Determination of sterols and stanol by high performance liquid chromatography with a pre-column derivatiation and fluorescent detection

2-(9-Carbazole) acetyl chloride, a novel fluorescence derivatization reagent, has been developed for the determination of sterols and stanol. The derivatization reaction was proceeded in benzene solution at 80 degrees C with triethylamine as a catalyst. The stable fluorescent derivatives of sterols and stanol were formed within 20 min. They were separated by reversed-phase high performance liquid chromatography and detected by a fluorescence detector at an excitation wavelength of 290 nm and an emission wavelength of 360 nm. The limit of detection is 12.6 similar to 29.5 nmol/L. The analytical method has been applied to the determination of,B-Sitosterol, Stigmasterol, Campesterol and beta-Sitotanol in rat serum samples. The accuracy of the proposed method was tested by recovery measurements on spiked samples and good recovery results (101.0% similar to 103.2%) were obtained.