Liquid chromatographic determination of triple therapy components in anti-HIV products: a review

Abstract

The reliable analytical determination of triple antiretroviral therapy (ART) fixed-dose combinations is critical to ensuring product quality, therapeutic efficacy, and regulatory compliance in global HIV-1 management. Compared with dual therapies, triple combinations pose substantially greater analytical challenges due to the simultaneous presence of chemically diverse agents spanning wide polarity ranges, overlapping pKa values, and disparate dose strengths. This review critically evaluates reported HPLC, UPLC, and HPTLC methods for WHO-defined triple ART regimens comprising NRTIs, NNRTIs, and INSTIs, with particular emphasis on how physicochemical properties (pKa, polarity, logP) govern chromatographic behavior and method design. Rather than providing descriptive method compilations alone, the review advances a physicochemistry-driven decision framework, linking analyte ionization state, mobile-phase pH, buffer selection, organic modifier strength, and column efficiency to achievable sensitivity and robustness. These relationships are synthesized into decision tables and simplified decision trees to guide analysts in rational method development and selection, consistent with ICH Q14 (Analytical Procedure Development) and ICH Q2(R2) principles. While UV-based HPLC/UPLC remains the dominant and most widely accepted approach for routine quality control and WHO prequalification, limitations associated with acidic, non-volatile buffers and restricted LC–MS/MS adoption are highlighted, particularly for low-dose, high-potency INSTI-containing regimens. HPTLC is critically appraised as a complementary technique, retaining relevance for resource-limited settings, high-throughput screening, and surveillance applications despite inherent sensitivity and reproducibility constraints. Overall, this review provides a regulatory-aligned, practically actionable roadmap for chromatographic analysis of triple ART combinations, supporting method harmonization, lifecycle adaptability, and equitable global access to reliable antiretroviral quality control.