The effect of Agolin Ruminant, a blend of essential oils, on methane (CH4) emissions were investigated in two in vivo experiments and in four in vitro experiments. In the in vivo experiments, four lactating dairy cows and four beef heifers were supplemented 0.2 g/d of the essential oils (ca. 2–4 ppm m/v) during an eight-weeks period, where the first two weeks served as control (no essential oils supplementation). In dairy cattle, essential oils tended to decrease the daily CH4 emissions (g/d) and CH4 relative to dry matter intake (g/kg DMI) by 15% and 14%, respectively, after 6 weeks of supplementation (P=0.07), but no difference was observed for CH4 relative to milk production (g/kg milk) (P=0.64) or CH4 relative to bodyweight (g/kg BW) (P=0.12). In the in vivo experiment with beef cattle daily CH4 emissions and CH4 relative to DMI did not change when supplemented the essential oils at a dose of 0.2 g/d (numerical decreases of 10 and 11% for g CH4/d and g CH4/kg DMI, respectively) but CH4 relative to body weight tended to decrease by 20% after 6 weeks of supplementation (P=0.07). The in vitro experiments were expected to replicate the results observed in vivo. However, no decrease in CH4 production was observed in 24 h batch incubations at concentrations up to 30 ppm (m/v). A longer contact time between the essential oils (15 and 30 ppm) and the feedstuff (essential oils added ca. 16 h prior the start of the incubation) did not elicit any effect on CH4 production and was not different from addition immediately prior to the start of the incubation. Longer incubation time (96 h and 14 d) and regular supply of both substrate and additive in a consecutive batch incubation system did not induce CH4 inhibition up to essential oils doses of 30 ppm (m/v) and hence, also were not able to replicate in vivo results. Using the gas production technique (GPT) methane was inhibited by 17% with an essential oils dose of 30 ppm after 24 h, but this decrease was not constant across all times during the 72 h incubation. The blend of essential oil was effective reducing daily emissions of methane in dairy cattle and emissions relative to body weight in beef cattle, interestingly, these effects were not observed in vitro regardless of the techniques used to replicate in vivo results. This might be due to differences in the mode of action of the essential oils in vitro and in vivo, which merits attention for future research.