Determination of Pharmacokinetics Differences of Ammuxetine Isomers in Rat Plasma Using On-Line Solid Phase Extraction Coupled with Liquid Chromatography-Tandem Mass Spectrometry

An on-line solid phase extraction (SPE) coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed to determine the novel chiral antidepressant, S/R-ammuxetine in rat plasma. The plasma sample pretreatment consisted of the following steps: protein precipitation using methanol and acetonitrile (50:50, V/V), an on-line SPE process to remove proteins and most matrices in plasma, and a separation step using a C sub(18) analytical column after elution of ammuxetine isomers enriched on SPE column. Then S/R-ammuxetine were determined by tandem mass spectrometry. The SPE column was a Retain PEP Javelin column (10 mm 2.1 mm, 5 mu m), while the chromatographic separation was achieved on a ZORBAX SB-C sub(18) (50 mm 2.1 mm, 3.5 mu m) analytical column. The multiple reaction monitoring mode of the positive ion was adopted in MS detection, and the precursors to the product ion transitions of m/z 292.1/154.0 and m/z 260.4/116.2 were used to measure S/R-ammuxetine and internal standard (propranolol). The method was linear over S/R-ammuxetine concentration range from 0.2 mu g L super(-1) to 1000 mu g L super(-1) with the correlation coefficients (R) of 0.9903 and 0.9951, respectively. The average intra-day precision values (RSD) were 1.2%-12% for S-ammuxetine and 0.4%-11.2% for R-ammuxetine, respectively. The average recovery values were 94.2%-101.6% for S-ammuxetine and 94.3%-109.4% for R-ammuxetine. This method exhibited a dramatically increased sensitivity compared to previous reports, thus could be used in the pharmacokinetic study of ammuxetine isomers in rat after intragastric administration.