The purpose of this study was to determine whether a difference in roughage intake could affect electrolyte and water balance in horses during endurance type exercise. In each of 2 experiments, 2 treatments (a high hay or limited hay diet) were assigned in a crossover design so that each horse performed 2 exercise tests, 1 on each treatment. In experiment 1, diet was controlled only the night before the exercise test. Water intake was lower (P<0.001) when horses received the limited hay diet. Exercise caused a decrease in plasma chloride and potassium concentration and an increase in plasma total protein concentration, but there were no diet differences (P>0.05). Weight loss was 2.8% of body weight for both treatments. In experiment 2, horses were adapted to the diets for 7 days prior to the exercise test. Again, water intake was lower when the horses received the limited hay diet (P<0.01). During exercise, both groups lost about 4% of bodyweight, but total protein concentration was lower (P<0.05) in the horses receiving the high hay diet. Plasma potassium concentration initially increased and then decreased during exercise (P<0.05), but the horses receiving the high hay diet maintained a higher potassium concentration during exercise (P<0.05). Sodium was not affected by diet or exercise (P>0.05). Plasma chloride concentration decreased during exercise (P<0.05). The increased water intake and lower plasma protein concentration associated with the hay diet may be beneficial for horses involved in endurance type exercise.