Silica particles with immobilized protein molecules and polymer brushes

We report on the synthesis of a thermosensitive polymer brush‐supported 4‐N,N‐dialkylaminopyridine catalyst and the comparison of its catalytic activity with the corresponding linear polymer‐supported version in the hydrolysis of p‐nitrophenyl acetate (NPA) as well as the effect of lower critical solution temperature (LCST) transition on catalytic activity. The polymer brushes were synthesized from initiator‐functionalized silica particles by surface‐initiated atom transfer radical polymerization of methoxytri(ethylene glycol) methacrylate and 2‐(N‐methyl‐N‐(4‐pyridyl)amino)ethyl methacrylate in the presence of a free initiator. Dynamic light scattering studies showed that the onset temperatures of the LCST transition of polymer brushes in pH 7.52 and 7.82 buffers were 42 and 38 °C, respectively. Under the same reaction conditions, the net initial rate of the hydrolysis of NPA catalyzed by hairy particles was 70–80% of that catalyzed by the free copolymer at the temperature below the LCST of polymer brushes. With further increasing the temperature above the LCST, the plot of logarithm of net initial rate versus inverse temperature exhibited a shift for the reactions catalyzed by hairy particles and leveled off or decreased slightly in the case of using the free copolymer as catalyst, presumably because the structures of the aggregates of hairy particles and free copolymer chains were different. © 2009 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 47: 2853–2870, 2009

Polymer brushes refer to polymer chains that are end-grafted to surfaces or interfaces at high grafting density. Polymer brushes are able to make surface self-assembly or surface coassembly with free block copolymer (BCP) chains, and surface nanostructures with different morphologies are formed on the surfaces. The formation of the surface nanostructures endows materials with new surface properties. To develop a simple and versatile approach to the synthesis of polymer brushes and surface nanostructures, the synthesis of polymer brushes and surface nanostructures on tannic acid (TA) coatings was investigated in this research. TA molecules were coated on the surfaces of amino-modified silica particles (SiO2-NH2) through noncovalent bonds. Quaternized poly(2-(dimethylamino)ethyl methacrylate)-block-polystyrene (qPDMAEMA-b-PS) chains were anchored onto the TA coatings through electrostatic interaction between qPDMAEMA blocks and TA coatings, and PS brushes are formed on the particle surfaces. Thermo gravimetric analysis demonstrates the grafting of the BCP chains on the TA coatings, and the calculation results indicate that the PS chains on the silica particles are in the polymer brush regime. PS brushes on TA coatings are able to make surface coassembly with free BCP chains, and surface nanostructures with different sizes and morphologies are produced on TA coatings. The grafting density of PS blocks and BCP concentration both exert significant influences on the formation of the surface nanostructures. The surface nanostructures can be removed from the particle surfaces in the aqueous solution of sodium chloride. This research provides a simple and versatile method for the fabrication of polymer brushes and removable surface nanostructures on any solid substrates, which have specific interactions with TA molecules.

We report on the design, investigation and comparison of the adhesive properties of poly-(N-isopropylacrylamide) and biocompatible polyethylene glycol-based thermoresponsive brushes. Specifically, a poly-(N-isopropylacrylamide), PNIPAM, poly-(oligo(ethylene glycol) methyl ether methacrylate-co-2-(2-methoxyethoxy)ethyl methacrylate), P(OEGMA-MEO2MA), and poly-(oligo(ethylene glycol) methyl ether methacrylate-co-oligo(propylene glycol) methacrylate), P(OEGMA-OPGMA) brushes were synthesised on planar silicon wafers and silica particles via the surface-initiated atom transfer radical polymerisation. Switching of swelling and surface charge were investigated using spectroscopic ellipsometric and zeta-potential measurements, respectively. Adhesion properties were studied in situ in aqueous solutions at different temperatures using three kinds of AFM probes with well defined tips geometries and radii. We found that: (1) adhesion properties of all polymer brushes reversibly switch with temperature—polymer brushes are completely non-adhesive below the Low Critical Solution Temperature (LCST) and become sticky above the LCST; (2) adhesive force scales nonlinearly with the probe radius; (3) adhesion energy, obtained using Derjaguin/Muller/Toporov approaches, decreases with the increase of the tip radius that is attributed to different depth of penetration of the probes in the polymer layer; (4) adhesion decreases in the sequence PNIPAM – P(OEGMA-MEO2MA) – P(OEGMA-OPGMA). Based on dynamic light scattering and cryo-TEM experiments, the latter effect is attributed to the formation of a thin oligoethylene glycol shell around the hydrophobic polymer core in the case of P(OEGMA-OPGMA). We expect that the P(OEGMA-OPGMA) system, which demonstrates less sticky adhesion properties, could be particularly promising for cell adhesion experiments and tissue engineering.

The potential hazard to humans exposed to nanomaterials such as silica and iron oxide was investigated using an in vitro macrophage cell culture system. Amorphous silica and iron oxide particles and nanomaterials (NMs) were dispersed in cell culture medium supplemented with either bovine serum albumin (BSA), lung lining fluid (LLF) or serum, in order to mimic the body fluids encountered during different routes of exposure in the body. End points investigated included macrophage viability and cytokine production. Silica NMs and particles (50 and 200 nm, respectively) were unmodified (plain) or aminated (NH2 ). Iron oxide NMs and particles, Fe3 O4 45 nm and Fe2 O3 280 nm were also used in this study. Silica particles and NMs induced a dose-dependent increase in cytotoxicity as measured by lactate dehydrogenase (LDH) release. Serum enhanced silica-induced interleukin (IL)-6, IL-10, IL-1β and MCP-1 release, whereas albumin partially inhibited MCP-1 release. Aminated silica, 50 nm was more potent than the 200-nm particles at inducing monocyte chemoattractant protein-1 (MCP-1) production when dispersed in medium or LLF, suggesting a size specific effect for these particles and this cytokine. Iron oxide particles were relatively inert compared with the silica particles and NMs; however, serum and albumin did affect cytokine release in some treatments. In conclusion, the data suggests that serum, compared with medium, BSA and LLF is very potent at enhancing macrophage responses to silica and iron oxide particles and NMs. Size was only influential in LLF for a limited number of parameters, whereas surface chemistry was not of consequence in this in vitro macrophage system.

A series of temperature-sensitive poly(N-isopropylacrylamide) (PNIPA) hydrogels with large pore size and fast response were prepared by carrying out polymerizations in aqueous sodium chloride solutions with different concentrations. In comparison with conventional PNIPA hydrogels, the PNIPA gels thus prepared have remarkably larger swelling ratios below their lower critical solution temperature (LCST), and exhibit much faster response rates as the temperature is raised above their LCST. The improved properties are due to the presence of inorganic salt, NaCl, which leads the phase separation and formation of a heterogeneous porous structure during the polymerizations. The study on the drug release properties shows the macroporous hydrogels exhibit modulated release in response to temperature. The model protein, bovine serum albumin (BSA), can be released completely from the porous hydrogels at the temperature lower than the LCST because the pore size of the hydrogels is larger than the protein molecules. However, the release of BSA from the gels almost stops after a "burst" release in the initial stage at the temperature higher than the LCST because the pores are closed at the high temperature.

In this study, cationic poly(2-(methacryloyloxy)ethyl) trimethylammonium chloride) (PMTAC) brush surfaces were prepared by surface-initiated atom transfer radical polymerization (SI-ATRP), and their properties were systematically investigated to discuss the factors affecting their bactericidal properties and interactions with proteins. Model equations for the analysis of electrophoretic behaviors were considered for accurate parameter estimation to indicate the charge density at the interface. The zeta potential dependency of the PMTAC brushes was successfully analyzed using Smolchowski's equation and the Gouy-Chapman model, which describes the diffusive electric double layer. The analysis of the quartz crystal microbalance with dissipation (QCM-D) indicated that the electrostatic interaction promoted protein adsorption, with a large quantity of a negatively charged protein, bovine serum albumin (BSA), being adsorbed. The bactericidal efficiency of the high-graft-density polymer brush (0.45 chains nm-2) was higher than that of the low-graft-density polymer brush (0.06 chains nm-2). To investigate the mechanism of this phenomenon, we applied the dissipation change (ΔD) of QCM-D analysis. The BSA was likewise adsorbed when the brush structure was changed; however, the negative ΔD indicated that the BSA-adsorbed, high-graft-density PMTAC brush became a rigid state. In the bacteria culture media, the behaviors were the same as BSA adsorption, and the high-graft-density polymer brush was also estimated to be more rigid than the low-graft-density polymer brush. Moreover, for S. aureus adhesion after incubating in TSB, a small slope of ΔD/ΔF plots considered initial adsorption of bacteria on the high-graft-density polymer brush strongly interacted compared to that of the low-graft-density polymer brush. The scattered value of the slope of ΔD/ΔF on the high-graft-density polymer brush was considered to be due to the dead bacteria between the bacteria and the polymer brush interface. These investigations for a well-defined cationic polymer brush will contribute to the design of antibacterial surfaces.

Addition of nonionic surfactant, C(12)E(9), to an aqueous dispersion of charge stabilized silica nanoparticles renders particle aggregation reversible. In contrast, aggregation of the same silica particles in aqueous solutions is irreversible. We use a combination of small-angle X-ray scattering (SAXS) and contrast matching small-angle neutron scattering (SANS) to investigate interparticle interactions and microstructure in dispersions of silica particles in aqueous nonionic surfactant solutions. We show that the silica particles interact through a screened Coulombic interaction in aqueous dispersions; interestingly, this interparticle interaction is hard-sphere-like in surfactant solutions. In surfactant solutions, we show that the final surfactant-particle structure can be modeled as 14 micelles adsorbed (on average) on the surface of each silica particle. This gives rise to the short-range interparticle repulsion that makes particle aggregation reversible, and results in the hard sphere interparticle interaction potential. Finally, we show that adsorption of polyethylene imine on the surface of the silica particles prevents adsorption of surfactant micelles on the particle surface.

Using Monte Carlo simulations, we investigate the protein adsorption behaviors on polymer brushes in the presence of hydrogen bonding between proteins and polymer brushes. It is interesting to observe in simulation that as a function of grafting density, the amount of protein adsorbed passes through a minimum, that is, a critical grafting density with the minimum amount of adsorbed protein exists. The existence of a critical grafting density uncovers two different driving forces of protein adsorption. The driving force of protein adsorption is mainly the hydrogen bonding between proteins and polymer brushes when the grafting density is above the critical value, whereas the driving force is the attraction between proteins and the hydrophobic substrate when the grafting density is below the critical value. It is extraordinary that at the critical grafting density, increasing the hydrophilicity of the polymer brushes will lead to an increase in protein adsorption, even when the formation of hydrogen bonding between the proteins and polymer brushes is suppressed by the increasing hydrophilicity of the polymer brushes. This mainly results from the conformational transition of the polymer chains from collapsed to stretched when the hydrophilicity is increased. It is noteworthy that the relationship between protein adsorption and grafting density reported in this study is quite consistent with our previous experimental work. Beyond that, the simulation results also provide detailed information about protein adsorption location and grafting chain conformation, which helps us to better understand the protein adsorption behaviors on polymer brushes induced by hydrogen bonding.

Brush-like block copolymers with poly(t-butyl methacrylate) (PBMA) and poly(N-isopropylacrylamide) (PNIPAAm) as side arms, PBMA-b-PNIPAAm, were designed and synthesized via a simple free radical polymerization route. The chemical structure and molecular weight of these polymer brushes were characterized and determined by nuclear magnetic resonance (1H NMR), Fourier transform infrared spectrometry (FTIR) and gel permeation chromatography (GPC). The micellar formation by these polymer brushes in aqueous solutions were detected by a surface tension technique, and the critical micelle concentration (CMC) ranged from 1.53 to 8.06 mg L−1. The morphology and geometry of polymer micelles were investigated by transmission electron microscope (TEM) and dynamic light scattering (DLS). The polymer micelles assume the regularly-spherical core-shell structure with well-dispersed individual nanoparticles, and the particle size was in the range from 36 to 93 nm. The PNIPAAm segments exhibited a thermoreversible phase transition, so the resulting block polymer brushes were temperature-sensitive and the low critical solution temperature (LCST) was determined by UV-vis spectrometer at about 28.82–29.40°C. The characteristic parameters of the polymer micelles such as CMC, micellar size and LCST values were affected by their compositional ratios and the length of hydrophilic or hydrophobic chains. The evaluation for caffeine drug release behavior of the block polymer micelles demonstrated that the self-assembled micelles exhibited thermal-triggered properties in controlled drug release.

Polymer brushes engineered to "specifically capture" and "release on demand" analytes such as dyes, proteins, and cells find biomedical applications ranging from protein immobilization to cell death. Utilizing a disulfide-linker-containing monomer as a building block enables the fabrication of a redox-responsive polymer brush platform with the "catch and release" attribute. Herein, thiol-reactive redox-responsive polymer brushes are fabricated using a pyridyl disulfide-based monomer, and their postpolymerization functionalization is demonstrated via thiol-disulfide exchange reaction with thiol-containing dyes, (bio)molecules, and cell adhesive ligands. After establishing reversible conjugation using a fluorescent dye and other model compounds, copolymer brushes postmodified with thiol-containing mannose demonstrated selective immobilization of concanavalin A in the presence of peanut agglutinin. In addition, a thiolated RGD peptide was conjugated to the side chain of polymer brushes to facilitate cell adhesion, followed by on-demand harvesting. To enable localized drug delivery to surface-adhered cells, orthogonal chain end and side chain functionalization using the thiol-Michael addition and thiol-disulfide exchange reaction, respectively, was used to conjugate the cell adhesive RGD peptide and the anticancer drug doxorubicin (DOX). On-demand DOX release and internalization by surface-bound cancer cells were demonstrated via cleavage of disulfide linkages in the presence of a reducing agent. This approach may provide an attractive methodology to deliver therapeutic agents precisely to specific cells.

Protein-repellent reactive surfaces that promote localized specific binding are highly desirable for applications in the biomedical field. Nonspecific adhesion will compromise the function of bioactive surfaces, leading to ambiguous results of binding assays and negating the binding specificity of patterned cell-adhesive motives. Localized specific binding is often achieved by attaching a linker to the surface, and the other side of the linker is used to bind specifically to a desired functional agent, as e.g. proteins, antibodies, and fluorophores, depending on the function required by the application. We present a protein-repellent polymer brush enabling highly specific covalent surface immobilization of biorecognition elements by strain-promoted alkyne-azide cycloaddition click chemistry for selective protein adhesion. The protein-repellent polymer brush is functionalized by highly localized molecular binding sites in the low micrometer range using polymer pen lithography (PPL). Because of the massive parallelization of writing pens, the tunable PPL printed patterns can span over square centimeter areas. The selective binding of the protein streptavidin to these surface sites is demonstrated while the remaining polymer brush surface is resisting nonspecific adsorption without any prior blocking by bovine serum albumin (BSA). In contrast to the widely used BSA blocking, the reactive polymer brushes are able to significantly reduce nonspecific protein adsorption, which is the cause of biofouling. This was achieved for solutions of single proteins as well as complex biological fluids. The remarkable fouling resistance of the polymer brushes has the potential to improve the multiplexing capabilities of protein probes and therefore impact biomedical research and applications.

This study demonstrates that the thermally induced collapse of end-grafted poly(N-isopropylacrylamide) (PNIPAM) above the lower critical solution temperature (LCST) of 32 degrees C depends on the chain grafting density and molecular weight. The polymer was grafted from the surface of a self-assembled monolayer containing the initiator (BrC(CH3)2COO(CH2)11S)2, using surface-initiated atom transfer radical polymerization. Varying the reaction time and monomer concentration controlled the molecular weight, and diluting the initiator in the monolayer altered the grafting density. Surface force measurements of the polymer films showed that the chain collapse above the LCST decreases with decreasing grafting density and molecular weight. At T > LCST, the advancing water contact angle increases sharply on PNIPAM films of high molecular weight and grafting density, but the change is less pronounced with films of low-molecular-weight chains at lower densities. Below the LCST, the force-distance profiles exhibit nonideal polymer behavior and suggest that the brush architecture comprises dilute outer chains and much denser chains adjacent to the surface.

Albumin-bound fatty acids induce mitochondrial oxidant stress and impair antioxidant responses in proximal tubular cells

Synthetic polymer single-chain nanoparticles (SCNPs) are an emerging new class of nanomaterials that possess similar folded structures as natural proteins. However, most SCNPs reported so far are p...

Reduction of protein adsorption on well-characterized polymer brush layers with varying chemical structures