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A fast and reliable reversed phase high performance liquid chromatography method for simultaneous determination of selected anti-retroviral and lumefantrine in human plasma

Heudi, Olivier and Maganda, Betty and Cortinovis, Agnes and Picard, Franck and Kretz, Olivier (2013) A fast and reliable reversed phase high performance liquid chromatography method for simultaneous determination of selected anti-retroviral and lumefantrine in human plasma. Journal of Chromatography B, 919/92. pp. 52-60. ISSN 1570-0232

Abstract

A fast and reliable high performance liquid chromatography (HPLC) method with UV diode array detection for the simultaneous quantitative analysis of the anti-retroviral drugs, nevirapine (NVP) and efavirenz (EFV) and the anti-malarial, lumefantrine (LUM) in human plasma has been developed and validated. The sample preparation consisted of a plasma protein precipitation with 0.5% acetic acid acetonitrile solution containing the internal standard halofantrine (HALO) prior the LC-analysis. Chromatographic separation was carried out on a Acclaim Polar® Advantage C16, column (150 x 4.6 mm, particle size, 3 µm) using a gradient of mobile phase made of 0.01% TFA in 0.1 M ammonium acetate (solvent A) and 0.1% TFA in acetonitrile (solvent B). The separation of NVP, EFV, LUM and HALO was achieved within 17 min at a flow rate of 1.0 ml min−1 and detections were performed at three wavelengths, 275 nm (NVP), 255 nm (EFV), and 300 nm (LUM). The selectivity of the method was demonstrated in 6 different human plasma batches. In addition, six potential concomitant drugs (zidovudine, pyrimethamine, trimethoprim lamivudine, sulfamethoxazole and fluconazole) were analyzed under our experimental conditions and none of them co-elute with EFV, NVP and LUM. Calibration graphs plotted with seven concentrations in duplicate of each compound were linear between the selected ranges with a regression coefficient (R2) greater than 0.998. Absolute extraction recovery for NVP, EFV and LUM were 99%, 98.6 and 102%, respectively. Inter- and intraday coefficients of variation for LUM, EFV and NVP were ≤10%. The lower limit of quantification was 0.125 μg/mL for LUM and 0.250 μg/mL for both EFV and NVP. Intra- and inter-assay relative standard deviation values were found to be less than 15% at the concentrations examined (0.125–10.0 μg/mL for LUM and 0.250 – 15.0 μg/mL for both EFV and NVP). This low cost method can be used for the simultaneous quantitative analysis of NVP, EFV and LUM concentrations within a clinically relevant concentration range using standard chromatography equipments. This makes our method particularly applicable and useful to resource-limited settings.

Item Type: Article
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Date Deposited: 13 Oct 2015 13:14
Last Modified: 13 Oct 2015 13:14
URI: https://oak.novartis.com/id/eprint/8713

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