Cyclosporine A/LC-MS/MS/Metabolites AM1/AM9/AM4N/ Renal transplant patients

A LC-MS/MS method was developed and validated for the determination of cyclosporine A (CsA) and its three phase l metabolites AM1, AM9, and AM4N in whole blood and lymphocates isolated on the Histopaque gradient. 200 micro of whole blood was preciitated with 10 mol/L zinc sulfute in acetonitrile/methanol (40:60, v/v) and lymphocatesisolated from 1,5 mL blood were wstracted with acetonitrile/methanol (40:60, v/v). The analytes and internal standard cyclosporine D were separated on RP column BEH C18, 2,1x50 mm, 1,7 micro using gradient LC-MS/MS analysis in positive electrospray mode. Time of analysis was 5 min. Linearity in blood was 5-2000 micro/L for CsA, AM1, and AM9; 2-500 micro/L for AM4N; and 2-500 micro/L for all substances in lymphocytes. Coefficient of variations was 1,8-9,8% and recovery was 92,0-110,0 %.