Simultaneous determination of quercetin,kaempferol and isorhamnetin in dog plasma by HPLC-MS/MS

AIM:A rapid,sensitive and highly selective liquid chromatography-tandem mass spectrometry method was developed and validated for simultaneous determination of quercetin,kaempferol and isorhamnetin in dog plasma.METHODS:Quercetin,kaempferol and isorhamnetin conjugates were hydrolysed chemically.The analytes were extracted from plasma samples by liquid-liquid extraction,separated on a Luna ODS-2 column(150 mm×2.1 mm I.D.,5 μm particle size),and detected by tandem mass spectrometry with a Finnigan IonSpray ionization interface.The mobile phase consisted of 0.1% aqueous formic acid(A)and gradient-grade acetonitrile(B).RESULTS:The calibration curves for quercetin,kaempferol and isorhamnetin were linear in concentration ranges of 0.5-100.0 ng/mL in dog plasma.The method has a lower limit of quantification(LLOQ)of 0.5 ng/mL for all the three flavonols.The intra-and inter-day precisions,expressed as the R.S.D.,were less than 7.3%,6.2% and 6.4% for quercetin,kaempferol and isorhamnetin,respectively,and the recovery was more than 70%,66% and 70%,respectively.The application of this assay was examined in a preliminary pharmacokinetic study of quercetin,kaempferol and isorhamnetin in beagle dogs after oral administration of 6 Ginkgo biloba tablets.CONCLUSION:The present method was suitable for determining quetcetin,kaempferol and isorhamnetin in dog plasma.