Silica derived from biocompatible silane precursors and containing covalently bound sugar moieties has recently been reported to be a much more biocompatible matrix for protein entrapment than any previously synthesized materials. To better understand the nature of these new materials, the steady-state and time-resolved fluorescence of human serum albumin (HSA) was used to examine the conformation, dynamics, accessibility, thermal stability, and degree of ligand binding after entrapment of the protein into sol−gel-processed glasses derived from either tetraethyl orthosilicate (TEOS) or diglycerylsilane (DGS), which in some cases contained covalently bound gluconamidylsilane (GLS) moieties. It was observed that the initial conformation, accessibility to external analytes, thermal stability, long-term stability, and degree of ligand binding to HSA were best in DGS-derived materials that contained covalently tethered GLS relative to unmodified DGS-derived materials, TEOS, or TEOS/GLS-derived materials. Measurement of protein rotational dynamics showed that entrapment led to an immediate loss of global motion in all materials. However, the restriction of motion was most dramatic in GLS-doped materials, suggesting preferential interactions of the protein with the sugar-coated surfaces. As aging proceeded, both protein dynamics and the degree of ligand binding decreased, with a gradual loss of segmental motion and a significant increase in local motion in the vicinity of the probe, consistent with unfolding and surface adsorption of the protein, leading to loss of function. Overall, our findings suggest that the use of a biocompatible precursor (DGS) and the addition of a covalently bound sugar both contribute to improved protein performance. However, of these two the use of a biocompatible precursor is the most important factor, and in such cases addition of sugars further improves protein performance. In contrast, the use of the sugar-based additive with a nonbiocompatible precursor such as TEOS imparted essentially no benefit, demonstrating the importance of biocompatible processing conditions.
Biocompatible silica derived from diglycerylsilane (DGS) and the sugar modified silane N-(3triethoxysilylpropyl)gluconamide (GLS) has been shown to have great utility for the entrapment of a number of delicate proteins. To further understand the nature of the DGS/GLS composite material, it is important to characterize the local microenvironment and properties of the silica surface within the DGS/ GLS monoliths. In this work, we have monitored both the steady-state and time-resolved anisotropy of the fluorescent probe rhodamine 6G (R6G) to determine the distribution of GLS within DGS/GLSderived materials where varying levels of GLS were added either to the sol or to previously formed DGS-based gels. The data suggest that the addition of GLS to an evolving sol results in preferential coating of silica nanoparticles, which remain present in the gelled material and slowly associate with the continuous polymer network. The preferential presence of GLS on the surface of free particles results in inefficient coating of the monolithic material with the sugarsilane and causes most of the added R6G to associate with the silica skeleton. On the other hand, the addition of GLS to a preformed monolith results in preferential modification of the monolithic silica surface and a much increased level of free dye. A significant amount of R6G-bound nanoparticles remain within these materials over an extended time, suggesting that the nanoparticles do not associate with the GLS-modified silica monolith. The activity of the enzyme horseradish peroxidase was evaluated in DGS derived materials that had GLS added to either the sol or the monolith, and it was determined that optimal activity was obtained in cases where GLS was added to the sol. These results highlight the importance of controlling the time of addition of organosilane additives to evolving sols and suggest that this factor might provide a means to control the properties of the resulting nanocomposite material.
The synthesis and characterization of a novel macroporous silica derived size exclusion chromatography (SEC) packing for quantitative analysis of high molecular weight (MW) polyacrylamide (PAM) are presented. Using this packing, a fast, sensitive and reproducible approach for quantitation of super high-MW PAM in demanding enhanced oil recovery (EOR) waters was developed and the effect of synthesis parameters on the properties of resultant materials was investigated. These parameters include salt addition, reaction temperature and duration, activation condition of functional groups on the silica surface, as well as the reaction cycles required for optimal silica modification. Moreover, SEC analysis conditions, such as mobile phase composition, flow rate, detection and sample preparation, were also explored and an optimal analysis protocol was developed. Under this optimized SEC analysis conditions, the synthesized macroporous materials proved satisfactory for quantification of PAM with average MW up to 22 million Daltons. An SEC analysis required less than few minutes with a detection limit of 1 ng, a linear response range of 0.1 to 75 mgIL with squared R value of 0.99 and reproducibility better than 9.2% RSD (relative standard deviation). The analysis of PAM in highly saline oilfield production water containing interfering high MW polymeric surfactants indicated the recovery ranges from 92.5% to 110.1% for 1.0 mgIL PAM and 94.2% to 103.8% for 50 mgIL PAM solution. This study presented for the first time that the reliable quantization of high MW PAM in highly demanding EOR waters can be achieved by SEC.
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