Back to the Drawing Board: A Unifying First-Principle Model for Correlating Sample UV-Vis Absorption and Fluorescence Emission.

The popular textbook and literature model I(λx,λm) = K(λx,λm)(1-10-Ax) or its variants for correlating the sample absorption and fluorescence often fails even for the simplest samples where the fluorophore is the only light absorber. Reported is a first-principle model I(λx,λm) = K(λx,λm)Ax,f10-(Ax,sdx+Am,sdm) for correlating the sample fluorescence measured with a conventional spectrofluorometer and its UV-vis absorbance quantified with a conventional UV-vis spectrophotometer. This model can be simplified or expanded for a variety of fluorescence analyses. First, it enables curve-fitting fluorescence intensity as a function of the fluorophore or sample absorbance over a sample concentration range impossible with existing models. Second, it provides the theoretical foundation for an inner-filter-effect (IFE)-correction method developed earlier and explains mathematically the linearity between the IFE-corrected fluorescence and the fluorophore concentration or absorbance. Third, this model can be expanded for quantitative mechanistic studies of fluorescence intensity variations triggered by stimuli treatments. One demonstrated example is to quantify temperature effects on the emission-wavelength-specific and total fluorescence quantum yield of anthracene. We expect that this first-principle model will be broadly adopted for both student education that promotes evidence-based learning and a variety of fluorescence applications where disentangling sample absorption and emission are critical for reliable data analysis.

Determination of colloidal gold nanoparticle surface areas, concentrations, and sizes through quantitative ligand adsorption.

Determination of the true surface areas, concentrations, and particle sizes of gold nanoparticles (AuNPs) is a challenging issue due to the nanoparticle morphological irregularity, surface roughness, and size distributions. A ligand adsorption-based technique for determining AuNP surface areas in solution is reported. Using a water-soluble, stable, and highly UV-vis active organothiol, 2-mercaptobenzimidazole (MBI), as the probe ligand, we demonstrated that the amount of ligand adsorbed is proportional to the AuNP surface area. The equivalent spherical AuNP sizes and concentrations were determined by combining the MBI adsorption measurement with Au(3+) quantification of aqua regia-digested AuNPs. The experimental results from the MBI adsorption method for a series of commercial colloidal AuNPs with nominal diameters of 10, 30, 50, and 90 nm were compared with those determined using dynamic light scattering, transmission electron microscopy, and localized surface plasmonic resonance methods. The ligand adsorption-based technique is highly reproducible and simple to implement. It only requires a UV-vis spectrophotometer for characterization of in-house-prepared AuNPs.

Studying the Effects of Cysteine Residues on Protein Interactions with Silver Nanoparticles.

Studies of protein and organothiol interactions with silver nanoparticles (AgNPs) are important for understanding AgNP nanotoxicity, antimicrobial activity, and material fabrications. Reported herein is a systematic investigation of the effects of both reduced and oxidized protein cysteine residues on protein interactions with AgNPs. The model proteins included wild-type and mutated protein GB3 variants that contain 0, 1, or 2 reduced cysteine residues, respectively. Bovine serum albumin (BSA) that contains a total of 34 oxidized (disulfide-linked) cysteine residues and one reduced cysteine residue was also included. Protein cysteine content has no detectable effect on the kinetics of protein/AgNP binding. However, only proteins that contain reduced cysteine residues induce significant AgNP dissolution. Proteins can slow down, but do not prevent the AgNP dissolution induced by subsequently added organothiols. The insights provided in this work are important to the mechanistic understanding of AgNP stability in biofluids that are rich in proteins and amino acid thiols.