Abstract
BACKGROUND: Methane (CH(4)) emissions from ruminants significantly contribute to greenhouse gas effects and energy loss in livestock production. Methyl-coenzyme M reductase (MCR) is the key enzyme in methanogenesis, making it a promising target for CH(4) mitigation. This study aimed to identify and validate plant-derived inhibitors by using molecular docking to screen compounds with strong binding affinity to the F430 active site of MCR and assessing their efficacy in reducing CH(4) emissions. RESULTS: Molecular docking analysis identified salvianolic acid C (SAC) as a potent inhibitor of MCR, showing a strong binding affinity to the F430 active site (binding energy: -8.2 kcal/mol). Enzymatic inhibition assays confirmed its inhibitory effect, with a half-maximal inhibitory concentration (IC(50)) of 692.3 µmol/L. In vitro rumen fermentation experiments demonstrated that SAC supplementation (1.5 mg/g DM) significantly reduced CH(4) production (P < 0.01) without negatively affecting major fermentation parameters. Microbial community analysis using 16S rRNA sequencing and metagenomics revealed that SAC selectively altered the rumen microbiota, increasing the relative abundance of Bacteroidota while significantly reducing Methanobrevibacter (P = 0.04). Moreover, metagenomic analysis showed the downregulation of key methanogenesis-related genes (mcrA and rnfC), suggesting a dual mechanism involving direct enzymatic inhibition and microbial community modulation. CONCLUSIONS: These findings indicate that SAC effectively reduces CH(4) production by inhibiting MCR activity and reshaping the rumen microbial community. As a plant-derived compound with strong inhibitory effects on methanogenesis, SAC presents a promising and sustainable alternative to synthetic CH(4) inhibitors, offering potential applications for mitigating CH(4) emissions in livestock production.