Abstract
An efficient, high-performance liquid-chromatographic method with diode-array detection (HPLC–DAD) has been established for simultaneous determination of retinol, α, (β + γ), and δ-tocopherols, and α, β, γ, and δ-tocotrienols in human serum. After deproteinization, the target vitamins in serum were extracted with n-hexane and the extract was evaporated under weak nitrogen flow. The residue was redissolved in methanol and the resulting solution was used for HPLC analysis. Retinol acetate and α-tocopherol acetate were used as internal standards. The internal standard calibration curves were linear over the range of 0.010–50.0 µg mL−1, with correlation coefficients >0.999. Mean recoveries of the method were 86.3–110 %, with intra-day and inter-day relative standard deviations less than 12.2 and 14.9 %, respectively. The detection limits of the method ranged from 0.001 to 0. 002 µg mL−1, and the quantification limits ranged from 0.002 to 0.008 µg mL−1. The method was successfully applied to analysis of the target vitamins in 50 human serum samples; all the analytes were detected at concentrations ranging from <0.002–23.0 µg mL−1.
Similar content being viewed by others
References
Gerald Combs F Jr (2009) The vitamins: fundamental aspects in nutrition and health, 3rd edn. Science, Beijing
Wolf G (1996) A history of vitamin A and retinoids. FASEB J 10:1102–1107
Berry DC, Noy N (2012) Signaling by vitamin A and retinol-binding protein in regulation of insulin responses and lipid homeostasis. BBA Mol Cell Boil L 1821:168–176
Blondin SA, Yeung EH, Mumford SL, Zhang C, Browne RW, Wactawski-Wende J, Schisterman EF (2013) Serum retinol and carotenoids in association with biomarkers of insulin resistance among premenopausal women. ISRN Nutrition, doi:10.5402/2013/619516
Jafari SM, Heidari G, Nabipour I, Amirinejad R, Assadi M, Bargahi A, Sanjdideh Z (2013) Serum retinol levels are positively correlated with hemoglobin concentrations, independent of iron homeostasis: a population-based study. Nutr Res 33:279–285
Mondul AM, Watters JL, Männistö S, Weinstein SJ, Snyder K, Virtamo J, Albanes D (2011) Serum retinol and risk of prostate cancer. Am J Epidemiol 173:813–821
Steiner M (1993) Vitamin E: more than an antioxidant. Clin Cardiol 16:16–18
Boscoboinik D, Szewczyk A, Hensey C, Azzi A (1991) Inhibition of cell proliferation by alpha-tocopherol. Role of protein kinase C. J Biol Chem 266:6188–6194
Zingg JM, Azzi A (2004) Non-antioxidant activities of vitamin E. Curr Med Chem 11:1113–1133
Guan F, Li G, Liu AB, Lee MJ, Yang Z, Chen YK, Yang CS (2012) δ- and γ-Tocopherols, but not α-tocopherol, inhibit colon carcinogenesis in azoxymethane-treated F344 rats. Cancer Prev Res 5:644–654
Lu G, Xiao H, Li GX, Picinich SC, Chen YK, Liu A, Yang CS (2010) A γ-tocopherol-rich mixture of tocopherols inhibits chemically induced lung tumorigenesis in A/J mice and xenograft tumor growth. Carcinogenesis 31:687–694
Li GX, Lee MJ, Liu AB, Yang Z, Lin Y, Shih WJ, Yang CS (2011) δ-Tocopherol is more active than α- or γ-tocopherol in inhibiting lung tumorigenesis in vivo. Cancer Prev Res 4:404–413
Wagner KH, Kamal-Eldin A, Elmadfa I (2004) Gamma-tocopherol—an underestimated vitamin? Ann Nutr Metab 48:169–188
Nesaretnam K, Guthrie N, Chambers AF, Carroll KK (1995) Effect of tocotrienols on the growth of a human breast cancer cell line in culture. Lipids 30:1139–1143
Yap WN, Chang PN, Han HY, Lee DTW, Ling MT, Wong YC, Yap YL (2008) γ-Tocotrienol suppresses prostate cancer cell proliferation and invasion through multiple-signaling pathways. Br J Cancer 99:1832–1841
Wada S, Satomi Y, Murakoshi M, Noguchi N, Yoshikawa T, Nishino H (2005) Tumor suppressive effects of tocotrienol in vivo and in vitro. Cancer Lett 229:181–191
Qureshi AA, Sami SA, Salser WA, Khan FA (2002) Dose-dependent suppression of serum cholesterol by tocotrienol-rich fraction (TRF25) of rice bran in hypercholesterolemic humans. Atherosclerosis 161:199–207
Khanna S, Roy S, Ryu H, Bahadduri P, Swaan PW, Ratan RR, Sen CK (2003) Molecular basis of vitamin E action tocotrienol modulates 12-lipoxygenase, a key mediator of glutamate-induced neurodegneneration. J Biol Chem 278:43508–43515
Das KK, Jargar JG, Hattiwale SH, Yendigeri SM, Das S, Dhundasi SA (2013) Serum vitamin E (α-tocopherol) estimation: a potential biomarker of antioxidant status evaluation on heavy metal toxicities. Recent Pat Biomark 3:36–43
Hussain N, Jabeen Z, Li YL, Chen MX, Li ZL, Guo WL, Shamsi IH, Chen XY, Jiang LX (2013) Detection of tocopherol in oilseed rape (Brassica napus L.) using gas chromatography with flame ionization detector. J Integr Agric 12:803–814
Aturki Z, D’Orazio G, Fanali S (2005) Rapid assay of vitamin E in vegetable oils by reversed-phase capillary electrochromatography. Electrophoresis 26:798–803
Khan MI, Khan A, Iqbal Z, Ahmad L, Shah Y (2010) Optimization and validation of RP-LC/UV–VIS detection method for simultaneous determination of fat-soluble anti-oxidant vitamins, all-trans-retinol and α-tocopherol in human serum: effect of experimental parameters. Chromatographia 71:577–586
Siluk D, Oliveira RV, Esther-Rodriguez-Rosas M, Ling S, Bos A, Ferrucci L, Wainer IW (2007) A validated liquid chromatography method for the simultaneous determination of vitamins A and E in human plasma. J Pharm Biomed Anal 44:1001–1007
Delgado-Zamarreño MM, Bustamante-Rangel M, Sánchez-Pérez A, Carabias-Martínez R (2004) Pressurized liquid extraction prior to liquid chromatography with electrochemical detection for the analysis of vitamin E isomers in seeds and nuts. J Chromatogr A 1056:249–252
Melchert HU, Pabel E (2000) Quantitative determination of α-, β-, γ- and δ-tocopherols in human serum by high-performance liquid chromatography and gas chromatography-mass spectrometry as trimethylsilyl derivatives with a two-step sample preparation. J Chromatogr A 896:209–215
De Leenheer AP, De Bevere VO, De Ruyter MG, Claeys AE (1979) Simultaneous determination of retinol and α-tocopherol in human serum by high-performance liquid chromatography. J Chromatogr B Biomed Sci Appl 162:408–413
Yap SP, Julianto T, Wong JW, Yuen KH (1999) Simple high-performance liquid chromatographic method for the determination of tocotrienols in human plasma. J Chromatogr B Biomed Sci Appl 735:279–283
Taibi G, Nicotra CMA (2002) Development and validation of a fast and sensitive chromatographic assay for all-trans-retinol and tocopherols in human serum and plasma using liquid–liquid extraction. J Chromatogr B 780:261–267
Cimadevilla HM, Hevia D, Miar A, Mayo JC, Lombo F, Sainz RM (2014) Development and validation of a single HPLC method for determination of α-tocopherol in cell culture and in human or mouse biological samples. Biomed Chromatogr 29(6):843–852
Pyka A, Sliwiok J (2001) Chromatographic separation of tocopherols. J Chromatogr A 935:71–76
Stöggl WM, Huck CW, Scherz H, Popp M, Bonn GK (2001) Analysis of vitamin E in food and phytopharmaceutical preparations by HPLC and HPLC-APCI-MS-MS. Chromatographia 54:179–185
Lee BL, New AL, Ong CN (2003) Simultaneous determination of tocotrienols, tocopherols, retinol, and major carotenoids in human plasma. Clin Chem 49:2056–2066
Irakli MN, Samanidou VF, Papadoyannis IN (2011) Development and validation of an HPLC Method for the simultaneous determination of tocopherols, tocotrienols and carotenoids in cereals after solid-phase extraction. J Sep Sci 34:1375–1382
Tsochatzis ED, Bladenopoulos K, Papageorgiou M (2012) Determination of tocopherol and tocotrienol content of greek barley varieties under conventional and organic cultivation techniques using validated reverse phase high-performance liquid chromatography method. J Sci Food Agr 92:1732–1739
Tsochatzis ED, Tzimou-Tsitouridou R (2014) Validated RP-HPLC method for simultaneous determination of tocopherols and tocotrienols in whole grain barley using matrix solid-phase dispersion. Food Anal Method 8:392–400
Abidin SZ, Ibahim MJ, Rajikin MH, Satar NA (2012) Determination of vitamin E isomers in plasma using ultra performance liquid chromatography. Malays J Anal Sci 16:71–78
Irakli MN, Samanidou VF, Papadoyannis IN (2012) Optimization and validation of the reversed-phase high-performance liquid chromatography with fluorescence detection method for the separation of tocopherol and tocotrienol isomers in cereals, employing a novel sorbent material. J Agr Food Chem 60:2076–2082
Grebenstein N, Frank J (2012) Rapid baseline-separation of all eight tocopherols and tocotrienols by reversed-phase liquid-chromatography with a solid-core pentafluorophenyl column and their sensitive quantification in plasma and liver. J Chromatogr A 1243:39–46
Górnaś P, Siger A, Czubinski J, Dwiecki K, Segliņa D, Nogala-Kalucka M (2014) An alternative RP-HPLC method for the separation and determination of tocopherol and tocotrienol homologues as butter authenticity markers: a comparative study between two European countries. Eur J Lipid Sci Tech 116:895–903
Shammugasamy B, Ramakrishnan Y, Manan F, Muhammad K (2014) Rapid reversed-phase chromatographic method for determination of eight vitamin E Isomers and γ-oryzanols in rice bran and rice bran oil. Food Anal Method 8:649–655
Wong YF, Makahleh A, Saad B, Ibrahim MNM, Rahim AA, Brosse N (2014) UPLC method for the determination of vitamin E homologues and derivatives in vegetable oils, margarines and supplement capsules using pentafluorophenyl column. Talanta 130:299–306
Olędzka I, Kaźmierska K, Plenis A, Kamińska B, Bączek T (2015) Capillary electromigration techniques as tools for assessing the status of vitamins A, C and E in patients with cystic fibrosis. J Pharm Biomed Anal 102:45–53
Chatzimichalakis PF, Samanidou VF, Papadoyannis IN (2004) Development of a validated liquid chromatography method for the simultaneous determination of eight fat-soluble vitamins in biological fluids after solid-phase extraction. J Chromatogr B 805:289–296
Franke AA, Murphy SP, Lacey R, Custer LJ (2007) Tocopherol and tocotrienol levels of foods consumed in Hawaii. J Agric Food Chem 55:769–778
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Conflict of interest
The authors declare no conflict of interests.
Rights and permissions
About this article
Cite this article
Yang, Y., Lu, D., Zhang, J. et al. Simultaneous HPLC–DAD Determination of Retinol and Eight Vitamin E Isomers in Human Serum. Chromatographia 78, 1359–1366 (2015). https://doi.org/10.1007/s10337-015-2951-6
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10337-015-2951-6