Abstract
A rapid, sensitive, selective and validated reverse phase high-performance liquid chromatography (RP-HPLC) method for the estimation of paclitaxel in micro-sample of rat plasma and in culture of cancer cells was performed in this study. The mobile phase consisted of an optimized mixture of methanol:water: trifluroacetic acid (80: 20: 0.1, v/v/v). Column elution at a flow rate of 1 mL/minute with UV detection at 225 nm at room temperature was used. The RP-HPLC method was successfully applied for the determination of paclitaxel in plasma samples and in culture of cancer cells with nano-quantity of estimation. The validation studies were performed in accordance with the International Conference on Harmonization (ICH) guidelines. The intra- and inter-day precision showed that the coefficients of variation ranged from 1.07% to 4.27% at different levels of concentrations. To the best of our knowledge, this study also reported for the first time the optimization of different solvents for effective extraction of paclitaxel wherein tert.-butyl methyl ether (TBME): diethyl ether (DEE) in 50: 50 v/v composition was found most efficient with extraction efficiency ranging between 77.99% and 91.74% and between 76.14 and 93.66% in the plasma and cell culture, respectively. This proposed method was successfully applied to study the pharmacokinetics of paclitaxel and the influence of verapamil and all-trans retinoic acid (atRA) on paclitaxel pharmacokinetics in rat models. This proposed method might emerge as a valuable aid in the laboratory monitoring of paclitaxel in a variety of in vitro as well as in vivo scenarios.
Highlights
This report describes a simple, robust and rapid reverse phase high-performance liquid chromatography (RP-high-performance liquid chromatography (HPLC)) method for routine quantitation of paclitaxel in standard quality control (QC) samples, rat plasma sample as well as in cancer cell lines. This method has been validated for its accuracy, precision, limit of detection (LOD) and limit of quantitation (LOQ) as per International Conference on Harmonization (ICH) guideline
Assay specificity was established in standard QC solutions as well as in plasma and cancer cell culture samples
Plasma as well as cell culture spiking showed no interference with the elution of paclitaxel as evinced from practically the same retention times of 4.359 and 4.35 minutes in case of plasma and cell culture spiked samples, respectively (Fig. 2d-e)
Summary
Several approaches have been used to develop assays for the determination of paclitaxel in biological fluids These include capillary electrophoresis[6], chromatography-mass spectrometry (LCMS)[7], immunoassay[8] and high-performance liquid chromatography (HPLC)[9,10,11]. Among these methods, HPLC methods have been most frequently used in the pharmacokinetic and toxicokinetic studies of paclitaxel due to their simplicity and sensitivity. This report describes a simple, robust and rapid RP-HPLC method for routine quantitation of paclitaxel in standard quality control (QC) samples, rat plasma sample as well as in cancer cell lines This method has been validated for its accuracy, precision, limit of detection (LOD) and limit of quantitation (LOQ) as per ICH guideline. The validated method has been administered to investigate the systemic exposure of paclitaxel following i.v. administrations in Sprague-Dawley rats
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