Rapid analysis of anthracycline antibiotics doxorubicin and daunorubicin by microchip capillary electrophoresis

doi:10.1016/j.microc.2009.03.005

Microchemical Journal

Volume 92, Issue 2, July 2009, Pages 170-173

https://doi.org/10.1016/j.microc.2009.03.005 Get rights and content

Abstract

A simple and rapid method has been developed for the analysis of anthracycline antibiotics doxorubicin (DOX) and daunorubicin (DAU) in human serum using mirochip-based capillary electrophoresis (CE) with laser-induced fluorescence (LIF) detection. In this study, method development included studies of the effect of buffer pH, buffer concentration, organic solvents and separation voltage on sensitivity and separation efficiencies for the CE separation of DOX and DAU. Acetonitrile was found to have significantly improved the sensitivity and separation efficiency. The method was validated with regard to reproducibilities, linearity and limit of detection (LOD). The optimum electrophoretic separation conditions were 10 mM sodium tetraborate buffer at pH 9.5 with 40% acetonitrile (V/V) and a separation voltage of 2.1 kV. DOX and DAU were separated in 60 s under the optimum separation conditions. Linear relationships were obtained between the concentration and peak area (or peak height) in the 1–75 µg mL^− 1 range and with the detection limits of 0.3 and 0.2 μg mL^− 1 for DOX and DAU, respectively. The stability of both migration time and peak height of the analytes showed relative standard deviations of less than 5% (n = 9). The potential of this method was verified by spiking a human serum sample with the two drugs and analyzing the recovery ratios.

Introduction

Anthracycline antibiotics represented by doxorubicin (DOX) and daunorubicin (DAU) are popularly used as anticancer drugs. DAU displays good properties against acute lymphocytic and acute granulocytic leukemias, while DOX has desirable curative effects on these diseases as well as a variety of neoplasmas. However, the clinical efficacy of DOX and DAU is related to their concentrations in the blood. When it surpasses a certain level, serious side-effects, such as cardiac toxicity, marrow suppression and oral ulcer are found. Due to the significant clinical importance of the anthracycline antibiotics, numerous analytical techniques have been developed to quantify them. Up till now, high-performance liquid chromatography (HPLC) [1], [2], [3], [4] and capillary electrophoresis (CE) [5], [6], [7], [8] are most frequently used for the determination of the anthracycline antibiotics.

Chip-based microfluidic systems have attracted broad interests in recent years as a major form for realizing the lab-on-a-chip, or micro total analysis system (µTAS) concept to achieve the integration of sample introduction, pretreatment, reaction, separation and detection in a miniaturized analysis system [9]. Compared to traditional analytical methods, microchip-based analysis provides inherent merits, such as fast analysis, very small amount of required reagents and samples. It provides possibilities for miniaturization, automation, portability, and real-time analysis. These unique characteristics make microchip CE suitable for the development of portable point-of-care medical diagnostic systems, especially in combination with laser-induced fluorescence detection which provides high sensitivity.

In this work, we describe the application of microchip CE with LIF detection to the analysis of DOX and DAU. The suitability of the proposed method for the evaluation of these drugs in biological samples was demonstrated by the analysis of a spiked human serum. The fact that this analysis takes less than 2 min showed a significant advantage over the conventional CE.

Section snippets

Chemicals and reagents

The stock solutions of DOX and DAU (0.5 mg mL^− 1) were prepared by dissolving the appropriate amount of drugs (provided by the Institute for Food and Drug control, Hangzhou, Zhejiang province, China) in acetonitrile–water (1:9, v/v) and stored in the dark at 4 °C. Working standards were freshly prepared by diluting stock solutions to desired concentrations with running buffer. A 10 mM sodium tetraborate containing 40% acetonitrile (pH 9.5) was used as a running buffer solution. All used

Fluorescence spectra of DOX and DAU

The chemical structures of DOX and DAU are shown in Fig. 1. They are anthracycline compounds having tetracycline ring and aromatic plane rigid structures. So they have strong native fluorescence and can be detected by direct fluorescence detection.

The excitation and emission spectra of DOX and DAU are illustrated in Fig. 2. It was confirmed that fluorescence signals were significantly sensitized by acetonitrile while a small shift in the maximum excitation and emission wavelengths was observed

Conclusions

The presented analysis methodology provided a simple, rapid and efficient way for the identification as well as real-time monitoring of DOX and DAU in human serum. The sample was directly introduced to the microfluidic chip after a simple pretreatment of protein precipitation. Under the optimum conditions, it took less than 2 min to analyze DOX and DAU in the human serum sample. This developed method has great potential for clinical application as a portable point-of-care medical diagnostic

Acknowledgement

This work was funded by the National Natural Science Foundation of China (project no. 20775068 and 20890020).

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Microchemical Journal

Abstract

Introduction

Section snippets

Chemicals and reagents

Fluorescence spectra of DOX and DAU

Conclusions

Acknowledgement

References (14)

Reversed-phased liqiud chromatographic determination of doxorubicin after on-line trace enrichment on iron(III)-loaded 8-hydroxyquiniline-bonded silica

Anal. Chim. Acta

Determination of doxorubicin and metabolites in murine specimens by high-performance liqiud chromatography

J. Chromatogr., B

Determination of doxorubicin and doxorubicinol in plasma of cancer patients by high-performance liquid chromatography

Anal. Biochem.

Determination of doxorubicin and its metabolites in rat serum and bile by LC: application to preclinical pharmacokinetic studies

J. Pharm. Biomed. Anal.

Bioanalysis of some anthracyclines in human plasma by capllary electrophoresis with laser-induced fluorescence detection

J. Chromatogr.

Determination of daunorubicin in human urine by capillary zone electrophoresis with amperometric detection

Anal. Chim. Acta

Quantification of noradrenaline and dopamine in Portulaca oleracea L. by capillary electrophoresis with laser-induced fluorescence detection

Anal. Chim. Acta

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