Conformational flexibility governs phosphate-induced solubility enhancement: a comparative study of cimetidine and famotidine by pH-dependent dissolution, NMR titration, DOSY, and DFT analysis.

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.