Simultaneous Determination of Glibenclamide, Metoprolol Tartrate, and Phenol Red Using HPLC‐PDA: Application in Rat Intestinal Permeability Studies and Its Greenness Assessment Using NEMI, Analytical Eco‐Scale, GAPI, MoGAPI, AGREE, AGREEprep, RAPI, B

ABSTRACT This study presents the development and validation of an environmentally friendly HPLC‐PDA method for the simultaneous determination of glibenclamide (GLB), metoprolol tartrate (MET), and phenol red (PR). The method was optimized to ensure high trueness, precision, and robustness, adhering to ICH guidelines. The pKa of GLB was determined using reverse‐phase HPLC under varying pH conditions, providing valuable insights into its physicochemical properties and enhancing its pharmaceutical formulation potential. Intestinal permeability studies of GLB, conducted via the in situ Single Pass Intestinal Perfusion (SPIP) technique, validated the method's applicability in biological systems. In addition to analytical performance, the environmental sustainability of the method was assessed using green metrics, including the Analytical Eco‐Scale, GAPI, MoGAPI, AGREE, AGREEprep RAPI, BAGI, and CACI. The method achieved high scores, reflecting its alignment with the principles of green analytical chemistry and its minimal ecological impact. This dual focus on analytical rigor and environmental responsibility emphasizes the method's potential for broader applications in pharmaceutical and biomedical research. The findings of this study contribute to the ongoing efforts in developing sustainable analytical methodologies. Future research could explore its applicability to other pharmaceutical compounds and expand its use in complex matrices, further promoting environmentally conscious practices in analytical chemistry.