Improved analysis HPLC-ESI/triple method for mapping the methotrexate by mass spectrometry

Methotrexate is a commonly utilized agent in pediatric oncology therapy. However, significant interindividual variability in its clearance can lead to delayed clearance and resultant severe toxicity. This underscores the urgent need for efficient and sensitive analytical methods to ensure patient safety. In this study, we aimed to establish a high-performance liquid chromatography-tandem mass spectrometry (HPLC-ESI/triple) method for the quantitative determination of methotrexate concentrations in plasma. This method is intended to facilitate therapeutic drug monitoring in pediatric patients, thereby allowing for a better understanding of the pharmacokinetics of methotrexate in vivo. The results indicate that the established HPLC-ESI/triple method can accurately and sensitively quantify methotrexate using only 10 μL of plasma. The recovery rates for all analytes exceeded 90 %, and matrix effects were minimal. Furthermore, our optimized method revealed that patient age significantly influences methotrexate blood concentration. Specifically, under identical dosage and administration intervals, an increase in patient age correlates with a decrease in measured blood concentration. Additionally, our findings suggest that measuring methotrexate concentrations within 24 h post-administration enhances the effectiveness of monitoring, thereby promoting rational drug use and ensuring optimal therapeutic dosing. In summary, we have conducted a comprehensive study establishing a robust method for determining methotrexate concentrations in patient plasma. The optimized HPLC-ESI/triple method is poised for widespread application in clinical practice to monitor methotrexate therapy in pediatric patients.