Linking Metabolites in Eight Bioactive Forage Species to Their in Vitro Methane Reduction Potential Across Several Cultivars and Harvests

An in vitro study was conducted to analyze the fermentation end-products from 17 cultivars of eight polyphenol containing forage species. The polyphenol composition and proanthocyanidin (PA) structural features of all cultivars were analyzed with UPLC-MS/MS in leaves of vegetative or generative plants. All samples were incubated with and without polyethylene glycol (PEG, a tannin-binding agent) to separate the tannin-effect on methane (CH 4 ) production from that of the forage quality. Sulla and big trefoil, two particularly PA rich species, were found to have the highest CH 4 reduction potential of up to 47%. However, they showed simultaneous and almost equal reductions in gas production (GP; a proxy for digestibility). The addition of PEG led to an increase in both GP and CH 4 production, confirming the role of tannins on CH 4 reduction. Moreover, PA structural features and concentration were found to be an important source of variation for CH 4 production from PA containing species. Despite having low polyphenol concentrations, chicory and plantain were found to reduce CH 4 production without reducing GP. Generally, the variation across cultivars from the same species was found to be lower than interspecies variability, and the results were found to be consistent across growth stages, indicating the findings representativeness.