method validation of methotrexate in human plasma by lc-ms technique in patients with brain tumor.

background and aims: a selective and sensitive high performance liquid chromatography-electrospray ionization mass spectrometry method has been established for determination of methotrexate in human plasma. methods: methotrexate was extracted from plasma with acetonitrile. the mobile phase consisted of acetonitrile-water-formic acid 74: 25: 1(v/v). twenty µl of sample was chromatographically analyzed using a repacked zorbax-xdb-ods c18 column (2.1mm×30mm, 3.5 micron). the mode of mass spectrometry was selected-ion monitoring (sim). results: standard curve was linear (r= 0.998) over the concentration range of 0.1–100.0 ng/ml and showed a suitable accuracy and precision. the limit of detection (lod) was 0.05ng/ml. conclusion: the method is quick, easy, very steady and precise for the partition, assignment and evaluation of methotrexate in human plasma.