Clinical Pharmacology Research Center, Peking Union Medical College Hospital, Beijing Key Laboratory of Clinical PK & PD Investigation for Innovative Drugs, NMPA Key Laboratory for Clinical Research and Evaluation of Drug, State Key Laboratory of Complex Severe and Rare Diseases, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100730, China.
Molecules. 2023 Feb 28;28(5):2254. doi: 10.3390/molecules28052254.
The new direct oral anticoagulants (DOACs) are increasingly used to treat and prevent thromboembolic disorders, and monitoring concentrations may be valuable in some special scenarios to prevent clinical adverse events. This study aimed to develop generic methods for the rapid and simultaneous analysis of four DOACs in human plasma and urine. Protein precipitation and one-step dilution were used to prepare the plasma and urine; the extracts were injected to ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) for analysis. Chromatographic separation was performed on an Acquity™ UPLC BEH C column (2.1 × 50 mm, 1.7 μm) with gradient elution of 7 min. A triple quadrupole tandem mass spectrometer with an electrospray ionization source was employed to analyze DOACs in a positive ion mode. The methods showed great linearity in the plasma (1500 ng/mL) and urine (1010,000 ng/mL) for all analytes (R ≥ 0.99). The intra- and inter-day precision and accuracy were within acceptance criteria. The matrix effect and extraction recovery were 86.597.5% and 93.5104.7% in the plasma, while 97.0101.9% and 85.199.5% in the urine. The stability of samples during the routine preparation and storage were within the acceptance criteria of less than ±15%. The methods developed were accurate, reliable, and simple for the rapid and simultaneous measurement of four DOACs in human plasma and urine, and successfully applied to patients and subjects with DOACs therapy for anticoagulant activity assessment.
新型直接口服抗凝剂（DOACs）越来越多地被用于治疗和预防血栓栓塞性疾病，在某些特殊情况下监测浓度可能有助于预防临床不良事件。本研究旨在开发用于快速同时分析人血浆和尿液中四种 DOACs 的通用方法。采用蛋白沉淀和一步稀释法处理血浆和尿液；提取液经超高效液相色谱-串联质谱法（UPLC-MS/MS）分析。色谱分离在 Acquity™ UPLC BEH C 柱（2.1×50mm，1.7μm）上进行，7 分钟梯度洗脱。三重四极杆串联质谱仪采用电喷雾源，正离子模式分析 DOACs。该方法在血浆（1500ng/mL）和尿液（1010,000ng/mL）中对所有分析物均表现出良好的线性关系（R≥0.99）。日内和日间精密度和准确度均符合要求。在血浆中，基质效应和提取回收率分别为 86.597.5%和 93.5104.7%，在尿液中分别为 97.0101.9%和 85.199.5%。在常规制备和储存过程中，样品的稳定性在允许偏差±15%以内。所建立的方法准确、可靠、简单，可用于快速同时测定人血浆和尿液中的四种 DOACs，并成功应用于接受 DOACs 治疗的患者和受试者，用于评估抗凝活性。
