Determination of depsipeptide (FR901228) in human plasma by liquid chromatography-tandem mass spectrometry

A sensitive method for the determination of depsipeptide (FR901228) in plasma was developed using high performance liquid chromatographic (HPLC) separation with tandem mass spectrometric detection. FR901228 was acidified by potassium acid phthalate (0.05 M, pH 4.0) and extracted with ethyl acetate (5 : 12, v/v); the supernatant of ethyl acetate extract was evaporated, reconstituted in 250 muL mobile phase, and then separated on a Keystone spherisorb C8, 5mu 2.1 x 100 mm(2) column with a mobile phase consisting of methanol-12 mM ammonium acetate (85: 15, v/v) at a flow rate of 0.2 mL/min. Detection was achieved by a PE SCIEX AP1365 LC/MS/MS System at unit (Q1) and low Q3) resolution in positive multiple reaction monitoring (MRM) mode, monitoring the transition of the FR901228 molecular ion m/z 541.0 to the product ion m/z 272.0, and of the internal standard (IS) (t-BOC-D-glutamic acid 1-benzyl ester, BGBE) molecular ion m/z 338.0 to the product ion m/z 91.0. The mean recovery for FR901228 was 60% with a lower limit of quantification (LLOQ) of 0.2 ng/mL using 0.5 mL plasma for extraction. This method was validated over a linear range of 1.0-1000 ng/mL, using BGBE as the IS. Results from a 5-day validation study demonstrated good within-day and between-day precision (CV% values were less than or equal to3.5% and less than or equal to5.5%, respectively) and accuracy (range from 99.7% to 112.5%) across the calibration range of 1.0-1000 ng/mL.