Abstract
1α‐Hydroxy‐23 carboxy‐24,25,26,27‐tetranorvitamin D~3~ (calcitroic acid) is known to be the major water‐soluble metabolite produced during the deactivation of 1,25‐(OH)~2~D~3~. This deactivation process is carried out exclusively by the multicatalytic enzyme CYP24 and involves a series of oxidation reactions at C~24~ and C~23~ leading to side‐chain cleavage and, ultimately, formation of the calcitroic acid. Like 1,25‐(OH)~2~D~3~, 1α,25‐1,25‐(OH)~2~D~2~ is also known to undergo side‐chain oxidation and side‐chain cleavage to form calcitroic acid (Zimmerman et al. [2001]. 1,25‐(OH)~2~D~2~ differs from 1,25‐(OH)~2~D~3~ by the presence of a double bond at C~22~ and a methyl group at C~24~. To date, there have been no studies detailing the participation of CYP24 in the production of calcitroic acid from 1,25‐(OH)~2~D~2~. We, therefore, studied the metabolism of 1,25‐(OH)~2~D~3~ and 1,25‐(OH)~2~D~2~ using a purified rat CYP24 system. Lipid and aqueous‐soluble metabolites were prepared for characterization. Aqueous‐soluble metabolites were subjected to reverse‐phase high‐pressure liquid chromatography (HPLC) analysis. As expected, 1,23(OH)~2~‐24,25,26,27‐tetranor D and calcitroic acid were the major lipid and aqueous‐soluble metabolites, respectively, when 1,25‐(OH)~2~D~3~ was used as substrate. However, when 1,25‐(OH)~2~D~2~ was used as substrate, 1,24(R),25‐(OH)~3~D~2~ was the major lipid‐soluble metabolite with no evidence for the production of either 1,23(OH)~2~‐24,25,26,27‐tetranor D or calcitroic acid. Apparently, the CYP24 was able to 24‐hydroxylate 1,25‐(OH)~2~D~2~, but was unable to effect further changes, which would result in side‐chain cleavage. These data suggest that the presence of either the double bond at C~22~ or the C~24~ methyl group impedes the metabolism of 1,25‐(OH)~2~D~2~ to calcitroic acid by CYP24 and that enzymes other than CYP24 are required to effect this process. J. Cell. Biochem. 88: 282–285, 2003. Published 2002 Wiley‐Liss, Inc.