Exploring Microenvironment Acidity Inside the Solvent-Filled Pores of Mesoporous Silica Thin Films via Single-Molecule Spectroscopy

Single-molecule (SM) spectroscopy was used to study the local acidity inside solution-filled silica mesopores. The dual emission, pH-sensitive dye C-SNARF-1 was used as the fluorescent probe. Mesoporous materials were prepared as thin films supported on glass substrates. A microfluidic device was used to seal the upper surface of the films and to allow for their exposure to flowing aqueous solutions of the dye under different pH and ionic strength conditions. Single-molecule data were collected by two-color wide-field fluorescence imaging in the pseudototal internal reflection fluorescence mode. Pairs of fluorescence images were acquired simultaneously in bands centered around 580 and 640 nm, each having a 40 nm bandwidth. The ratio of C-SNARF-1 emission in these two bands (I₅₈₀/I₆₄₀) is highly sensitive to pH in bulk solution. The SM data show that C-SNARF-1 remains sensitive to pH in mesoporous materials, but that the emission ratios obtained are often larger than in bulk solution. Results obtained as a function of the ionic strength and interpreted in conjunction with simulations are consistent with perturbation of the dye response by Coulombic interactions with charged sites (i.e., deprotonated silanol groups) on the pore surfaces. When the mesoporous films are exposed to low-pH solutions or high ionic strength buffers, the SM emission ratios are similar to those obtained in bulk solution, whereas those recorded at intermediate pH are highly variable, due to the aforementioned Coulombic interactions. When considered together with the results of simulations, the SM results reveal that the pH inside the mesopores is largely independent of the solution pH under low ionic strength conditions and is instead determined by the ionization of the surface silanols. The pH within the mesopores responds most strongly to changes in the solution pH at high ionic strength and at low or high pH. The silanol group pK_a is estimated to fall between 7.1 and 8.7 from the SM results. Overall, these studies provide new information on the factors affecting the pH within the confined environments of solution-filled silica mesopores. They also provide a better understanding of the efficacy of pH measurements in porous materials made using organic probe dyes.