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Eleven individual additives were incubated with either perennial ryegrass or with grass silage+barley grain (50 : 50) and the in vitro methane output was assessed using the gas production technique (GPT). Additives were: fatty acids (lauric, oleic, linoleic and linolenic acids), halogenated methane analogues (bromoethanesulfonate and bromochloromethane), pyromellitic diimide, statins (mevastatin and lovastatin), a probiotic (Saccharomyces cerevisiae) and an unsaturated dicarboxylic acid (fumaric acid). Each additive was included at a range of concentrations. Effects on methane output per gram of feed dry matter (DM) incubated (CH4/DMi) and disappeared (CH4/DMd), as well as other fermentation variables, were evaluated after 24 h of incubation. The addition of increased concentrations of individual fatty acids, bromoethanesulfonate and pyromellitic diimide caused a dose-dependent decline in methane output (CH4/DMi, CH4/DMd), when incubated with either perennial ryegrass or grass silage+barley grain. No methane output was detected for either feed with the addition of ≥5 µM bromochloromethane. The statins were ineffective inhibitors of methane output regardless of feed type. For perennial ryegrass, S. cerevisiae caused a dose-dependent decline in CH4/DMd and fumaric acid a dose-dependent decline in CH4/DMi and CH4/DMd. The effectiveness of lauric, oleic, linoleic and linolenic acids and bromoethanesulfonate to reduce methane output was more pronounced when incubated with grass silage+barley grain than with perennial ryegrass, and therefore the type of feed is an important component for any future in vitro and in vivo studies to be undertaken with these additives. Thus, incorporating different feed types in the initial in vitro screening protocols of all new additives is recommended.

Reducing in vitro rumen methanogenesis for two contrasting diets using a series of inclusion rates of different additives