Metabolism of [3α-3H] 25-hydroxyvitamin D2 in kidneys isolated from normal and vitamin D2-intoxicated rats

With the availability of A-ring labelled 25OHD(2), [3 alpha-H-3] 25OHD(2), we have performed the present study to examine the metabolism of 25OHD(2) using physiological substrate concentrations in perfused kidneys isolated from both normal and vitamin D-2-intoxicated rats. Our results indicate that [3 alpha-H-3] 25OHD(2) is metabolized into both 24(S),25,28-trihydroxyvitamin D-2 [24(S),25,28(OH)(3)D-2] and 24(R),25,26-trihydroxyvitamin D-2 [24(R), 25,26(OH)(3)D-2], and the amounts of these two metabolites produced in the kidney of vitamin D-2-intoxicated rat were about 3-5 times higher than those produced in the kidney of normal rat, Similar results were also obtained with rat kidney homogenates incubated with [3 alpha-H-3] 25OHD(2). Furthermore, we noted that the production of both 24(S),25,28(OH)(3)D-2 and 24(R),25,26(OH)(3)D-2 in the kidney homogenates of vitamin D-2-intoxicated rats increased with the time of incubation and then subsequently decreased, The decrease in both 24(S),25,28(OH)(3)D-2 and 24(R),25,26(OH)(3)D-2 coincided with an increase in the fraction of total radioactivity distributed in the aqueous phase of the kidney homogenates. This finding suggested the possibility of further metabolism of 24(S),25,28(OH)(3)D-2 and 24(R), 25,26(OH)(3)D-2 into polar water-soluble metabolite(s). We then measured the radioactivity in the aqueous phase of kidney homogenates of both normal and vitamin D-2-intoxicated rats incubated with [3 alpha-H-3] 25OHD(2). It was noted that the amount of radioactivity in the aqueous phase of kidney homogenates of vitamin D-2-intoxicated rats is higher than that present in the aqueous phase of kidney homogenates of normal rats, Thus, our study provides evidence for the first time for the formation of both 24(S),25,28(OH)(3)D-2 and 24(R),25, 26(OH)(3)D-2 under physiological conditions, and the possibility of their further metabolism into as yet unidentified polar water-soluble metabolite(s). As the formation of all these metabolites is increased in the kidney of vitamin D-2-intoxicated rats when compared to normal rats, it appears that the increased rate of metabolism of 25OHD(2) during hypervitaminosis D-2 plays a significant role in the deactivation of 25OHD(2).