Comparison of LC-MS3 and LC-MRM strategy for quantification of methotrexate in human plasma and its application in therapeutic drug monitoring

Lei Yin; Zhengchao Ji; Haiwei Cao; Yanyan Li; Jing Huang

doi:10.1016/j.jpba.2021.114345

Comparison of LC-MS³ and LC-MRM strategy for quantification of methotrexate in human plasma and its application in therapeutic drug monitoring

J Pharm Biomed Anal. 2021 Oct 25:205:114345. doi: 10.1016/j.jpba.2021.114345. Epub 2021 Aug 30.

Authors

Lei Yin¹, Zhengchao Ji², Haiwei Cao², Yanyan Li³, Jing Huang⁴

Affiliations

¹ Department of Laboratory Medicine, The First Hospital of Jilin University, Jilin University, Xinmin Street, Changchun 130061, PR China; School of Life and Pharmaceutical Sciences, Dalian University of Technology, Panjin 124221, PR China. Electronic address: leiyin@dlut.edu.cn.
² Department of Laboratory Medicine, The First Hospital of Jilin University, Jilin University, Xinmin Street, Changchun 130061, PR China.
³ Department of Laboratory Medicine, The First Hospital of Jilin University, Jilin University, Xinmin Street, Changchun 130061, PR China. Electronic address: huangj@jlu.edu.cn.
⁴ Department of Laboratory Medicine, The First Hospital of Jilin University, Jilin University, Xinmin Street, Changchun 130061, PR China. Electronic address: yanyanli@jlu.edu.cn.

PMID: 34492453
DOI: 10.1016/j.jpba.2021.114345

Abstract

A simple, highly selective and high throughput liquid chromatography tandem mass spectrometry cubed (LC/MS³) method was developed and validated for quantification of methotrexate in human plasma. The MS³ detection is a scanning mode of QTrap MS systems or ion trap MS systems. After simple protein precipitation with methanol, methotrexate and methotrexate-d₃ were separated on an Agilent Poroshell 120 SB-C18 column (4.6 × 50 mm, 2.7 µm) using isocratic elution with a mobile phase consisting of 60% 0.1% formic acid in water and 40% 0.1% formic acid in acetonitrile. The flow rate is 0.8 mL/min. MS³ detection in positive ion mode used the MRM³ transitions at m/z 455.2→308.2→175.1 for quantification of methotrexate and m/z 458.2→311.2→175.1 for quantification of methotrexate-d₃. The total run time was only 3 min for each sample. The LC/MS³ assay was linear in the concentration range 10-3000 ng/mL(R² ≥ 0.995) and the intra- and inter-day accuracies were< 3.72% and precisions were< 7.78% at all concentrations. The absolute recoveries (%) and matrix effect (%) for methotrexate in human plasma were between 92.6 and 114.3. The novelty of the presented methodology is the MS³ technique resulting in enhanced selectivity and sensitivity. The application of this LC-MS³ method was successfully completed on 46 human plasma samples and the quantitative results of identical human plasma samples were compared with another LC-MRM based method. Passing-Bablok regression coefficients demonstrated that there is no significant difference between the LC-MS³ method and LC-MRM method. Bland-Altman plots showed a concordant results, supporting the developed LC-MS³ method is a reliable and accurate assay for determination of methotrexate in human plasma. This work is also a proof of concept for using LC-MS³ technique to determination of chemicals in biological samples.

Keywords: High throughput; LC-MS(3); Methotrexate; Therapeutic drug monitoring.

MeSH terms

Chromatography, Liquid
Drug Monitoring
Humans
Methotrexate*
Reproducibility of Results
Tandem Mass Spectrometry*

Substances

Methotrexate