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Determination of vitamin D in fortified and nonfortified milk powder and infant formula using a specific radioassay after purification by high-performance liquid chromatography

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Author: Berg, H. van den · Boshuis, P.G. · Schreurs, W.H.P.
Type:article
Date:1986
Institution: Instituut CIVO-Toxicologie en Voeding TNO
Source:Journal of Agricultural and Food Chemistry, 2, 34, 264-268
Identifier: 230262

Abstract

A reliable and sensitive radioassay is described for the determination of vitamin D in milk powder and infant formula. After saponification of the sample interfering compounds like sterols are removed by digitonin precipitation and chromatography on small columns packed with alumina. [3H] Vitamin D is added to the sample as an internal standard, to correct for losses due to previtamin D formation, as well as other procedural losses. Vitamin D is quantitated by a competitive protein binding (CPB) assay after final cleanup of the extract on a straight-phase HPLC column. Diluted sheep serum is used as the source of the binding protein, having equal affinity for both vitamins D2 and D3. With this HPLC-CPB method vitamin D can be determined with high specificity in concentrations as low as 0.1 ng/tube (ca. 0.01 IE/g). Recovery was between 90 and 110%. The coefficients of variation were 2.4 (within run) and 7.5 (between run), respectively.