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Sensitive and specific chromatographic methods for the pharmacokinetic evaluation of carboplatin in young patients Burns, Robbin Bruce

Abstract

Carboplatin, c/s-diamminedichloro-1,1-cyciobutanedicarboxylatoplatinum II, is a second generation platinum antineoplastic agent that cross-links DNA. Dosing of carboplatin in adults is frequently based on formulas that relate renal elimination to AUC. Pharmacokinetic studies evaluating these dosing schemes are complicated by the assay methods utilized. In particular, substitution of platinum for carboplatin measurements in pharmacokinetic studies has occurred due to the poor sensitivity of existing HPLC assays for the parent drug. The objectives of this study were (1) to develop and validate a sensitive and specific HPLC assay method for determination of carboplatin in plasma ultrafiltrate, and (2) to compare carboplatin AUC estimates determined using free (non protein-bound) carboplatin versus estimates using free platinum. Assay methods for quantitation of carboplatin were developed utilizing HPLC with direct UV detection and with UV detection following post-column (PC) derivatization. Both methods yielded quantitation limits that were approximately 10-fold more sensitive than previous HPLC-UV methods. For the HPLC-PC method, sodium bisulfite was used to derivatize carboplatin to products possessing enhanced UV absorbance at 290 nm. The greater selectivity of this analytical wavelength removed the need for sample extraction, resulting in a simpler and more rapid assay procedure. MethCBDCA (bis(methylamine)cyclobutanedicarboxylatoplatinum II) and MethMal (bis(methylamine)malonatoplatinum II) were synthesized as internal standards for the HPLC-UV and HPLC-PC methods, respectively, the chemical structures being confirmed by HPLC-MS under positive electrospray ionization conditions. Both HPLC-UV and HPLC-PC methods were validated for carboplatin concentrations from 0.05 to 40 |ag/mL. Validated assay parameters included limits of detection and quantitation, specificity, precision, accuracy, linearity, and ruggedness. Recovery and stability were examined using the HPLC-UV method only. Assay variability was much higher for the HPLC-PC method due to time-dependent changes in signal response caused by the degradation of the post-column reagent, sodium bisulfite. The more precise HPLC-UV method was used in the pharmacokinetic study. The pharmacokinetic behaviour of carboplatin during eight cycles of chemotherapy in two young patients was investigated. Carboplatin concentrations in plasma ultrafiltrate were determined by the HPLC-UV method, while plasma ultrafiltrate platinum concentrations were determined by atomic absorbance. Based on visual examination of the concentration-time profiles, free carboplatin and free platinum elimination were clearly different. To better characterize the magnitude of the differences, AUC estimates derived from free platinum data were compared to those derived from free carboplatin data. The free platinum AUC estimates were as much as two-fold greater than their free carboplatin counterparts. However, the observed differences were much smaller in one patient than in the other. Further study is needed to more accurately characterize the extent of these differences across a larger patient population. Currently, we recommend that HPLC and AA assay methods not be used interchangeably for determination of carboplatin pharmacokinetic parameters.

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