Manganese-modified biochar enhances white-rot fungi-mediated lignocellulose humification: insights from molecular transformation and multi-omics analyses

Humic substance formation is essential for soil health, yet natural humification is slow, and synthetic production is often inefficient. White-rot fungi (WRF)-mediated lignocellulose humification shows potential but is limited by slow substrate degradation. Given the high biocatalytic potential of Mn(II) and its compatibility with the enzyme system of WRF, manganese-modified biochar (MBC) was utilized to enhance the degradation and conversion efficiency of lignocellulose. A 35-day solid-state fermentation experiment showed that 5% MBC addition increased lignin degradation by 93.4%, humic acid content by 47.7%, and polymerization degree by 112.4% compared with CK. Mechanistically, first, MBC enhanced extracellular lignocellulose-degrading enzymes and Mn-associated Fenton-like oxidation, thereby improving lignocellulose depolymerization. Second, interpretable machine learning and mass difference-based association analysis identified key molecular reactivity traits, suggesting that MBC facilitated humification by promoting the selective generation and conversion of reactive precursor molecules. In particular, enhanced DOM reactivity and increased precursor formation through deamination, dealkylation, and delignification were associated with the generation of more aromatic, oxygen-functionalized, and unsaturated humic-like substances, likely through polyphenol-dominated pathways. Finally, transcriptomic and metabolomic analyses indicated that MBC upregulated the expression of CAZyme-related genes and metabolic pathways associated with precursor generation, including phenolic/quinone biosynthesis, indicating that enhanced substrate depolymerization and subsequent precursor transformation jointly contributed to lignocellulose humification. Overall, this study provides mechanistic insights into how MBC enhances WRF-mediated lignocellulose humification and offers a theoretical basis for optimizing the bioconversion of lignocellulose-rich organic wastes.