CHARACTERIZATION OF THE METABOLIC PATHWAY OF 1,25-DIHYDROXY-16-ENE VITAMIN-D-3 IN RAT-KIDNEY BY ONLINE HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY-ELECTROSPRAY TANDEM MASS-SPECTROMETRY
B. Yeung et al., CHARACTERIZATION OF THE METABOLIC PATHWAY OF 1,25-DIHYDROXY-16-ENE VITAMIN-D-3 IN RAT-KIDNEY BY ONLINE HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY-ELECTROSPRAY TANDEM MASS-SPECTROMETRY, Biochemical pharmacology, 49(8), 1995, pp. 1099-1110
1,25-Dihydroxy-16-ene vitamin D-3 is a synthetic analog of 1,25-dihydr
oxyvitamin D-3, the most physiologically active metabolite of vitamin
D-3. The renal metabolism of 1,25-dihydroxy-16-ene vitamin D-3 had bee
n studied previously using a perfused rat kidney system [Reddy et al.,
Bioorg Med Chem Lett 3: 1879-1884, 1993], and its C-24 oxidative meta
bolic pathway had been found to be different from that of 1,25-dihydro
xyvitamin D-3 by HPLC. To further delineate the differences between th
e C-24 oxidative metabolic pathways of 1,25-dihydroxyvitamin D-3 and 1
,25-dihydroxy-16-ene vitamin D-3 in this present study we investigated
the C-24 oxidation pathway of 1,25-dihydroxy-16-ene vitamin D-3 using
a novel detection approach based on on-line capillary liquid chromato
graphy coupled to electrospray tandem mass spectrometry. Two types of
tandem mass spectrometric detection were employed to characterize the
metabolites in the kidney perfusate: (a) the preliminary screening of
metabolites by parent scan, which led to the tentative discovery of th
e production of 1,23,25-trihydroxy-24-oxo-16-ene vitamin D-3, a new me
tabolite of 1,25-dihydroxy-16-ene vitamin D-3, and (b) the pharmacokin
etic studies of; the substrate, 1,25-dihydroxy-16-ene vitamin D-3 and
its metabolites by multiple reaction monitoring. In the latter, the ma
ss spectrometric sensitivity for quantification was found to be about
20-fold better than UV detection. The current work concluded that the
C-24 oxidative metabolic pathway of 1,25-dihydroxy-16-ene vitamin D-3
closely mimicked that of its natural counterpart. Furthermore, the use
of mass spectrometry permitted the clearance rate of the starting sub
strate to be studied at a more physiological level (ng/mL or submicrom
olar level), which had not been possible previously by HPLC-UV detecti
on.