This study aimed to identify the optimal doses of the macroalgae Asparagopsis taxiformis and Oedogonium sp., individually and in combination, which would decrease the in vitro production of methane while minimizing adverse effects on fermentation, using rumen inoculant from Bos indicus steers. The dose-response experiment evaluated ten doses of Asparagopsis [ranging from 0 to 16.7 % of the organic matter (OM) incubated] and seven doses of Oedogonium (ranging from 0 to 100 % OM) using Rhodes grass hay as a basal substrate. Asparagopsis was highly effective in decreasing the production of methane with a reduction of 99 % at doses as low as 2 % OM basis. However, a dose of 2 % OM also decreased the production of volatile fatty acids (VFA). Oedogonium was less effective with doses ≥50 % OM significantly decreasing the production of methane. A combination of Asparagopsis (2 % OM) and Oedogonium (25 and 50 % OM) continued to suppress the production of methane, independent of the inclusion rate of Oedogonium. The effectiveness of Asparagopsis demonstrates its potential for the mitigation of methane emissions from ruminants at inclusion rates of ≤2 % OM. Oedogonium is a potential feed supplement due to its nutritional value, but supplements ≤25 % OM are recommended to avoid adverse effects on apparent in vitro fermentation.