A rapid and sensitive liquid chromatography-tandem mass spectrometric assay for duloxetine in human plasma: Its pharmacokinetic application

Abstract

This paper describes a simple, rapid and sensitive liquid chromatography-tandem mass spectrometry assay for the determination of duloxetine in human plasma. A duloxetine stable labeled isotope (duloxetine d₅) was used as an internal standard. Analyte and the internal standard were extracted from 100 μL of human plasma via solid phase extraction technique using Oasis HLB cartridges. The chromatographic separation was achieved on a C₁₈ column by using a mixture of acetonitrile-5 mM ammonium acetate buffer (83:17, v/v) as the mobile phase at a flow rate of 0.9 mL/min. The calibration curve obtained was linear (r²≥0.99) over the concentration range of 0.05-101 ng/mL. Multiple-reaction monitoring mode (MRM) was used for quantification of ion transitions at m/z 298.3/154.1 and 303.3/159.1 for the drug and the internal standard, respectively. Method validation was performed as per FDA guidelines and the results met the acceptance criteria. A run time of 2.5 min for each sample made it possible to analyze more than 300 plasma samples per day. The proposed method was found to be applicable to clinical studies.

Keywords: Duloxetine in human plasma; Liquid chromatography–tandem mass spectrometry; Method validation; Pharmacokinetic studies; Solid-phase extraction (SPE).

Fig. 1

Chemical structures of duloxetine hydrochloride and duloxetine d5 oxalate (IS).

Fig. 2

Typical MRM chromatograms of duloxetine (left panel) and IS (right panel) in human blank plasma (A), human plasma spiked with IS (B), and an LLOQ sample along with IS (C).

Fig. 3

MRM chromatograms resulting from the analysis of subject blank plasma sample (A) and 5.67 h subject plasma sample (B), after the administration of a 60 mg oral single dose of duloxetine. The sample concentration was determined to be 48.2 ng/mL.

Fig. 4

Mean plasma concentration–time profile of duloxetine in human plasma following oral administration of duloxetine hydrochloride (60 mg capsule) to healthy volunteers (n=6).