Supersaturation of Dissolved Hydrogen and Methane in Rumen of Tibetan Sheep

Hydrogen (H(2)) is an essential substrate for methanogens to produce methane (CH(4)), and also influences pathways of volatile fatty acids (VFA) production in the rumen. Dissolved H(2) (H(2 (aq))) is the form of H(2) available to microbes, and dissolved CH(4) (CH(4 (aq))) is important for indicating methanogens activity. Rumen H(2 (aq)) concentration has been estimated by assuming equilibrium with headspace gaseous H(2) (H(2 (g))) concentration using Henry's law, and has also been directly measured in the liquid phase in some in vitro and in vivo experiments. In this in vivo study, H(2 (aq)) and CH(4 (aq)) concentration measured directly in rumen fluid and their corresponding concentrations estimated from their gaseous phase concentrations, were compared to investigate the existence of equilibrium between the gas and liquid phases. Twenty-four Tibetan sheep were randomly assigned to two mixed diets containing the same concentrate mixed with oat grass (OG diet) or barley straw (BS diet). Rumen gaseous phase and contents were sampled using rumenocentesis and oral stomach tubing, respectively. Rumen H(2 (aq)) and CH(4 (aq)) concentration and VFA profile differed between sheep fed OG and BS diets. Measured H(2 (aq)) and CH(4 (aq)) concentration were greater than H(2 (aq)) and CH(4 (aq)) concentrations estimated using gas concentrations, indicating lack of equilibrium between gas and liquid phase and supersaturation of H(2) and CH(4) in rumen fluid. As a consequence, Gibbs energy changes (ΔG) estimated for various metabolic pathways were different when calculated using dissolved gases concentrations directly measured and when using dissolved gases concentrations assuming equilibrium with the gaseous phase. Dissolved CH(4), but not CH(4 (g)), was positively correlated with H(2 (aq)). Both H(2 (aq)) and H(2 (g)) concentrations were positively correlated with the molar percentage of butyrate and negatively correlated with the molar percentage of acetate. In summary, rumen fluid was supersaturated with both H(2) and CH(4), and H(2 (aq)) was closely associated with the VFA profile and CH(4 (aq)) concentration. The assumption of equilibrium between dissolved gases and gaseous phase affected ΔG estimation.