Development of an LC–MS/MS assay forquantification of intact INSL3 in rat plasma: supplementary data

Background: Relatively large disulfide-linked polypeptides can serve as signaling molecules for a diverse

array of biological processes and may be studied in animal models to investigate their function in vivo.

The aim of this work was to develop an LC–MS/MS assay tomeasure a model peptide, INSL3, in rat plasma.

Results: A dual enrichment strategy incorporating both protein precipitation and solid phase extraction

was utilized to isolate INSL3 from rat plasma, followed by targeted LC–MS/MS detection. The method was

able to measure full-length INSL3 (6.1 kDa) down to 0.2 ng/ml with acceptable accuracy and precision.

Conclusion: The final assay was applied to support an exploratory pharmacokinetic study to evaluate

steady-state concentrations of dosed INSL3 in rat plasma.