High performance liquid chromatographic determination of cholecalciferol (vitamin D3) in food--a comparison with a bioassay method.

The vitamin D3 levels in fortified milk (0.5% fat) and other low fat products, such as infant formulas, porridges and gruels were determined chemically and biologically. The chemical method was based on a semi-preparative straight phase clean-up procedure, followed by an analytical reversed phase quantifying step. The biological tests were carried out using the curative x-ray method. The agreement between the results of the two method was good. The overall recoveries for the HPLC-method were 95% (SD = 2.8%, n = 13) at concentrations of 10 micrograms per 100 g and 5 micrograms per litre for fortified products and milk, respectively. The minimum detectable amount was 250 pg vitamin D3. The pre-vitamin D3 formation was studied during saponification conditions and the isomer ratio was estimated.