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[Display omitted] • The roles of biochar on microbial growth and activity were systematically analyzed. • Biochar improved the microbial community diversity and population. • Biochar enhanced cellular enzyme activity and accelerated electron transfer. • Biochar promoted functional gene expression and regulated metabolic pathway. • Biochar targeted design to achieve high microbial activity in AD were suggested. The biochar is a well-developed porous material with various excellent properties, that has been proven with excellent ability in anaerobic digestion (AD) efficiency promotion. Current research is usually focused on the macro effects of biochar on AD, while the systematic review about the mechanisms of biochar on microbial behavior are still lacking. This review summarizes the effects and potential mechanisms of biochar on microorganisms in AD systems, and found that biochar addition can provide habitats for microbial colonization, alleviate toxins stress, supply essential nutrients, and accelerate interspecies electron transferring. Moreover, it also improves microbial community diversity, facilitates EPS secretion, enhances functional enzyme activity, promotes functional genes expression, and inhibits the antibiotic resistance genes transformation. Future research directions including biochar targeted design, in-depth microbial mechanisms revelation, and modified model development were suggested, which could promote the widely practical application of of biochar-amended AD technology. [ABSTRACT FROM AUTHOR]

• Magnetic biochar addition caused a loosed structure of protein for boosting CH 4 yield. • Fe 2 O 3 and FeO of magnetic biochar first bound with proteins via hydrogen bond. • The C=O, C N and N H of protein were subsequently destroyed by magnetic biochar. • Hydrolysis-acidification and syntrophic metabolisms were promoted by magnetic biochar. The addition of composite conductive materials is being increasingly recognized as a promising strategy to enhance anaerobic digestion (AD) performance. However, the influence of these materials on protein hydrolysis has been poorly documented. Here, a novel magnetic biochar derived from oil sludge and straw was synthesized using different iron sources and successfully applied in sludge AD. Experimental results revealed that magnetic biochar modified by Fe2+ exhibited excellent electron transfer capacity, moderate magnetization, diverse functional groups (e.g. C=O, C-O=O-), and abundant iron distribution. These characteristics significantly enhanced the hydrolysis of tryptophan-like components, leading to increased methane production (144.44 mL gVS−1 vs 79.72 mL gVS−1 in the control test). Molecular docking analysis revealed that the binding of magnetic biochar related Fe2+ and Fe3+, onto sludge proteins via hydrogen bond played a key role in promoting subsequent protein hydrolysis. Additionally, the noteworthy conservation of protein structures from α-helix and β-sheet to random coil, along with the breakdown of the amide I-associated C=O group and amide III-related C N and N H bonds following the addition of magnetic biochar, accelerated the degradation of sludge protein. Observation of variations in protease activity, coenzyme F420, electron transfer system (ETS), and conductivity within the AD systems, particularly the enrichment of Methanospirillum and Methanosaeta archaea , as well as the Petrimonas, Comamonas, and Syntrophomonas bacteria, suggested that magnetic biochar facilitated a conducive environment by improving hydrolysis-acidification and the direct interspecies electron transfer (DIET) process for acetoclastic methanogens. Moreover, metabolic pathways further proved that tryptophan metobalism and acetoclastic methanogenesis were both facilitated by magnetic biochar. This study provides an in-depth understanding of the impact of magnetic biochar on protein hydrolysis in sewage sludge AD. [Display omitted] [ABSTRACT FROM AUTHOR]