Highly effective antimicrobial nanocomposites based on hydrogel matrix and silver nanoparticles: long-lasting bactericidal and bacteriostatic effects.

Antimicrobial nanocomposites (NCs) are being used as an alternative antibacterial therapy for killing antibiotic-resistant pathogenic bacteria. The NCs are made of Ag nanoparticles (AgNPs) inside biocompatible hydrogel matrixes. The NCs were synthesized by the absorption of AgNO3 solution into a hydrogel matrix, followed by UV light irradiation, without using additional toxic reactants. The hydrogels used as matrixes are based on N-isopropylacrylamide (NIPAM) and copolymers with different functional groups: 2-acrylamide-2-methylpropanesulfonic acid (AMPS), N-hydroxyethylacrylamide (HEAA) and (3-acrylamidepropil)trimethylammonium chloride (APTMAC). Neutral, anionic and cationic groups were added to the matrixes in order to study their effects on the release of antibacterial species. The NCs were characterized by UV-visible spectroscopy and transmission electronic microscopy. The kinetics of the release of Ag+ ions from the NCs were followed by UV-visible spectroscopy at 300 nm. Biological experiments were based on the plate count method and agar diffusion testing against Pseudomonas aeruginosa. The bacterial death rate using the NCs is higher than when PNIPAM and nanoparticles in solution are used and seems to be related to the large amount of AgNPs contained inside the gels. In all cases, inhibition and diffusion halos were observed upon the exposure of bacterial cultures on agar to NC discs. The presence of both halos confirmed the bactericidal and bacteriostatic effects of the NCs. The reusability (prolonged use) of the materials was demonstrated until the Ag-NP content was exhausted. The NCs with a higher antibacterial capacity are based on a PNIPAM-co-6%APTMAC matrix. It was demonstrated that these NC materials have the capacity to maintain an aseptic/antiseptic zone for 7 to 15 days.

Synthesis of polyaniline (PANI) and functionalized polyaniline (F-PANI) nanoparticles with controlled size by solvent displacement method. Application in fluorescence detection and bacteria killing by photothermal effect.

Polyaniline nanoparticles (PANI-NPs) were easily obtained applying the solvent displacement method by using N-methylpyrrolidone (NMP) as good solvent and water as poor solvent. Different polymers such as polyvinylpyrrolidone (PVP), chondroitin sulfate (ChS), polyvinyl alcohol (PVA), and polyacrylic acid (PAA) were used as stabilizers. Dynamic light scattering and scanning electron microscopy corroborated the size and morphology of the formed NPs. It was demonstrated that the size of nanoparticles could be controlled by setting the concentration of PANI in NMP, the NMP to water ratio, and the stabilizer's nature. The functionalization and fluorescence of NPs were checked by spectroscopic techniques. Since polyaniline show only weak intrinsic luminescence, fluorescent groups were linked to the polyaniline chains prior to the nanoparticle formation using a linker. Polyaniline chains were functionalized by nucleophilic addition of cysteamine trough the thiol group thereby incorporating pendant primary aliphatic amine groups to the polyaniline backbone. Then, dansyl chloride (DNS-Cl), which could act as an extrinsic chromophore, was conjugated to the amine pendant groups. Later, the functionalized polyaniline was used to produce nanoparticles by solvent displacement. The optical and functional properties of fluorescent nanoparticles (F-PANI-NPs) were determined. F-PANI-NPs in the conductive state (pH < 4) are able to absorb near infrared radiation (NIR) creating a photothermal effect in an aqueous medium. Thus, multifunctional nanoparticles are obtained. The application of NIR on a F-PANI-NPs dispersion in contact with Pseudomonas aeruginosa causes bacterial death. Therefore, the F-PANI-NPs could be tracked and applied to inhibit different diseases caused by pathogenic microorganisms and resistant to antibiotics as well as a new disinfection method to surgical materials.

Smart polyaniline nanoparticles with thermal and photothermal sensitivity.

Conductive polyaniline nanoparticles (PANI NPs) are synthesized by oxidation of aniline with persulfate in acid media, in the presence of polymeric stabilizers: polyvinilpyrrolidone (PVP), poly(N-isopropylacrylamide) (PNIPAM), and hydroxylpropylcellulose (HPC). It is observed that the size of the nanoparticles obtained depends on the polymeric stabilizer used, suggesting a mechanism where the aggregation of polyaniline molecules is arrested by adsorption of the polymeric stabilizer. Indeed, polymerization in the presence of a mixture of two polymers having different stabilizing capacity (PVP and PNIPAM) allows tuning of the size of the nanoparticles. Stabilization with biocompatible PVP, HPC and PNIPAM allows use of the nanoparticle dispersions in biological applications. The nanoparticles stabilized by thermosensitive polymers (PNIPAM and HPC) aggregate when the temperature exceeds the phase transition (coil to globule) temperature of each stabilizer (Tpt = 32 °C for PNIPAM or Tpt = 42 °C for HPC). This result suggests that an extended coil form of the polymeric stabilizer is necessary to avoid aggregation. The dispersions are reversibly restored when the temperature is lowered below Tpt. In that way, the effect could be used to separate the nanoparticles from soluble contaminants. On the other hand, the PANI NPs stabilized with PVP are unaffected by the temperature change. UV-visible spectroscopy measurements show that the nanoparticle dispersion changes their spectra with the pH of the external solution, suggesting that small molecules can easily penetrate the stabilizer shell. Near infrared radiation is absorbed by PANI NPs causing an increase of their temperature which induces the collapse of the thermosensitive polymer shell and aggregation of the NPs. The effect reveals that it is possible to locally heat the nanoparticles, a phenomenon that can be used to destroy tumor cells in cancer therapy or to dissolve protein aggregates of neurodegenerative diseases (e.g. Alzheimer). Moreover, the long range control of aggregation can be used to modulate the nanoparticle residence inside biological tissues.

Biointerfacial behavior of stallion spermatozoa adhered to hydrogel surfaces: Impact of the hydrogel chemical composition and the culture medium.

Novel soft materials based on hydrogel are proposed to enhance the selection of high-quality stallion sperm based on their adhesion capacity. The hydrogel surfaces are derived from polyacrylamide (PAAm), which is copolymerized with neutral and ionic co-monomers to modify the interfacial properties. The hydrogels undergo characterization through FTIR spectroscopy, assessment of swelling capacity, and wettability under various experimental conditions. Sperm adhesion capacity on the hydrogels is examined through several parameters including the percentage of bound sperm (%Sp) to hydrogels, tail oscillation intensity and flagellar movement. The biointerfacial properties of sperm-hydrogel systems vary based on the chemical composition of hydrogel as well as the components present in the culture medium. High %Sp and excellent metabolic activity of the spermatozoa are observed on hydrogel surfaces that possess moderate hydrophilicity. Specifically, a cationic hydrogel in BGM3 culture medium and a neutral surface in BGM3 medium supplemented with BSA exhibit favorable outcomes. Scanning Electron Microscopy (SEM) reveals the normal morphology of the head and tail in spermatozoa adhered to the hydrogel. Therefore, these hydrogel surfaces are potential materials for selecting stallion sperm with high quality, and their application could be extended to the study of other mammalian reproductive cells.

Electromagnetic radiation driving of volume changes in nanocomposites made of a thermosensitive hydrogel polymerized around conducting polymer nanoparticles.

Polymeric nanocomposites were obtained by the formation of a thermosensitive hydrogel matrix around conducting polymer (CP) nanoparticles. The CP is able to absorb electromagnetic radiation which is converted into heat and induces the phase transition of the surrounding hydrogel. The method chosen to form the hydrogel is the free radical polymerization of a copolymer (N-isopropylacrylamide (NIPAM) and 2-acrylamide-2-methylpropano sulfonic acid (AMPS), PNIPAM-co-2% AMPS) in the presence of bisacrylamide as the crosslinker. The nanoparticles are polypyrrole nanospheres (PPy NP), polyaniline nanofibers (PANI NF), and polyaniline nanospheres (PANI NP). The morphology of the composites was studied using SEM microscopy and the percentage composition of each material was evaluated by thermogravimetric analysis (TGA). The swelling equilibrium capacity and rate are clearly affected by the nanoparticle shape and nature. However, the nanocomposites LCST are similar to that of the matrix. Upon RF irradiation, the three nanocomposites increase the temperature and reach the LCST after 320 seconds of irradiation (320 kJ). Upon MW application, the local temperature reaches the LCST after only 30 s (21 kJ), resulting in a MW more effective than RF to drive the transition. These results demonstrate that the proposed materials are useful as a remotely driven actuator.

Mechanical and physicochemical behavior of a 3D hydrogel scaffold during cell growth and proliferation.

Some of the essential properties for cellular scaffolding are the capability to maintain the three-dimensional (3D) structure, good adhesion, and adequate elastic modulus during cell growth, migration, and proliferation. Biocompatible synthetic hydrogels are commonly used as cellular scaffolds because they can mimic the natural extracellular matrices (ECMs). However, it is possible that the physicochemical and mechanical behavior of the scaffold changes during cell proliferation and loses the scaffold properties but this is rarely monitored. In this work, the physicochemical and mechanical properties of a macroporous soft material based on poly(N-isopropyl acrylamide) (PNIPAM) have been studied during a period of 75 days at culture condition while bovine fetal fibroblasts (BFF) were grown within the matrix. The interconnected macroporous hydrogel was obtained by cryogelation at -18 °C. The swelling capacity of the scaffold was not altered during cell proliferation but changes in the mechanical properties were observed, beginning with the high elastic modulus (280 kPa) that progressively decreased until mechanical stability (40 kPa) was achieved after 20 culture days. It was observed that the matrix-cell interactions together with collagen production favor normal cellular processes such as cell morphology, adhesion, migration, and proliferation. Therefore, the observed behavior of macroporous PNIPAM as a 3D scaffold during cell growth indicates that the soft matrix is cytocompatible for a long time and preserves the suitable properties that can be applied in tissue engineering and regenerative medicine.

Synthesis of a Smart Conductive Block Copolymer Responsive to Heat and Near Infrared Light.

A method for the synthesis of a linear block copolymer (PNIPAM-b-PANI), containing a thermoresponsive block (poly(N-isopropylacrylamide), PNIPAM) and a Near Infrared (NIR) light-absorbing block (polyaniline, PANI), is reported. The synthetic approach involves a two-step successive polymerization reaction. First, the radical polymerization of NIPAM is done using 4-aminothiophenol as a chain transfer agent for the obtention of thermosensitive block terminated with an aniline (ANI) moiety. Second, the oxidative polymerization of ANI is initiated in ANI moiety of thermosensitive block to grow the second conductive PANI block. 1H nuclear magnetic resonance (NMR) and FT-IR spectroscopy shows the characteristics peaks of both polymeric blocks revealing the successful copolymerization process. Static Light Scattering (SLS) and UV-Visible combined measurements allowed the determination of the Mw for PNIPAM-b-PANI macromolecule: 5.5 × 105 g mol-1. The resulting copolymer is soluble in water (8.3 g L-1) and in non-aqueous solvents, such as ethanol, formic acid, acetonitrile, and others. Both polymer blocks chains show the properties of the polymer chains. The block copolymer shows a lower critical solution temperature (LCST) at the same temperature (32-34 °C) than PNIPAM, while the copolymer shows pH dependent UV-vis-NIR absorption similar to PANI. The PNIPAM block suffers a coil to globule transition upon NIR light irradiation (785 nm, 100 mW), as shown by turbidimetry and Atomic Force Microscopy (AFM), due to local heating (more than 9 °C in 12 min) induced by the NIR absorption at the PANI block. Furthermore, the electrical conductivity of PNIPAM-b-PANI thin films is demonstrated (resistivity of 5.3 × 10-4 Ω-1 cm-1), indicating that the PANI block is present in its conductive form.

Improving the retention and reusability of Alpha-amylase by immobilization in nanoporous polyacrylamide-graphene oxide nanocomposites.

Alpha-amylase was immobilized inside three different polymeric matrices: polyacrylamide hydrogel (PAAm), polyacrylamide-graphene oxide nanocomposite (PAAm-GO) and alginate in order to study and compare the effect of the matrix on the catalytic performance. The morphology, swelling, mechanical properties, retention efficiency, and the catalytic behavior of these newly supported biocatalysts were studied. Nanocomposite made of PAAm-GO matrix incorporated 98% of the enzyme, likely through a cooperative effect, while alginate gels incorporated only 30%. Moreover, the enzyme retention using PAAm-GO reached a value of 97.5%. Starch hydrolysis catalyzed by the immobilized enzyme in PAAm-GO matrix showed similar kinetics profiles up to 5 cycles suggesting that the enzymatic activity is retained. These results compare very favorably with conventional immobilization in alginate where almost no activity was observed after 3 cycles. All results suggest that the PAAm matrices protect the biocatalyst allowing its reusability. Moreover, the improvements in enzyme catalytic properties via immobilization made this system as an excellent candidate in bio-industrial applications such as bioethanol production. Furthermore, the synthesized catalyst could produce a high yield of bioethanol by using enzymes and yeast immobilized in the same PAAm matrix. In this way, it is possible to produce sequential or simultaneous saccharification and fermentation.

Cytotoxicity and bioadhesive properties of poly-N-isopropylacrylamide hydrogel.

Several hydrogel surfaces present properties that simulate the mechanical and physicochemical features of extracellular matrix (ECM), providing a platform that mimic the native cellular milieus. Poly-N-isopropylacrylamide (PNIPAM) hydrogels are receiving attention in biomedical field due to their thermosensibility and soft texture. However, more extensive biocompatibility and cellular interactions studies with cell lines are needed. Therefore, the aim of this study is focus on evaluating the biocompatibility of PNIPAM through cytotoxicity, genotoxicity, and proliferation tests in murine preadipose cells (3T3-L1), human embryonic kidney cells (HEK293) and human carcinoma-derived cells (A549) in presence of hydrogel surfaces. Bioadhesive capacity above PNIPAM surfaces was also analyzed. MTT and neutral red uptake assays shown non-cytotoxic effect of PNIPAM in the studied cell lines. Genotoxicity was evaluated by the single-cell gel electrophoresis assay, where DNA damages were not detected. [3H]-thymidine staining allowed to corroborate that cell proliferation had progressed correctly. Adopted morphologies for each cell line over PNIPAM were similar to cell growing observed on polystyrene, indicating that the surfaces favor the cell attachment during 5 days' culture. The good biocompatibility of PNIPAM surfaces make it an interesting scaffold with clinical potential in tissue regeneration engineering, and a possible adipose and kidney tissue-engineered construct.

Photoreduction of Ru(bpy)(3)2+ by amines in aqueous solution. Kinetics characterization of a long-lived nonemitting excited state.

The photochemistry of Ru(bpy)(3)+2 in the presence of amines was investigated in water by laser flash photolysis. N,N'-Dimethylaniline and p-phenylenediamine quench the luminescent metal to ligand charge transfer (MLCT) excited state of the complex by an electron transfer reaction that produces the semireduced form Ru(bpy)3+ in relatively high yields. On the other hand, triethylamine (TEA) and aniline do not quench the MLCT. Nevertheless, when laser flash irradiation at 532 nm is carried out in the presence of these amines, the formation of Ru(bpy)3+ is clearly detected by its transient absorption at 510 nm. These results are interpreted by an electron transfer reaction with the participation of a nonemitting excited state of the complex, formed independently of the MLCT from the Franck-Condon or the relaxed singlet excited state. The rate constants for the quenching of this state by TEA and aniline and the quantum yields for Ru(bpy)(3)+ were determined. The new state is formed in a very fast process and has a lifetime of ca 4 micros in water.

Bioethanol production by reusable Saccharomyces cerevisiae immobilized in a macroporous monolithic hydrogel matrices.

Performance of yeasts on industrial processes can be dramatically improved by immobilization of the biocatalyst. The immobilization of Saccharomyces cerevisiae inside monolithic macroporous hydrogels were produced by in-situ polymerization of acrylamide around a live yeast suspension under cryogelation conditions. Preculture of the yeasts was not necessary and this innovative and simple procedure is amenable to scaling-up to industrial production. The yeasts were efficiently retained in monolithic hydrogels, presenting excellent mechanical properties and high cell viability. Macroporous hydrogels showed a fast mass transport allowing the hydrogel-yeast complexes achieved similar ethanol yield and productivity than free yeasts, which is larger than those reached with yeasts immobilized in compact hydrogels. Moreover, the same yeasts were able to maintain its activity by up to five reaction cycles with a cell single batch during fermentation reactions.

Distribution of amines in water/AOT/n-hexane reverse micelles: influence of the amine chemical structure.

The distribution of different aliphatic and aromatic amines: n-butylamine (n-BA), isobutylamine (i-BA), tert-butylamine (t-BA), piperidine (PIP), N,N-dimethylaniline (DMA) and N-methylaniline (MA) in water/sodium 1,4-bis(2-ethylhexyl)sulfosuccinate(AOT)/n-hexane reverse micelles was investigated by steady-state fluorescence measurements. The partition constants were measured by an indirect method based on the effect that amine partitioning exert on the bimolecular rate of the reaction between a microphase incorporated fluorophore (Ru(bpy)2+(3)) and the quencher, (Fe(CN)3-(6)). For MA, that can act as a quencher of the fluorophore a direct method was used. The results show that primary amines have larger partition constants than the secondary ones. For tertiary amines the distribution constants were practically negligible. Laser flash photolysis experiments confirmed that tertiary amines, both aliphatic and aromatic, are not incorporated to the micellar pseudophase. The effect of the amine structure on the partition constant was analyzed through linear solvation free energy relationships (LSER) using solute parameters and compared with those obtained for alcohols. Hydrogen bond interactions with the AOT polar heads appear to be the main driving force for the distribution of amines between the organic and micellar pseudophases, whereas the size of the alkyl or aromatic group tends to hinder it.

Functionalised conjugated materials as building blocks of electronic nanostructures.

Two different approaches towards conjugated material (carbon nanotubes, conjugated polymers) functionalisation are presented: covalent bonding of functional groups and covalent interaction with soluble polymers. Covalent functionalisation of carbon nanotubes is made by reaction of the aromatic ring with aryl radicals, produced by reduction of diazonium ions. In the case of conducting polymers, covalent functionalisation is brought about by reaction of polyanilines with diazotized aromatic amines (including amino terminated azo dyes). The non covalent functionalisation of carbon nanotubes is made by wrapping the nanotubes with soluble conducting polyanilines. The functionalised materials are characterised by FTIR spectroscopy, X-ray diffraction, dynamic light scattering, ultraviolet-visible absorption and emission spectroscopy, transmission electron microscopy, cyclic voltammetry, differential electrochemical mass spectroscopy and conductivity measurements. The materials are to build ionic self assembled multilayers using a layer-by-layer deposition process. The charge transport and electrocatalytic behaviour of the assemblies, relevant to the application of the assemblies in nanostructured electrochemical biosensors, are evaluated using different redox molecules and/or its intrinsic electroactivity as probes.

The quenching of indolic compounds by monosubstituted benzenes and the photoreaction with chlorobenzene.

The quenching of the excited singlet of indole and its methyl derivatives by monosubstituted benzenes has been studied in three solvents of different polarities and bimolecular quenching rate constants have been determined. Below the diffusion limit the rate constants decrease when solvent polarity increases, with the exception of those for the quenching by benzonitrile. For the latter the rate constants are near the diffusion limit in all cases. The decrease may be understood in terms of a lower energy of the indole excited state in the more polar solvents. In cyclohexane the remnant emission in the presence of high concentration of the quencher is clearly red shifted, indicating the presence of a new emitting species that can be ascribed to the presence of an exciplex. In some cases the emission of the exciplex can be clearly separated from the molecular fluorescence. In the quenching by chlorobenzene in ethyl acetate and acetonitrile a new highly fluorescent product is formed. This photoreaction is observed with all the indole derivatives and the kinetics of the reaction was followed by the increase in fluorescence intensity at a wavelength were the reactants do not emit. Three major products were identified as phenyl substituted indoles by GC-MS and their fluorescence emission and excitation spectra.