Oral supplementation of clay has been reported to function as buffer in dairy cows. However, its effects on rumen, blood, and fecal pH have been variable among studies. Our objective was to determine the effects of 3 levels of dietary clay (EcoMix) supplementation after a grain challenge. Ten multiparous rumen-cannulated Holstein cows (BW = 648 kg ± 12kg), 142 (60–502) DIM, were assigned to 1 of 5 treatments in a completely randomized replicated 5 X 5 Latin Square. Periods consisted of an 18-d adaptation period followed by a 3-d (d 19–21) measurement period. Cow average DMI from d 15–17 was used to restrict feed to 75% on d 18 and to give 20% wheat flour (grain challenge) on d 19, via cannula. Treatments were: CON, no EcoMix and no grain challenge; POS, no EcoMix with a grain challenge; 0.5%, 1%, and 2%, EcoMix as percentages of dietary DMI all with a grain challenge. Statistical analysis was performed using the MIXED procedure of SAS. Two contrasts CONT1 (POS vs. CON), CONT2 (POS vs. average of 0.5%, 1%, and 2%) were compared along with the linear and quadratic treatment effects. Rumen, fecal, and blood pH along with blood metabolites were measured at 0, 4, 8, 12, 16, 20, 24, 36, and 48 h relative to grain challenge. Rumen pH (P = 0.003) and fecal pH were lower (CONT1, P < 0.001) for POS (6.03 ± 0.06; 6.14 ± 0.04) than CON (6.20 ± 0.06; 6.38 ± 0.04), respectively. There was a linear treatment effect for rumen pH (P = 0.001) and fecal pH (P = 0.05). Fecal pH (6.22 ± 0.04, P < 0.0001) was higher for cows that received EcoMix then POS (6.14 ± 0.04, CONT2). There was a difference for negative incremental area under the curve (Rumen pH below 5.6/24h; 0.5% = 7.93 1% = 8.56, 2% = 7.79, SEM = 0.8, P = 0.005) when compared with POS (11.0, CONT2). Cows fed EcoMix had higher milk yield, (0.5% = 28.8 kg, 1% = 30.2 kg, 2% = 29.1 kg, SEM = 3.4), 3.5% FCM (0.5% = 29.9 kg, 1% = 34.1 kg, 2% = 33.1 kg, SEM = 3.4, P = 0.02) and ECM (0.5% = 29.1 kg, 1% = 32.8 kg, 2% = 31.6 kg, SEM = 3.3 P = 0.01) than cows in POS (CONT2, 27.72 kg, 28.0 kg, 27.71kg, respectively). In conclusion, cows that received EcoMix had higher rumen pH, milk yield, ECM and FCM than cows in POS.