Influence of the Sequence Motive Repeating Number on Protein Folding in Spider Silk Protein Films.

Like multiblock copolymers, spider silk proteins are built of repetitive sequence motives. One prominent repetitive motif is based on the consensus sequence of spidroin 4 of the spider Araneus diadematus ADF4. The number x of the repeating sequence motives (C) determines the molecular weight of the recombinant ADF4-based, engineered spider silk protein denoted as eADF4(Cx). eADF4(Cx) can be used as a model for intrinsically disordered proteins (IDP) and to elucidate their folding. Herein, the influence of the variation of the sequence motive repeating number x (x = 1, 2, 4, 8, 16) on the protein folding within eADF4(Cx) films was investigated. eADF4(Cx) films were cast from 1,1,1,3,3,3-hexafluoropropan-2-ol (HFIP) solutions onto planar silicon model substrates, revealing mainly helical or random coil structure. Upon treatment with methanol vapor (ptm), the formation of crystalline ß-sheets was triggered. Dichroic Fourier-transform infrared (FTIR) spectroscopy, circular dichroism, spectroscopic ellipsometry, atomic force microscopy, grazing-incidence small-angle X-ray scattering (GISAXS), grazing-incidence wide-angle X-ray scattering (GIWAXS), and electrokinetic and contact angle measurements were used to get information concerning the secondary structure and folding kinetics, orientation of ß-sheets, the ratio of parallel/antiparallel ß-sheets, domain sizes and distributions, surface topography, surface potential, hydrophobicity and the film integrity under water. Significant differences in the final ß-sheet content, the share of antiparallel ß-sheet structures, film integrity, surface potential, and isoelectric points between eADF4(Cx) with x = 1, 2 and eADF4(Cx) with x = 4, 8, 16 gave new insights in the molecular weight-dependent structure formation and film properties of IDP systems. GISAXS and kinetic measurements confirmed a relation between ß-sheet crystal growth rate and final ß-sheet crystal size. Further, competing effects of reduced diffusibility hindering accelerated crystal growth and enhanced backfolding promoting accelerated crystal growth with increasing molecular weight were discussed.

Polarity and functionality tailored conjugated microporous polymer coatings on silica microspheres for enhanced pollutant adsorption.

Many sources of pollution that are generated by modern society are not addressable by conventional methods. Especially organic compounds, like pharmaceutics, are particularly hard to remove from waterbodies. Herein, a new approach is presented using conjugated microporous polymers (CMPs) to coat silica microparticles yielding specifically tailored adsorbents. The CMPs are generated with three different monomers: 2,6-dibromonaphthalene (DBN), 2,5-dibromoaniline (DBA) and 2,5-dibromopyridine (DBPN) respectively coupled to 1,3,5-triethynylbenzene (TEB) via Sonogashira coupling. By optimizing the polarity of the silica surface, all three CMPs were converted into microparticle coatings. The resulting hybrid materials feature the advantages of being adjustable in polarity and functionality, as well as morphology. Sedimentation allows facile removal of the coated microparticles after the adsorption. Further, the expansion of the CMP to a thin coating increases the accessible surface area compared to the bulk material. These effects were demonstrated by the adsorption of the model drug diclofenac. Thereby, the aniline-based CMP proved to be most advantageous due to a secondary crosslinking mechanism of amino and alkyne functionalities. An outstanding adsorption capacity of 228 mg diclofenac per gram of the aniline CMP within the hybrid material was achieved. This represents a five-fold increase compared to the value obtained by the pure CMP material underlining the advantages of the hybrid material.

SiO2 nanospheres as surfactant and template in aqueous dispersion polymerizations yielding highly nanoporous resin particles.

HYPOTHESIS: High nitrogen containing resins such as poly(melamine-co-formaldehyde) (PMF) are known for their very good adsorption properties. Until now, using an ecofriendly hard-templating approach with SiO2 nanospheres in water for synthesis, only yielded either highly porous particles with diameters up to 1 µm or non-porous particles with diameters above 1 µm. Small particles cannot be used as fixed bed adsorbents in columns because of the very high pressure occurring. EXPERIMENTS: To yield particles with high porosity and larger diameters for the use as fixed bed adsorbent, we investigated the influence of several synthesis parameters on porosity and particle morphology. FINDINGS: From all variations, we proposed a mechanism for the complex interplay between the PMF prepolymer and resin species with SiO2 nanoparticles acting both as Pickering-like surfactant and template particle. With this knowledge we were able to produce a suitable column material with high specific surface area up to 260 m2/g. We then proved the application of this material for aqueous dichromate adsorption in batch, yielding a maximum capacity of 138 mg/g with recyclability. In column experiments, the contamination of 5 mg/L dichromate in water was reduced to drinking water safe levels for an influent volume equal to over 160 bed volumes.

Tuning the pore structure of templated mesoporous poly(melamine-co-formaldehyde) particles toward diclofenac removal.

The increasing demand and implementation of pharmaceutics poses severe risk to different aquatic species as detectable contaminant in almost every surface water worldwide. Diclofenac (DCF) as one of the most common used analgesics was investigated as contaminant to be removed by adsorption onto nanoporous poly(melamine-co-formaldehyde) (PMF) particles featuring a very high amount of nitrogen functionalities. To achieve a high specific surface area (up to 416 m2/g) and a tunable pore system by hard templating, four different SiO2 nanoparticles were used as template. Differences in the pore formation and achieved pore structure were elucidated. For the first time, the adsorption of DCF onto PMF was tested. In batch adsorption experiments, impactful adsorption capacities as high as 76 µmol/g were achieved and complete removal at initial concentrations of 2 mg/L DCF. Differences in the connectivity and the micropore structure were decisive for uptake in low concentrations and the achieved adsorption capacity, respectively. As the presented PMF particles can be easily synthesized with the monomers formaldehyde and melamine combined with colloidal silica as sacrificial template and water as green solvent, this material presents a viable adsorbent for the removal of DCF at a larger scale. Our study further indicates a high potential for the removal of other pharmaceuticals.

Probing Crowdedness of Artificial Organelles by Clustering Polymersomes for Spatially Controlled and pH-Triggered Enzymatic Reactions.

Most sophisticated biological functions and features of cells are based on self-organization, and the coordination and connection between their cell organelles determines their key functions. Therefore, spatially ordered and controllable self-assembly of polymersomes to construct clusters to simulate complex intracellular biological functions has attracted widespread attention. Here, we present a simple one-step copper-free click strategy to cross-link nanoscale pH-responsive and photo-cross-linked polymersomes (less than 100 nm) to micron-level clusters (more than 90% in 0.5-2 µm range). Various influencing factors in the clustering process and subsequent purification methods were studied to obtain optimal clustered polymeric vesicles. Even when polymeric vesicles separately loaded with different enzymes (glucose oxidase and myoglobin) are coclustered, the overall permeability of the clusters can still be regulated through tuning the pH values on demand. Compared with simple blending of those enzyme-loaded polymersomes, the rate of enzymatic cascade reaction increased significantly due to the interconnected complex microstructure established. The connection of catalytic nanocompartments into clusters confining different enzymes of a cascade reaction provides an excellent platform for the development of artificial systems mimicking natural organelles or cells.

Laser-Assisted Direct Grafting of Poly(ethyleneimine) on Poly(methyl methacrylate).

Demand for direct chemical modification of functional material on a surface is increasing in various fields. A new approach for a functionalized surface is investigated by applying a conventional laser in order to generate chemical activation by photothermal energy. Poly(ethyleneimine) (PEI), with a high density of amino groups, is chemically grafted on poly(methyl methacrylate) (PMMA) by irradiation of a CO2 laser (10.6 µm). Laser parameters such as power, scan rate, and focal length are observed to play an important role in order to introduce effective photothermal energy for the chemical reaction between PEI and PMMA. By optimization of laser parameters, the amide compound is produced as a result of the reaction of amine from PEI and the ester of PMMA successfully. The PMMA surface modified with PEI is analyzed by XPS and TOF-SIMS to identify the functional groups. Furthermore, the surface is characterized in terms of wettability, adhesion force, and surface charge for various applications. Finally, reaction with dye and metal on the amine-terminated PMMA shows promising results in supplying a selective and reliable functional substrate.

Mesoporous Poly(melamine-co-formaldehyde) Particles for Efficient and Selective Phosphate and Sulfate Removal.

Due to the existence-threatening risk to aquatic life and entire ecosystems, the removal of oxyanions such as sulfate and phosphate from anthropogenic wastewaters, such as municipal effluents and acid mine drainage, is inevitable. Furthermore, phosphorus is an indispensable resource for worldwide plant fertilization, which cannot be replaced by any other substance. This raises phosphate to one of the most important mineral resources worldwide. Thus, efficient recovery of phosphate is essential for ecosystems and the economy. To face the harsh acidic conditions, such as for acid mine drainage, an adsorber material with a high chemical resistivity is beneficial. Poly(melamine-co-formaldehyde) (PMF) sustains these conditions whilst its very high amount of nitrogen functionalities (up to 53.7 wt.%) act as efficient adsorption sides. To increase adsorption capacities, PMF was synthesized in the form of mesoporous particles using a hard-templating approach yielding specific surface areas up to 409 m2/g. Different amounts of silica nanospheres were utilized as template and evaluated for the adsorption of sulfate and phosphate ions. The adsorption isotherms were validated by the Langmuir model. Due to their properties, the PMF particles possessed outperforming maximum adsorption capacities of 341 and 251 mg/g for phosphate and sulfate, respectively. Furthermore, selective adsorption of sulfate from mixed solutions of phosphate and sulfate was found for silica/PMF hybrid particles.

A Complementary and Revised View on the N-Acylation of Chitosan with Hexanoyl Chloride.

The modification of the biobased polymer chitosan is a broad and widely studied field. Herein, an insight into the hydrophobization of low-molecular-weight chitosan by substitution of amino functionalities with hexanoyl chloride is reported. Thereby, the influence of the pH of the reaction media was investigated. Further, methods for the determination of the degree of substitution based on 1H-NMR, FTIR, and potentiometric titration were compared and discussed regarding their accuracy and precision. 1H-NMR was the most accurate method, while FTIR and the potentiometric titration, though precise and reproducible, underlie the influence of complete protonation and solubility issues. Additionally, the impact of the pH variation during the synthesis on the properties of the samples was investigated by Cd2+ sorption experiments. The adjusted pH values during the synthesis and, therefore, the obtained degrees of substitution possessed a strong impact on the adsorption properties of the final material.

A Comparative Study on the Flocculation of Silica and China Clay with Chitosan and Synthetic Polyelectrolytes.

Flocculation is still one of the most important and efficient processes for water treatment. However, most industrial processes, such as in water treatment plants, still use huge amounts of synthetic polyelectrolytes for the flocculation process. Here we compare the flocculation of two different suspended particles, i.e., silica particles and china clay, with the biopolymer chitosan and two common strong synthetic polyelectrolytes. As a flocculant, chitosan featured a minimum uptake rate of 0.05 mg/g for silica and 1.8 mg/g for china clay. Polydiallyldimethylammonium chloride (PDADMAC) for comparison possessed a minimum uptake rate of 0.05 mg/g for silica and 2.2 mg/g for china clay. Chitosan as an environmentally friendly biopolymer competes with the synthetic polyelectrolytes and thus represents a beneficial economic alternative to synthetic flocculants.

Characterization of chitosan with different degree of deacetylation and equal viscosity in dissolved and solid state - Insights by various complimentary methods.

In recent years, chitosan has attracted considerable interest in many fields due to its sufficient charge density under biological, non-hazardous conditions. Since chitosan originates from natural resources and has two different monomer units, its characterization must be carried out in a goal-oriented and precise manner. This work focuses on the characterization of chitosans most important parameters - solubility, crystallinity, degree of deacetylation (DD) and molecular weight - in a simple and convenient way. The DD was determined using Nuclear Magnetic Resonance spectroscopy (NMR), Particle Charge Detection (PCD), Fourier Transform Infrared spectroscopy (FTIR), CHN elemental analysis (CHN-EA) and conductometric/potentiometric titration with special attention to its physical state as solid or liquid. Investigation of DD by FTIR was successfully determined by calculating peak heights, peak areas and peak deconvolution from a linear combination of Gaussian and Lorentzian functions. Asymmetrical flow field flow fractionation with light scattering detection (AF4-LS) was applied in order to calculate molar masses and radii. In addition, pH-potentiometric titrations demonstrated a reproducible displacement of the point of zero charge (PZC) in form of a hysteresis depending on the titration direction. The DD affects the crystallinity, which was determined by deconvolution of the crystalline and amorphous domains.

Removal and Separation of Heavy Metal Ions from Multicomponent Simulated Waters Using Silica/Polyethyleneimine Composite Microparticles.

Composite solid surfaces with high content of functional groups (FGs) are useful materials in different types of applications requiring stimuli-responsive "hard/soft" architectures, their improved properties rising from the combination of organic-inorganic parts. Among different types of weak polyelectrolytes, poly(ethyleneimine) (PEI) is of great interest in the construction of composite systems with thin layer-by-layer (LbL) organic films due to the large number of amino groups per unit mass of polymer. Herein, the spherical silica microparticles were modified with linear (L) or branched (B) PEI chains using LbL deposition of a copper complex (PEIL-Cu2+ or PEIB-Cu2+) and poly(acrylic acid) (PAA), glutaraldehyde selective cross-linking, followed by copper and PAA extraction from the multilayer. The newly formed silica/(PEIL)10 and silica/(PEIB)10 composites were used in batch and column sorption/desorption experiments of four heavy metal ions (Cu2+, Ni2+, Co2+, and Cd2+). In noncompetitive conditions ([FG]/Σ[M2+] > 9), all heavy metal ions were retained on composites, demonstrating the potential application of the prepared functional microparticles in surface water treatment. However, in competitive conditions ([FG]/Σ[M2+] < 9), only Cu2+ is sorbed in high amount (∼2.5 mmol·g-1 PEI) on composites, with simultaneous displacement of already sorbed ions, demonstrating the solid-phase extraction and chromatographic properties of the synthesized silica/(PEIL)n and silica/(PEIB)n composites.

Solubility and Selectivity Effects of the Anion on the Adsorption of Different Heavy Metal Ions onto Chitosan.

The biopolymer chitosan is a very efficient adsorber material for the removal of heavy metal ions from aqueous solutions. Due to the solubility properties of chitosan it can be used as both a liquid adsorber and a solid flocculant for water treatment reaching outstanding adsorption capacities for a number of heavy metal ions. However, the type of anion corresponding to the investigated heavy metal ions has a strong influence on the adsorption capacity and sorption mechanism on chitosan. In this work, the adsorption capacity of the heavy metal ions manganese, iron, cobalt, nickel, copper, and zinc were investigated in dependence on their corresponding anions sulfate, chloride, and nitrate by batch experiments. The selectivity of the different heavy metal ions was analyzed by column experiments.

Hollow Au@TiO2 porous electrospun nanofibers for catalytic applications.

Catalytically active porous and hollow titania nanofibers encapsulating gold nanoparticles were fabricated using a combination of sol-gel chemistry and coaxial electrospinning technique. We report the fabrication of catalytically active porous and hollow titania nanofibers encapsulating gold nanoparticles (AuNPs) using a combination of sol-gel chemistry and coaxial electrospinning technique. The coaxial electrospinning involved the use of a mixture of poly(vinyl pyrrolidone) (PVP) and titania sol as the shell forming component, whereas a mixture of poly(4-vinyl pyridine) (P4VP) and pre-synthesized AuNPs constituted the core forming component. The core-shell nanofibers were calcined stepwise up to 600 °C which resulted in decomposition and removal of the organic constituents of the nanofibers. This led to the formation of porous and hollow titania nanofibers, where the catalytic AuNPs were embedded in the inner wall of the titania shell. The catalytic activity of the prepared Au@TiO2 porous nanofibers was investigated using a model reaction of catalytic reduction of 4-nitrophenol and Congo red dye in the presence of NaBH4. The Au@TiO2 porous and hollow nanofibers exhibited excellent catalytic activity and recyclability, and the morphology of the nanofibers remained intact after repeated usage. The presented approach could be a promising route for immobilizing various nanosized catalysts in hollow titania supports for the design of stable catalytic systems where the added photocatalytic activity of titania could further be of significance.

Preparation of water soluble hydrochloric chitosan from low molecular weight chitosan in the solid state.

In this study, low molecular weight chitosan salt (LMWC-HCl) highly soluble in water was prepared from low molecular weight chitosan (LMWC) in the solid state exposed to hydrogen chloride gas as a reagent. The effects of chitosan particle size, exposure conditions, reaction temperature and reaction time were investigated on the solubility and the molecular weight of obtained products. The formation of the chloride salt was observed after 3 h in a range of temperatures from 4 to 50 °C. The solubility of prepared LMWC-HCl was over 98% for all samples, much higher than that of the original LMWC. The average molecular weight of the LMWC-HCl was about 20-90 kDa with a quite narrow distribution and lower compared to the LMWC. LMWC-HCl and LMWC showed the same high antibacterial activity against Bacillus cereus and Vibrio parahaemolyticus. This facile and efficient process for solubilization of LMWC has potential for industrial application of chitosan.

Biorelated Polyelectrolyte Coatings Studied by in Situ Attenuated Total Reflection-Fourier Transform Infrared Spectroscopy: Deposition Concepts, Wet Adhesiveness, and Biomedical Applications.

In this conceptual contribution, thin functional coatings consisting of either pure polyelectrolytes (PELs) or complexes between oppositely charged PELs at model and applied substrates are outlined. Latter PEL/PEL complexes were deposited by two concepts. In a first well-known concept, PEL multilayers (PEM) were consecutively deposited according to the layer-by-layer (LbL) technique. In a second less known concept, PEL complex (PEC) nanoparticles (NPs) preformed by mixing polycation (PC) and polyanion (PA) solutions were deposited in one step. Both concepts based on binary oppositely charged PELs are compared to one based on a single polycation system. Examples shall be given on adhesiveness, nanostructure, and biomedical applications of PEM and PEC NP coatings. In situ attenuated total reflection (ATR) infrared (IR) spectroscopy, circular dichroism (CD), and scanning force microscopy (SFM) were used for molecular, optical, and microscopic characterization. At first, results on the adsorbed amount and wet adhesiveness of pure (single-component) PEL coatings as a function of charge density are given to motivate coatings of mixed oppositely charged PELs. Second, the wet adhesiveness of PEM and PEC NP coatings of identical PEL compounds in aqueous media varying the molar charge ratio ( n-/ n+) and the deposition step z, respectively, is compared. Upon comparing the three PEL deposition concepts, it is suggested that the lack or absence of excess charge at the PEL/surface interface is one of the main factors for the wet adhesiveness of all pure PEL, PEM, and PEC NP coatings. Finally, the potential of PEM and PEC NP coatings for biomedical applications is outlined. Concerning biopassivation, PEM coatings excessed or terminated by PA repel proteins with low isoelectric points. Concerning bioactivation, PEM coatings loaded with antibiotics as well as PEC NP coatings loaded with therapeutic bisphosphonates showed retarded, optionally temperature responsive drug release for applications in acute surgery and bone healing, and immunoglobulin/PEL complex coatings might open theranostic applications.

The Influence of Salt Anions on Heavy Metal Ion Adsorption on the Example of Nickel.

The biodegradable polysaccharide chitosan possesses protonated and natural amino groups at medium pH values and has therefore been used as an adsorbing material for nickel salts in water treatment. Nickel is a problematic heavy metal ion which can cause various diseases and disorders in living organisms. Here, we show the influence of oxyanions (e.g., nitrate and sulfate) to the adsorption of nickel ions. Hence, simultaneously we are addressing the increasing global problem of nitrate and sulfate ion pollution in groundwater and surface water. A series of adsorption experiments was carried out in order to determine (i) the adsorption equilibrium, (ii) the adsorption capacity in dependence on the initial nickel ion concentration, and (iii) the influence of the anion presented in solution for the adsorption capacity. Surface morphology of chitosan flakes before and after the adsorption process has been studied with SEM-EDX analysis. The chitosan flakes exhibited promising adsorption capacities of 81.9 mg·g-1 and 21.2 mg·g-1 for nickel (sulfate) and nickel (nitrate), respectively. The calculated values of Gibbs free energy change ΔG° confirm the higher adsorption of nickel ions in presence of sulfate ions. Hence, higher anion valence leads to a higher adsorption capacity.

Complex calcium carbonate/polymer microparticles as carriers for aminoglycoside antibiotics.

Composite microparticles of CaCO3 and two pectin samples (which differ by the functional group ratio) or corresponding nonstoichiometric polyelectrolyte complexes with different molar ratios (0.5, 0.9 and 1.2) are obtained, characterized and tested for loading and release of streptomycin and kanamycin sulphate. The synthesized carriers were characterized before and after drug loading in terms of morphology (by SEM using secondary electron and energy selective backscattered electron detectors), porosity (by water sorption isotherms) and elemental composition (by elemental mapping using energy dispersive X-ray and FTIR spectroscopy). The kinetics of the release mechanism from the microparticles was investigated using Higuchi and Korsmeyer-Peppas mathematical models.

Surface Functionalization by Stimuli-Sensitive Microgels for Effective Enzyme Uptake and Rational Design of Biosensor Setups.

We highlight microgel/enzyme thin films that were deposited onto solid interfaces via two sequential steps, the adsorption of temperature- and pH-sensitive microgels, followed by their complexation with the enzyme choline oxidase, ChO. Two kinds of functional (ionic) microgels were compared in this work in regard to their adsorptive behavior and interaction with ChO, that is, poly(N-isopropylacrylamide-co-N-(3-aminopropyl)methacrylamide), P(NIPAM-co-APMA), bearing primary amino groups, and poly(N-isopropylacrylamide-co-N-[3-(dimethylamino) propyl]methacrylamide), P(NIPAM-co-DMAPMA), bearing tertiary amino groups. The stimuli-sensitive properties of the microgels in the solution were characterized by potentiometric titration, dynamic light scattering (DLS), and laser microelectrophoresis. The peculiarities of the adsorptive behavior of both the microgels and the specific character of their interaction with ChO were revealed by a combination of surface characterization techniques. The surface charge was characterized by electrokinetic analysis (EKA) for the initial graphite surface and the same one after the subsequent deposition of the microgels and the enzyme under different adsorption regimes. The masses of wet microgel and microgel/enzyme films were determined by quartz crystal microbalance with dissipation monitoring (QCM-D) upon the subsequent deposition of the components under the same adsorption conditions, on a surface of gold-coated quartz crystals. Finally, the enzymatic responses of the microgel/enzyme films deposited on graphite electrodes to choline were tested amperometrically. The presence of functional primary amino groups in the P(NIPAM-co-APMA) microgel enables a covalent enzyme-to-microgel coupling via glutar aldehyde cross-linking, thereby resulting in a considerable improvement of the biosensor operational stability.

Autotemplate Microcapsules of CaCO3/Pectin and Nonstoichiometric Complexes as Sustained Tetracycline Hydrochloride Delivery Carriers.

New types of composites were obtained by an autotemplate method for assembling hollow CaCO3 capsules by using pH-sensitive polymers. Five pectin samples, which differ in the methylation degree and/or amide content, and some nonstoichiometric polyelectrolyte complex dispersions, prepared with the pectin samples and poly(allylamine hydrochloride), were used to control the crystal growth. The morphology of the composites was investigated by scanning electron microscopy, and the polymorphs characteristics were investigated by FTIR spectroscopy. The presence of the polymer in the composite particles was evidenced by X-ray photoelectron spectroscopy, particle charge density, and zeta-potential. The new CaCO3/pectin hollow capsules were tested as a possible matrix for a tetracycline hydrochloride carrier. The kinetics of the drug release mechanism was followed using Higuchi and Korsmeyer-Peppas mathematical models.

The removal of stickies with modified starch and chitosan--highly cationic and hydrophobic types compared with unmodified ones.

The removal of dissolved and colloidal substances (DCS) in paper cycling water, so called stickies, with tailored natural polymers, having cationic as well as hydrophobic groups, was investigated using model suspensions made by the recycling of paper. The sticky content, characterized by the turbidity, the anionic charge, and the total organic carbon content (TOC) was increased by the addition of latex. The dynamic surface tension was established as useful tool for the characterization of the sticky content. The sticky removal using the starch derivatives (with benzyl- as well as ethyl-substituents), from very low up to high cationic charge and N-(benzyl)chitosan was compared. Depending on the properties of the derivatives two possible mechanisms can be found: "charge dominated removal" or "removal dominated by hydrophobicity." It seems that turbidity and TOC are lowered especially due to charge interaction whereas the increase of the surface tension is mainly caused by the hydrophobic character of the modified natural polymers.