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Sirolimus (rapamycin) has a macrolide structure and is under clinical investigation as an immunosuppressant after organ transplantation. An HPLC/mass spectrometry assay to quantify sirolimus in blood was developed. 28-O-Acetyl sirolimus was used as internal standard. Blood samples were extracted with C18 columns. The extracts were injected into an HPLC system and isocratically eluted with methanol/1% formic acid (90/10 by vol) from a 150 X 4 mm C18 analytical column. The HPLC system was connected to a triple-stage quadrupole mass spectrometer with an electrospray interface and positive ions were detected. The limit of quantification in 1 mL of blood was 0.25 microgram/L and the calibration curve in blood was linear up to 250 microgram/L. The recovery from blood was 88 +/- 26% and interassay variation at 1 microgram/L was 19% and at 15 microgram/L 9.3%. Hydroxy, dihydroxy, demethyl, and didemethyl sirolimus as well as sirolimus were detected in blood of kidney graft patients.

clinical chemistry

Sensitive and specific quantification of sirolimus (rapamycin) and its metabolites in blood of kidney graft recipients by HPLC/electrospray-mass spectrometry