Conformational flexibility governs phosphate-induced solubility enhancement: a comparative study of cimetidine and...

Inorganic phosphate buffers are widely used in pharmaceutical dissolution testing, yet the direction of their effect on drug solubility is not uniform across solutes: for some basic drugs, phosphate decreases apparent solubility through insoluble salt formation or ionic-strength effects, while for others, our prior work has documented concentration-dependent solubility changes inconsistent with either mechanism. We report here that phosphate selectively increases the apparent cationic solubility of cimetidine (CIM) up to 2.6-fold but has a negligible effect on famotidine (FAM), two structurally distinct H₂-receptor antagonists, across a range of pH values and buffer compositions. This solubility-enhancing effect is mechanistically distinct from the well-documented solubility-decreasing effects of phosphate reported for other basic drugs via insoluble salt formation or ionic-strength suppression; those mechanisms predict a concentration-dependent decrease in solubility, which is thermodynamically incompatible with the concentration-dependent increase observed here for CIM. The differential response between CIM and FAM cannot be explained by classical Henderson-Hasselbalch ionization models or by differences in pH or buffering capacity. Three independent experimental approaches converge on a consistent interpretation. pH-dependent solubility measurements showed that phosphate selectively increases the cationic solubility (Si) of CIM while leaving the neutral-form solubility (S₀) unchanged; FAM showed no significant Si response. ¹H NMR titration revealed progressive downfield shifts of CIM imidazole resonances (∼1.0 ppm over 10-100 mM phosphate) with vicinal coupling constants consistent with a shift toward the cis/gauche (Zusammen) conformation; FAM and histamine responded minimally (∼0.5-0.6 ppm). DOSY diffusion measurements showed that phosphate decreases CIM diffusion disproportionately relative to bulk viscosity, indicating stronger solute-solvent coupling for the conformationally flexible molecule. DFT-based Boltzmann analysis reveals that CIM possesses a broad low-energy conformer ensemble (partition function Z = 2.62), whereas FAM is dominated by a single rigid geometry (Z = 1.10). The correlation between conformational entropy and phosphate-induced solubility enhancement suggests that molecular flexibility is a key determinant of ion-specific buffer effects, with implications for BCS solubility classification using phosphate-buffered media.