MSACL 2019 EU Abstract

Fully Automated LC-MS/MS Analysis of Anticoagulants Using a Novel Reagent Kit

Toshikazu Minohata (1, 2) , Sigrid Baumgarten (3), Franck Chevalier (1), Fanny Dayot (1), Jean-François Hoeffler (1)
(1) Alsachim SAS, Illkirch, France (2) Shimadzu Corporation, Kyoto, Japan (3) Shimadzu Europa GmbH, Germany

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Franck Chevallier (Presenter) Alsachim >> POSTER (PDF)

Relevant Financial Disclosures (within past 24 months) Conflict mitigation NOT REQUIRED.

Introduction
Novel oral anticoagulants (NOACs) are, as an alternative therapy to vitamin K antagonists, used frequently to treat and prevent thromboembolism. Their precise quantitation is necessary to identify the presence/absence of an anticoagulant effect or to determine the concentration of drug that may be helpful for patient management.
Such analysis is mainly done by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). To streamline the workflow, we have developed a complete reagent kit including stable isotope labeled standards for better precision and accuracy. Furthermore, we demonstrate here the use of a fully automated sample preparation system (CLAM-2000) coupled online with LC-MS/MS.

Methods
To demonstrate that this multi-analyte approach, with a fully automated system LC-MS/MS, can be used as a walk-away unit, we have used a novel kit for anticoagulants analysis called DOSINACOTM (Alsachim SAS). The kit includes 9 analytes (Acenocoumarol, Apixaban, Argatroban, Betrixaban, Dabigatran, Edoxaban, Fluindione, Rivaroxaban and Warfarin) and their corresponding Stable Isotope Labeled Standards. CLAM-2000 (Shimadzu, Japan) was programmed to perform protein precipitation followed by filtration and sample collection. The sample is transported from CLAM-2000 to HPLC without human intervention for LC-MS/MS analysis in 5 min.

Results
A panel analysis of 9 Anticoagulants using an automated sample preparation system, seamlessly integrated on-line with LC-MS/MS, and combined with a ready to use reagent kit, demonstrates the capability to use a standardized platform for therapeutic drug monitoring even for non-expert users of Mass Spectrometry. We carried out concurrent analysis over a range of concentrations in 10 μg /L to 500 μg /L for the Novel Oral Anticoagulants. The calibration curves that were generated had linear regression values of r2 >0.99 for each curve. The classical LC-MS method limitations are thus dramatically decreased and it is eliminating potential errors traditionally associated with manual sample handling.

Conclusions & Discussion
The novel system workflow results in easier and safer operation for users without Chromatography and Mass Spectrometry experience, thus reducing risk of exposure and improving management of patients.