Simultaneous quantification of cyclosporine, tacrolimus, sirolimus and everolimus in whole blood by liquid chromatography-electrospray mass spectrometry

Clin Biochem. 2008 Jun;41(9):728-35. doi: 10.1016/j.clinbiochem.2008.02.014. Epub 2008 Mar 12.


Objectives: The aim of this work was to develop a selective method for the simultaneous quantification of cyclosporine, tacrolimus, sirolimus and everolimus in whole blood.

Design and methods: An automated on-line solid-phase extraction system coupled with liquid chromatography-mass spectrometry (LC-MS) was used. After a simple protein precipitation, the supernatant was load on a C8 column with a mobile phase composed of MeOH/H(2)O (5/95 v/v), supplemented with formic acid 0.02% and sodium formate 1 muM. After column-switching, the analytes were transferred in the back-flush mode on a C18 column with MeOH/H(2)O (65/35). The valve was then commuted to its initial position and the chromatographic separation was performed with a gradient of MeOH/H(2)O (65/35-95/5). The sodium adducts [M+Na](+) were monitored for quantification with an electrospray ionization-single quadrupole MS.

Results: The LC-MS assay was fully validated on a concentration range of 2.5-30 ng/mL for tacrolimus, sirolimus and everolimus and of 50-1500 ng/mL for cyclosporine, allowing a quantification of cyclosporine 2 h post-dose without sample dilution. Trueness, repeatability and intermediate precision were found to be satisfactory.

Conclusion: This method provided a selective, rapid and automated procedure that can be easily used for routine quantification of immunosuppressive drugs in most clinical laboratories.

Publication types

  • Comparative Study

MeSH terms

  • Chromatography, Liquid / methods
  • Chromatography, Liquid / standards
  • Cyclosporine / blood*
  • Cyclosporine / chemistry
  • Everolimus
  • Humans
  • Sirolimus / analogs & derivatives*
  • Sirolimus / blood*
  • Sirolimus / chemistry
  • Spectrometry, Mass, Electrospray Ionization / methods
  • Spectrometry, Mass, Electrospray Ionization / standards
  • Tacrolimus / blood*
  • Tacrolimus / chemistry


  • Cyclosporine
  • Everolimus
  • Sirolimus
  • Tacrolimus