Development and validation of an analytical method for the determination of direct oral anticoagulants (DOAC) and the direct thrombin-inhibitor argatroban by HPLC-MS/MS

开发并验证通过 HPLC-MS/MS 测定直接口服抗凝剂 (DOAC) 和直接凝血酶抑制剂阿加曲班的分析方法

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作者:Lea Brückner, Jan Beyer-Westendorf, Oliver Tiebel, Jörg Pietsch

Abstract

Since direct oral anticoagulants (DOAC) are administered frequently to an elderly, co-morbid population, medical emergencies including trauma, acute bleeding or organ failure are not uncommon. In these situations, the type, dosage or the time of last intake of anticoagulants is often unknown and single substance analysis by functional tests is only possible if the substance contained in the sample is known. A reliable and validated toxicology screen of DOAC and argatroban would be helpful inform not only attending physicians in the emergency department but also law enforcement and courts of justice. After precipitation with acetone, HPLC separation was achieved on a Phenomenex Luna Pentafluorophenyl Colum using acetonitrile-water (90:10, v/v) as mobile phase system. Detection was performed using a 3200 Q Trap mass spectrometer (AB Sciex). For analysis MRM Scans (MS/MS) with positive ionization were chosen. The method was validated for blank serum as the matrix of choice. Limits of detection are between 0.5 and 1.0 ng/mL, limits of quantification are between 1.9 and 3.6 ng/mL and recoveries are above 60%. The applicability of the method was demonstrated by the determination of DOAC in body fluids from forensic cases and in therapeutic drug monitoring. The rapid simultaneous detection and quantification of apixaban, argatroban, dabigatran etexilate, dabigatran, edoxaban and rivaroxaban in body fluids by HPLC-MS/MS closes an important gap in emergency toxicology.

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