Collaborative study: determination of retinol and carotene by high-performance liquid chromatography

Seventeen laboratories participated in the analysis of 10 products for carotene and retinol by high-performance liquid chromatography. Test materials were saponified and the nonsaponifiables were extracted with petroleum ether. The extract was injected into the liquid chromatograph for determination of carotene at 450 nm/436 nm, using a C18 reversed-phase column with a mobile phase of acetonitrile:methylene chloride:methanol:water (70:20:8:2, v/v). Retinol was determined at 325 nm/313 nm by using a C18 reversed-phase column with a mobile phase of methanol:water (90:10, v/v). The biological activities of retinol and carotene expressed in international units were summed to give total activity. The repeatability (within laboratory) coefficients of variation ranged from 3.46–15.65%, and the reproducibility (among laboratories) coefficients of variation ranged from 5.34–15.77%.