High-pressure liquid chromatographic method for the determination of 25-hydroxycholecalciferol in cow plasma

✍ Scribed by K.T. Koshy; A.L. VanDerSlik

• Publisher: Elsevier Science
• Year: 1976
• Tongue: English
• Weight: 496 KB
• Volume: 74
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(76)90209-8

✦ Synopsis

A high-pressure liquid chromatographic (hplc) procedure was developed for the determination of 2%hydroxycholecalciferol(25-OH-DJ in cow plasma or serum. The procedure involved extraction with an ethanol-ethyl ether mixture, separation of the aqueous phase, solvent partitions, column chromatography on silica gel, and, finally, determination by reversed phase hplc on a C&bonded microparticulate silica column. The identity of the drug in the extract was confirmed by comparison with a standard by liquid, thin-layer, and gas-liquid chromatography as the free steroid and the heptafluorobutyrate and by the uv spectra and also from the mass spectrum of the heptafluorobutyrate.

Twenty-four samples from cows on normal diet (dry, lactating, and pregnant) were analyzed. The normal circulating levels of 25OH-D3 ranged from 40 to 58 r&g; mean 48 -C 5.0 rig/g. The procedure was used to analyze a limited number of human and hog samples. Human serum contained lo-20 rig/g which was in agreement with literature values. Hog serum contained 18 t&g.