Nonaqueous reverse phase liquid chromatographic determination of vitamin D2 in multivitamin tablets, using vitamin D3 as internal standard.

The technique of nonaqueous reverse phase chromatography has been successfully applied to the separation of lipophilic vitamins D2 and D3. Separation is achieved with a Zorbax-ODS column and a mobile phase of 10% methanol in acetonitrile. A predetermined amount of vitamin D3 internal standard is added to a portion of a powered tablet sample, which is then extracted and partitioned with a mixture of dimethyl sulfoxide, water, methanol, and hexane. A portion of the hexane extract is concentrated, and purified by passing it through a disposable silica cartridge to remove interfering materials. The method should also be applicable to formulations containing vitamin D3 where vitamin D2 can then be used as the internal standard. A system suitability test is included in the method.