Temperature-programmed non-aqueous electrochromatographic separation of retinyl esters

Abstract

An in-house assembled electrochromatograph was used for temperature-controlled non-aqueous electrochromatography. Reversed-phase separations of retinyl esters were performed on continuous bed columns. The continuous bed columns were prepared by sol-gel bonding of C30 material in 180 μm internal diameter fused silica capillaries. The mobile phase consisted of 2.5 mM lithium acetate in N,N-dimethylformamide-acetonitrile-methanol (2 + 7 + 1, v/v). The use of temperature programming increased the resolution of earlier-eluting compounds as well as improved the peak shape and decreased the retention time of later-eluting compounds. Separations of retinyl esters (all-trans-retinyl acetate, palmitate, heptadecanoate, stearate, oleate, and linoleate) were completed in 12 minutes. The within-day and between-day variations of retention times of all-trans-retinyl palmitate were <0.6% (n = 6) relative standard deviation (RSD) and 2.3% (n = 3) RSD, respectively. Further, the within-day and between-day variations of peak areas were < 2.7% (n = 6) RSD, and 3.4% (n = 3) RSD, respectively. The column examined was stable for more than two weeks of continuous injections of standard solutions and real samples. Lipid extracts of bearded seal (Erignathus barbatus) liver were analyzed.