LC-MS/MS methods for the detection of isoprostanes (iPF2alpha-III and 8,12-iso-iPF2alpha-VI) as biomarkers of CCl4-induced oxidative damage to hepatic tissue.
Isoprostanes are formed after peroxidation of arachidonic acid and are promising biomarkers for reactive oxygen species. A LC-MS/MS based method was developed for the quantitation of two isoprostanes (iPF2alpha-III and 8,12-iso-iPF2alpha-VI) in hepatocytes, tissue and urine samples of rats. A column switching method was used to reduce sample preparation to a minimum. Precision was 9.4% and accuracy was between 96 and 114% for free iPF2alpha-III in tissue at concentrations from 1.9 to 6.1 ng/g. Treatment of rats with CCl4 to induce oxidative stress resulted in a dose-dependent increase (two- to three-fold) of iPF2alpha-III and 8,12-iso-iPF2alpha-VI in liver and kidney. For both isoprostanes an increase of four- to five-fold was observed in CCl4 treated hepatocytes and six- to eight-fold in CCl4 treated and glutathione depleted hepatocytes. In conclusion, the presented method is sensitive, specific and precise to be applied for the quantitation of iPF2alpha-III and 8,12-iso-iPF2alpha-VI which are shown to increase by CCl4 treatment in vitro and in vivo.
Sicilia
Tina
T
Institut für Toxikologie, Universität Würzburg, Versbacher Strasse 9, 97078 Würzburg, Germany.
Mally
Angela
A
Schauer
Ute
U
Pähler
Axel
A
Völkel
Wolfgang
W
D000818:Animals; D015415:Biomarkers / Q000032:analysis / Q000378:metabolism; D002251:Carbon Tetrachloride / Q000633:toxicity*; D002853:Chromatography, Liquid / Q000295:instrumentation / Q000379:methods*; D028421:Isoprostanes / Q000032:analysis* / Q000378:metabolism; D008099:Liver / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D008297:Male; D015394:Molecular Structure; D010084:Oxidation-Reduction / Q000187:drug effects; D051381:Rats; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry / Q000295:instrumentation / Q000379:methods*
D015415:Biomarkers; D028421:Isoprostanes; D002251:Carbon Tetrachloride