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Synthesis and lectin recognition of polystyrene core-glycopolymer corona nanospheres.

Polymeric nanospheres with a polystyrene core and a glucosyloxyethyl methacrylate (GEMA) oligomer corona were synthesized by the free radical coplymerization of styrene (M(1)) plus a GEMA macromonomer (M(2)) at various molar ratios (M(1)/M(2) = 50-150) in the presence of AIBN (1 mol % to the total monomer) in an ethanol/water (3/2, v/v) solvent. The size of the nanospheres was controlled from 300 to 620 nm by altering the monomer ratio. The size distributions were significantly narrow. The amount of glucose conjugated per unit surface area of the nanosphere, which was analyzed by the anthron-sulfuric acid method, was 1.01-2.28 microg cm(-1), which increased with an increase in size. The transmittance of a solution of dispersed nanospheres (the corresponding glucose concentration was 73 microM) increased by the addition of the glucose-binding protein concanavalin A (Con A) (1-50 microM), indicating that the nanospheres were being precipitated by the cross-linking of ConA. An enzyme-linked lectin assay (ELLA) revealed that Con A bound to the glucose on the nanospheres 250-700-fold more than to monomeric glucose. The binding activity to the nanospheres was less than that to a GEMA oligomer, and decreased with an increase in the amount of GEMA oligomer grafted onto the nanosphere, possibly because of steric hindrance of the lectin binding to the glucose on the nanospheres. The polystyrene core-glycopolymer corona nanosphere is a useful material for studying sugar-biomolecule recognition.

Concanavalin A↗

Novel thymine-functionalized polystyrenes for applications in biotechnology. 2. Adsorption of model proteins.

This paper investigates the adsorption of bovine serum albumin (BSA) and bovine hemoglobin (BHb) model proteins onto novel thymine-functionalized polystyrene (PS-VBT) microspheres, in comparison with polystyrene (PS) microspheres. Maximum adsorption was obtained for both proteins near their corresponding isoelectric points (pI at pH = 4.7 for BSA and 7.1 for BHb). FTIR and adsorption isotherm analysis demonstrated that, although both proteins were physisorbed onto PS through nonspecific hydrophobic interactions, adsorption onto the functionalized copolymers occurred by both physisorption and chemisorption via hydrogen bonding. FTIR analysis also indicated conformational changes in the secondary structure of BSA and BHb adsorbed onto PS, whereas little or no conformation change was seen in the case of adsorption onto PS-VBT. Atomic force microscopy (AFM), consistent with the isotherm results, also demonstrated monolayer adsorption for both proteins. AFM images of BSA adsorbed onto copolymers with 20 mol % surface VBT loading showed exclusively end-on orientation. Adsorption onto copolymers with lower functionality showed mixed end-on and side-on orientation modes of BSA, and only the side-on orientation was observed on PS. The AFM results agreed well with theoretically calculated and experimentally obtained adsorption capacities. AFM together with calculated and observed adsorption capacity data for BHb indicated that this protein might be highly compressed on the copolymer surface. Adsorption from a binary mixture of BSA and BHb onto PS-VBT showed good separation at pH=7.0; approximately 90% of the adsorbed protein was BHb. The novel copolymers have potential applications in biotechnology.

Adsorption↗

Screening and characterization of affinity peptide tags specific to polystyrene supports for the orientated immobilization of proteins.

Dodecapeptides that exhibit a high affinity specific to a polystyrene surface (PS-tags) were screened using an Escherichia coli random peptide display library system, and the compounds were used as a peptide tag for the site-specific immobilization of proteins. The various PS-tags obtained after 10 rounds of biopanning selection were mainly composed of basic and aliphatic amino acid residues, most of which were arranged in close proximity to one another. Mutant-type glutathione S-transferases (GSTs) fused with the selected PS-tags, PS19 (RAFIASRRIKRP) and PS23 (AGLRLKKAAIHR) at their C-terminus, GST-PS19 and GST-PS23, when adsorbed on the PS latex beads had a higher affinity than the wild-type GST, and the specific remaining activity of the immobilized mutant-type GSTs was approximately 10 times higher than that of the wild-type GST. The signal intensity detected for GST-PS19 and GST-PS23 adsorbed on hydrophilic and hydrophobic PS surfaces using an anti-peptide antibody specific for the N-terminus peptide of GST was much higher than that for the wild-type GST. These findings indicate that the mutant-type GSTs fused with the selected peptide tags, PS19 and PS23, could be site-specifically immobilized on the surface of polystyrene with their N-terminal regions directed toward the solution. Thus, the selected peptide tags would be useful for protein immobilization in the construction of enzyme-linked immunosorbent assay (ELISA) systems and protein-based biochips.

Absorption↗

Polytetrahydrofuran cross-linked polystyrene resins for solid-phase organic synthesis.

Currently, divinylbenzene cross-linked polystyrene (DVB-PS) is the polymer of choice for use in solid-phase organic synthesis (SPOS). While much research has been directed toward the optimization of linker groups for the attachment of compounds to the polymer, the development of new polymers themselves has been relatively neglected. In an attempt to overcome the shortcomings of DVB-PS and to develop new polymers with optimum properties for use in organic synthesis, we have prepared a series of polystyrene polymers that incorporate flexible polytetrahydrofuran (PTHF) based cross-linkers. The objective of incorporating PTHF into the polymers was to slightly increase the overall polarity of the polymer and thus render the resins more organic solvent-like. Since the degree to which a resin swells in and absorbs a particular solvent correlates to how well substrates attached to the polymer are solvated, we compared the swelling of our new resins to commercially available DVB-PS resins. In all cases, we found that our resins swelled to a much greater extent than do DVB-PS resins, and their use should therefore allow for SPOS reaction conditions that more closely mimic homogeneous solution-phase conditions. It was also found that the PTHF chain length of the cross-linker does not affect the level of swelling since all of our cross-linkers afford resins with comparable levels of increased swelling. Furthermore, we have examined the utility of our resins in directed ortho-metalation reactions and found that the increased swelling of our resins allows for isolation of reaction products in yields comparable to what is achieved using standard solution-phase conditions.

Combinatorial Chemistry Techniques↗

Tablets of functionalized polystyrene beads alone and in combination with solid reagents or catalysts. Preparation and applications in parallel solution and solid phase synthesis.

Pretreatment of polystyrene beads with a nonpolar organic solvent is the key for the generation of mechanically robust tablets consisting of neat functionalized polystyrene beads, both alone and in combination with solid reagents or catalysts. The novel dosing methodology provides accurately preweighed tablets in virtually any shape and size and with excellent disintegration properties, speeding up parallel solution and solid phase synthesis. The use of tablets is demonstrated in parallel Mitsunobu and acylation reactions.

Catalysis↗

Development of cross-linked polystyrene-supported chiral amines featuring a fluorinated linker for gel-phase 19F NMR spectrometry monitoring of reactions.

Ten cross-linked polystyrene-supported, protected chiral amines featuring both a spacer, comprising from 5 to 15 atoms, and a fluorinated linker have been successfully prepared. The development of the monitoring technique by gel-phase 19F NMR spectrometry on cross-linked polystyrene derivatives proved to be of high value in four steps of the process, as shown by the comparison of data gathered from both a classic NMR spectrometer and elemental analysis. Gel-phase 19F NMR spectrometry, thus, constitutes a useful technique that complements IR and 13C NMR spectrometries for the qualitative monitoring of reactions. In addition, quantitative determination of the conversion in a given transformation is possible, provided that 19F chemical shifts of the substrate and the product be different enough (Deltadelta>base width), as illustrated by the Mitsunobu coupling process (16-->17). The technique is nondestructive, and the samples used to monitor the reactions may be returned to the reaction medium. Deprotection of the above amines was achieved and furnished eight of the final resins in good to acceptable purity for future applications.

Amines↗

Polynuclear aromatic hydrocarbon and particulate emissions from two-stage combustion of polystyrene: the effects of the secondary furnace (afterburner) temperature and soot filtration.

Laboratory experiments were conducted in a two-stage horizontal muffle furnace in order to monitor emissions from batch combustion of polystyrene (PS) and identify conditions that minimize them. PS is a dominant component of municipal and hospital waste streams. Bench-scale combustion of small samples (0.5 g) of shredded styrofoam cups was conducted in air, using an electrically heated horizontal muffle furnace, kept at Tgas = 1000 degrees C. Upon devolatilization, combustion of the polymer took place in a diffusion flame over the sample. The gaseous combustion products were mixed with additional air in a venturi and were channeled to a secondary muffle furnace (afterburner) kept at Tgas = 900-1100 degrees C; residence time therein varied between 0.6 and 0.8 s. At the exits of the primary and the secondary furnace the emissions of CO, CO2, O2, NOx, particulates as well as volatile and semivolatile hydrocarbons, such as polycyclic aromatic hydrocarbons (PAH), were monitored. Online analyzers, gravimetric techniques, and gas chromatography coupled to mass spectrometry (GC-MS) were used. Experiments were also conducted with a high-temperature barrier filter, placed just before the exit of the primary furnace to prevent the particulates from entering into the secondary furnace. Results demonstrated the beneficial effect of the afterburner in reducing PAH concentrations, including those of mutagenic species such as benzo[a]pyrene. Concentrations of individual PAH exhibited a pronounced after burner temperature dependence, typically ranging from a small decrease at 900 degrees C to a larger degree of consumption at 1100 degrees C. Consumption of PAH was observed to be the dominant feature at 900 degrees C, while significant quantities of benzene and some of its derivatives, captured by means of carbosieve/Carbotrap adsorbents, were formed in the afterburner at a temperature of 1000 degrees C. In the primary furnace, about 30% of the mass of the initial polystyrene was converted into soot, while the total mass of PAH represented about 3% of the initial mass of combustible. The afterburner reduced the particulate (soot) emissions by only 20-30%, which indicates that once soot is formed its destruction is rather difficult because its oxidation kinetics are slow undertypical furnace conditions. Moreover, increasing the afterburnertemperature resulted in an increasing trend of soot emissions therefrom, which might indicate competition between soot oxidation and formation, with some additional formation occurring at the higher temperatures. Contrary to the limited effect of the afterburner, high-temperature filtration of the combustion effluent prior to the exit of the primary furnace allowed for effective soot oxidation inside of the ceramic filter. Filtration drastically reduced soot emissions, by more than 90%. Limited soot formation in the afterburner was again observed with increasing temperatures. The yields of both CO and CO2 were largely unaffected by the temperature of the afterburner but increased at the presence of the filter indicating oxidation therein. A previously developed kinetic model was used to identify major chemical reaction pathways involving PAH in the afterburner. The experimental data at the exit of the primary furnace was used as input to these model computations. A first evaluation of the predictive capability of the model was conducted for the case with ceramic filter and a temperature of 900 degrees C. The afterburner was approximated as a plug-flow reactor, and model predictions at a residence time of 0.8 s were compared to experimental data collected at its exit. In agreement with the experimental PAH concentration, only a minor impact of the afterburner treatment was observed for most species at 900 degrees C. OH was deduced to be the major reactant with a mole fraction about 4 orders of magnitudes higher than that of hydrogen radicals. Evidence for the need of further work on the quantitative assessment of oxidation of PAH and their radicals is given.

Air Pollutants↗

Adsorption of a cationic dye molecule on polystyrene microspheres in colloids: effect of surface charge and composition probed by second harmonic generation.

Nonlinear optical probe, second harmonic generation (SHG), of the adsorption of the dye molecule malachite green (MG), in cationic form at pH < or = 5, on polystyrene microspheres in aqueous solution is used to study the effect of surface charge and composition on molecular adsorption. Three types of polystyrene microspheres with different surface composition are investigated: (1) a sulfate terminated, anionic surface, (2) a neutral surface without any functional group termination, and (3) an amine terminated, cationic surface. The cationic dye was found to adsorb at all three surfaces, regardless of surface charge. The adsorption free energies, DeltaG's, measured for the three surfaces are -12.67, -12.39, and -10.46 kcal/mol, respectively, with the trend as expected from the charge interactions. The adsorption density on the anionic surface, where attractive charge-charge interaction dominates, is determined by the surface negative charge density. The adsorption densities on the neutral and cationic surfaces are on the other hand higher, perhaps as a result of a balance between minimizing repulsive charge interaction and maximizing attractive molecule-substrate and intermolecular interactions. The relative strength of the SH intensity per molecule, in combination of a model calculation, reveals that the C(2) axis of the MG molecule is nearly perpendicular to the surface on the anionic surface and tilts away from the surface norm when the surface is neutral and further away when cationic. Changing the pH of the solution may alter the surface charge and subsequently affect the adsorption configuration and SH intensity.

Adsorption↗

Photochemical interaction of polystyrene nanospheres with 193 nm pulsed laser light.

The photochemical interaction of 193 nm light with polystyrene nanospheres is used to produce particles with a controlled size and morphology. Laser fluences from 0 to 0.14 J/cm2 at 10 and 50 Hz photofragment nearly monodisperse 110 nm spherical polystyrene particles. The size distributions before and after irradiation are measured with a scanning mobility particle sizer (SMPS), and the morphology of the irradiated particles is examined with a transmission electron microscope (TEM). The results show that the irradiated particles have a smaller mean diameter ( approximately 25 nm) and a number concentration more than an order of magnitude higher than nonirradiated particles. The particles are formed by nucleation of gas-phase species produced by photolytic decomposition of nanospheres. A nondimensional parameter, the photon-to-atom ratio (PAR), is used to interpret the laser-particle interaction energetics.

Lasers↗

Adsorption of an endoglucanase from the hyperthermophilic Pyrococcus furiosus on hydrophobic (polystyrene) and hydrophilic (silica) surfaces increases protein heat stability.

The interaction of an endoglucanase from the hyperthermophilic microorganism Pyrococcus furiosus with two types of surfaces, that is, hydrophobic polystyrene and hydrophilic silica, was investigated, and the adsorption isotherms were determined. The adsorbed hyperthermostable enzyme did not undergo loss of biological activity. A model was proposed for the mechanism of interaction of the enzyme with the surface based on the shape of the adsorption isotherm, the morphological characteristics of the enzyme, and the thermodynamic parameters of the system. The enzyme was irreversibly immobilized at the solid/liquid interface even at high temperatures, and most interestingly, it acquired further heat stabilization upon adsorption. The denaturation temperature increased from 108 degrees C in solution to 116 degrees C upon adsorption on hydrophilic silica particles. Adsorption on the hydrophobic polystyrene surface even shifted the denaturation temperature to 135 degrees C, the most extreme experimentally determined protein denaturation temperature ever reported. Maintenance of the biological function particularly at high temperatures is important for the development of solid substrate immobilized enzymes for applications in biocatalysis and biotechnology. This also presents an additional stabilization mechanism employed by nature where the extracellular hyperthermostable enzyme remains folded and active at the extreme temperatures of its natural environment by adsorption on the surface of rocks and other materials appearing in the surroundings of the microorganism.

Adsorption↗

Surface structural characterization of protein- and polymer-modified polystyrene microspheres by infrared-visible sum frequency generation vibrational spectroscopy and scanning force microscopy.

Structural investigations of bare and surface-modified polystyrene microspheres (beads) have been carried out by infrared-visible sum frequency generation (SFG) vibrational spectroscopy and scanning force microscopy (SFM). Bead surfaces have been modified by either the covalent linking of immunoglobulin G (IgG) and bovine serum albumin (BSA) or the nonspecific adsorption of a Pluronic surfactant. After surface modification with protein, SFG signals in the aliphatic CH-stretch region are detected at both the buffer/bead and air/bead interfaces, indicating that some amino acid residues in proteins adopt preferred orientations. SFG results indicate that the hydrophobic poly(propylene glycol) moieties in the Pluronic order when adsorbed onto the bead, at both the buffer/bead and air/bead interfaces, whereas hydrophilic poly(ethylene glycol) groups align to a lesser extent. SFG spectra also show that the phenyl rings of bare polystyrene beads in contact with air or buffer are ordered, with a dipole component directed along the surface normal, but become less ordered after the adsorption of either proteins or the polymer. Molecular orientation and ordering at the bead surface affect its hydrophobicity and aggregation behavior. SFM results reveal the formation of nonuniform islands when bare beads with more hydrophobic character are spun-cast onto a silica substrate. In the presence of adsorbed protein, a hexagonal packing of beads, with some defects, is observed, depending on the bulk pH and the type of attached protein. Adsorbed Pluronic causes the beads to aggregate in a disordered fashion, as compared to the behavior of bare and protein-modified beads.

Air↗

Encapsulation of polystyrene latex with temperature-responsive poly(N-isopropylacrylamide) via a self-assembling approach and the adsorption behaviors therein.

Temperature-responsive poly(N-isopropylacrylamide) (PNIPAm)-encapsulated polystyrene latex was prepared via a layer-by-layer self-assembly of cationic and anionic polyNIPAms alternately. Studies showed that the size of PNIPAm-encapsulated polystyrene particles (m-PS) increased with the encapsulation manipulation, as verified by both transmission electron microscopy and dynamic light scattering measurements. The m-PS underwent a dramatic volume decrease at the lower critical solution temperature (LCST) of PNIPAm, with the onset temperature shifting to a higher temperature range as the number of encapsulating layers increased. A qualitative study on the adsorption behavior of Ag nanoparticles revealed that while the pristine (p) Ag nanoparticles were predominantly adsorbed on the m-PS below the LCST, the hydrophobically modified one (m-Ag) was preferentially adsorbed on the same PS above the LCST, which corresponded to the hydrophilic to hydrophobic transition of the m-PS at the LCST.

Acrylic Resins↗

Periodic ZnO nanorod arrays defined by polystyrene microsphere self-assembled monolayers.

We demonstrate a low-cost and effective method to fabricate hexagonally patterned, vertically aligned ZnO nanorod arrays. Selective wet-etching is used to develop the catalyzing gold particle hexagonal pattern with the aid of a polystyrene microsphere self-assembled monolayer. The gold particles have tunable sizes independent of the polystyrene microsphere's diameter and are inherently round in shape. Each ZnO rod is grown individually from a catalyzing site via catalyst-initiated epitaxy, and the original hexagonal periodicity is well-preserved. The rods have flat ends, and the diameters of the rods can be controlled well by the amount of source materials. This method provides a promising way to create ZnO one-dimensional nanostructures for applications as two-dimensional photonic crystal, sensor arrays, nanolaser arrays, and optoelectronic devices.

Crystallization↗

Simple synthesis of sialyllactose-carrying polystyrene and its binding with influenza virus.

Glycoconjugate polystyrenes bearing sialyllactose moieties were prepared via a simple method from a mixture of alpha2-6 and a2-3 linked sialyllactose isomers of bovine milk origin. The reducing end of sialyllactose was converted to an amino function with ammonium hydrogen carbonate and then coupled with p-vinylbenzoyl chloride. The resulting styrene derivative substituted with sialyllactose via an amide linkage was polymerized with ammonium peroxodisulfate and N,N,N',N-tetramethylethylenediamine in water at 30 degrees C. The interaction of the glycopolymer with influenza A and B viruses was investigated by three different methods. The glycopolymer inhibited the hemagglutination of influenza A virus (PR/8/34) and its activity was 10(3) times higher than that of the oligosaccharide itself. The cytopathic effect of virus-infected MDCK (Madine-Darby canine kidney) cells was inhibited by the glycopolymer. The homopolymer showed 10(2) times higher inhibitory activity than naturally-occurring fetuin. It was also found that various viruses could be trapped by the glycopolymer adsorbed on a polystyrene surface. The inhibitory and trapping activities of the glycopolymers were correlated with the sialyl linkage specificities of the virus strains.

Animals↗

Syntheses, characterization and application of cross-linked polystyrene-ethyleneglycol acrylate resin (CLPSER) as a novel polymer support for polypeptide syntheses.

Cross-linked polystyrene-ethyleneglycol acrylate resin (CLPSER) was developed for the solid-phase synthesis of peptide by introducing a cross-linker, O,O'-bis(2-acrylamidopropyl)polyethylene glycol(1900) (Acr(2)PEG), into polystyrene. The cross-linker was prepared by treating acryloyl chloride with O,O'-bis(2-aminopropyl) polyethylene glycol(1900) [(NH(2))(2)PEG] in the presence of diisopropylethylamine. The copolymer was prepared either by bulk or inverse suspension copolymerization of Acr(2)PEG(1900) and styrene using sorbitan monolaurate as the suspension stabilizer, and a mixture of ammonium peroxodisulfate and benzoyl peroxide as the radical initiators. The resin was characterized using gel-phase (13)C NMR, infrared (KBr) spectroscopic techniques and the morphological features of the resin were investigated using scanning electron microscopy photographs. CLPSER showed excellent swelling in a broad range of solvents and was found to be chemically inert to various reagents and solvents used in solid-phase peptide synthesis. To demonstrate the usefulness of the new resin in polypeptide synthesis, the support was derivatized with an 'internal reference' amino acid (norleucine) and a handle 4-(4-hydroxymethyl-3-methoxy)butyric acid. The new resin was compared with commercial supports such as Merrifield and Sheppard resins by synthesizing an acyl carrier protein (65-74) fragment under the same experimental conditions. HPLC profiles revealed the high efficiency of the newly developed support. Resin capability in peptide synthesis was further demonstrated by the solid phase synthesis of a 25-residue peptide from the E2/NS1 region hepatitis C viral polyprotein.

Acrylates↗

Towards biodegradable polyolefins: strategy of anchoring minute quantities of monosaccharides and disaccharides onto functionalized polystyrene, and their effect on facilitating polymer biodegradation.

A hypothesis was developed, and successfully tested, to greatly increase the rates of biodegradation of polyolefins, by anchoring minute quantities of glucose, sucrose or lactose, onto functionalized polystyrene (polystyrene-co-maleic anhydride copolymer) and measuring their rates of biodegradation, which were found to be significantly improved.

Bacillus↗

Evidence for autolysin-mediated primary attachment of Staphylococcus epidermidis to a polystyrene surface.

Biofilm formation on a polymer surface which involves initial attachment and accumulation in multilayered cell clusters (intercellular adhesion) is proposed to be the major pathogenicity factor in Staphylococcus epidermidis foreign-body-associated infections. We have characterized two distinct classes of biofilm-negative Tn917 mutants in S. epidermidis affected in initial attachment (class A) or intercellular adhesion (class B). mut1 (class A mutant) lacks five surface-associated proteins with molecular masses of 120, 60, 52, 45 and 38 kDa and could be complemented by transformation with a 16.4 kb wild-type DNA fragment. The complemented mutant was able to attach to a polystyrene surface, to form a biofilm, and produced all of the proteins missing from mut1. Subcloning experiments revealed that the 60 kDa protein is sufficient for initial attachment. Immunofluorescence microscopy using an antiserum raised against the 60 kDa protein showed that this protein is located at the cell surface. DNA-sequence analysis of the complementing region revealed a single open reading frame which consists of 4005 nucleotides and encodes a deduced protein of 1335 amino acids with a predicted molecular mass of 148kDa. The amino acid sequence exhibits a high similarity (61% identical amino acids) to the atl gene product of Staphylococcus aureus, which represents the major autolysin; therefore the open reading frame was designated atlE. By analogy with the S. aureus autolysin, AtlE is composed of two bacteriolytically active domains, a 60 kDa amidase and a 52 kDa glucosaminidase domain, generated by proteolytic processing. The 120 kDa protein missing from mut1 presumably represents the unprocessed amidase and glucosaminidase domain after proteolytic cleavage of the signal- and propeptide. The 45 and 38kDa proteins are probably the degradation products of the 60 and 52 kDa proteins, respectively. Additionally, AtlE was found to exhibit vitronectin-binding activity, indicating that AtlE plays a role in binding of the cells not only to a naked polystyrene surface during early stages of adherence, but also to plasma protein-coated polymer surfaces during later stages of adherence. Our findings provide evidence for a new function of an autolysin (AtlE) in mediating the attachment of bacterial cells to a polymer surface, representing the prerequisite for biofilm formation.

Adhesins, Bacterial↗

Development of a rapid polymerase chain reaction-ELISA assay using polystyrene beads for the detection of Toxoplasma gondii DNA.

AIMS: To develop a rapid colourimetric assay for the detection of Toxoplasma gondii DNA using polystyrene beads as solid support. METHODS AND RESULTS: A nested-polymerase chain reaction (PCR)-ELISA assay for the detection of T. gondii DNA was standardized by optimizing the hybridization time and probe concentration. Its detection threshold was then determined and compared with Southern blotting hybridization. These were found to be equivalent, but the PCR-ELISA-beads test is easier to perform and the turnaround time is much shorter than with Southern blot. CONCLUSIONS: The PCR-ELISA-beads assay is a valuable tool for the detection of T. gondii DNA. SIGNIFICANCE AND THE IMPACT OF THE STUDY: Our results demonstrate that this PCR-ELISA assay, using polystyrene beads, can be used as a routine diagnostic test for the detection of T. gondii in clinical laboratories.

Animals↗