Ruminal methane (CH(4)) production results in the loss of up to 12% of gross energy intake and contributes nearly 20% of the United States' annual emission of this greenhouse gas. We report the effects of select nitrocompounds on ruminal fermentation after 22 h in vitro incubation (39 degrees C) with or without additions of hydrogen (H(2)), formate or both. In incubations containing no added reductant, CH(4) production was inhibited 41% by 2-nitro-1-propanol (2NPOH) and >97% by 3-nitro-1-propionic acid (3NPA), nitroethane (NE) and 2-nitroethanol (2NEOH) compared to non-treated controls and H(2) did not accumulate. With formate as the sole added reductant, nitro-treatment reduced CH(4) production by >99% and caused 42% to complete inhibition of formate catabolism compared to controls, and the accumulation of H(2) increased slightly. Nitro-treatment decreased CH(4) production 57-98% from that of controls when supplied H(2) or formate plus H(2). Formate catabolism was decreased 42-84% from that in controls by all nitro-treatments except 3NPA with both formate and H(2). Greater than 97% of the added H(2) was catabolized within controls; >84% was catabolized in nitro-treated incubations. Acetate, propionate and butyrate accumulations were unaffected by nitro-treatment irregardless of reductant; however, effects on ammonia and branched chain fatty acid accumulations varied. These results suggest that nitro-treatment inhibited formate dehydrogenase/formate hydrogen lyase and hydrogenase activity.