Approximately, 45 mL of rumen fluid was taken from the rumen by a stomach tube connected to a vacuum pump at 0and 4 h after feeding on the last day of each period. Ruminal pH and temperature were determined using a portable pH and temperature meter (HANNA Instruments HI 8424 microcomputer, Singapore). Ruminal NH3–N concentration was analyzed using Kjeltech Auto 1030 Analyzer (AOAC, 1995; ID 973.18).