright &y& Elsevier]

Abstract: This study investigated the effect of EDTA concentration in medium on the growth, hydrogen production and nitrogenase activity of Rhodopseudomonas faecalis RLD-53 by batch cultures. Experimental results indicated that bacterial growth and hydrogen production were strongly inhibited with EDTA concentration increasing to 0.6–0.7 g/L. However, the lag time of hydrogen production and the trends of biomass at EDTA concentration of 0–0.5 g/L were similar. The maximum cumulative hydrogen volume of 3325 ml H2/L culture, hydrogen production rate of 27.6 ml H2/L/h and hydrogen yield of 2.97 mol H2/mol acetate were obtained when EDTA concentration was at 0.3 g/L, and the maximum OD660 attained 3.83. And the nitrogenase activity also reached a maximum value of 1331.9 μl-C2H4/h/mg dry weight in the medium containing Fe2+ and EDTA. These results showed that a proper concentration of EDTA can promote the availability of iron, thereby further enhancing the activity of nitrogenase and photo-hydrogen production. [Copyright &y& Elsevier]

Abstract: In this work, a novel bio-carrier was used to immobilize photo-fermentative bacteria for hydrogen production. The results showed that the bacterial immobilization and hydrogen production were strongly affected by particle size, amount of bio-carrier and light intensity. Controlling the proper size of bio-carrier not only prevented light shading effect from each other, but also made solid carriers better fluidized during operation. The scanning electronic microscopy revealed that the biofilm formed by photo-fermentative bacteria on the surface of bio-carrier enhanced hydrogen production. Because of bio-carrier fluidization, each immobilized bacterium could receive light energy and produce hydrogen. With the optimal particle size (2 × 2 mm), amount (3% weight to volume ratio) and light intensity (6000 lux), the maximum hydrogen yield of 3.24 mol H2/mol acetate and production rate of 36.06 ml/l/h were obtained in continuous operation stage. Bio-carrier was an effective solid carrier to immobilize photo-fermentative bacteria for improving hydrogen production. [Copyright &y& Elsevier]

Abstract: In this study, some key factors, for example, diluted ratio of effluents, the ratio of dark-photo bacteria, light intensity and light/dark cycle influencing hydrogen production by combining Clostridium butyricum and immobilized Rhodopseudomonas faecalis RLD-53 in batch culture, were investigated. Experimental results showed the photo-hydrogen yield decreased when increasing diluted ratio from 1:0.5 to 1:3, and it reached the maximum value of 4368ml-H2/l-effluents at the ratio of 1:0.5. When the ratio of dark-photo bacteria was at 1:2, the hydrogen yield reached highest value of 4.946mol-H2/mol-glucose and cumulative hydrogen volume was 5357ml-H2/l-culture during the combination process. When the light intensity was at 10.25W/m2, the hydrogen volume of photo-fermentation and the combination process reached maximum value of 4260ml-H2/l-effluents and 5892ml-H2/l-culture, respectively. During the combination process, maximum total hydrogen yield was 5.374mol-H2/mol-glucose. Meanwhile, hydrogen production under light/dark cycle was evaluated. [Copyright &y& Elsevier]

Abstract: Clostridium butyricum and Rhodopseudomonas faecalis RLD-53 were employed to produce hydrogen in mixed culture with glucose as sole substrate. Due to the great difference on growth rate and acid-resistant capacity between photo-fermentative bacteria and dark-fermentative bacteria, directly mixed culture of the two kinds of bacteria in different ratio was studied in this work. Hydrogen yield, volatile acids, pH and biomass in different periods were evaluated. Acetic acid and butyric acid produced by C. butyricum were dominant terminal fermentation products, and they were effective substrates for photo-fermentative bacteria. The cooperation was formed in a way like food chain. But compared to the production rate of volatile acids produced by C. butyricum, the utilization rate by photo-fermentative bacteria was far slower. The results demonstrated that the growth of photo-fermentative bacteria was limited when pH decreased sharply. The best ratio of C. butyricum to R. faecalis RLD-53 was 1:600. The maximum yield of hydrogen reached 122.4ml-H2/vessel and hydrogen production rate was 0.5ml-H2/ml-culture/day. [Copyright &y& Elsevier]

Abstract: In order to obtain the high H2 yield, photo-hydrogen production by Rhodopseudomonas faecalis strain RLD-53 in fed-batch culture using acetate as the sole carbon was studied. In repeated fed-batch culture, biomass increased rapidly from 0.05 to 0.65g/l within 120h, with the specific maximal growth rate estimated 0.68×10−3 g/h. After 120h, biomass increased slowly and after each time feeding biomass increased slightly. The specific cumulative H2 volumes in each phase were 2791.3, 1161.7, 1445.5 and 840.6mlH2/l-culture, respectively. The average H2 yield was 3.17mol H2/mol acetate based on the whole process while the average substrate conversion efficiency reached 79.3%. Specific maximum H2 production rate and H2 content was 37.2mlH2/l/h and 95.5%, respectively. The results demonstrated the repeated fed-batch mode obtained higher efficiency for hydrogen production, feeding acetate concentration and control of pH were important to fed-batch culture hydrogen production. [Copyright &y& Elsevier]

Abstract: In this work, the effects of different parameters on co-culture hydrogen production using Clostridium Butyricum and immobilized Rhodopseudomonas faecalis RLD-53 were investigated. The maximum hydrogen yield of 4.134mol H2/mol glucose was obtained using 6g/l glucose, 50mmol/l phosphate buffer, initial pH of 7.5, the ratio of dark to photo bacteria of 1:10 and light intensity of 8000lux. The maximum hydrogen production rate was 33.85ml H2/l/h. Phosphate buffer concentration was the most important parameter influencing hydrogen production in this co-culture. The ratio of acetate to butyrate increased from 0.74 to 1.82 in the soluble metabolites from C. butyricum with phosphate buffer concentration of 10–50mmol/l. Experimental results could be of great significance for further pilot studies of co-culture hydrogen production. [Copyright &y& Elsevier]

Abstract: In the present study, the effect of Ni2+ (0–10μmol/l), Fe2+ (0–200μmol/l) and Mg2+ (0–15mmol/l) concentration on photo-hydrogen production from acetate was investigated by batch culture. Results showed that under a proper concentration range, Ni2+ was able to enhance the hydrogen production rate and the hydrogen yield; Fe2+ was able to increase the hydrogen yield, and hydrogen production rate was enhanced only when the culturing time was 24–72h. Ni2+ and Fe2+ at a higher concentration inhibited cell growth. When Ni2+ and Fe2+ concentrations were 4μmol/l and 80μmol/l, respectively, maximal hydrogen yield of 2.87 and 2.78mol H2/mol acetate was obtained when batch culturing at 35°C with initial pH 7.0. Mg2+ did not significantly affect hydrogen production and hydrogen yield which maintained at about 2.45mol H2/mol acetate, but it was favorable to cell growth. [Copyright &y& Elsevier]

The Rhodopseudomonas faecalis strain RLD-53 was isolated from freshwater pond sludge and was demonstrated it could produce hydrogen. This study to investigate their ability of hydrogen production under some conditions in batch culture experiments. At pH 7.0, temperature 35°C and light intensity of 4000lux, the H2 yield was 2.64mol-H2/mol-acetate, 2.34mol-H2/mol-propionate, 1.75mol-H2/mol-lactate and 3.55mol-H2/mol-malate, respectively. The maximal H2 production rate of 32.62ml-H2/l/h and H2 yield of 2.84mol-H2/mol-acetate were achieved when beef extract was used as nitrogen source. Light intensity is the most important factor for H2 production, H2 production yield and rate decreased with increasing light intensity and reached highest under light intensity of 3000–5000lux. Result indicated the strain RLD-53 was a high efficient bacteria for hydrogen production. [Copyright &y& Elsevier]

Abstract: The effect of butyrate addition on H2 production from acetate by purple non-sulphur photosynthetic bacteria Rhodopseudomonas faecalis RLD-53 was studied in this work. Experimental results showed that butyrate was not used as carbon source for H2 production, but different concentration (12.5–100mmol/l) of butyrate added in acetate medium has obviously affected H2 yield and maximum H2 production rate. H2 yield and maximum H2 production rate increased with the increasing of butyrate concentration when butyrate concentrations at 12.5–25mmol/l and then gradually decreased when the butyrate concentration by further increase to 100mmol/l; this indicated that the increase of butyrate would lead to the inhibition of H2 production. H2 yield (3.43mol-H2/mol-acetate) and maximum H2 production rate (32.54ml/l/h) were the highest when acetate was at 25mmol/l and butyrate at 25mmol/l. These are higher than most reported values in the literature. [Copyright &y& Elsevier]

Hydrogen recovery through solar-driven biomass conversion by photo-fermentative bacteria (PFB) has been regarded as a promising way for sustainable energy production. However, a considerable fraction of organic substrate was consumed for the growth of PFB as biocatalysts, furthermore, these PFB were continuously washed out from the photobioreactor in continuous operation because of their poor flocculation. In this work, PFB bioaggregate induced by L-cysteine was applied in a sequencing batch photobioreactor to enhance continuous hydrogen production and reduce biomass washout. The effects of the hydraulic retention time (HRT), influent concentration and light intensity on hydrogen production of the photobioreactor were investigated. The maximum hydrogen yield (3.35 mol H2/mol acetate) and production rate (1044 ml/l/d) were obtained at the HRT of 96 h, influent concentration of 3.84 g COD/l, and light intensity of 200 W/m2. With excellent settling ability, biomass accumulated in the photobioreactor and reached 2.15 g/l under the optimum conditions. Structural analysis of bioaggregate showed that bacterial cells were covered and tightly linked together by extracellular polymeric substances, and formed a stable structure. Therefore, PFB bioaggregate induced by L-cysteine is an efficient strategy to improve biomass retention capacity of the photobioreactor and enhance hydrogen recovery efficiency from organic wastes. [ABSTRACT FROM AUTHOR]

Abstract: Fermentative H2 production in continuous stirred tank reactor (CSTR) system with bacteria attached onto granular activated carbon (GAC) was designed to produce H2 continuously. The H2 production performances of CSTR with suspended and attached-sludge from molasses were examined and compared at various organic loading rates (8–40g COD/L/d) at hydraulic retention time of 6h under mesophilic conditions (35°C). Both reactor systems achieved ethanol-type fermentation in the pH ranges 4.5–4.8 and 3.8–4.4, respectively, while ORP ranges from −450 to −470mV and from −330 to −350mV, respectively. The hydrogen production rate in the attached system was higher compared to that of the suspended system (9.72 and 6.65L/d/L, respectively) while specific hydrogen production rate of 5.13L/g VSS/d was higher in the suspended system. The attached-sludge CSTR is more stable than the suspended-sludge CSTR with regard to hydrogen production, pH, substrate utilization efficiency and metabolic products (e.g., volatile fatty acids and ethanol) during the whole test. [Copyright &y& Elsevier]

[Display omitted] • Pretreatment is a critical process for biohydrogen production from biomass. • A distinction should be made between the initial pH and the pH of the process. • Dissolved bio-H 2 concentrations are not always in equilibrium with the gas phase. • The combined fermentation methods offer advantages compared to dark fermentation. • Iron catalysts are attractive due to high abundance and relatively low toxicity. Biohydrogen is a green energy carrier with the potential to reduce our dependency on fossil fuels. The production of biohydrogen from lignocellulosic materials may be low cost, but commercial production has not been realized. The issues to be addressed include screening and construction of microorganisms that meet industrial requirements, optimization of bioreactor designs, and novel hybrid fermentation systems such as combining dark fermentation (DF) with methanogenic systems and doping nanomaterials to increase biohydrogen yield. This paper focuses on the biohydrogen production, highlighting various fermentative bioreactor types, influential factors in dark fermentation, and pretreatment techniques. Besides, current strategies to produce biohydrogen, including synthetic biology approaches and metabolic engineering, are assessed for their potential to contribute to net zero carbon emission. Pretreatment of biomass using dilute acids is the most common technique to destroy the complicated structure of lignocellulosic biomass. Mesophilic temperatures (37 °C), HRT of 72 h, and pH near 7.0, are recurrent parameters of biomass fermentation. The combined approach of electro-fermentation and dark fermentation produces a higher yield of biohydrogen compared to the single dark fermentation. Furthermore, additives (mainly iron-based materials) have been investigated to improve the efficiency of biomass pretreatment as well as biohydrogen production. Finally, this paper points out that the vigorous development of novel engineered strains is an important part of achieving commercial-scale production of biohydrogen. Overall, considerable efforts are needed from both technical and managing aspects to achieve a full-scale application of biohydrogen production to make this technology economical, efficient and sustainable. [ABSTRACT FROM AUTHOR]

[Display omitted] • Corn-stalk derived biochar was used in dark fermentation and chain elongation. • Corn-stalk derived biochar showed the promotion effect in dark fermentation. • Dark fermentation metabolites can produce caproate without extra electron donor. • Corn-stalk derived biochar increased the caproate production in chain elongation. This study proposes a new model in which ethanol and acetate produced by dark fermentation are processed by Clostridium kluyveri for chain elongation to produce caproate with an addition of biochar prepared from cornstalk residues after acid pretreatment and enzymatic hydrolysis (AERBC) in the dark fermentation and chain elongation processes. The results show a 6–25% increase in hydrogen production in dark fermentation with adding AERBC, and the maximum concentration of caproate in the new model reached 1740 mg/L, 61% higher than that in the control group. In addition, caproate was obtained by dark fermentation, using liquid metabolites as substrates with an initial pH range of 6.5–7.5. Finally, the electron balance and electron transfer efficiency in the new model were analyzed, and the role of AERBC in dark fermentation and chain elongation was investigated. This study provides a new reference for the use of dark-fermented liquid metabolites and cornstalk residue. [ABSTRACT FROM AUTHOR]

Abstract In this work, the synergistic effects of L-cysteine and biofilm on the performance of reactor were investigated in the photo-fermentative biofilm reactor. Both L-cysteine and biofilm could enhance the performance of reactor for producing hydrogen. The optimal L-cysteine concentrations in the biofilm reactor (BR) and control reactor (CR, reactor with no biofilm) were 0.5 g/L and 0.75 g/L, respectively. In addition, the maximum cumulative hydrogen volume (3500 ± 170 mL/L), substrate conversion efficiency (79.4 ± 4.0%) and hydrogen yield 867 ± 43 mL/g (acetate) were obtained in the BR at L-cysteine concentration of 0.5 g/L. It is intriguing that extracellular DNA (eDNA) occupied 50% of EPS, which could play a key role in hydrogen production. Biofilm could decrease EPS concentration of cells in the BR at each L-cysteine concentration to divert more energy into hydrogen. The moderate L-cysteine concentration could improve bacterial growth and regulate the EPS to promote the biofilm formation and enhance hydrogen production. Thus, the regulation of L-cysteine concentration will be a practical method to enhance hydrogen production by the biofilm reactor. Highlights • Both the L-cysteine and biofilm could enhance photo-H 2 production. • H 2 production in the BR is regulated by biofilm bacteria as "leader wolf". • Lower EPS concentration was favorable to produce more hydrogen. • Protein and eDNA were the important compositions of EPS and construct the biofilm. [ABSTRACT FROM AUTHOR]

Synergistic system of dark fermentation and algal culture was initially operated at batch mode to investigate the energy production and nutrients removal from molasses wastewater in butyrate-type, ethanol-type and propionate-type fermentations. Butyrate-type fermentation was the most appropriate fermentation type for the synergistic system and exhibited the accumulative hydrogen volume of 658.3 mL L −1 and hydrogen yield of 131.7 mL g −1 COD. By-products from dark fermentation (mainly acetate and butyrate) were further used to cultivate oleaginous microalgae. The maximum algal biomass and lipid content reached 1.01 g L −1 and 38.5%, respectively. In continuous operation, the synergistic system was stable and efficient, and energy production increased from 8.77 kJ L −1  d −1 (dark fermentation) to 17.3 kJ L −1  d −1 (synergistic system). Total COD, TN and TP removal efficiencies in the synergistic system reached 91.1%, 89.1% and 85.7%, respectively. This study shows the potential of the synergistic system in energy recovery and wastewater treatment. [ABSTRACT FROM AUTHOR]

To achieve stable and efficient photo-fermentative hydrogen production, this work investigated photo-fermentative hydrogen production by forming biofilm on the surface of carrier in the biofilm reactor (BR). Results showed the hydrogen production performance was greatly improved by formed biofilm. The time of hydrogen production and efficiency of substrate utilization were enhanced obviously compared to the control reactor (CR). When the CR was used, hydrogen production stopped at 7th day and maximum cumulative hydrogen volume and hydrogen yield were 1730 ± 87 mL/L and 1.44 ± 0.07 mol H 2 /mol acetate, respectively. However, in the BR hydrogen production volume of 3028 ± 150 mL/L and hydrogen yield of 2.52 ± 0.13 mol H 2 /mol acetate were obtained, which were enhanced about 75% compared to that of the CR. The time of hydrogen production extended from 7 days of CR to 12 days of BR and the substrate conversion efficiency increased from 36% of CR to 63% of BR. It was worth noting at 8th day that substrate was almost utilized completely but hydrogen production still lasted for 4 days. This suggested that the formation of biofilm in BR was favorable to continuous hydrogen production and substrate utilization with high efficiency. Results demonstrated the BR can get a more stable and consistent operating process and it was a proper and potential way to produce hydrogen by photo-fermentative bacteria (PFB). [ABSTRACT FROM AUTHOR]

[Display omitted] • High biohydrogen performance of different simulated human feces (SHF) concentrations achieved by pure culture. • Characterization of human fecal biochar as a promotor in the hydrogen production process was studied. • The best H 2 production was found for 40 wt%-SHFs with 128.54 mmol H 2 /L in 180 h. • The highest H 2 production of 148.75 mmol H 2 /L was obtained with 6 g/L HFBC-800. Human feces contain a lot of nutrients and organic matter, which can be used in many fields. In this study, biochar (HFBC) was prepared at 300, 500 and 800 °C using human feces as substrate, and the characteristics of HFBC prepared at different pyrolysis temperatures were investigated. It was confirmed that HFBC prepared at 800 ℃ (HFBC-800) had a better pH buffer capacity, more proportion of π-π shake-up satellites, better graphitization degree and higher conductivity, which could promote the efficiency of biohydrogen production of bacteria. Furthermore, the biohydrogen production of Thermoanaerobacterium thermosaccharolyticum M18 using different concentrations of simulated human feces (SHF) was further investigated. The best biohydrogen production was found for 40 wt%-SHF, with 128.54 mmol H 2 /L produced in 180 h. The highest biohydrogen production of 148.75 mmol H 2 /L was obtained by adding 6 g/L HFBC-800 to the biohydrogen production process. This study provides a feasibility for high-performance hydrogen production from human feces and presents an insight into the resource utilization of human waste. [ABSTRACT FROM AUTHOR]

In this study, a semi-continuous culture mode was used for photo-hydrogen production by Rhodopseudomonas sp. nov. strain A7 and 3 key operating parameters (the interval feeding time, initial feeding time and the feeding volume ratio) were optimized to improve hydrogen production performance. The rate and yield of hydrogen production decreased from 19.7 ml-H 2 /l/h to 4.63 ml-H 2 /l/h with the interval feeding time increased from 1 day to 3 days, and the yield of hydrogen decreased gradually and reached the maximum value of 2.11 mol-H 2 /mol acetate when the interval feeding time is at 1 day. The initial feeding time was extended from 3 days to 4 days, the rate of hydrogen production decreased from 24.8 ml-H 2 /l/h to 23 ml-H 2 /l/h, and the rate of hydrogen production decreased to 22.2 ml-H 2 /l/h when it was extended to 5 days. And specific yield of hydrogen production of 2.65, 2.48 and 2.38 mol-H 2 /mol-acetate was obtained, respectively. The yield of hydrogen increased with the increase of feeding volume ratio from 10% to 40% and reached a maximum value of 3.22 mol-H 2 /mol acetate at feeding volume ratio of 40%, the specific yield and rate of hydrogen production gradually decreased with the feeding volume ratio further increased to 60%. The optimal parameters of semi-continuous operation are as following: the initial feeding time of 3 days, interval feeding time of 1 day and feeding volume ratio of 40%. In these conditions, the maximum yield (3.22 mol-H 2 /mol acetate) and rate (30 ml-H 2 /l/h) of hydrogen production and light energy conversion efficiency of 1.9% were obtained. Compared to other hydrogen production mode (batch, continuous and fed-batch culture), the semi-continuous culture mode for photo-hydrogen production exhibited great potential due to following advantages: less loss of biomass, the well stability of system, easy operation and control. Therefore it is the best operation mode for photo-hydrogen production with high yield of hydrogen production and light energy conversion efficiency. [ABSTRACT FROM AUTHOR]

• Effects of microplastics (MPs) on the semi-continuous H 2 production were investigated. • Mesophilic and thermophilic H 2 production were enhanced in MPs supplemented groups. • MPs can favor the H 2 -producing bacterial growth and optimize the microbial community. • The highest semi-continuous H 2 production rate reached 4.85 L L−1 d−1. In this study, the influences of polyethylene terephthalate microplastics (PET MPs) on the semi-continuous hydrogen (H 2) production and microbial community under mesophilic and thermophilic fermentation were investigated. Results indicated that PET MPs can enhance the semi-continuous H 2 production performance, and the highest H 2 production rates in mesophilic and thermophilic MPs supplemented groups were 4.85 and 3.6 L L−1 d−1, which were 21.6% and 63.6% higher than the control groups, respectively. Moreover, it was found that PET MPs could favor the growth of H 2 -producing bacteria and optimize the microbial community structure. In mesophilic MPs supplemented groups, the abundance of genus Lactococcus and Ethanoligenens increased from 20.77% and 6.7% to 38.69% and 10.78%, respectively. The abundance of genus Thermoanaerobacterium became the dominant bacteria in thermophilic fermentation, and its abundance increased from 9.02% to 69.38%. This work provides novel insights into the function of MPs in fermentative H 2 production system. [ABSTRACT FROM AUTHOR]

Abstract: Biological mycelia pellets, which are formed spontaneously in the process of Aspergillus niger Y3 fermentation, were explored as carrier for immobilization of Clostridium sp. T2 to improve hydrogen production. Batch fermentation tests showed that optimal dosage and size of mycelia pellets for hydrogen production were 0.350 g 150 ml−1 medium and 1.5 mm. Under these conditions, hydrogen production with immobilized cells on mycelia pellets was further investigated in continuous stirred-tank reactor (CSTR) with hydraulic retention time (HRT) ranging from 12 to 8 h. It obtained that the maximum hydrogen production rate reached 2.76 mmol H2 L−1 h−1 at 10 h HRT, which was 40.8% higher than the carrier-free process, but slightly lower than the counterpart immobilized in sodium alginate with the value of 3.15 mmol H2 L−1 h−1. SEM observation showed that abundant cells were closely adhered to mycelia pellets. The present results indicate the potential of using mycelia pellets as biological carrier for enhancing hydrogen production. [Copyright &y& Elsevier]

Abstract: In the process of producing H2 from lignocellulosic materials, inhibitory compounds could be potentially formed during pre-treatment. This work experimentally investigated the effect of lignocellulose-derived inhibitors on growth and hydrogen production by Thermoanaerobacterium thermosaccharolyticum W16. Representative compounds presented in corn stover acid hydrolysate were added in various concentrations, individually or in various combinations and subsequently inhibitions on growth and H2 production were quantified. Acetate sodium was not inhibitory to T. thermosaccharolyticum W16, rather than it was stimulatory to the growth and H2 production. Alternatively, furfural, hydroxymethylfurfural (HMF), vanillin and syringaldehyde were potent inhibitors of growth and hydrogen production even though these compounds showed inhibitory effect depending on their concentrations. Synergistic inhibitory effects were exhibited in the introduction of combinations of inhibitors to the medium and in hydrolysate with concentrated inhibitors. Fermentation results from hydrolysates revealed that to increase the efficiency of this bioprocess from corn stover hydrolysate, the inhibitory compounds concentration must be reduced to the levels present in the raw hydrolysate. [ABSTRACT FROM AUTHOR]

Abstract: A novel mesophilic hydrogen-producing bacterium was isolated from cow dung compost and designated as Clostridium sp. HR-1 by 16S rRNA gene sequence. The optimum condition for hydrogen production by strain HR-1 was pH of 6.5, temperature of 37 °C and yeast extract as nitrogen sources. The strain HR-1 has the ability to utilize kinds of hexose and pentose as carbon sources for growth and H2 production. Cell growth and hydrogen productivity were investigated for batch fermentation on media containing different ratios of xylose and glucose. Glucose was the preferred substrate in the glucose and xylose mixtures. The high glucose fraction had higher cell biomass production rate. The rate of glucose consumption was higher than xylose consumption, and remained essentially constant independent of xylose content of the mixture. The rate of xylose utilization was decreased with increasing of the glucose fraction. The average H2 yield and specific H2 production rates with xylose and glucose are 1.63 mol-H2/mol xylose and 11.14-H2 mmol/h g-cdw, and 2.02 mol-H2/mol-glucose and 9.37 mmol-H2/h g-cdw, respectively. Using the same initial substrate concentration, the maximum average H2 yield and specific H2 production rates with the mixtures of 9 g/l xylose and 3 g/l glucose was 2.01 mol-H2/mol-mixed sugar and 12.56 mmol-H2/h g-cdw, respectively. During the fermentation, the main soluble microbial products were ethanol and acetate which showed trends with the different ratios of xylose and glucose. [ABSTRACT FROM AUTHOR]

Abstract: 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 11.2gL−1 glucose, 3.4gL−1 xylose and 0.5gL−1 arabinose under the optimum condition of cellulase dosage 25Ug−1 substrate with supplement xylanase 30Ug−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. [Copyright &y& Elsevier]

Abstract: Understanding how a bioreactor functions is a necessary precursor for successful reactor design and operation. This paper describes a two-dimensional computational fluid dynamics simulation of three-phase gas–liquid–solid flow in an expanded granular sludge bed (EGSB) reactor used for biohydrogen production. An Eulerian–Eulerian model was formulated to simulate reaction zone hydrodynamics in an EGSB reactor with various hydraulic retention times (HRT). The three-phase system displayed a very heterogeneous flow pattern especially at long HRTs. The core-annulus structure developed may lead to back-mixing and internal circulation behavior, which in turn gives poor velocity distribution. The force balance between the solid and gas phases is a particular illustration of the importance of the interphase rules in determining the efficiency of biohydrogen production. The nature of gas bubble formation influences velocity distribution and hence sludge particle movement. The model demonstrates a qualitative relationship between hydrodynamics and biohydrogen production, implying that controlling hydraulic retention time is a critical factor in biohydrogen-production. [Copyright &y& Elsevier]

The design of an optimum and cost-efficient medium for high-level production of hydrogen by Ethanoligenens harbinense B49 was attempted by using response surface methodology (RSM). Based on the Plackett–Burman design, Fe2+ and Mg2+ were selected as the most critical nutrient salts. Subsequently, the optimum combination of the selected factors and the sole carbon source glucose were investigated by the Box–Behnken design. Results showed that the maximum hydrogen yield of 2.21mol/molglucose was predicted when the concentrations of glucose, Fe2+ and Mg2+ were 14.57g/L, 177.28mg/L and 691.98mg/L, respectively. The results were further verified by triplicate experiments. The batch reactors were operated under an optimized condition of the respective glucose, Fe2+ and Mg2+ concentration of 14.5g/L, 180mg/L and 690mg/L, the initial pH of 6.0 and experimental temperature of 35±1 oC. Without further pH adjustment, the maximum hydrogen yield of 2.20mol/molglucose was obtained based on the optimized medium with further verified the practicability of this optimum strategy. [Copyright &y& Elsevier]

Abstract: The biofilm-based expanded granular sludge bed (EGSB) reactor was developed to treat starch-containing wastewater and simultaneously recovery hydrogen by mixed microbial culture. Granular activated carbon (GAC) was used as the support media. Operating at the temperature of 30°C for over 400 days (data not shown), the EGSB reactor presented high efficiency in hydrogen production and COD removal ability. The maximum hydrogen production rate (HPR) was found to be 1.64L/L.d under the organic loading rate (OLR) of 1.0g-starch/L.d, pH of 4.42 and HRT of 4h. The hydrogen yield (HY) peaked at 0.11L/g-COD, under the OLR of 0.5g-starch/L.d, pH of 3.95 and HRT of 8h. Hydrogen volume content was estimated to be 35–65% of the total biogas. The average COD removal rate was 31.1% under the OLR of 0.125g-starch/L.d and HRT of 24h. The main dissolved fermentation products were ethanol, acetate and butyrate. The average attached biofilm concentration was estimated to be 8.26g/L, which favored hydrogen production and COD removal. It is speculated that the low pH operation in the present system would contribute significantly to lower the cost of alkaline amount required for pH control in the continuous operation, especially in the scale-up biohydrogen producing system. A model, built on the back propagation neural network (BPNN) theory and linear regression techniques, was developed for the simulation of EGSB system performance in the biodegradation of starch synthesis-based wastewater and simultaneous hydrogen production. The model well fitted the laboratory data, and could well simulate the removal of COD and the production of hydrogen in the EGSB reactor. [Copyright &y& Elsevier]

Abstract: In order to enrich hydrogen producing bacteria and to establish high-efficient communities of the mixed microbial cultures, inoculum needs to be pretreated before the cultivation. Four pretreatment methods including heat-shock pretreatment, acid pretreatment, alkaline pretreatment and repeated-aeration pretreatment were performed on the seed sludge which was collected from a secondary settling tank of a municipal wastewater treatment plant. In contrast to the control test without any pretreatment, the heat-shock pretreatment, acid pretreatment and repeated-aeration pretreatment completely suppressed the methanogenic activity of the seed sludge, but the alkaline pretreatment did not. Employing different pretreatment methods resulted in the change in fermentation types as butyric-acid type fermentation was achieved by the heat-shock and alkaline pretreatments, mixed-acid type fermentation was achieved by acid pretreatment and the control, and ethanol-type fermentation was observed by repeated-aeration pretreatment. Denaturing gradient gel electrophoresis (DGGE) profiles revealed that pretreatment method substantially affected the species composition of microbial communities. The highest hydrogen yield of 1.96mol/mol-glucose was observed with the repeated-aeration pretreatment method, while the lowest was obtained as the seed sludge was acidified. It is concluded that the pretreatment methods led to the difference in the initial microbial communities which might be directly responsible for different fermentation types and hydrogen yields. [Copyright &y& Elsevier]

The difficulty and long duration of start-up wastes numerous costs, labors and time and a little fluctuate during the process might fail it. However, studies dealing with the problem hindering accelerated start-up are still insufficient. Current research focused to develop a method for accelerated start-up in an efficient way. This work outlined a novel alternative for accelerated start-up. This joint method, adding waste effluent with applying biofilm reactor, could successfully start up hydrogen production in the first 24 h via increasing ability of hydrogen producers while the control group produced little hydrogen. The two factors, biofilm formation and addition of waste effluent, expressed the combined effects on accelerated start-up. This study suggested that little molecules like quorum sensing system factors and indoles might be the crucial regulating and stimulating factors and express the accelerated start-up ability only in biofilm reactors. • Adding waste effluent in biofilm reactor could rapidly start up the reactor. • This joint method started and enhanced the bacterial activity in the early stage. • Quorum sensing system factors and indoles acted as regulation factors. • The regulation factors only stimulated hydrogen production with biofilm formation. [ABSTRACT FROM AUTHOR]

Cysteine can participate in the essential metabolism of photo-fermentation and plays an important role in biological hydrogen production. However, D-cysteine is usually ignored in current researches despite its potential amounts and unknown effects. This study firstly investigated the photo-fermentative cell growth and hydrogen production with two chiralities of cysteine (L-cysteine and D-cysteine). This and previous studies illustrated proper L-cysteine concentration could improve hydrogen production performance of Rhodopseudomonas sp. nov. strain A7. However, D-cysteine inhibited the cells proliferation and hydrogen production. Groups of initial D-cysteine concentration above 0.3 g/L postponed reactor start-up for several days and obtained little hydrogen production and bacterial biomass. Groups of initial D-cysteine concentration under 0.3 g/L could operate reactor normally in time and obtained about 1000 mL/L hydrogen and 2.0 g/L biomass. Addition time was another key factor, and groups of adding D-cysteine on day 1 influenced little on cells growth but decreased hydrogen production. Other groups with adding time on day 2–5 obtained normal biomass and hydrogen production. Besides, 0.3 g/L functioned as the limit concentration in different initial biomass concentrations for D-cysteine. This work concluded whether the cells could grow and produce hydrogen depended on the addition time and initial D-cysteine concentrations. D-cysteine determined the duration of adapt phase and was regarded as the quorum sensing factor. Furthermore, although biofilm reactors could improve hydrogen production compared to control reactor, biofilm showed little influence on D-cysteine limit concentration. This study would pave the way for the future photo-fermentative hydrogen production. [ABSTRACT FROM AUTHOR]

• Formed biofilm could enhance the H 2 production by regulating proteomic and EPS. • The expression level of key protein greatly influenced the ability of H 2 production. • Structure and concentration of EPS were critical to stable biofilm and releasing H 2. Photo-fermentative hydrogen production, the new energy production alternative, was greatly enhanced by formed biofilm. To understand the mechanism of enhancement, the intracellular proteome and extracellular polymeric substance (EPS) i i EPS: extracellular polymeric substanceiiPFHP: Photo-fermentative hydrogen productioniiiPFB: photo-fermentative bacteriaivQS: quorum sensingvCLSM: Confocal laser scanning microscopyviCR: Control reactorviiBR: Biofilm reactorviiiTEM: Transmission electron microscopeixXPS: X-ray photoelectron spectroscopyxFTIR: Fourier transform infrared spectroscopyxiSM: Supplementary materialsxiiDEPs: Differentially expressed proteinsxiiiTCA cycle: Tricarboxylic acid cyclexivGO: Gene ontologyxvKEGG: Kyoto encyclopedia of genes and genomes during biofilm formation were investigated in this work. Experimental results indicated that a possible and effective altered system could transfer light to hydrogen. Proteins were significantly regulated, for example those related with nitrogenase, flagellin, EPS transportation and DNA duplication were up-regulated while those concerned photosystem were down-regulated. It revealed these changes of proteins contributed to positive activity of key enzymes, improved communication system and increased total light utilization efficiency thus leading to enhanced capacity of hydrogen production. Besides above metabolic changes inside the cells, EPS secreted by the bacteria played an important role in hydrogen production and its yield decided the release of hydrogen. When EPS descended to a lower concentration during biofilm formation, it meant carbon source for EPS synthesis was reduced, and more energy and reducing power could be transferred into hydrogen energy. More importantly, this work found that composition and structure of EPS were efficiently influenced by the formation of biofilm, such as benzene and O-H structure, secondary protein structure and the kinds of protein, which were important to stable biofilm and efficient hydrogen production. Therefore, final hydrogen yield was improved by altered protein and EPS resulted from biofilm formation. This study demonstrated that formation of biofilm is an efficient, ecological and attracting way to the future bio-hydrogen production. [ABSTRACT FROM AUTHOR]