Alkaline Biochar Promoted Stable and Efficient Thermophilic Hydrogen Production from Lignocellulose by Restoring Coculture System Homeostasis

The construction of coculture systems for enhancing hydrogen production from lignocellulose has been widely applied, but they frequently suffer from instability caused by environmental factors such as pH fluctuations and high substrate concentrations, leading to reduced hydrogen yields. In this study, the optimal biochar, AB-10, was prepared by treating sugar cane bagasse with 20 g/L NaOH at 60 °C for 4 h, followed by pyrolysis at 600 °C for 2 h. The addition of AB-10 biochar increased hydrogen production in the coculture system to 5092 mL/L, representing a 205.09% improvement compared to the control group. After alkaline modification, the surface area and micropore area of the biochar increased by 2.13-fold and 2.50-fold, respectively, facilitating microbial colonization. Moreover, AB-10 biochar maintained the proportion of Acetivibrio thermocellus DSM 1313 above 20% in the coculture system, effectively preventing system collapse under high substrate loading. The observed increases in enzyme activities, redox capacity, electron-transfer efficiency, and the concentrations of electron bifurcation-related components elucidate the mechanisms by which AB-10 biochar enhances dark fermentation performance at elevated substrate concentrations. Overall, this study provides valuable insights into maintaining coculture system homeostasis and improving hydrogen production during dark fermentation.