Comparison of gas chromatography-mass spectrometry and liquid chromatography-tandem mass spectrometry methods to quantify alpha-tocopherol and alpha-tocopherolquinone levels in human plasma.
Two mass spectrometric methods were established for the quantitative analyses of alpha-tocopherol (TH) and its oxidation product alpha-tocopherolquinone (TQ) in human plasma. Both methods make use of isotopically labeled internal standards of different levels of deuteration (d3-TH and d6-TQ). Plasma (100 microl) was saponified in the presence of a mixture of antioxidants, and then TH and TQ were extracted with hexane. With the GC-MS method, the analytes were first converted into O-trimethylsilyl derivatives before analysis in the selective ion monitoring mode. The derivatization procedure led to the quantitative conversion of TQ into the O-trimethylsilyl derivative of tocopherolhydroquinone, giving rise to a more stable molecule with less fragmentation than for TQ. The increased stability of the molecule resulted in an enhanced contribution of the base peak to the total observed ions and therefore an increased sensitivity of the base peak for quantification. With the liquid chromatography-tandem mass spectrometry (LC-MS/MS) method, TH and TQ were detected by multiple reaction monitoring after positive electrospray ionization. The GC-MS and LC-MS/MS methods showed nearly the same accuracy (>95%) and the same within-day precisions, with less than 5 and 10% for TH and TQ, respectively. The between-day precision and the limit of quantification for TQ in plasma were better by LC-MS/MS (4%; 3 nM) than by GC-MS (21%; 10 nM). Analysis and method validation were carried out with plasma samples obtained from a male volunteer pre- and postexercise. Both techniques showed that the ratio of TQ/TH was elevated by 35% immediately after exercise and had returned to basal levels when measured 24 h later.
Mottier
Pascal
P
Nestlé Research Center, Nestec Ltd., Vers-chez-les-Blanc, Lausanne 26, 1000, Switzerland. pascal.mottier@rdls.nestle.com
Gremaud
Eric
E
Guy
Philippe A
PA
Turesky
Robert J
RJ
D002853:Chromatography, Liquid / Q000379:methods; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D006801:Humans; D008297:Male; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D014810:Vitamin E / Q000031:analogs & derivatives* / Q000097:blood*; D024502:alpha-Tocopherol / Q000097:blood*
D014810:Vitamin E; C002421:tocopherylquinone; D024502:alpha-Tocopherol