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In vivo solid-phase microextraction for monitoring intravenous concentrations of drugs and metabolites

Nature Protocols volume 6, pages 896–924 (2011)Cite this article

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Abstract

This protocol for in vivo solid-phase microextraction (SPME) can be used to monitor and quantify intravenous concentrations of drugs and metabolites without the need to withdraw a blood sample for analysis. The SPME probe is inserted directly into a peripheral vein of a living animal through a standard medical catheter, and extraction occurs typically over 2–5 min. After extraction, the analytes are removed from the sorbent and analyzed by, for example, liquid chromatography–tandem mass spectrometry. It has been validated in comparison with conventional blood analysis, and we describe here the in vitro experiments typically conducted during method development. The new-generation biocompatible SPME probes are designed specifically for extraction of semi-volatiles and nonvolatiles directly from aqueous samples and can be steam sterilized. Sorbents are coated on fine-gauge surgical steel wire (200-μm diameter), which is more rugged and biocompatible than conventional fibers (100-μm fused silica fiber). They incorporate a binding agent that resists fouling by the biological matrix and does not cause an immune response in the experimental animal. The sorbents used (coating thickness of ∼50 μm) are selected for their affinity for the types of small molecules of interest. The procedure is illustrated by the analysis of benzodiazepines with polypyrrole-coated wires inserted into peripheral blood vessels of beagles, although it can be adapted for use in smaller animals. The in vivo sampling can require as little as 1 min, in which case the entire procedure from sampling to instrumental analysis can take as little as 30 min.

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Figure 1: A schematic setup of a flow system to simulate animal circulatory blood system.
Figure 2: The in vivo assembly with the SPME coating retracted and exposed.
Figure 3: Schematic of injection tee for manual probe desorption.
Figure 4: Amounts recovered from successive re-extractions from the same probe and effects of reconditioning on inter-batch reproducibility.
Figure 5: Setup for coating PPY wires.
Figure 6: Comparison of PPY probe extraction calibrations (binding isotherms) in buffer versus plasma versus whole blood.
Figure 7: Representative chromatograms for four benzodiazepines after PPY extractions from a dog plasma sample.
Figure 8: Static equilibration time profiles.
Figure 9
Figure 10: Pharmacokinetic profiles from PPY devices.
Figure 11: Pharmacokinetic profiles from PEG-C18 devices.
Figure 12: In vivo SPME sampling in rats.
Figure 13: Pharmacokinetic profiles of carbamazepine (CBZ) in mice.

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Authors and Affiliations

  1. Department of Chemistry, University of Waterloo, Waterloo, Ontario, Canada

    Heather L Lord, Dajana Vuckovic & Janusz Pawliszyn

  2. Department of Biology, University of Waterloo, Waterloo, Ontario, Canada

    Xu Zhang

  3. Albany College of Pharmacy and Health Sciences, Albany, New York, USA

    F Marcel Musteata

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  1. Heather L Lord
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  3. F Marcel Musteata
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H.L.L. wrote and edited the manuscript. X. Z., F. M.M., D.V. and J.P. participated in editing the manuscript.

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Correspondence to Janusz Pawliszyn.

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Lord, H., Zhang, X., Musteata, F. et al. In vivo solid-phase microextraction for monitoring intravenous concentrations of drugs and metabolites. Nat Protoc 6, 896–924 (2011). https://doi.org/10.1038/nprot.2011.329

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