Determination of vitamin A and vitamin E in human serum by ultra-high performance liquid chromatography-tandem triple quadrupole mass spectrometry

Abstract

To establish a method for the determination of vitamin A(retinol) and four active forms of vitamin E(α-tocopherol, β-tocopherol, γ-tocopherol and δ-tocopherol) in human serum by ultra-high performance liquid chromatography-tandem triple quadrupole mass spectrometry(UPLC-MS/MS). The sample was deproteinized by methanol, then extracted by n-hexane, dryness under nitrogen and followed by a reconstitution with methanol. The analysis was performed on a C_(30) column(3 mm×150 mm, 2. 6 μm), and isometric elution using 0. 1% formic acid in methanol and 5 mmol/L ammonium formate in 0. 1% formic acid as the mobile phase. The samples were determined by mass spectrometry in the positive ion mode with the multiple reaction monitoring mode, quantified by the internal standard method. Vitamin A and four active forms of vitamin E were separated within 42 minutes, and β-tocopherol and γ-tocopherol can be distinguished. The linear was good for retinol in the range of 0. 0050-2. 5 μg/mL, 0. 036-20 μg/mL for α-tocopherol, 0. 042-8. 0 μg/mL for β-tocopherol and 0. 020-10 μg/mL for the other tocopherols. The limits of detection for retinol and tocopherols were in the range of 5. 76-31. 6 ng/mL. Recoveries of retinol and tocopherols at different levels were in range of 84. 4%-118. 6%, with the relative standard deviations were 1. 22%-8. 50%(n=6). This method is fast, accurate and sensitive and the preprocessing is simple, which can be used for determination of vitamin A and four active forms of vitamin E in human serum effectively.