The objective of the present study was to investigate whether feeding barley grain steeped in lactic acid (LA) and treated with heat would affect rumen fermentation patterns, milk production and composition in lactating dairy cows. Eight rumen-cannulated Holstein cows were assigned to a 2 ×2 crossover design and fed once daily a total mixed ration containing rolled barley grain (319.6 g/kg, dry matter basis) steeped for 48 h in equal quantity of tap water (CTR) or in 10 g/L of LA and heat at 55◦C (LAH). Results demonstrated that cows fed the LAH diet had rumen pH values above 5.8 during most intensive fermentation phases (i.e., 8–12 h) post-feeding (P0.05) postprandial concentrations of volatile fatty acids (VFA) but lowered (P