Gas chromatography-mass spectrometry analysis of vitamin E and its oxidation products.
To facilitate studies of vitamin E ( alpha-tocopherol) antioxidant actions in tissues, we have developed a stable isotope dilution capillary gas chromatography-mass spectrometry assay for alpha-tocopherol and its three principal oxidation products, alpha-tocopherolquinone, 5,6-epoxy-alpha-tocopherolquinone, 2, 3-epoxy-alpha-tocopherolquinone, and for alpha-tocopherolhydroquinone, a reduction product of alpha-tocopherolquinone. Deuterium-labeled internal standards (5, 7-[2H3-methyl]-alpha-tocopherol, 2,6-[2H3-methyl]- alpha-tocopherolquinone, 2,6-[2H3-methyl]-5,6-epoxy- alpha-tocopherolquinone, 2,6-[2H3-methyl]-2,3-epoxy- alpha-tocopherolquinone, and 2-[2H3-methyl]- alpha-tocopherolhydroquinone are added to cell or tissue homogenates. The products then are extracted and converted to O-trimethylsilyl derivatives. Products are analyzed by capillary gas chromatography with on-column injection and detected by selected ion monitoring of characteristic fragment ions in electron ionization mode. Standard curves were linear from 25 fmol to 2 pmol for products and from 25 fmol to 4 pmol for alpha-tocopherol. The use of 2H6- and 2H3-internal standards for alpha-tocopherolquinone and alpha-tocopherolhydroquinone permits simultaneous analysis of both products despite possible redox interconversion during sample workup. alpha-Tocopherol oxidized in microsomes treated with azo-bis(amidinopropane HCl) was quantitatively accounted for as the epoxyquinones, alpha-tocopherolquinone, and alpha-tocopherolhydroquinone. However, over half of the oxidation products were present in microsomes as acid-labile tocopherone precursors. This method permits comprehensive assessment of vitamin E status in tissues and quantitative studies of vitamin E antioxidant actions and turnover.
Liebler
D C
DC
Department of Pharmacology and Toxicology, University of Arizona, Tucson, 85721, USA.
Burr
J A
JA
Philips
L
L
Ham
A J
AJ
D000818:Animals; D000975:Antioxidants / Q000032:analysis / Q000737:chemistry; D003903:Deuterium; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D008297:Male; D008862:Microsomes, Liver / Q000378:metabolism; D010084:Oxidation-Reduction; D051381:Rats; D017207:Rats, Sprague-Dawley; D014810:Vitamin E / Q000032:analysis* / Q000737:chemistry
D000975:Antioxidants; D014810:Vitamin E; D003903:Deuterium