Determination of melphalan in human plasma by UPLC–UV method
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It is desirable to develop a fast method for quantification of melphalan due to its instability. Here we report a method for quantification of melphalan (MPL) in human plasma using a UPLC–PDA system. Briefly, 50 µL plasma sample was mixed with 25 µL internal standard (2500 ng/mL acetylmelphalan in methanol) and 25 µL 20% trichloroacetic acid, and centrifuged at 21,000 g (15,000 rpm) at 4 °C for 3 min. The supernatant (5 µL) was injected onto an Acquity™ BEH C18 LC column (2.1 × 50 mm, 1.7 µm) and eluted with 25 mM NH4AC (pH 4.7)—acetonitrile in a gradient mode at a flow rate of 0.6 mL/min. The column kept at 40 ± 5 °C and the autosampler kept at 4 ± 5 °C. The detector set at 261 nm, and sampling rate was 40points/sec. The retention times were typically 2.11 min for melphalan and 2.38 min for the internal standard. Total run time is 4 min per sample. Calibration range was 100–40,000 ng/mL. The lower limit of quantification was 100 ng/mL. The method was validated based on the FDA guidelines, and applied to a clinical pharmacokinetic study in pediatric patients.
KeywordsMelphalan UPLC–UV Plasma Pediatric Pharmacokinetic.
This work was supported by the National Center for Research Resources and the National Center for Advancing Translational Sciences, National Institutes of Health, through UCSF-CTSI Grant Number UL1 RR024131 (a shared instrument award to Dr. Huang) and UCSF-CTSI Grant number KL2 TR000143 (Dr. Long-Boyle). Its contents are solely the responsibility of the authors and do not necessarily represent the official views of the NIH.
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Conflict of interest
The authors declare that they have no conflict of interest.
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