The objective of our study was to assess the effect of dietary starch level on reticulorumen pH and determine the subsequent effects on blood indicators of copper (Cu) status when fed without or with additional sulfur (S) and molybdenum (Mo). Sixty Holstein-Friesian dairy cows (33 ± 2.5 DIM) yielding 41 (±0.9) kg/d were blocked and randomly allocated to 1 of 4 dietary treatments in a 2 × 2 factorial design experiment for a duration of 98 d. The treatment diets contained 15 mg Cu/kg DM, were isonitrogenous and isoenergetic, and had a grass silage-to-corn silage ratio of 1:1 on a DM basis. Dietary starch levels were 225 g/kg DM (HS) or 148 g/kg DM (LS), and were either unsupplemented (−) or supplemented (+) with 1.2 g of S/kg DM and 4.1 mg Mo/kg DM. Reticulorumen pH was measured using pH boluses inserted in 6 cows per treatment, and blood samples were collected at wk 0, 1, 2, 4, 6, 10 and 14 of the study. Performance and blood parameters were analyzed as a 2 × 2 factorial design using a repeated measures ANOVA, with main effects of starch level (S), Cu antagonists (A) and their interaction (S × A) using Genstat (v18). Mean reticular pH was 0.2 higher in cows receiving LS diets. The addition of dietary antagonists had no effect (P > 0.05) on milk yield but reduced (P < 0.001) DMI by 1.9 kg/d. We observed no effect (P > 0.05) of dietary treatment on plasma Cu, iron, or zinc with mean values of 15.3, 42.1, and 14.4 µmol/L respectively. In contrast, we found that ceruloplasmin activity was 3.9 mg/dL higher (P < 0.05) in cows fed HS diets, and there was an interaction (P < 0.05) on plasma Mo, whereby cows fed additional S and Mo had a 0.25 µmol/L higher concentration in cows fed HS+ compared with LS+. Our results demonstrate altered Cu-antagonist metabolism resulting from feeding different starch levels that altered rumen pH, and highlight the need to take into consideration the pH modulatory potential of diets when selecting appropriate Cu supplementation strategies for lactating dairy cows.