Development of a chiral HPLC method to evaluate in vivo enantiomeric inversion of an unstable, polar radiosensitizer in plasma

Abstract

A chiral HPLC method to quantify in vivo enantiomeric inversion of prodrug CI-1010 ( I R ) or its drug II R (PD 146923), a radiosensitizer, upon X-irradiation of dosed rats was developed. These polar enantiomers were separated only by using normal-phase chiral HPLC. A Chiralpak AS column provided the best separation. Isolation of analytes from plasma employed solid-phase extraction (SPE), and required conditions that were compatible with normal-phase HPLC. Options for SPE were restricted by the chemically reactive nature of both prodrug and drug, which produced analyte losses as high as 100%. Acceptable recoveries using SPE required evaluation of conditions for analyte chemical stability. The validated method gave a lower-limit of quantitation (LLOQ) of 200 ng/ml for each enantiomer extracted from 0.15 ml of plasma. The LLOQ of the inverted enantiomer could be detected in the presence of 10 000 ng/ml of the dosed enantiomer. Precision (RSD) ranged from 14.2 to 4.4%, and from 24.2 to 5.1% for II S and II R , respectively. Accuracy (RE) was ±13.1 and ±13.2%, respectively. Recoveries ranged from 44.3 to 71.4%, and from 40.7 to 67.9%, for II S and II R , respectively.