Effects of monensin withdrawal and subspecies of cattle on the utilization of bermudagrass hay (13.7% CP) were evaluated using ruminally cannulated steers (5 Bos indicus, BI and 5 Bos taurus, BT; 398 kg BW). Subspecies were concurrently subjected to a 2 period, 2 treatment crossover design. Treatments were withdrawal from either 0 (CON) or 200 mg·hd⁻¹·d⁻¹ (MON) monensin (Rumensin 90; Elanco Animal Health, Greenfield, IN) in 0.91 kg DDGS, fed for 42 d. Withdrawal was evaluated for a 28 d period. Rumen fluid was collected 2 h after feeding on d 1, 4, 7, 14, and 21 after the cessation of MON feeding for determination of pH, VFA, ruminal ammonia-N, rate of ammonia production and methane-producing activity. No subspecies × treatment or subspecies × treatment × day interactions were observed (P ≥ 0.16). Treatment × day interactions (P ≤ 0.01) were observed for acetate:propionate ratio and molar percentages of acetate and propionate. There was a marked decrease in molar % propionate between d 1 and 4 from 19.1 to 18.0%; however, molar % propionate remained greater (P ≤ 0.09) for MON than CON through d 7. Molar percentage of acetate increased with MON withdrawal from 68.8 to 69.8 between d 0 and 4. Acetate:propionate ratio was lower (P ≤ 0.01) on d 0 for MON than CON (3.4 vs. 4.0). By d 4, MON had increased to 3.8, and was not different (P = 0.14) from CON. By d 14, no differences (P ≥ 0.88) between MON and CON remained for acetate, propionate, and acetate:propionate ratio. Steers previously fed monensin had similar (P ≥ 0.12) total VFA concentrations by d 4. Greater (P < 0.01) ruminal ammonia-N concentrations (1.88 vs. 1.73 mM) were observed in CON than MON steers. Monensin had no effect (P ≥ 0.69) on rate of ammonia production or methane-producing activity. A treatment × day interaction occurred (P ≤ 0.05) for pH with MON steers having higher pH than CON on d 1, 7, and 14. Bos indicus steers tended to have higher (P = 0.08) pH than BT (6.53 vs 6.46); no other subspecies effects were observed. Results indicate that the monensin induced reduction in acetate:propionate ratio persists for at least 7 d post-withdrawal.