Abstract
An effective analytical method is requisite to ensure the accurate identification and quantification of drug(s), either in bulk material or in complex matrices, which form part of finished pharmaceutical products. For the purpose of a pharmaceutical formulation study, it became necessary to have a simple, yet robust and reproducible reversed-phase HPLC method for the simultaneous detection and quantification of lamivudine (3TC), tenofovir disoproxil fumarate (TDF), and dolutegravir sodium (DTG) in bulk form, complex polymeric matrices, and during drug release studies. A suitable method was developed using a Kinetex® C18, 250 × 4.6 mm column as stationary phase and a mobile phase consisting of 50 : 50 v/v methanol and water with 1 mL orthophosphoric acid, with a flow rate of 1.0 mL/min and column temperature maintained at 35°C. A detection wavelength of 260 nm and an injection volume of 10 μL were used. The method was validated according to the International Conference on Harmonization (ICH) guideline Q2 (R1), and the parameters of linearity and range, accuracy, precision, specificity, limit of detection (LOD), limit of quantification (LOQ), robustness, and stability were all determined. Acceptable correlation coefficients for linearity (R2) of >0.998 for each of the three drugs were obtained. The LOD was quantified to be 56.31 μg/mL, 40.27 μg/mL, and 7.00 μg/mL for 3TC, TDF, and DTG, respectively, and the LOQ was quantified as 187.69 μg/mL, 134.22 μg/mL, and 22.5 μg/mL for 3TC, TDF, and DTG, respectively. In relation to all the determined validation parameters, this method proves to be suitable for the accurate identification and quantification of the three ARVs, either alone or in combination, as well as when incorporated into polymeric matrices. Furthermore, the method proves to be suitable to detect degradation of the compounds.
Highlights
Lamivudine (3TC) is a nucleoside analog reverse transcriptase inhibitor (NRTI), used for the treatment of HIV-1, HIV-2, and hepatitis B infection (Figure 1(a)) [1, 2]
Linearity and Range. e linearity of the method was established from a regression plot of peak response area against the concentration level of each drug. e linearity was demonstrated across the range of 150.0–1200.0 μg/mL for 3TC and Tenofovir disoproxil fumarate (TDF) and 1.5–210 μg/ml for DTG, which was evident from the correlation coefficients (R2) of >0.998 (Table 1), proving that there exists a good correlation between method responses across the concentration range
Accuracy. e accuracy of the proposed method was conducted through recovery studies, which were performed by preparing samples at three concentration levels, 50%, 100%, and 150% as outlined in Table 1, falling within the concentration range of 300–900 μg/mL for 3TC and TDF and 53–158 μg/mL for DTG. ese solutions were analysed against a reference standard solution of known concentration, and the recovered concentration was quantified and reported
Summary
Lamivudine (3TC) is a nucleoside analog reverse transcriptase inhibitor (NRTI), used for the treatment of HIV-1, HIV-2, and hepatitis B infection (Figure 1(a)) [1, 2]. Tenofovir disoproxil fumarate (TDF) was the first nucleotide analog reverse transcriptase inhibitor (NtRTI) (Figure 1(b)), approved for the treatment of HIV infection [3]. Dolutegravir (DTG) is a unique second-generation integrase strand transfer inhibitor (INSTI) (Figure 1(c)), developed as a result of the limitations of the first-generation INSTIs, which includes potency, resistance by the virus, dosing frequency, dosing weight, and drug genetic barrier [5,6,7]. It is effective against numerous HIV-1 and HIV-2 clinical isolates [8]. The combination of 3TC, TDF, and DTG is formulated as a fixed-dose combination (FDC) oral tablet
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