Analysis of fat-soluble vitamins. XXI. High pressure liquid chromatographic assay methods for vitamin D in vitamin D concentrates.

Two high pressure liquid chromatographic methods, a straight and a reverse phase system, were developed and compared with the official (chemical) AOAC method for vitamin D concentrates. The effects of the systematic error and the reproducibility of using an internal or external standard were studied, as well as the effect of using peak height or peak height X retention time for calculating the potential vitamin D content. A method is given for determining the conversion factor to calculate previtamin D as vitamin D. Based on the results of the comparison, the following conditions were selected for collaborative study: straight phase, amyl alcohol-hexane mobile phase, external standard, and calculation of potency by peak height.