Dark fermentative hydrogen production from lignocellulosic residues is often limited by inefficient electron distribution and competing metabolic pathways. In this study, a lignin-derived humic substance-biochar (LHS-BC) was developed to enhance thermophilic hydrogen production from lignocellulosic residues. Among the tested materials, the lignin-derived humic-like substance obtained via alkaline oxidative humification and subsequently combined with biochar (HSH@BC) exhibited the best performance, increasing hydrogen production (mL/L) by 29.76% compared with the control. Kinetic analysis showed that HSH@BC significantly increased hydrogen production potential (1113.98 mL/L) and reduced lag time. The composite promoted cellulose degradation, enhanced cellulase and hydrogenase activities, and increased intracellular NAD+/NADH levels. Metabolic analysis revealed a shift from ethanol-type fermentation to acetate-butyrate pathways, leading to higher hydrogen yield. Electrochemical characterization suggests that cytochrome c may be involved in electron exchange with the quinone functional groups in LHS-BC. PICRUSt-based functional prediction suggested potential enrichment of central metabolic pathways, including glycolysis, pyruvate metabolism, the tricarboxylic acid (TCA) cycle, and the pentose phosphate pathway (PPP). Overall, LHS-BC improved hydrogen production by simultaneously regulating microbial community structure, metabolic pathways, and electron transfer processes, providing an effective strategy for thermophilic biohydrogen production from lignocellulosic biomass.
Biochar enhances dark fermentative biohydrogen production (BHP), yet conventional biochar is limited by low porosity and few active sites. While nitrogen doping and chemical activation can individually upgrade biochar, the synergistic effect of urea doping combined with sodium bicarbonate (NaHCO3) activation, and its consequence for intracellular metabolic networks, remains unclear. Herein, material characterization, 16S rRNA sequencing, and non-targeted metabolomics were integrated to elucidate how urea-doped NaHCO3-activated rice-straw biochar (UBC-A) enhances cellulolytic BHP. UBC-A achieved the highest hydrogen production of 192.52 mL·g-1, representing a 6.6-fold (561.35% relative improvement) of the control; the hydrogen production lag period was shortened to 13.93 h, and the energy conversion efficiency was 14.19%. UBC-A exhibited enhanced graphitization and hierarchical porous structure. Microbiome analysis revealed selective enrichment of hydrogen-producing taxa (Clostridia, Thermoanaerobacterium) and cellulolytic microbes, alongside suppression of competitors. Metabolomics identified 113 significantly differential metabolites (P < 0.05), revealing system-wide metabolic rewiring centered on three interconnected hubs: (i) L-glutamate-driven TCA cycle activation and GABA-mediated acid stress alleviation; (ii) 2-hydroxyglutarate as a novel indicator of enhanced NADH regeneration capacity; and (iii) glycerophospholipid-mediated membrane restructuring facilitating extracellular electron transfer. Correlation analysis established significant associations between these hydrogen producers and key upregulated metabolites, indicating that UBC-A optimizes BHP by synchronizing community assembly with metabolic pathway redirection. These findings advance a structure-microbiome-metabolism framework for agricultural-waste valorization and biohydrogen industrialization.
Tao Sheng, Jiaxing Meng, Chengwei Song et al.· Environmental Research· 0 citations
Treating lignocellulosic agricultural wastes with cellulase generates monosaccharides for biofuel production and saccharification residues that can be converted into biochar with better performance than those from untreated materials. However, while cellulase pretreatment is highly effective for gramineous straws, its efficacy on hardwood-like lignocellulosic materials remains unknown. This study explored the applicability of cellulase pretreatment to tobacco stalks (hardwood-like) for biochar preparation. Results showed cellulase pretreatment enhanced biochar stability and subsequent optimization via thermal air oxidation and colloid modification further improved its performance, confirming it as a high-quality precursor biochar. Cellulase pretreatment primarily increased raw material crystallinity, which in turn improved biochar crystallinity and structural stability. Compared with untreated biochar (BC-O), colloid-modified biochar (BC-SAC) exhibited a rise in specific surface area from 4.9 to 344.1 m2/g, alongside rapid phenol adsorption reaching equilibrium within 12 h and enhanced removal performance toward various phenolic contaminants. Environmental and economic evaluations indicated that BC-SAC achieved a carbon emission reduction of 0.68 t CO2 per year per ton of feedstock. Additionally, its iodine value, a key pricing index for carbon-based adsorbents, rose from 503 mg/g to 881 mg/g. This study broadens the feedstock scope of cellulase pretreatment for biochar production and is of great significance for efficient multi-product utilization of lignocellulosic biomass resources.
Kai Yang, Kang Wang, Wenyan Xie et al.· Bioresource Technology· 0 citations
Introduction Lignin recalcitrance, together with its complex interactions with cellulose and hemicelluloses within the plant cell wall, remains a major barrier to efficient lignocellulosic biomass valorization. Mild alkaline pretreatment partially addresses this challenge by solubilizing lignin and acetate into a lignin-rich alkaline liquor (AL). However, the recovered lignin is largely oligomeric, limiting its direct microbial conversion. Methods To evaluate strategies for improving AL bioconversion, alkaline liquor was subjected to thermochemical depolymerization under different severities, generating liquors with compositions ranging from oligomer-rich to monomer-rich profiles. The resulting streams were evaluated through two bioconversion routes: aerobic metabolism by Pseudomonas putida KT2440 and anaerobic digestion for biomethane production. Results Mild to moderate depolymerization conditions (≤240 °C) improved the growth of P. putida, whereas the highest depolymerization severity strongly inhibited bacterial growth. This inhibition was associated with increased concentrations of aromatic monomers that are poorly metabolized by P. putida, including phenol and alkyl-substituted phenols. In anaerobic digestion assays, mild to moderate depolymerization yielded the highest specific methane productions, corresponding to a 10%–20% increase relative to non-depolymerized AL, while severe depolymerization conditions negatively affected methane production. Discussion These findings demonstrate that controlled lignin depolymerization can enhance both aerobic and anaerobic bioconversion by balancing chemical accessibility with biological compatibility. This approach provides a promising strategy for improving lignin utilization within integrated biorefinery concepts.
Fabrícia Farias de Menezes, F. M. Kashiwagi, J. J. Silva et al.· Frontiers in Bioengineering...· 0 citations
n-Butanol is a promising advanced biofuel and versatile platform chemical. However, its fermentative production by solventogenic clostridial strains remains economically limited by reliance on costly edible feedstocks of corn and sugarcane. Lignocellulosic biomass provides an abundant, non-food alternative, but its effective conversion necessitates pretreatment which inevitably generates weak acids, furan derivatives, and lignin-derived phenolic compounds. These by-products synergistically inhibit clostridial metabolism by disrupting pH balance, depleting NADH/NADPH, and compromising membrane integrity. To address this bottleneck, this review systematically examines the formation and inhibitory effects of these compounds, and then summarizes the recent strategies of metabolic engineering and co-culture for enhancing the tolerance of strains against various pretreatment-derived inhibitors. In addition, the review traces the progress of pretreatment technologies from conventional acid, alkaline, and physicochemical methods to emerging ionic liquids and deep eutectic solvents, and compares their influence on butanol fermentation performance. By linking lignocellulose pretreatment, inhibition mechanisms, metabolic engineering, and bioprocess engineering, this review provides a systems-level framework for designing more efficient lignocellulosic butanol pathways. Future research interests are also provided, including predictive modeling to control inhibitor generation, strain improvement for lignin-derived phenolic tolerance, techno-economic analysis and life-cycle assessment, and integrated lignin valorization, all aimed at advancing economically viable and sustainable biorefinery processes.
Hongzhen Luo, Wenwen Zhang, Tingting Liu et al.· Frontiers in Chemical Engine...· 0 citations
Lignocellulosic fruit peel waste represents an abundant, carbon-neutral feedstock for green biohydrogen production via dark fermentation, yet its rigid compact structure and high cellulose crystallinity severely restrict saccharification and fermentative hydrogen yield. In this study, a hydrothermal acidification pretreatment strategy was proposed to boost dark fermentative biohydrogen generation from navel orange peel waste, and systematic investigations were conducted to reveal the regulating mechanisms of key pretreatment parameters (hydrochloric acid concentration, pretreatment temperature, duration) on reducing sugar release and hydrogen-producing performance. Multiscale characterizations including SEM, XRD, FTIR, and TG were integrated to unravel the microstructural and chemical compositional evolution of raw and pretreated substrates. The results demonstrated that hydrothermal acidification effectively disrupted the dense lignocellulosic network of navel orange peel, lowered cellulose crystallinity, and greatly improved substrate accessibility for hydrolytic reactions and microbial adhesion. Under the optimal pretreatment condition (1.0 mol/L HCl, 120 °C, 1 h), the concentration of released reducing sugars reached 10.2 g/L, which was 67.2% higher than that of untreated raw peel. The corresponding maximum cumulative hydrogen yield attained 36.5 mL H2/g TS, representing a 67.4% improvement relative to the untreated control group. Pearson correlation analysis verified that pretreatment temperature, acid concentration, and duration exhibited strong positive correlations with hemicellulose and cellulose removal efficiencies, while excessive pretreatment (HCl > 1.0 mol/L, temperature > 120 °C, duration > 1 h) generated inhibitory by-products that suppressed microbial hydrogen evolution. This study comprehensively clarifies the structural modification and biohydrogen promotion mechanism of hydrothermal acidification pretreatment on pectin-rich biomass, and delivers a cost-effective, facile technical route for high-value energy valorization and harmless disposal of fruit processing solid wastes.
Cong Zhan, Qin Li, Li Wu et al.· Energies· 0 citations
Lignocellulosic biomass is a renewable and abundant resource for producing high-value chemicals as sustainable alternatives to petroleum-derived products. Among these, 5-hydroxymethylfurfural (HMF) and furfural (FA), generated through catalytic dehydration of biomass-derived sugars, serve as pivotal intermediates for enzymatic synthesis of valuable platform chemicals like 2,5-furandicarboxylic acid (FDCA). In recent studies, glyoxal oxidases (GlyOx, EC 1.2.3.15) have emerged as promising candidates for the biotransformation of furan derivatives. This study focused on the recombinant production and characterization of a GlyOx from the medicinal basidiomycete Ganoderma lucidum capable of oxidizing HMF and FA with 51.5% and 23.0% conversion, respectively, after 48 h. Furthermore, the enzyme oxidized HMF derived from hexose-rich wheat straw hydrolysates, leading to 32.4% conversion after 72 h, as well as FA derived from beechwood hemicellulosic hydrolysates, leading to furoic acid with 17.0% conversion after 48 h, marking the first time such enzymatic activity has been demonstrated on furans originating from genuine lignocellulosic biomass sugar streams.
Maria-Konstantina Karonidi, Κoar Chorozian, A. Marianou et al.· Journal of Agricultural and...· 1 citation