Determination of cyclophosphamide enantiomers in plasma by LC‐MS/MS: Application to pharmacokinetics in breast cancer and lupus nephritis patients

Abstract

This article describes the enantioselective analysis of cyclophosphamide (CPA) in human plasma using LC-MS/MS. CPA enantiomers were extracted from plasma using a mixture of ethyl acetate and chloroform (75:25, v/v). The enantiomers were separated on a Chiralcel(R) OD-R column, with the mobile phase consisting of a mixture of acetonitrile and water (75:25, v/v) plus 0.2% formic acid. The protonated ions and their respective product ions were monitored using two functions, 261 > 141 for CPA enantiomers and 189 > 104 for the internal standard (antipyrine). Recovery rates were higher than 95% and the quantification limit was 2.5-ng/ml plasma for both enantiomers. The coefficients of variation and the relative errors obtained for the validation of intra- and interassay precision and accuracy were less than 10%. The method was applied for the investigation of the enantioselective pharmacokinetics of CPA in a lupus nephritis patient treated with 1 g CPA infused over 2 h and in a breast cancer patient treated with 0.9 g infused over 1 h. No stereoselectivity in the pharmacokinetic parameters was observed for either patient. Clearance values of 2.63 and 2.93 l/h and of 3.36 and 3.61 l/h for (-)-(S) and (+)-(R)-CPA were obtained for the breast cancer and lupus nephritis patient, respectively.

A review of bioanalytical methods for chronic lymphocytic leukemia drugs and metabolites in biological matrices

“…Ifosfmaide and CP were extracted using chloroform and separated by achiral‐chiral coupled column chromatography (Masurel & Wainer, ). An isocratic method consisting of acetonitrile and 0.2% formic acid in water at a flow rate of 0.5 mL/min by Chiralcel OD‐R column was used for enantiomeric separation of CP (De Miranda Silva et al, ). Recovery was 75% with liquid–liquid extraction using ethyl acetate and chloroform mixture.…”

“…Recovery was 75% with liquid–liquid extraction using ethyl acetate and chloroform mixture. The detection and quantification of CP enantiomers was done with multiple reaction monitoring mode (de Miranda Silva et al, ). A simultaneous chiral estimation of CP, carboxyethylphosphoramide mustard enantiomers (Attié de Castro, Scatena Gdos, Cass, Simões, & Lanchote, ), and 4‐OHCP (de Castro et al, ) was developed for the quantification in patient plasma samples.…”

Enantioselectivity in the Metabolism of Cyclophosphamide in Patients With Multiple or Systemic Sclerosis

“…Although the (R)‐(+) and (S)‐(–) enantiomers may differ in their therapeutic responses and toxic effects, the clinical consequences of these differences have not been adequately established. The results of clinical studies investigating enantioselectivity in the kinetic disposition of cyclophosphamide are inconclusive, do not evaluate the individual enantiomers of the active metabolite 4‐hydroxycyclophosphamide, do not evaluate the individual enantiomers of the metabolite carboxyethylphosphoramide mustard, which exposure is related to outcome disease, and involve a small number of patients in different disease states and treated with different combinations of drugs …”

“…The results of clinical studies investigating enantioselectivity in the kinetic disposition of cyclophosphamide are inconclusive, do not evaluate the individual enantiomers of the active metabolite 4-hydroxycyclophosphamide, do not evaluate the individual enantiomers of the metabolite carboxyethylphosphoramide mustard, which exposure is related to outcome disease, and involve a small number of patients in different disease states and treated with different combinations of drugs. 13,[18][19][20][21][22][23] The pharmacokinetics of cyclophosphamide and its metabolites, 4-hydroxycyclophosphamide and carboxyethylphosphoramide mustard, as an enantiomeric mixture is well defined for the cyclophosphamide intravascular administration in high dose (continuous or daily dosing) to patients with cancer. 11,13,[24][25][26][27][28] However, there are no data on the enantioselective kinetic disposition and metabolism of cyclophosphamide in patients with chronic inflammatory autoimmune diseases such as multiple sclerosis and systemic sclerosis.…”