Fluorometric Determination of Vitamin Constituents in Human Plasma Using Ultra Performance Liquid Chromatography
Edward C Bell*, Mathew John, Rodney J Hughes and Thu Pham
Department of Pharmaceutical Sciences, College of Pharmacy and Health Sciences, Southern University, 3100 Cleburne Street, Houston, TX 77004, USA
- *Corresponding Author:
- Edward C Bell
3100 Cleburne Street, Houston
TX 77004, USA
E-mail: [email protected]
Received date: September 19, 2012; Accepted date: October 16, 2012; Published date: October 20, 2012
Citation: Bel EC, John M, Hughes RJ, Pham T (2012) Fluorometric Determination of Vitamin Constituents in Human Plasma Using Ultra Performance Liquid Chromatography. J Chromat Separation Techniq 3:143. doi:10.4172/2157-7064.1000143
Copyright: © 2012 Bel EC, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Purpose: A rapid, selective and sensitive ultra performance liquid chromatography method has been developed for the detection and quantification of several vitamins in human plasma. Methods: Alpha tocopherol, gamma tocopherol, and retinol are assayed using fluorescence detection. Excitation/emission wavelengths are 295 nm/330 nm and 325 nm/470 nm for the analysis of both tocopherols and retinol, respectively. Retinol acetate is employed as the internal standard. The reversed phase method incorporates gradient elution with a mobile phase consisting of methanol and acetonitrile. Separation of vitamin compounds is achieved using a bridged ethyl-hybrid C18 column. Results: The retention times for retinol, retinol acetate, gamma tocopherol and alpha tocopherol are 1.6 minutes, 1.8 minutes, 3.9 minutes and 4.3 minutes, respectively. The limits of quantification for retinol, gamma tocopherol and alpha tocopherol, were 0.02 μg/ml, 0.02 μg/ml, and 0.1 μg/ml, respectively. Conclusion: The assay method is suitable for the analysis of these vitamins in human plasma following the ingestion of foods fortified with these fat soluble vitamins.