Simultaneous Quantification of Lamivudine, Zidovudine and Related Impurities in Fixed Oral Dosage Combination Using RP-HPLC with DAD detection

Abstract

A simple and fast isocratic Reverse Phase High Performance Liquid Chromatography (RP-HPLC) method has been developed and validated for the simultaneous determination of Lamivudine, zidovudine and their related impurities in tablets. The method consists of a mobile phase combination of Acetonitrile (HPLC grade) and Buffer (0.0680 g of Potassium Dihydrogen Orthophosphate, 0.3 ml of Triethylamine, pH adjusted to 8.0 with Orthophosphoric acid to a final volume preparation of 100 ml) in the ratio 10:90. Phenomenex Luna 5-µm C18 (2)-250 x 4.6-mm, 5-µm) was used as the stationary phase. The column oven was set to a temperature of 30±1^oC. Quantification was achieved with a DAD detector set at 270 nm.

Resolution was achieved at a short run time of 25 minutes. Zidovudine related impurity C, Lamivudine, salicylic acid, Zidovudine and Zidovudine related impurity B eluted at 3.749±0.004, 4.862±0.013, 15.332±0.064, 21.201±0.076 and 23.682±0.117 respectively. Relative retention times (RRT) for lamivudine unknown related impurities with respect to Zidovudine were 0.15, 0.17, 0.30 and 0.59. RRT for Zidovudine unknown related impurities with respect to Zidovudine were 0.39 and 0.63.  The method was found to be specific, robust, accurate and precise for the estimation of Zidovudine related impurity C, Lamivudine, salicylic acid, Zidovudine and Zidovudine related impurity B in fixed oral dosage tablets over the concentration ranges of 0.0204 mg/mL-0.0088 mg/mL, 0.0962 mg/mL-0.7699 mg/mL, 0.1929 mg/mL-1.5410 mg/mL and 0.0088 mg/mL-0.024 mg/mL respectively. The Correlation Coefficient (r²) for Zidovudine related impurity C, Lamivudine, salicylic acid, Zidovudine and Zidovudine related impurity B were greater than 0.998. The LOD were found to be between 1.9x10^-4 mg/mL to 2.69 x10^-4 mg/mL.  The proposed method is precise, specific, accurate and robust for the simultaneous estimation of Zidovudine related impurity C, Lamivudine, salicylic acid, Zidovudine, Zidovudine related impurity B and other related impurities in dosage forms.

Keywords: Zidovudine related impurity C, Lamivudine, salicylic acid, Zidovudine, Zidovudine related impurity B, Lamivudine related impurities, Zidovudine related impurities RP-HPLC, Validation.