A Chromatographic Approach for Simultaneous Quantification of Multiple Anticoagulants in Human Plasma Samples

Area of Science:

• Analytical Chemistry
• Pharmacology

Background:

• Anticoagulants are crucial for managing thromboembolic disorders.
• Warfarin requires strict monitoring due to its narrow therapeutic window and adverse event risk.
• Direct oral anticoagulants (DOACs) offer improved safety but may need monitoring in specific clinical scenarios.

Purpose of the Study:

• To develop and validate a high-performance liquid chromatography (HPLC) method for simultaneous quantification of warfarin, dabigatran, rivaroxaban, and apixaban in human plasma.
• To establish a reliable analytical tool for monitoring these anticoagulants in clinical practice.

Main Methods:

• Simultaneous quantification of four anticoagulants using HPLC with UV/Vis detection.
• Method development involved optimizing column type (monolithic phenyl column), elution conditions (gradient), and injection volume.
• Validation included assessing selectivity, linearity, accuracy, precision, limit of detection (LOD), and limit of quantification (LOQ).

Main Results:

• The developed HPLC method achieved efficient separation of the four anticoagulants within 12 minutes.
• The method demonstrated excellent linearity, accuracy (85.7%-115.0%), and precision (CV ≤ 9.4%).
• Analyte stability was confirmed at room temperature for 24 hours, with minor stability issues noted for dabigatran after freeze-thaw cycles.

Conclusions:

• A rapid, selective, and validated HPLC method for simultaneous quantification of four major anticoagulants in human plasma was successfully developed.
• This method provides a valuable tool for therapeutic drug monitoring, particularly in complex clinical situations requiring precise anticoagulant level assessment.