HPLC method development for the quantitative analysis of Lamivudine, Tenofovir Disoproxil Fumarate and Nevirapine

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MAY, 2016
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An accurate and precise reverse phase HPLC method has been developed for the simultaneous quantification of Lamivudine, Nevirapine and Tenofovir disoproxil fumarate. Phenomenex Synergi C18 (250mm×4.6, 4μm) with methanol: ammonium acetate buffer (adjusted to pH 2.80): acetonitrile in a ratio 50:40:10 v/v was found to achieve suitable separation of the antiretroviral drugs. A flow rate of 1.0ml/min, temperature at 25°C and UV detection at 270nm were used. The run time was 10minutes with retention times of 3.26, 5.42 and 7.55 minutes for lamivudine, nevirapine and tenofovir disoproxil fumarate respectively. The HPLC method developed showed good linearity within the ranges of 10-59µg/ml, 7-42µg/ml and 15-90µg/ml with correlation coefficient of 0.9973, 0.9951 and 0.9968 for tenofovir disoproxil fumarate, nevirapine and lamivudine respectively. The limits of detection were 5.50µg/ml, 3.15µg/ml and 3.93µg/ml for lamivudine, nevirapine and tenofovir disoproxil fumarate respectively. The limits of quantification were 16.68µg/ml, 9.54µg/ml and 10.04µg/ml for lamivudine, nevirapine and tenofovir disoproxil fumarate respectively. The method is accurate in the range 90%- 110% recovery and precise (all %RSD values for inter-day and intraday studies within acceptable criteria). This method detected the antiretroviral drugs in the various formulations within the same retention times as the pure powder samples which informed the specificity of the method. Quantitative analysis of formulations containing the antiretroviral drugs under study were carried out. The percentage content of Tenofovir, lamivudine and Nevirapine in Tenofovir/ Lamivudine fixed-dose combination co-blistered with Nevirapine were 98.55%± 0.17, 105.33%± 0.85 and 99.20%± 1.17 respectively.
Dissertation submitted to The Department of Pharmaceutical Chemistry of The Faculty of Pharmacy and Pharmaceutical Sciences, KNUST in partial fulfilment of the requirements for the award of Master of Philosophy Degree in Pharmaceutical Chemistry.