Date Published: February 7, 2017
Publisher: Public Library of Science
Author(s): Priscilla Bento Matos Derogis, Livia Rentas Sanches, Valdir Fernandes de Aranda, Marjorie Paris Colombini, Cristóvão Luis Pitangueira Mangueira, Marcelo Katz, Adriana Caschera Leme Faulhaber, Claudio Ernesto Albers Mendes, Carlos Eduardo dos Santos Ferreira, Carolina Nunes França, João Carlos de Campos Guerra, Pablo Garcia de Frutos.
Rivaroxaban is an oral direct factor Xa inhibitor, therapeutically indicated in the treatment of thromboembolic diseases. As other new oral anticoagulants, routine monitoring of rivaroxaban is not necessary, but important in some clinical circumstances. In our study a high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method was validated to measure rivaroxaban plasmatic concentration. Our method used a simple sample preparation, protein precipitation, and a fast chromatographic run. It was developed a precise and accurate method, with a linear range from 2 to 500 ng/mL, and a lower limit of quantification of 4 pg on column. The new method was compared to a reference method (anti-factor Xa activity) and both presented a good correlation (r = 0.98, p < 0.001). In addition, we validated hemolytic, icteric or lipemic plasma samples for rivaroxaban measurement by HPLC-MS/MS without interferences. The chromogenic and HPLC-MS/MS methods were highly correlated and should be used as clinical tools for drug monitoring. The method was applied successfully in a group of 49 real-life patients, which allowed an accurate determination of rivaroxaban in peak and trough levels.
Rivaroxaban (Xarelto, Janssen Pharmaceuticals, Titusville, New Jersey) is an oral anticoagulant that directly inhibits activated factor X (FXa). It is indicated for the prevention of thromboembolism in patients with atrial fibrillation , the treatment of deep-vein thrombosis (DVT)  and the prevention of recurrent DVT and pulmonary embolism (PE) following an acute DVT in adults [3, 4], in prevention of venous thromboembolism (VTE) in patients undergoing total knee or hip replacement surgery [5–7].
HPLC-MS/MS is becoming an important tool for clinical laboratories because it is a very specific, selective and sensitive technique. Additionally, permits a wide range of applications and it is possible to obtain a large number of quantitative or qualitative results [24, 25]. Diverse compounds, as anticoagulant agents, can be directly measured by an HPLC-MS/MS method. This technique has much better selectivity than coagulation activity-based assays, enabling specific detection and quantification of coagulation inhibitors .
In the present work it was developed and validated a fast, accurate and precise HPLC-MS/MS method for rivaroxaban determination. A simple sample preparation and a relative short chromatographic run time were used. The method was successfully applied to determine rivaroxaban concentrations two hours before and two hours after drug intake. Hemolytic, icteric or lipemic plasma samples can be used in rivaroxaban measurement by HPLC-MS/MS without interferences. The chromogenic and HPLC-MS/MS methods were highly correlated and should be used as clinical tools for drug monitoring.