Improved HPLC method for doxorubicin quantification in rat plasma to study the pharmacokinetics of micelle‐encapsulated and liposome‐encapsulated doxorubicin formulations

Abstract

An improved simple, rapid and accurate HPLC method for quantification of doxorubicin derived from micelle-encapsulated or liposome-encapsulated doxorubicin formulation in rat plasma was described. The mobile phase consisting of a mixture of methanol-water [containing 0.1% formic acid anhydrous and 0.1% ammonia solution (25%), pH 3.0], 60:40, was delivered at a flow rate of 1.0 mL/min. Sample preparation for micelle- or liposome-encapsulated doxorubicin in rat plasma were achieved directly by protein precipitation with acetonitrile. Doxorubicin and daunorubicin (internal standard, IS) were separated on a C(18) reversed-phase HPLC column and quantified by a fluoresence detection with an excitation wavelength of 475 nm and an emission wavelength of 580 nm. The linearity was obtained over the range of 5.0-1000.0 ng/mL and 1.0-200.0 microg/mL for doxorubicin and the lower limit of quantitation was 5.0 ng/mL. For each level of quality control samples, inter- and intra-assay precision was less than 9.6 and 5.1% (relative standard deviation), respectively, and percentage error was within +/-2.6%. The extraction recoveries of doxorubicin in the range of 10 ng/mL to 100 microg/mL in rat plasma were between 94.1 and 105.6%. This method was successfully applied to the pharmacokinetic study of doxorubicin formulations after i.v. administration to rats.

HPLC quantification of doxorubicin in plasma and tissues of rats treated with doxorubicin loaded poly(alkylcyanoacrylate) nanoparticles

“…Blood was collected on heparinized tubes and centrifuged at 2000 × g for 10 min. To preserve Dox from degradation [24], all samples were stored at −20 • C until analysis.…”

“…Several RP-HPLC methods of quantification of Dox in rodent plasma were described during the last decade [10][11][12][13][14][15][16][17][18][19][20][21][22][23][24]. Sample preparation in general includes either a single step of protein precipitation [10][11][12] or solid phase extraction [13,14] or liquid-liquid extraction (LLE) [15] before injection into the chromatograph.…”

“…Sample preparation in general includes either a single step of protein precipitation [10][11][12] or solid phase extraction [13,14] or liquid-liquid extraction (LLE) [15] before injection into the chromatograph. The separation of Dox from its metabolites and daunorubicin, used as internal standard was achieved on various stationary phases [10,[12][13][14][15][16][18][19][20][21][22][23][24]. Detection methods include UV [14,16], tandem mass spectrometry [17][18][19][20], electrochemistry [13], chemiluminescence [21] and fluorescence [11,12,[22][23][24].…”

Quantitative liquid chromatographic analysis of anthracyclines in biological fluids

“…In an attempt to overcome their toxicity or drug resistance, prodrugs and special pharmaceutical formulations have been developed. Since these changes often require a different analytical approach, the interested reader is referred to the individual methods regarding the analysis of peptide-conjugated [29][30][31] or polymer-bound [32] prodrugs and micellar [33], pegylated liposomal [33][34][35], liposomal [36,37] or embolizing [38][39][40] formulations. Here we present an overview of 35 original methods published since 1990 for the determination of doxorubicin, epirubicin, daunorubicin, idarubicin and metabolites in biological fluids.…”

A Novel Strategy for the Delivery of Doxorubicin to Reduce Cardiotoxicity

“…The extracts were separated by centrifugation at 1200rpm for 10minutes. One ml of this was filtered through 0.45-μm-membrane filter and the amount of drug present in each organ was determined by HPLC method as reported previously 19 with UV detector at 475 nm. HPLC consist of C-18 column and mobile phase as a mixture of methanol-water [containing 0.1% formic acid anhydrous and 0.1% ammonia solution (25%), p H 3.0], 60:40, with a flow rate of 1.0 mL/min.…”