Investigation of microenvironmental heterogeneity in nanoporous silica by quantitative confocal imaging and computer simulation
Creators: name: Lee, Hong Bok
Publisher: name: University of Iowa
Year: 2025
Subjects: Fluorescence Imaging
Fluorescence spectroscopy
Nanoporous silica
Single-molecule spectroscopy
public abstract
Confocal Microscopy
Liquid Chromatography

description: Nanoporous silica particles have been widely used in chemical and biomedical engineering and sciences such as catalysis, chemical separations, and biosensors. Although many experimental and computational studies have unraveled transport kinetics and diffusivity through the pores, there is still a room for improvement of understanding microscale interfacial area between mobile phase and stationary phase in liquid chromatography. Heterogeneous distribution of adsorption and desorption sites of silanol group on the pore surfaces has accounted for low resolving power in the chemical separation: peak tailing and band broadening.
description: Our methods based on fluorescence and confocal microscopy visualized the distribution of effective diffusion and kinetics at adsorption sites through the intraparticle condition in the liquid chromatography. The hypothesis of the study is that the molecular dynamics inside chromatographic silica particle pores are affected by both Brownian motion and adsorption kinetics because the diffusing molecules inside the pores will have chances to adsorb and desorb at hydrophobic layers or silanol groups on the pore surfaces. Those populations are expected to be heterogenous through the silica particle because the porous structure has been observed as highly heterogeneous by many other electron microscopy imaging studies. To test this hypothesis, the translational diffusion and desorption rates of fluorescent probe in the silica particle were imaged using fluorescence correlation spectroscopy (FCS) with a confocal imaging strategy. The FCS with a confocal microscopy can reveal the hidden heterogeneity in molecular populations through the accumulation of single molecule signals at high spatial-temporal resolution. In this contribution, quantitative imaging of diffusion and adsorption kinetics in the nanoporous silica particle were constructed using our home-built Imaging FCS and computer simulation works. The result of quantitative images showed us the heterogeneity of those kinetics over the silica particle which implies the origin of the dispersive chromatographic peak resolution. The mathematical model for fitting and interpreting the autocorrelated fluctuation data was tested by generating Monte Carlo simulation combining Brownian motion and random stopping-departing motion.
description: Ratiometric intensity imaging can provide chemical information of equilibrium constant which describes the attraction force of a wet nanoporous silica particle. Fast scanning can be achieved by laser scanning confocal microscopy (LSCM). Based on LSCM, we constructed quantitative Gibbs free-energy (∆G) images of the C18 silica particles in a packed column. The heterogeneity of interparticle and intraparticle particles were visualized by analyzing the ∆G images, and the result provided a potential origin of the problematic band broadening issue.
description: Eddy diffusion and longitudinal diffusion are crucial parameters that dominantly affect the chromatographic column efficiency. Because the eddy diffusion is caused by irregular interstitial space through a packed column, 3-D visualization of the packed bed structure is an essential source for constructing interstitial diffusion simulations or any theoretical models to unravel the eddy diffusion which is one of the most unpredictable parameters in van Deemter equation. Expansion of 3-D visualization was demonstrated by combining a lateral sequent imaging strategy and a useful function of image analysis software.
description: Overall, the imaging strategies of transport kinetics and diffusivity, thermodynamic free energy, structural images through the heterogeneous chromatographic environment were independently developed, and these studies will be essential towards improving the understanding of the origin of resolution limit in chemical separation science. Ultimately, these scientific approaches, knowledge and insight will be required to design a better chemical plant for more efficient separations.