The effect of endogenous increases in plasma lactate concentration on substrate mobilization during prolonged submaximal leg work was studied in four male subjects who performed supramaximal arm work on an intermittent basis. For this experiment (L-A), the arms were worked four times for 1-min periods at 5-min intervals beginning 60 min after the start of exercise. A ride of similar duration (120 min) and intensity (59% Vo2max) served as the control (L). The progressive rise in plasma lactate (2.1 to 7.2 mM) following arm work in the L-A condition coincided with a significant depression (P<0.05) in plasma free fatty acids (0.938 to 0.720 mM) during the same time period. In contrast, free fatty acids continued to rise in the L experiment. Blood glucose, pyruvate, and glycerol concentrations were also different between the two experiments, with blood glucose showing the most pronounced (5.2 to 8.2 mM) effect of the L-A condition. During the 30-min period of submaximal exercise following the arm work (L-A) the values of these blood constituents tended to approach the control (L) experiment. In the cases of blood glycerol and plasma free fatty acids no difference was found between the two experiments following 120 min of work. Metabolite concentrations in the vastus lateralis muscle were not different for ATP and glucose 6-phophate; however, a higher lactate (P<0.05) and a lower creatine phosphate (P<0.05) were observed following the arm work. A greater glycogen utilization (P<0.05) occurred during the L-A condition, but no difference between the two experiments was apparent in the glycogen depletion pattern of the fast- and slow-twitch fibers of the vastus lateralis muscle.