Research Article TheValidationofaSimple,Robust,Stability-IndicatingRP-HPLC MethodfortheSimultaneousDetectionofLamivudine,Tenofovir DisoproxilFumarate,andDolutegravirSodiuminBulkMaterial and Pharmaceutical Formulations Omobolanle Ayoyinka Omoteso , 1 MarnusMilne , 2 andMariqueAucamp 1 1 School of Pharmacy, University of the Western Cape, Bellville, Cape Town, 7530, South Africa 2 School of Pharmacy, Sefako Makgatho Health Sciences University, Ga-Rankuwa, Pretoria 0208, South Africa Correspondence should be addressed to Marique Aucamp; maucamp@uwc.ac.za Received 20 September 2021; Revised 30 December 2021; Accepted 8 January 2022; Published 4 February 2022 Academic Editor: Mohamed Abdel-Rehim Copyright © 2022 Omobolanle Ayoyinka Omoteso et al. is is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. 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 ® C 18 ,250 × 4.6mmcolumnasstationaryphaseandamobilephaseconsistingof50:50v/vmethanolandwaterwith1mL 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. e method was validated according to the International Conference on Harmonization (ICH) guideline Q2 (R 1 ), 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 (R 2 )of >0.998 for each of the three drugs were obtained. e 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. 1.Introduction 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 dis- oproxil fumarate (TDF) was the first nucleotide analog re- verse transcriptase inhibitor (NtRTI) (Figure 1(b)), approved for the treatment of HIV infection [3]. A combination of TDF with other NRTIs and different classes of antiretroviral drugs (ARVs) causes synergistic effects showing activity against all subtypes of HIV-1 and certain strains of HIV-2 [4]. 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 [5–7]. It is effective against numerous HIV-1 and HIV-2 clinical isolates [8]. e 2016 WHO Consolidated Guidelines recommended the combination of the mentioned three ARVs as the first-line regimen mainstay of HIV treatment. Currently, the combi- nation of 3TC, TDF, and DTG is formulated as a fixed-dose combination (FDC) oral tablet. Hindawi International Journal of Analytical Chemistry Volume 2022, Article ID 3510277, 11 pages https://doi.org/10.1155/2022/3510277