A gas-chromatographic method for the determination of vitamin D in fortified non-fat dried milk

Abstract

A gas-chromatographic method with temperature programming has been adapted for determining vitamin D in fortified non-fat dried milk by using an acetone-ethanol mixture for protein precipitation and ascorbic acid as an anti-oxidant, followed by extraction with chloroform, with a butylated hydroxyanisole-2,6-di-t-butyl-p-cresol mixture as the second anti-oxidant.

Sample purification is accomplished by using multi-bore silicic acid column chromatography with gradient elution, followed by antimony trichloride treatment and 2-stage silicic acid micro-column chromatography.

Vitamins D₂ and D₃ are differentiated and determined by gas-liquid chromatography, with one vitamin as internal standard for the other. An over-all average recovery of 104 per cent. of vitamin D₂ was obtained when vitamin D₂ was used as the internal standard on samples fortified at levels of 10 to 50 µg per 100 g of non-fat dried milk. Approximate total analysis time was 7 hours.