A Fluorescent Probe‐Based Platform for Precise Pharmaceutical Analysis and Quality Assessment of Tirzepatide

Abstract

Tirzepatide, a first-class dual glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-1 (GLP-1) receptor agonist, has emerged as a breakthrough in the management of type 2 diabetes and obesity. Given its recent approval and expanding clinical applications, the development of accurate, sensitive, and efficient analytical methods for its determination in pharmaceutical formulations and biological matrices is critically important. In this study, a novel and efficient analytical probe was developed for the selective and sensitive determination of tirzepatide in pharmaceutical dosage form and spiked plasma. Acriflavine was employed as a fluorescent probe due to its strong native fluorescence, excellent stability, and high sensitivity to molecular interactions. The method was based on the formation of a stable 1:1 acriflavine-tirzepatide complex at pH 8.0, resulting in static fluorescence quenching measured at 505 nm following excitation at 451 nm. Validation in accordance with ICH guidelines confirmed the method's excellent linearity over 0.1-20 μg/mL (r = 0.9999), with a detection limit of 0.012 μg/mL and a quantitation limit of 0.036 μg/mL, alongside outstanding precision (RSD < 1%) and accuracy. Furthermore, greenness assessment using multiple evaluation tools confirmed the environmentally friendly nature of the method, highlighting its suitability for routine quality control, pharmacokinetic studies, and potential application to future GIP/GLP-1-based therapies.