23-carboxy-24,25,26,27-tetranorvitamin D3 (calcioic acid) and 24-carboxy-25,26,27-trinorvitamin D3 (cholacalcioic acid): end products of 25-hydroxyvitamin D3 metabolism in rat kidney through C-24 oxidation pathway.
During the past two and half decades the elucidation of the metabolic pathways of 25OHD(3) and its active metabolite 1alpha,25(OH)(2)D(3) progressed in parallel. In spite of many advances in this area of vitamin D research, the unequivocal identification of the end products of 25OHD(3) metabolism through C-24 oxidation pathway has not been achieved. It is now well established that both 25OHD(3) and 1alpha,25(OH)(2)D(3) are metabolized through the same C-24 oxidation pathway initiated by the enzyme 24-hydroxylase (CYP24A1). Based on the information that the end product of 1alpha,25(OH)(2)D(3) metabolism through C-24 oxidation pathway is 1alpha-OH-23- COOH-24,25,26,27-tetranor D(3) or calcitroic acid; the metabolism of 25OHD(3) into 23-COOH-24,25,26,27-tetranor D(3) has been assumed. Furthermore, a previous study indicated 24-COOH-25,26,27-trinor D(3) as a water soluble metabolite of 24R,25(OH)(2)D(3) produced in rat kidney homogenates. Therefore, 24-COOH-25,26,27-trinor D(3) was also assumed as another end product of 25OHD(3) metabolism through C-24 oxidation pathway. We embarked on our present study to provide unequivocal proof for these assumptions. We first studied the metabolism of 25OHD(3) at low substrate concentration (3x10(-10)M) using [1,2-(3)H]25OHD(3) as the substrate in the perfused rat kidneys isolated from both normal and vitamin D(3) intoxicated rats. A highly polar water soluble metabolite, labeled as metabolite X was isolated from the kidney perfusate. The amount of metabolite X produced in the kidney of a vitamin D intoxicated rat was about seven times higher than that produced in the kidney of a normal rat. We then produced metabolite X in a quantity sufficient for its structure identification by perfusing kidneys isolated from vitamin D intoxicated rats with high substrate concentration of 25OHD(3) (5x10(-6)M). Using the techniques of electron impact and thermospray mass spectrometry, we established that the metabolite X contained both 23-COOH-24,25,26,27-tetranor D(3) and 24-COOH-25,26,27-trinor D(3) in a ratio of 4:1. The same metabolite X containing both acids in the same ratio of 4:1 was also produced when 24R,25(OH)(2)D(3) was used as the starting substrate. Previously, the trivial name of cholacalcioic acid was assigned to 24-COOH-25,26,27-trinorvitamin D(3). Using the same guidelines, we now assign the trivial name of calcioic acid to 23-COOH-24,25,26,27-tetranor D(3). In summary, for the first time our study provides unequivocal evidence to indicate that both calcioic and cholacalcioic acids as the end products of 25OHD(3) metabolism in rat kidney through C-24 oxidation pathway.
Reddy
G Satyanarayana
GS
Epimer LLC, 4 Richmond Square, Suite 500, Providence, RI 02906, USA. Satya_Reddy@Brown.edu
Omdahl
John L
JL
Robinson
Matthew
M
Wang
Guochun
G
Palmore
G Tayhas R
GT
Vicchio
Domenick
D
Yergey
Alfred L
AL
Tserng
Kou-Yi
KY
Uskokovic
Milan R
MR
D000818:Animals; D053898:Biosynthetic Pathways; D002112:Calcifediol / Q000737:chemistry / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D006887:Hydroxycholecalciferols / Q000737:chemistry / Q000378:metabolism*; D066298:In Vitro Techniques; D007668:Kidney / Q000378:metabolism*; D008055:Lipids / Q000737:chemistry; D008297:Male; D008956:Models, Chemical; D015394:Molecular Structure; D010084:Oxidation-Reduction; D010477:Perfusion; D051381:Rats; D017207:Rats, Sprague-Dawley; D012995:Solubility; D014867:Water / Q000737:chemistry
D006887:Hydroxycholecalciferols; D008055:Lipids; D014867:Water; C043003:24,25,26,27-tetranor-23-hydroxyvitamin D3; D002112:Calcifediol