An acid-tolerant Clostridium sp. BLY-1 strain with high biohydrogen production rate.

Screening and isolating acid-tolerant bacteria capable of efficient hydrogen production can mitigate the inhibitory effects on microbial activity caused by rapid pH drops during fermentation. In this study, we isolated an acid-tolerant and highly efficient hydrogen-producing bacterium, named Clostridium sp. BLY-1, from acidic soil. Compared to the model strain Clostridium pasteurianum DSM 525, BLY-1 demonstrates a faster growth rate and superior hydrogen production capabilities. At an initial pH of 4.0, BLY-1's hydrogen production is 7.5 times greater than that of DSM 525, and under optimal conditions (pH=5.0), BLY-1's hydrogen production rate is 42.13% higher than DSM 525. Genomic analysis revealed that BLY-1 possesses a complete CiaRH two-component system and several stress-resistance components absent in DSM 525, which enhance its growth and hydrogen production in acidic environments. These findings provide a novel avenue for boosting the hydrogen production capabilities of Clostridium strains, offering new resources for advancing the green hydrogen industry.

Bioconversion of industrial wastes to hydrogen: A review on waste-to-wealth technologies.

Energy plays a significant role in attaining the sustainable growth of the industrial sector of any nation. The resources for getting energy are limited and cannot fulfill the huge demand for energy supply in the near future. Generating fuels from various waste materials and biomass is widely viewed as a sustainable energy source and a viable option for the future. Currently, researchers are particularly interested in synthesizing hydrogen (H2) without emitting CO2 and other greenhouse gases (GHGs). Hydrogen is recognized as a pristine and environmentally friendly energy source, presenting an optimal substitute for fossil fuels due to its high energy content of 122 kJg-1. The traditional methods for the production of H2 are cost-intensive and heavy input requirements are needed. Thus, the synthesis of H2 through biological approaches is cost-effective and eco-friendly alternating with easy operational requirements with ambient reaction conditions. The most common drawback of the biological production of H2 is the low yield and production rates of gas during scale-up conditions. This review is focused on different processes used to convert the wastes into H2 energy along with their pattern of utilization and the effect on the environment.

Influences of bisphenol A on hydrogen production from food waste by thermophilic dark fermentation.

The extensive use of plastic products in food packaging and daily life makes them inevitably enter the treatment process of food waste (FW). Plasticizer as a new pollutant is threatening the dark fermentation of FW. Our study showed that bisphenol A (BPA) at > 250 mg/L had a significant inhibition on hydrogen production from FW by thermophilic dark fermentation. The endogenous ATP content and lactate dehydrogenase (LDH) release showed that high level of BPA not only inhibited the growth of hydrogen-producing consortium, but also led to cell death. In addition, BPA mainly affects the hydrogen-producing consortium by reducing cell membrane fluidity, damaging cell membrane integrity and reducing cell membrane potential, resulting in cell death. This study provides some new insights into the mechanism of the effect of BPA on hydrogen production from FW by thermophilic dark fermentation, and lays the foundation on the utilization of FW.

Insight into furfural-tolerant and hydrogen-producing microbial consortia: Mechanism of furfural tolerance and hydrogen production.

Furfural-tolerant and hydrogen-producing microbial consortia were enriched from soil, with hydrogen production of 259.84 mL/g-xylose under 1 g/L furfural stress. The consortia could degrade 2.5 g/L furfural within 24 h in the xylose system, more efficient than in the sugar-free system. Despite degradation of furfural to furfuryl alcohol, the release of reactive oxygen species and lactate dehydrogenase was also detected, suggesting that furfuryl alcohol is also a potential inhibitor of hydrogen production. The butyrate/acetate ratio was observed to decrease with increasing furfural concentration, leading to decreased hydrogen production. Furthermore, microbial community analysis suggested that dominated Clostridium butyricum was responsible for furfural degradation, while Clostridium beijerinckii reduction led to hydrogen production decrease. Overall, the enriched consortia in this study could efficiently degrade furfural and produce hydrogen, providing new insights into hydrogen-producing microbial consortia with furfural tolerance.

Recent advancements in wastewater treatment via anaerobic fermentation process: A systematic review.

This manuscript delves into the realm of wastewater treatment, with a particular emphasis on anaerobic fermentation processes, especially dark, photo, and dark-photo fermentation processes, which have not been covered and overviewed previously in the literature regarding the treatment of wastewater. Moreover, the study conducts a bibliometric analysis for the first time to elucidate the research landscape of anaerobic fermentation utilization in wastewater purification. Furthermore, microorganisms, ranging from microalgae to bacteria and fungi, emphasizing the integration of these agents for enhanced efficiency, are all discussed and compared. Various bioreactors, such as dark and photo fermentation bioreactors, including tubular photo bioreactors, are scrutinized for their design and operational intricacies. The results illustrated that using clostridium pasteurianum CH4 and Rhodopseudomonas palustris WP3-5 in a combined dark-photo fermentation process can treat wastewater to a pH of nearly 7 with over 90% COD removal. Also, integrating Chlorella sp and Activated sludge can potentially treat synthetic wastewater to COD, P, and N percentage removal rates of 99%,86%, and 79%, respectively. Finally, the paper extends to discuss the limitations and future prospects of dark-photo fermentation processes, offering insights into the road ahead for researchers and scientists.

Effects of carbon source variability on enhanced Bio-hydrogen production.

This study investigated how glucose, starch, and rapeseed oil, three common food waste components with diverse molecular and physicochemical characteristics, influenced hydrogen production and microbial communities in dark fermentation under varying carbon/nitrogen (C/N) ratios. The results indicated that glucose and starch groups, significantly increased hydrogen yields to 235 mL H2/gVS (C/N = 40) and 234 mL H2/gVS (C/N = 40), respectively, while rapeseed oil, with a lower yield of 30 mL H2/gVS (C/N = 20), demonstrated a negative impact. Additionally, an accumulation of propionate was observed with increasing carbon source complexity, suggesting that simpler carbon sources favored hydrogen production and bacterial growth. Conversely, lipid-based materials required rigorous pre-treatment to mitigate their inhibitory effects on hydrogen generation. Overall, this study underscores the importance of carbon source selection, especially glucose and starch, for enhancing hydrogen production and microbial growth in dark fermentation, while highlighting the challenges posed by lipid-rich substrates that require intensive pre-treatment to optimize yields.

Enhancing biohydrogen production from xylose at low temperature (20 °C) using natural FeS2 Ore: Thermodynamic analysis and mechanistic insights.

This study investigates the efficacy of pyrite in enhancing biohydrogen production from xylose at low temperature (20 °C). Higher hydrogen yield rates (Rm) and reduced lag time (λ) were achieved across initial xylose concentrations ranging from 2-10 g/L. At an optimal xylose concentration of 5 g/L, pyrite reduced λ by 2.5 h and increased Rm from 1.3 to 2.7 mL h-1. These improvements are attributed to pyrite's ability to enhance the secretion of extracellular polymeric substance and flavins, facilitate NADH and NAD+ generation and transition, and favor biohydrogen production. Thermodynamic analyses and Gibbs free energy calculations further elucidated pyrite's role in the full reaction process and rate-limiting steps at low temperature. This study offers valuable insights into improving the efficiency of biohydrogen production at low temperature, with significant implications for energy conservation.

Biohydrogen production by lactate-driven dark fermentation of real organic wastes derived from solid waste treatment plants.

This study evaluated the hydrogen production potential through lactate-driven dark fermentation (LD-DF) of organic wastes from solid waste treatment plants, including the organic fraction of municipal solid waste (OFMSW), mixed sewage sludge, and two OFMSW leachates. In initial batch fermentations, only OFMSW supported a significant hydrogen yield (70.1 ± 7.7 NmL-H2/g-VS added) among the tested feedstocks. Lactate acted as an important hydrogen precursor, requiring the presence of carbohydrates for sequential two-step lactate-type fermentation. The impact of operational pH (5.5-6.5) and initial total solids (TS) concentration (5-12.5 % w/w) was also evaluated using OFMSW as substrate, obtaining hydrogen yields ranging from 6.6 to 55.9 NmL-H2/g-VSadded. The highest yield occurred at 6.5 pH and 7.5 % TS. The LD-DF pathway was indicated to be present under diverse pH and TS conditions, supported by employing a specialized microbial consortium capable of performing LD-DF, along with the observed changes in lactate levels during fermentation.

Novel Insights into Alkyl Polyglucoside Biosurfactant Promoting Anaerobic Dark Fermentation for Hydrogen Production in Sludge.

Anaerobic fermentation of excess sludge (ES) for hydrogen production is a crucial strategy for resource utilization and environmentally friendly treatment. However, the low hydrolysis efficiency of ES and the depletion of produced hydrogen have become the limiting factors for low hydrogen yield. This study innovatively applied the bio-based surfactant alkyl polyglucoside (APG) to enhance the efficiency of dark fermentation for hydrogen production from ES. When the APG content was 100 mg/g (calculated based on total suspended solids), the maximum hydrogen production reached 17.8 mL/g VSS, approximately 3.7 times that in the control group. Mechanistic analysis revealed that APG promoted the release of organic matter from ES. APG also facilitated the release of soluble protein and soluble polysaccharide, increasing the organic matter reduction rate to 34.8%, significantly higher than other groups. APG enhanced the accumulation of volatile fatty acids and promoted the proportion of small molecular carboxylic acids. Enzyme activity analysis revealed that APG promoted the activity of hydrolytic enzymes but inhibited the activity of hydrogen-consuming enzymes. The research results provide a green and environmentally friendly strategy for the efficient resource utilization of ES.

Biomass acid pretreatment impacts on metabolic routes and bacterial composition of dark fermentation process.

Complex organic matter represents a suitable substrate to produce hydrogen through dark fermentation (DF) process. To increase H2 yields, pretreatment technology is often required. The main objective of the present work was to investigate thermo-acid pretreatment impact on sugar solubilization and biotic parameters of DF of sorghum or organic fraction of municipal solid waste (OFMSW). Biochemical hydrogen potential tests were carried out without inoculum using raw or thermo-acid pretreated substrates. Results showed an improvement in sugar solubilization after thermo-acid pretreatments. Pretreatments led to similar DF performances (H2 and total metabolite production) compared to raw biomasses. Nevertheless, they were responsible for bacterial shifts from Enterobacteriales towards Clostridiales and Bacillales as well as metabolic changes from acetate towards butyrate or ethanol. The metabolic changes were attributed to the biomass pretreatment impact on indigenous bacteria as no change in the metabolic profile was observed after performing thermo-acid pretreatments on irradiated OFMSW (inactivated indigenous bacteria and inoculum addition). Consequently, acid pretreatments were inefficient to improve DF performances but led to metabolic and bacterial community changes due to their impact on indigenous bacteria.

Enhancing biohydrogen production from xylose through natural FeS2 ore: Mechanistic insights.

In this study, we investigated the advantages of utilizing natural FeS2 ore in the context of dark fermentative hydrogen production within a fermentation system employing heat-treated anaerobic granular sludge with xylose as the carbon source. The results demonstrated a significant improvement in both hydrogen production and the maximum rate, with increases of 2.58 and 4.2 times, respectively. Moreover, the presence of FeS2 ore led to a reduction in lag time by more than 2-3 h. The enhanced biohydrogen production performance was attributed to factors such as the intracellular NADH/NAD+ ratio, redox-active components of extracellular polymeric substances, secreted flavins, as well as the presence of hydrogenase and nitrogenase. Furthermore, the FeS2 ore served as a direct electron donor and acceptor during biohydrogen production. This study shed light on the underlying mechanisms contributing to the improved performance of biohydrogen production from xylose during dark fermentation through the supplementation of natural FeS2 ore.

Salt and heat stress enhances hydrogen production in cyanobacteria.

Cyanobacteria are among the most suitable organisms for the capture of excessive amounts of CO2 and can be grown in extreme environments. In our research we use the single-celled freshwater cyanobacteria Synechococcus elongatus PCC7942 PAMCOD strain and Synechocystis sp. PCC6714 for the production of carbohydrates and hydrogen. PAMCOD strain and Synechocystis sp. PCC6714 synthesize sucrose when exposed to salinity stress, as their main compatible osmolyte. We examined the cell proliferation rate and the sucrose accumulation in those two different strains of cyanobacteria under salt (0.4 M NaCl) and heat stress (35 0C) conditions. The intracellular sucrose (mol sucrose content per Chl a) was found to increase by 50% and 108% in PAMCOD strain and Synechocystis sp. PCC6714 cells, respectively. As previously reported, PAMCOD strain has the ability to produce hydrogen through the process of dark anaerobic fermentation (Vayenos D, Romanos GE, Papageorgiou GC, Stamatakis K (2020) Photosynth Res 146, 235-245). In the present study, we demonstrate that Synechocystis sp. PCC6714 has also this ability. We further examined the optimal conditions during the dark fermentation of PAMCOD and Synechocystis sp. PCC6714 regarding H2 formation, increasing the PAMCOD H2 productivity from 2 nmol H2 h- 1 mol Chl a- 1 to 23 nmol H2 h- 1 mol Chl a- 1. Moreover, after the dark fermentation, the cells demonstrated proliferation in both double BG-11 and BG-11 medium enriched in NaNO3, thus showing the sustainability of the procedure.

Functional link hybrid artificial neural network for predicting continuous biohydrogen production in dynamic membrane bioreactor.

Conventional machine learning approaches have shown limited predictive power when applied to continuous biohydrogen production due to nonlinearity and instability. This study was aimed at forecasting the dynamic membrane reactor performance in terms of the hydrogen production rate (HPR) and hydrogen yield (HY) using laboratory-based daily operation datapoints for twelve input variables. Hybrid algorithms were developed by integrating particle swarm optimized with functional link artificial neural network (PSO-FLN) which outperformed other hybrid algorithms for both HPR and HY, with determination coefficients (R2) of 0.97 and 0.80 and mean absolute percentage errors of 0.014 % and 0.023 %, respectively. Shapley additive explanations (SHAP) explained the two positive-influencing parameters, OLR_added (1.1-1.3 mol/L/d) and butyric acid (7.5-16.5 g COD/L) supports the highest HPR (40-60 L/L/d). This research indicates that PSO-FLN model are capable of handling complicated datasets with high precision in less computational timeat 9.8 sec for HPR and 10.0 sec for HY prediction.

Investigating the efficiency of a two-stage anaerobic-aerobic process for the treatment of confectionery industry wastewaters with simultaneous production of biohydrogen and polyhydroxyalkanoates.

The scope of the current study was to investigate the efficiency of a two-stage anaerobic-aerobic process for the simultaneous treatment and valorization of selective wastewater streams from a confectionary industry. The specific wastewater (confectionary industry wastewater, CIW) was a mixture of the rinsing eluting during washing of the cauldrons in which jellies and syrups were produced, and contained mainly readily fermentable sugars, being thus of high organic load. The first stage of the process was the dark fermentation (DF) of the CIW in continuous, attached-biomass systems, in which the effect on hydrogen yields and distribution of metabolites were studied for different packing materials (ceramic or plastic), hydraulic retention times, HRTs (12 h-30 h) and feed substrate concentration (20 g COD/L- 50 g COD/L). In the second stage, the effectiveness of the aerobic treatment of the DF effluents was evaluated in terms of the reduction of the organic load and the production of polyhydroxyalkanoates (PHAs) through an enriched mixed microbial culture (MMC). The MMC was developed in a continuous draw and fill system, in which the accumulation potential of PHAs was studied. It was shown that the hydrogen production rates decreased for increasing substrate concentration and HRTs, with a maximum of 12.70 ± 0.35 m3 H2/m3 initial CIW achieved for the lowest HRT and feed concentration and using ceramic beads as packing material. Butyrate, acetate and lactate were the main metabolites generated in all cases, in different ratios. The distribution of metabolites during DF was shown to highly affect the efficiency of the second process in terms of both the reduction of organic load and the PHAs yields. The highest removal of organic load achieved after 48 h of aerobic treatment was 84.0 ± 0.9 %, whereas the maximum PHAs yield was 21.46 ± 0.13 kg PHAs/m3 initial CIW.

Effects of food wastes based on different components on digestibility and energy recovery in hydrogen and methane co-production.

This study was conducted for four organic fractions (carbohydrates, proteins, cellulose, lipids) at an inoculum concentration of 30 % and a total solid (TS) of 8 % to investigate the effect of the main components of food waste on the performance of the two-stage anaerobic digestion. The results showed that the gas phase products were closely related to the composition of the substrate, with the carbohydrate and lipid groups showing the best hydrogen (154.91 ± 2.39mL/gVS) and methane (381.83 ± 12.691mL/gVS) production performance, respectively. However, the increased protein content predisposes the system to inhibition of gas production, which is mutually supported by changes in the activity of dehydrogenase and coenzyme F420. Butyric acid (53.19 %) dominated the liquid phase products in both stages, indicating that all four organic fractions were butyric acid-based fermentation and that the final soluble chemical oxygen demand degradation reached 72.97 %-82.86 %. The carbohydrate and cellulose groups achieved the best energy recovery performance, with conversion rates exceeding 65 %. The above results can provide a useful reference for the resource utilization of food waste.

Comparison of calcium magnesium ferrite nanoparticles for boosting biohydrogen production.

Dark fermentation (DF) is an eco-friendly process that simultaneously achieves organic matter degradation and obtains hydrogen (H2). Nonetheless, low H2 yield mainly caused by poor activity of key microbes, is still a problem that requires being resolved. In this work, MgFe2O4 and Ca0.5Mg0.5Fe2O4 nanoparticles (NPs) were synthetized and served as additives to boost H2 form from DF. H2 productivity gradually increased with the rise of NPs, and declined when NPs exceeded their optimal dosages. The highest H2 yield was 183.6 ± 3.2 mL/g glucose at 100 mg/L of MgFe2O4 NPs, being 35.2 % higher than that of the control yield (135.8 ± 3.1 mL/g glucose). However, the highest H2 yield of 171.9 ± 2.5 mL/g glucose occurred at 400 mg/L of Ca0.5Mg0.5Fe2O4 NPs, increasing by 26.6 % over the control. Interestingly, the two NPs favored the butyric acid pathway for H2 synthesis. This provides guidance for multi-element oxide NPs used in DF.

Production of biological hydrogen from Quinoa residue using dark fermentation and estimation of its microbial diversity.

Although they are one of the world's environmental problems, agricultural wastes or residues are carbohydrate-rich and low-cost, so they are used as raw materials for the manufacture of biohydrogen (bio-H2). Among biological hydrogen manufacture methods, the dark fermentation method is suitable for processing waste or residues. In this regard, no study has been found in the literature on determining the potential of biological hydrogen manufacture from quinoa residue by the dark fermentation method. This work was carried out in a dark room at 36 ± 1 °C under different operating conditions in anaerobic batch bio-reactors fed with thermally pretreated anaerobic mixed bacteria + raw quinoa or quinoa extract liquid + nutrients. In the study, gas analyses were performed and biohydrogen production was detected in all the bio-reactors. Besides, taxonomic content analyses and organic acid analyses were executed. Maximum bio-H2 production was found as follows: at pH 4.5, 14,543.10-4 mL in the bio-reactor fed with 1.00 g quinoa/L and 1880.10-4 mL in the bio-reactor fed with 0.50 g quinoa extract/L, and at pH 4.0, 61,537.10-4 mL in the bio-reactor fed with 1.00 g quinoa/L and 1511.10-4 mL in the bio-reactor fed with 0.75 g quinoa extract/L. In the bio-reactors fed with raw quinoa residue, Clostridium butyricum and Hathewaya histolytica were detected as the most dominant bacteria at pH 4.5 and 4.0, respectively, whereas in the bio-reactors fed with quinoa extract liquid, Fonticella tunisiensis were detected as the most dominant bacteria at both pH 4.5 and pH 4.0.

Evaluation of short-circuited electrodes in combination with dark fermentation for promoting biohydrogen production process.

Short-circuited electrodes, in combination with dark fermentation, were evaluated in a biohydrogen production process. The system is based on an innovative design of a non-compartmented electromicrobial bioreactor with a conductive tubular membrane as cathode and a graphite felt as anode. In particular, the electrode specialization occurred when the bioreactor was inoculated with manure as the whole medium and when a vacuum was applied in the tubular membrane, for allowing continuous extraction of gaseous species (H2, CH4, CO2) from the bioreactor. This specialization of the electrodes as anode and cathode was further confirmed by microbial ecology analysis of biofilms and by cyclic voltammetry measurements. In these experimental conditions, the potential of the electrochemical system (short-circuited electrodes) reached values as low as -320 mV vs. SHE, associated with a significant bioH2 production. Moreover, a higher bioH2 production occurred and a potential of the electrochemical system as low as -429 mV vs SHE was temporarily observed, when additional heat treatments of the whole manure were applied in order to remove methanogen microorganisms (i.e., hydrogen consumers). In the bioreactor, the higher production of bioH2 would be promoted by electrofermentation from the current flow observed between short-circuited anode and cathode.

Combining pretreatments and co-fermentation as successful approach to improve biohydrogen production from dairy cow manure.

The present paper is focused on enhancing the production of biohydrogen (bioH2) from dairy cow manure (DCM) through dark fermentation (DF). Two enhancement production strategies have been tested: i) the combination of H2O2 with sonification as pretreatment and ii) the co-fermentation with cheese whey as co-substrate. Concerning the pretreatment, the best combination was investigated according to the response surface methodology (RSM) by varying H2O2 dosage between 0.0015 and 0.06 g/gTS and ultrasonic specific energy input (USEI) between 35.48 and 1419.36 J/gTS. The increase of carbohydrates concentration was used as target parameter. Results showed that the combination of 0.06 g/gTS of H2O2 with 1419.36 J/gTS of USEI maximized the concentration of carbohydrates. The optimized conditions were used to pretreat the substrate prior conducting DF tests. The use of pretreatment resulted in obtaining a cumulative bioH2 volume of 51.25 mL/L and enhanced the bioH2 production by 125% compared to the control test conducted using raw DCM. Moreover, the second strategy, i.e. co-fermentation with cheese whey (20% v/v) as co-substrate ended up to enhancing the DF performance as the bioH2 production reached a value of 334.90 mL/L with an increase of 1372% compared to the control DF test. To further improve the process, dark fermentation effluents (DFEs) were valorized via photo fermentation (PF), obtaining an additional hydrogen production aliquot.

Integrated dark-fermentation-microbial electrosynthesis for efficient wastewater treatment and bioenergy recovery.

High ammonia concentration in wastewater can hinder methane production rate in anaerobic digestion (AD)-microbial electrosynthesis systems (ADMES). To address this issue, a dual-chamber reactor was fabricated using an anion exchange membrane (AEM) to separate the dark-fermentation (DF) and ADMES process, preventing ammonia migration from the DF chamber to the ADMES chamber. As a result, the DF-ADMES achieved a high methane yield based on chemical oxygen demand (COD) of 0.35 L CH4/gCOD compared to control operation AD (0.23 L CH4/gCOD) and ADMES (0.30 L CH4/gCOD). Additionally, hydrogen could be recovered from the DF chamber which improved the energy efficiency of the DF-ADMES reactor (91.7 %) as compared to control AD (53.4 %) and ADMES (71.9 %). Thus, a dual-chamber DF-ADMES with an AEM separator could be a feasible design for scalable treatment of high nitrogen-containing wastewater and high bioenergy recovery.