An innovative UHPLC-ESI-MS/MS method for the analysis of prostaglandin-E2, 8-iso-prostaglandin-F2α and 8-iso-prostaglandin-E2 in dried blood spots is proposed. The non-invasive and painless monitoring of these inflammation and oxidative stress mediators in preterm newborns suffering from Patent Ductus Arteriosus could assess their potential role as predictors of response to therapy. Analytes were extracted from dried blood spot (50 µL) with 70:30 v/v methanol:water mixture. The extracts were purified by a semi-automated micro-extraction by packed sorbent (MEPS) before injection in the UHPLC-MS/MS instrument. Analytes were separated by a reversed-phase Polaris-C18 column and a gradient elution with aqueous formic acid (0.1%) and 50:50 v/v methanol:acetonitrile mixture. Triple quadrupole mass spectrometer operated in ESI(-) and MRM mode. The fast and easy sample clean-up by MEPS guaranteed a high reproducibility (RSD < 5%) and extraction yield (90-105%) in less than ten minutes, and limited the required volume of both solvents (30-100 µL) and sample (100 µL), with respect to conventional solid phase extraction. Limits of detection below 20 pg/mL, linearity (R2 > 0.99) over three orders of magnitude and satisfactory overall intra- and inter-day precisions (RSD < 10%) were obtained. The deuterated internal standard was successfully laid on the filter paper instead of the common addition to the extraction solvent, in order to control both storage and extraction steps. Particular attention was paid to stability issues. To the best of our knowledge, this is the first analytical method combining a fast and reproducible MEPS procedure and a highly sensitive and selective UHPLC-ESI-MS/MS analysis for the determination of prostaglandin-E2, 8-iso-prostaglandin-F2α and 8-iso-prostaglandin-E2 in dried blood spots. The method resulted suitable for the monitoring of preterm newborns (<32 weeks' gestation) with Patent Ductus Arteriosus.
Micro extraction by packed sorbent coupled to liquid chromatography-tandem mass spectrometry for the analysis of prostanoids and isoprostanoids in dried blood spots of preterm newborns suffering from patent ductus arteriosus
Denise Biagini;Shaula Antoni;Silvia Ghimenti;Tommaso Lomonaco;Armando Cuttano;Rosa T. Scaramuzzo;Massimiliano Ciantelli;Fabio Di Francesco;Roger Fuoco
2019-01-01
Abstract
An innovative UHPLC-ESI-MS/MS method for the analysis of prostaglandin-E2, 8-iso-prostaglandin-F2α and 8-iso-prostaglandin-E2 in dried blood spots is proposed. The non-invasive and painless monitoring of these inflammation and oxidative stress mediators in preterm newborns suffering from Patent Ductus Arteriosus could assess their potential role as predictors of response to therapy. Analytes were extracted from dried blood spot (50 µL) with 70:30 v/v methanol:water mixture. The extracts were purified by a semi-automated micro-extraction by packed sorbent (MEPS) before injection in the UHPLC-MS/MS instrument. Analytes were separated by a reversed-phase Polaris-C18 column and a gradient elution with aqueous formic acid (0.1%) and 50:50 v/v methanol:acetonitrile mixture. Triple quadrupole mass spectrometer operated in ESI(-) and MRM mode. The fast and easy sample clean-up by MEPS guaranteed a high reproducibility (RSD < 5%) and extraction yield (90-105%) in less than ten minutes, and limited the required volume of both solvents (30-100 µL) and sample (100 µL), with respect to conventional solid phase extraction. Limits of detection below 20 pg/mL, linearity (R2 > 0.99) over three orders of magnitude and satisfactory overall intra- and inter-day precisions (RSD < 10%) were obtained. The deuterated internal standard was successfully laid on the filter paper instead of the common addition to the extraction solvent, in order to control both storage and extraction steps. Particular attention was paid to stability issues. To the best of our knowledge, this is the first analytical method combining a fast and reproducible MEPS procedure and a highly sensitive and selective UHPLC-ESI-MS/MS analysis for the determination of prostaglandin-E2, 8-iso-prostaglandin-F2α and 8-iso-prostaglandin-E2 in dried blood spots. The method resulted suitable for the monitoring of preterm newborns (<32 weeks' gestation) with Patent Ductus Arteriosus.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
