Development and validation of a rapid HPLC-MS/MS method for simultaneous determination of cyclosporine A and tacrolimus in whole blood for routine therapeutic drug monitoring in organ transplantation

Background: Therapeutic drug monitoring is an integral part of organ transplantation. A rapid, simple, economical, and robust high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method for simultaneously determining the immunosuppressants cyclosporine A and tacrolimus might increase detection efficiency.

Methods: In this study, we developed and validated a rapid HPLC-MS/MS method. Whole blood samples of 100 μL were prepared by protein precipitation with acetonitrile and 0.5 mol. L^-1 ZnSO₄. Chromatography was performed on a pre-column using a gradient elution with 20 mmol. L^-1 ammonium formate and 0.1% (v/v) formic acid in water (mobile phase A) and 0.1% (v/v) formic acid in methanol (mobile phase B) at a flow rate of 1.5 mL.min^-1. The analysis time was 2.2 min. Electrospray ionization and multiple reaction monitoring were performed. The lower limit of quantification was set at 1 ng. L^-1 for tacrolimus and 50 ng. L^-1 for cyclosporine A.

Results: The method showed adequate accuracy and precision with a sufficient linear range. The calibration curve range of tacrolimus and cyclosporine A was 1-30 and 50-1500 ng·mL-¹, respectively. All correlation coefficients were >0.99.

Conclusions: The developed HPLC-MS/MS is rapid and can be used for simultaneous monitoring of tacrolimus and cyclosporine A.