Quantitative determination of 25‐hydroxyvitamin D3 3‐sulphate in human plasma using high performance liquid chromatography

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Abstract
The quantitative determination of 25-hydroxyvitamin D3 3-sulphate in human plasma was completed using reversed phase high performace liquid chromatography with UV detection (265 nm) and an internal standard method. The vitamin D sulphate fraction was obtained from a plasma specimen with the combined use of a Bond Elut C18 cartridge for solid-phase extraction and a piperidinohydroxypropyl Sephadex LH-20 column for lipophilic ion-exchange chromatography. Separation of the compounds was performed on a YMC-Pack ODS-AM column. The limit of quantitation was 5 ng/mL and the assay was linear from 5 to 50 ng/mL. The proposed method is satisfactory in its accuracy and precision.