LC-MS/MS Method Validation for Quantification of Nirmatrelvir in Human Plasma

Journal article

Nirmatrelvir, a key antiviral agent in the treatment of COVID-19, requires accurate and reliable monitoring of drug levels to optimize therapeutic efficacy. In this study, we developed and validated a sensitive and specific LC-MS/MS method for the quantification of nirmatrelvir in human plasma. The method includes nirmatrelvir-D9 as an internal standard, with quantification achieved using selected reaction monitoring in positive electrospray ionization mode, targeting m/z 500.3 ⟶ 110.1 for nirmatrelvir and m/z 509.3 ⟶ 110.1 for nirmatrelvir-D9. Sample preparation involved a simple phospholipid removal step using 96-well plate and automated liquid handler, which improved efficiency in a high-throughput process. The validated method, following international bioanalytical guidelines, demonstrated a linear range from 10.9 to 3013 ng/mL. Intra- and interassay precisions were both below 15%. All validation tests meet the criteria for matrix effect, carryover, dilution integrity, and stability. The method offers a rapid analysis time of 2 min per sample and provides highly accurate, reproducible results, making it a valuable tool for evaluating the pharmacokinetics of nirmatrelvir in clinical settings.Trial registrationClinicalTrials.gov identifier: NCT05041907.

Authors: Kaewkhao, N; Tarning, J; Blessborn, D

• Publication status: Published
• Peer review status: Peer reviewed
• Publisher: Wiley
• Journal: International Journal of Analytical Chemistry
• Volume: 2025
• Issue: 1
• Pages: 6625833
• Publication date: 2025-11-17
• Acceptance date: 2025-09-19
• DOI: 10.1155/ianc/6625833
• EISSN: 1687-8779
• ISSN: 1687-8760
• Pmid: 41321979
• Language: English
• Keywords: Nirmatrelvir; Validation; Covid-19; LC-MS/MS; Plasma