Direct pharmacokinetic analysis of puqietinone by in vivo microdialysis sampling and turbulent-flow chromatography coupled with liquid chromatography–mass spectrometry

Sample pretreatment is a key step in bioanalytical process because of possible interference and matrix effects in mass spectrometry analysis. In this work, a novel strategy towards high speed and sensitivity was developed combining in vivo microdialysis (MD) sampling, turbulent-flow chromatography (TFC), and liquid chromatography–mass spectrometry (LC–MS). The procedures of cleanup, preconcentration, and separation were completed on-line in one step within 10 min. During the MD optimization procedure, 1% hydroxypropyl-β-cyclodextrin (HP-β-CD) was used to improve the relative recovery of the analyte. Untreated MD samples were directly injected, and a TFC precolumn was flushed for 1 min with aqueous phase of 4 mL/min flow rate to desalt and concentrate biosamples. The retained analytes were then back-flushed by a switching valve onto a fast LC column (4.6 mm × 50 mm, 1.8 μm) for separation. Another diverter valve was employed to prevent the HP-β-CD that interferes with the ESI process from entering the MS. Puqietinone, a lipophilic alkaloid from Fritillaria puqiensis, was used as a case for validation. Results showed that the limit of quantification for puqietinone was 0.10 ng/mL, and good linearity (R2 = 0.9993) was maintained over the range of 1.02–200.02 ng/mL. Accuracy and precision were satisfactory within the range of the standard curve. This approach was able to effectively eliminate the influences of matrix effect and carry-over as the injection volume increased up to 20 μL. The developed method was successfully applied to pharmacokinetic study of puqietinone after intravenous administration to rat. Results demonstrate the potential of using MD with TFC-LC/MS for in vivo monitoring experiments.