Thin-layer chromatographic separation of a number of bile acids with mobile phases based on surfactants

Abstract

The aim of this study was the separation of cholic, ursodeoxycholic, chenodeoxycholic, deoxycholic, and lithocholic bile acids by thin-layer chromatography (TLC) with mobile phases based on surfactants. The effect of surfactant type and concentration, the pH of the mobile phase, the type and amount of the mobile phase modifier, and the effect of temperature and type of material backed silica gel TLC layers on the investigated bile acid separation were studied. The separation was carried out on normal-phase TLC plates Sorbfil PTLC-P-A-UV-254 and Sorbfil PTLC-AF-A-UV-254. The acid zones after separation were visualized by using sulfuric acid solutions followed by heating the TLC plate and acid spots observing under 365-nm ultraviolet (UV) light. Mobile phases based on surfactants, namely, cationic cetylpyridinium chloride, anionic sodium dodecyl sulfate, and nonionic Tween-80, were used. Aliphatic alcohols 1-propanol, 1-butanol, and 1-pentanol as well as the nonionic surfactant Tween-80 were used as modifiers of the mobile phase. The optimal separation of the studied acids was achieved using the two-dimensional TLC method with sequential elution with mobile phases based on cetylpyridinium chloride, first with the addition of 1-butanol and then, perpendicular to the initial direction, with the addition of 1-pentanol. The proposed method for the bile acid separation can be used in the analysis of human biological fluids and drugs.