Enteric methane production in response to direct ruminal infusion of synthetic bromoform in cattle

Methane inhibitors in cattle diets are a promising strategy to reduce enteric methane (CH₄) emissions. Bromoform (CHBr₃) is effective but not well understood in terms of mechanism, dose response, and side effects such as reduced feed intake. Studies using CHBr₃-containing seaweed are inadequate to isolate the effect of CHBr₃ from other factors. This study aimed to determine the dose-dependent effects of synthetic CHBr₃, delivered via ruminal infusion, on DMI, enteric gas emissions, temporal patterns of gas emissions, and ruminal fermentation in cattle. Four nonlactating, nonpregnant, rumen-fistulated Holstein-Friesian cows (12 yr of age; 781 ± 33 kg BW; mean ± SD) were randomly assigned to 1 of 4 treatment sequences in a 4 × 4 Latin square design. Treatments consisted of 0 (CNTL), 59 (LOW), 192 (MED), or 592 (HIGH) mg CHBr₃/d infused into the rumen as an aqueous solution via the rumen fistula. The daily CHBr₃ dose was divided into 3 equal infusions every 8 h over a 24-h period (0545, 1345, and 2145 h) for 14 consecutive d during experimental periods, followed by a 7-d washout period. Cows were fed grass hay ad libitum, and additional concentrate was supplied through a GreenFeed (C-Lock Inc.), which was also used for enteric gas measurements. Total DMI did not respond linearly or quadratically to increasing doses of CHBr₃ but was numerically 6.2 kg/d lower for HIGH compared with CNTL. Both CH₄ production (g/d) and yield (g/kg DMI) decreased quadratically with increasing doses of CHBr₃. Conversely, hydrogen (H₂) production (g/d) and yield (g/kg DMI) increased quadratically with increasing dose of CHBr₃. Total VFA concentration did not respond linearly or quadratically to increasing doses of CHBr₃ but was numerically 27.8 mM lower for HIGH compared with CNTL. Molar proportions of propionate, butyrate, valerate, and branched-chain VFA increased quadratically, and the molar proportion of acetate decreased quadratically with increasing dose of CHBr₃. Separate 4-parameter logistic functions were fitted to CH₄ and H₂ production (g/d) over the infusion and washout periods, using data from the HIGH treatment. The decrease in CH₄ production and increase in H₂ production stabilized after 56 and 49 h of CHBr₃ infusions, respectively (defined as the time at which 95% of the asymptotic change in gas production was attained). After cessation of the CHBr₃ infusions, the increase in CH₄ production and decrease in H₂ production stabilized after 117 and 99 h, respectively. In conclusion, synthetic CHBr₃ supplementation reduced enteric CH₄ production in a nonlinear, dose-dependent manner, with minor effects at 59 and 192 mg CHBr₃/d doses, but ~90% reduction at 592 mg CHBr₃/d. The HIGH treatment numerically reduced DMI, indicating a potential trade-off between efficacy and feed intake. Temporal modeling showed that the CH₄-suppressing effect of CHBr₃ infusion at 592 mg/d emerged within 2 to 3 d and reversed more gradually over 4 to 5 d after the last infusion. These findings highlight the importance of both dose and temporal dynamics in evaluating CHBr₃ for CH₄ mitigation in ruminants.