Comparison of dihydrofolate reductase activities for folic acid in pigs and rats using in vivo and in vitro evaluation techniques

The activity of dihydrofolate reductase (DHFR) for folic acid (PteGlu) was evaluated in pigs by in vivo and in vitro experiments. The results were com pared with those of rats, Since bile secretion of reduced folates reflects the activity of DHFR for PteGlu in the body, the bile secretion rates of re duced folates including tetrahydrofolate (H(4)PteGlu), 5-methyltetrahydrofo late, 5,10-methylenetetrahydrofolate, and 10-formyltetrahydrofolate were de termined by using high-performance liquid chromatography with electrochemic al detection, after the intravenous injection of PteGlu at 1 mg/kg body wei ght to pigs and rats. Although the PteGlu injection raised the total secret ion rate of reduced folates, the total increased amount of reduced folates secreted into bile from 0 h to 2.5 h after PteGlu injection in pigs was abo ut one-tenth of that in rats. The enzyme kinetics of DHFR for PteGlu was ex amined at the physiological condition (pH 7.4 and 37 degreesC), Affinity ch romatography was applied to liver homogenates of pigs and rats to obtain DH FR. The final product of the enzyme reaction, H(4)PteGlu, was measured. The Km for pig enzyme was similar to that for rat enzyme, whereas the Vmax for the pig enzyme was less than 1/5 of that for the rat's. The comparison of the ratio of Vmax to Km between pig and rat enzymes suggests that PteGlu is a much less efficient substrate for pig liver DHFR. In short, these result s from in vivo and in vitro experiments suggest that the role of DHFR for P teGlu in pigs is physiologically much less important than that in rats.