Electronic Effects of Ligand Substitution in a Family of Co^II ₂ PARACEST pH Probes

We report three new Co₂-based paramagnetic chemical exchange saturation transfer (PARACEST) probes with the ability to ratiometrically quantitate pH. A Co^II ₂ complex, [LCo₂(etidronate)]^-, featuring tetra(carboxamide) and OH-substituted etidronate ligands with opposing pH-dependent CEST peak intensities, was previously shown to exhibit a linear correlation between log(CEST_OH/CEST_NH) and pH in the pH range 6.5-7.6 that provided a sensitivity of 0.99(7) pH unit^-1 at 37 °C. Here, we demonstrate through a series of CF₃-functionalized Co^II ₂ complexes [(^XL′)Co₂(etidronate)]^- (X = NO₂, F, Me), that modest changes in the electronic structure of Co^II centers through remote ligand substitution can significantly affect the NMR and CEST properties of Co₂-based PARACEST probes. Variable-pH NMR and CEST analyses reveal that the chemical shifts of the ligand protons are highly affected by the nature of the X substituent. The ratios of OH and NH CEST peak intensities at 115 and 88, 93 and 79, and 88 and 76 ppm for X = NO₂, F, and Me, respectively, afford pH calibration curves with remarkably high sensitivities of 1.49(9), 1.48(7), and 2.04(5) pH unit^-1 across the series. The 1.5-2-fold enhancement in pH sensitivity for the CF₃-functionalized Co₂ probes stems from the complete separation of the OH and NH CEST peaks. Furthermore, incorporation of electron-withdrawing CF₃ groups shifts the detection window to a more acidic range of pH 6.2-7.4. Finally, the Co^II ₂ complexes are found to be extremely robust toward substitution and oxidation in aqueous solutions. Taken together, these results highlight the unique ability of transition metal-based PARACEST probes to provide a highly sensitive concentration-independent measure of pH and demonstrate that modest ligand modifications can be a powerful tool for optimizing the pH sensing performance of these probes.