Determination of vitamin A and vitamin E in human serum by high performance liquid chromatography with fluorescence detection

OBJECTIVE To establish a method for the simultaneous and rapid determination of vitamin A and vitamin E of different configurations in human serum by high performance liquid chromatography(HPLC) with multi-wavelength fluorescence detector. METHODS The serum was mixed after adding internal standard, and acetonitrile was added for protein precipitation. The mixture was centrifuged after extraction with n-hexane. The n-hexane layer was dried by N_2 flow, the residue was dissolved with methanol. The HPLC system was consisted of WATERS Symmetry C_(18) column(4. 6 mm × 250 mm, 5 μm) and the mobile phase was methanol. The column temperature was 30 ℃ and fluorescence detector with online wavelength conversion method was carried out for the quantitative detection. RESULTS The liner range of determination of vitamin A, α-vitamin E, β-vitamin E and δ-vitamin E were 0. 050-2. 0 μg/mL, 0. 50-50 μg/mL, 0. 050-5. 0 μg/mL and 0. 050-5. 0 μg/mL, respectively(r≥0. 996). The minimum detection limits of the method for vitamin A and vitamin E were all 0. 02 μg/mL. The intraday and interday relative standard deviations(RSDs) were less than 3% at high, medium and low concentrations. The recoveries of the samples at the three concentrations were 91. 2%-107. 5%, and the RSDs were less than 10%. CONCLUSION This method is simple and accurate, with higher sensitivity than using UV detector and can be used for the simultaneous determination of vitamin A and vitamin E of different configurations in serum, and is suitable for rapid detection of batch serum samples.