Low Degree Of Modification Research Articles

Photopolymerization of L-DOPA-Alginate Immobilized Cell Wall Laccase for Textile Dye Decolorization

Alginate is a naturally occurring polymer derived from brown algae biomass, which has numerous applications in various fields. Chemical modification of alginate is widely used to improve alginate’s physicochemical properties and provide new potential for multiple applications. In this article, we modified alginate with L-DOPA, using periodate oxidation and reductive amination, to obtain more suitable biopolymer for biocatalyst immobilization and hydrogel formation. Obtained modified alginate was used for the immobilization of laccase on cell walls. For this purpose, laccase from Streptomyces cyaneus was expressed on the surface of Saccharomyces cerevisiae EBY100 cells. The obtained cell wall laccase was immobilized within L-DOPA-alginate beads by crosslinking the L-DOPA-alginate with calcium ions and laccase. The effect of additional crosslinking of beads by green light-induced photopolymerization with eosin Y was investigated. The immobilized laccase systems were used for dye decolorization and investigated in multiple treatment processes. Beads with L-DOPA-alginate with a higher degree of modification (5.0 mol%) showed higher enzymatic activity and better decolorization efficiency than those with a lower degree of modification (2.5 mol%). Obtained immobilized biocatalysts are suitable for decolorizing dye Evans Blue due to their high efficiency and reusability.

Metal-organic framework (UiO-66 and UiO-66-NH2)-based adsorbents for bilirubin removal used in hemoperfusion.

Excessive accumulation of bilirubin in patients with hyperbilirubinemia can lead to tissue and organ damage and neurological diseases, and is even life-threatening in severe cases. Hemoperfusion is an effective method for removing bilirubin, but clinically used hemoperfusion adsorbents have unsatisfactory adsorption capacity and kinetics. In order to obtain a safe and efficient bilirubin adsorbent, Zr-based Metal-Organic Framework (MOF) material UiO-66 with high specific surface area and aqueous medium stability was prepared and modified with varying degrees of amination to improve its adsorption capacity. According to adsorption experiments in aqueous solution and simulated plasma, it was confirmed that the unsaturated coordinated zirconium in UiO-66 can effectively induce the aggregation and precipitation of free bilirubin unbound to albumin and the amino group on UiO-66-NH2 has a strong affinity for albumin bound bilirubin. The adsorption effect of UiO-66-NH2 with a high degree of amino modification is significantly stronger than that of UiO-66-NH2 with a low degree of modification. In simulated plasma with a bilirubin concentration of 40 mg dL-1, the adsorption capacities of UiO-66 and UiO-66-NH2-1.9 can reach 69.08 mg g-1 and 81.13 mg g-1. The adsorption isotherm fitting and adsorption kinetics fitting results also show that UiO-66 and UiO-66-NH2 are good adsorbents for bilirubin. In dynamic adsorption, the adsorbents also showed good performance and did not affect the protein in the plasma. The hemolysis test, coagulation time test, and cytotoxicity test confirmed that the bilirubin adsorbents based on UiO-66 and UiO-66-NH2 have good blood compatibility and biocompatibility. This study provides new ideas for the development of a novel bilirubin adsorbent and a theoretical basis for the study of bilirubin adsorption mechanisms.

Tunable Cross-Linking and Adhesion of Gelatin Hydrogels via Bioorthogonal Click Chemistry.

Engineering cytocompatible hydrogels with tunable physico-mechanical properties as a biomimetic three-dimensional extracellular matrix (ECM) is fundamental to guide cell response and target tissue regeneration or development of in vitro models. Gelatin represents an optimal choice given its ECM biomimetic properties; however, gelatin cross-linking is required to ensure structural stability at physiological temperature (i.e., T > Tsol-gel gelatin). Here, we use a previously developed cross-linking reaction between tetrazine (Tz)- and norbornene (Nb) modified gelatin derivatives to prepare gelatin hydrogels and we demonstrate the possible tuning of their properties by varying their degree of modification (DOM) and the Tz/Nb ratio (R). The percentage DOM of the gelatin derivatives was tuned between 5 and 15%. Hydrogels prepared with higher DOM cross-linked faster (i.e., 10-20 min) compared to hydrogels prepared with lower DOM (i.e., 60-70 min). A higher DOM and equimolar Tz/Nb ratio R resulted in hydrogels with lower weight variation after immersion in PBS at 37 °C. The mechanical properties of the hydrogels were tuned by varying DOM and R by 1 order of magnitude, achieving elastic modulus E values ranging from 0.5 (low DOM and nonequimolar Tz/Nb ratio) to 5 kPa (high DOM and equimolar Tz/Nb ratio). Human dental pulp stem cells were embedded in the hydrogels and successfully 3D cultured in the hydrogels (percentage viable cells >85%). An increase in metabolic activity and a more elongated cell morphology was detected for cells cultured in hydrogels with lower mechanical properties (E < 1 kPa). Hydrogels prepared with an excess of Tz or Nb were successfully adhered and remained in contact during in vitro cultures, highlighting the potential use of these hydrogels as compartmentalized coculture systems. The successful tuning of the gelatin hydrogel properties here developed by controlling their bioorthogonal cross-linking is promising for tissue engineering and in vitro modeling applications.

MaDHS: Many‐objective discrete harmony search to improve existing package design

AbstractRecently, many computational intelligence algorithms have been proposed to address software remodularization problem. Unfortunately, it has been observed that the performance of optimizers degrades with the optimization problem containing more than three objectives. In this paper, we propose a many‐objective discrete harmony search (MaDHS) to address the software remodularization problem having more than three objectives. The basic idea of MaDHS is that it uses the quality indicator Iϵ + and external archive to rank and store the nondominated solutions. Along with MaDHS, five remodularization objectives, ie, low coupling, high cohesion, low modification degree, quality of class distribution, and low package instability have also been adapted to improve the package structure of existing object‐oriented software systems. To improve the accuracy of modularization solution, the coupling and cohesion objectives are formulated in terms of various dimensions of direct coupling relationships. To test the supremacy of the proposed approach, it is evaluated over eight real‐world object‐oriented software systems. Simulation results show that the proposed approach outperforms the other existing approaches in terms of couplings, cohesion, modularization quality, modularization merit factor, rate per refactoring of achieved improvement, and external developers view.

Hyaluronan hydrogels with a low degree of modification as scaffolds for cartilage engineering

In the field of cartilage engineering, continuing efforts have focused on fabricating scaffolds that favor maintenance of the chondrocytic phenotype and matrix formation, in addition to providing a permeable, hydrated, microporous structure and mechanical support. The potential of hyaluronan-based hydrogels has been well established, but the ideal matrix remains to be developed.This study describes the development of hyaluronan sponges-based scaffolds obtained by lysine methyl-ester crosslinking. The reaction conditions are optimized with minimal chemical modifications to obtain materials that closely resemble elements in physiological cellular environments. Three hydrogels with different amounts of crosslinkers were produced that show morphological, water-uptake, mechanical, and stability properties comparable or superior to those of currently available hyaluronan-scaffolds, but with significantly fewer hyaluronan modifications. Primary human chondrocytes cultured with the most promising hydrogel were viable and maintained lineage identity for 3 weeks. They also secreted cartilage-specific matrix proteins. These scaffolds represent promising candidates for cartilage engineering.

The emulsification properties of octenyl- and dodecenyl- succinylated inulins

The emulsification properties of a series of inulin samples modified using varying amounts of octenyl- and dodecenyl-succinic anhydride have been studied by determining the droplet size as a function of time, temperature and in the presence of electrolyte using laser diffraction. It was found that ∼2% modified inulin was required to stabilise 15% w/v medium chain triglyceride emulsions and that the minimum droplet size was achieved when the % hydrophobe content was >8 mol%. The dodecenyl derivatives produced emulsions with a smaller droplet size than the octenyl derivatives. Apart from OSA-inulin with a low degree of modification, stability was maintained when stored for a period of up to 21 days at room temperature and at 50 °C. The zeta potential of the emulsion droplets was determined by Laser Doppler Velocimetry and was found to increase from ∼5 mv to ∼60 mv as the pH increased from 2 to 7 due to the presence of carboxylate ions present in the linkage between the inulin and alkenyl chains. The droplet size was found to increase on the addition of electrolyte indicating that the adsorbed polymer layer was insufficient to provide a steric repulsive barrier and that stabilisation was mainly due to electrostatic repulsive forces.

Morphologically Tunable Coassembly of Double Hydrophilic Block Polyelectrolyte with Oppositely Charged Fluorosurfactant

We report on the formation and structure development of polyelectrolyte–surfactant complexes, PE–S, of double hydrophilic block copolymers poly(sodium 2-sulfamate-3-carboxylate isoprene)-block-poly(ethylene oxide), PSCI–PEO, and cationic fluorosurfactant, N-(3,3,4,4,5,5,6,6,7,7,8,8,9,9,10,10,10-heptadecafluorodecyl)pyridinium chloride, HFDPCl. We compare the behavior of four PSCI–PEO samples differing in comonomer composition and polyisoprene block modification degree. Coassembled core–shell nanoparticles with the core formed by the PSCI/HFDPCl complex and the shell of PEO blocks were characterized by microscopic techniques (cryogenic transmission electron microscopy, atomic force microscopy) and by small-angle neutron scattering. Interactions between PSCI–PEO and HFDPCl were studied by isothermal titration calorimetry. We show that the bulky fluorosurfactant ions drive the coassembly toward structures with less curved interfaces such as cylindrical and wormlike micelles or vesicles. We also demonstrate the role of hydrophobic interactions in the system induced by the presence of unmodified polyisoprene units which in the case of the PSCI–PEO copolymer with a low degree of modification lead to formation of PSCI–PEO micelles and prevent the copolymer from the coassembly with HFDPCl.

Sorptive ceramic membranes functionalized with HS-groups

Sol–gel method based on the reaction of hydrolytic copolycondensation of tetraethoxysilane (TEOS) or 1,2-bis(triethoxysilyl)ethane (BTESE) with 3-mercaptopropyltrimethoxysilane (MPTMS) was used to functionalize ceramic membranes with Si(CH2)3SH groups. The obtained samples were studied using FTIR spectroscopy, SEM, contact angle measurements, and tested for heavy metals ions retention in the Amicon stirred ultrafiltration cell using model Ag(I) nitrate solutions. It was determined that membrane functionalization is accompanied with the decreasing water flux through the membrane, confirming the deposition of polysiloxane and polysilsesquioxane layers. According to SEM, such layer is about 2.5μm thick and is composed of SH-containing nanoparticles of 60–70nm in diameter. The gaps between nanoparticles determine porosity of the functionalized layer. According to filtration studies, the sample with low degree of modification (DM) possesses high flux (550L/m2h), but its retention of silver(I) ions drops sharply with permeate collection, suggesting sorption mechanism of Ag(I) removal. Whereas, samples with higher DM values are characterized by low flux, but high retention, which increases (up to 70–90%) with permeate collection, suggesting that both sorption and size-exclusion mechanisms of heavy metal removal are involved.

Biocompatible acid-labile polymersomes from PEO-b-PVA derived amphiphilic block copolymers

A family of amphiphilic block copolymers with pendent ortho ester groups were synthesized by modifying the double hydrophilic block copolymer PEO114-b-PVA240 with 2-ethylidene-4-methyl-1,3-dioxolane (EMD) under mild conditions (30 °C). The degree of modification (DM) of the ortho ester groups can be tuned by simply varying the feed ratio of EMD to the hydroxyl groups in the PVA block. These block copolymers are stable in an anhydrous environment. Laser light scattering (LLS) and transmission electron microscopy (TEM) measurements revealed that in weakly basic aqueous buffer, these amphiphilic block copolymers self-assembled into aggregates with different size and morphology, ranging from solid-like spherical nanoparticles to polymersomes as the DM increased. The acid-triggered dissociation behaviour of the aggregates were studied by LLS, nile red (NR) fluorescence and TEM. The copolymer aggregates dissociated faster in a buffer with the lower pH; the dissociation rate of the aggregates became faster for the copolymers with lower DM. The polymersomes can load both hydrophilic biomacromolecules like lysozyme and hydrophobic anticancer drug doxorubicin (DOX). The drug-loaded polymersomes were stable in neutral phosphate buffer for at least 6 h with a payload leakage of less than 25% in 12 h at 37 °C; however, significant acid-triggered payload release was accomplished even at a mildly acidic pH (6.0). Finally, the DOX-loaded polymersomes exhibited a concentration-dependent toxicity to MCF-7 and HeLa cells while the copolymers themselves are non-toxic.

Acetylated and native corn starch blend films produced by blown extrusion

Thermoplastic starch (TPS) materials were developed from mixtures of native and acetylated corn starches with glycerol. To optimize the formulations an experimental design for multicomponent mixtures was used and the assayed formulations were determined by statistical software. Blends and pellets humidity content increased with glycerol concentration. Starch destructuration during the extrusion process was studied by thermal analysis. Films presented homogenous structure, rough surfaces and certain stickiness. They presented different properties, related mainly to the differential characteristics of native and acetylated starches and to hydrophilic character of glycerol. Their mechanical behavior indicated that they are a good option as a food packaging materials since TPS films resulted enough resistant to protect the product and flexible to resist moderate deformations. Besides, the use of acetylated starch in the formulations enhanced film resistance and reduced their WVP, despite of its low modification degree. The storage of the films under controlled conditions increased their stiffness, while their flexibility and WVP were reduced. Plasticizer migration towards the matrix surface was observed in stored films. Films resulted stable till aw=0.7 and due to their selective gaseous permeability they are useful to package products susceptible to oxidation or to control vegetable respiration and senescence.

Gel Properties of Hyaluronic Acid Dermal Fillers

Most of the hyaluronic acid (HA)-based dermal fillers currently on the market are chemically modified with cross-linkers to improve the mechanical properties and duration in vivo. To investigate differences in the properties of dermal fillers that can be related to the respective cross-linking and manufacturing methods used. Thirteen commercially available HA fillers were analyzed. Two different measures of gel strength were used: the elastic modulus (G') determined by rheology and a measure of the swelling capacity of the gel (c(min)). The degree of modification was determined using nuclear magnetic resonance spectroscopy, and the cross-linking ratio was determined using size exclusion chromatography coupled with mass spectrometry. There was a wide variation in gel strength, and the degree of modification varied between 1% and 8% for the HA fillers investigated. Both measures of gel strength, G* and c(min), can be used because the results from the two methods are well correlated. No differentiation in filler properties could be seen as a result of manufacturing process used, except that the nonanimal stabilized HA stabilization process resulted in products with high gel strength and a low degree of modification.

Labeled Nanoparticles Based on Pharmaceutical EUDRAGIT® S 100 Polymers

The pharmaceutically important polymer P(MAA-r-MMA)(1:2) (EUDRAGIT(®) S100) was investigated concerning its behavior to form nanoparticles via nanoprecipitation. The particles obtained were characterized regarding their size, shape, and characteristics using DLS, SEM, and AUC. Furthermore, the P(MAA-r-MMA)(1:2) copolymer was modified with different markers in order to achieve polymer-based nanocarrier systems, which are detectable and may be useful for controlled drug delivery devices to monitor the drug pathways. The particles were labeled by physical entrapment as well as by covalent attachment of various markers, e.g., radicals, fluorescent-, and near-infrared dyes, to the polymer. Physical entrapment of radicals into the polymeric units was performed by co-nanoprecipitation of P(MAA-r-MMA)(1:2) and a radical marker. By means of covalent binding of the markers to the polymer, a stable and more defined labeling of the particles was also performed, leading only to a low degree of modification of the pharmaceutical polymer. After nanoprecipitation, the resulting labeled particles were characterized by SEM and DLS, whereas their biocompatibility was proven by in vitro studies. In order to ensure the possibility of detection of the particles inside the body for drug delivery-, sensor-, and imaging applications, the polymeric carriers were also investigated by electron spin resonance, fluorescence, as well as near-infrared spectroscopy.

Batch purification of high-purity lysozyme from egg white and characterization of the enzyme modified by PEGylation

PEGylation is one of the most promising and extensively studied strategies for improving the pharmacological properties of proteins as well as their physical and thermal stability. Purified lysozyme obtained from hen egg white by batch mode was modified by PEGylation with methoxypolyethyleneglycol succinimidyl succinato (mPEG-SS, MW 5000). The conjugates produced retained full enzyme activity with the substrate glycol chitosan, independent of degree of enzyme modification, although lysozyme activity with the substrate Micrococcus lysodeikticus was altered according to the degree of modification. The conjugate with a low degree of modification by mPEG-SS retained 67% of its enzyme activity with the M. lysodeikticus substrate. The mPEG-SS was also shown to be a highly reactive polymer. The effects of pH and temperature on PEGylated lysozymes indicated that the conjugate was active over a wide pH range and was stable up to 50°C. This conjugate also showed resistance to proteolytic degradation, remained stable in human serum, and displayed greater antimicrobial activity than native lysozyme against Gram-negative bacteria.

Catalytic activity of molecular imprinted vinylpyridine/acrylonitrile/divinylbenzene terpolymers with guanidyl ligands-Cu(II) inside the active centres

Summary — A series of novel molecularly imprinted catalysts was prepared by suspension polymerization of acrylonitrile. N-vinylpyridine and divinylbenzene in the presence of Cu(II) ions and various templates: acetophenone, dibenzoylmethane, 4-methoxybenzyl alcohol or 4-hydroxyacetophenone. To improve the catalytic properties, the catalysts were additionally modified inside the active centres by indirect aminolysis of nitrile groups using ethylenediamine and thiourea/ethyl iodide. The catalytic activity of initial samples as well as the modified ones was tested in the model oxidation of hydroquinone to p-benzoquinone using hydrogen peroxide. The catalysts prepared with Cu(II) ions and 4-methoxybenzyl alcohol as the template exhibit the highest activity. In spite of low modification degree, catalysts with guanidyl ligands inside the active centres show higher reaction yield and selectivity than unmodified samples.

Surface Flocculation as a New Tool for Controlling Adsorption Processes

The dyeing of modified pulp, particularly with regard to the mechanism involved, was studied. The reaction mechanism for the modification of cellulose fibres in aqueous solution by cationic agents was also described. The results obtained were expressed in the form of adsorption isotherms. The capture of molecules, colloidal particles, etc. during the adsorption processes involving modified pulp could be explained by two possible mechanisms: (i) aggregation (coagulation or flocculation) in solution followed by adsorption and (ii) surface aggregation (coagulation or flocculation). Surface aggregation appeared to be the most likely for pulp with a low degree of modification. The typical Langmuir adsorption isotherm correlated the data obtained for unmodified pulp very well.

Synthesis and preliminary characterisation of charged derivatives and hydrogels from scleroglucan

The synthesis of negatively and positively charged polyelectrolytes from scleroglucan is described. Polycarboxylates were synthesised through nucleophilic substitution with chloroacetic acid or through a selective 2,2,6,6-tetramethyl-1-piperidinyloxy (TEMPO)-mediated oxidation of the primary alcohol groups. Amine groups were introduced through nucleophilic substitution with 2-chloroethylamine or 3-chloropropylamine. Reaction conditions were varied to obtain insight into the influence of variables on the degree of substitution. The conformational behaviour of the obtained polyelectrolytes was studied as a function of pH, temperature and solvent. For the products with a low degree of modification, evidence of an ordered conformation was found, whereas the polymers with a higher degree of modification behaved as random coils in solution. The negatively charged polymers were reticulated using the Ugi four-component condensation, obtaining negatively charged hydrogels. The positively charged polymers were reticulated using diethyl squarate (3,4-diethoxy-3-cyclobutene-1,2-dion, DES) to obtain positively charged hydrogels.

Tolerogenic capacity of poly(ethylene glycol) (PEG)-modified ovalbumins in relation to their immunoreactivity towards anti-ovalbumin antibody

Ovalbumin (OVA) was chemically modified with poly(ethylene glycol) (PEG) derivatives: activated PEG2, 2,4-bis[O-methoxypoly(ethylene glycol)]-6-chloro-s-triazine and activated PM13, copolymer of poly(oxyethylene) allyl methyl diether and maleic anhydride. Pretreatment of BALB/c mice with PEG2- or PM13-OVA suppressed the production of both IgG- and IgE-class anti-OVA antibodies induced by subsequent immunizations with unmodified OVA. PEG2-OVA had higher potency to suppress OVA-specific immune response than PM13-OVA. Although extensive modification (62%) of amino groups in the OVA molecule with activated PEG2 was required to diminish most of its immunoreactivity towards anti-OVA antibodies, only a low degree of modification (27%) was sufficient to induce tolerogenicity to OVA. Thus, the loss of immunoreactivity of PEG2-OVA was not prerequisite for its tolerogenic capacity. Moreover, the use of completely denatured PEG2-OVA immunogen lead to the loss of its ability to induce immune tolerance to native OVA. These observations are discussed in relation to the regulatory mechanism of immune response.

Recovery of serum proteins using cellulosic affinity membrane modified by immobilization of Cu2+ ion

An affinity membrane was prepared from a porous cellulose membrane, and adsorption and recovery of serum proteins were investigated from the viewpoint that affinity membranes are efficacious against separation and purification of biomaterials. Into the cellulose membrane, iminodiacetate (IDA) group that acts as a ligand to metal ions was introduced (Cell–IDA membrane), and then Cu2+ ion was immobilized (Cell–IDA–Cu membrane). Bovine serum albumin (BSA) and γ-globulin (BγG), which are the major proteins in blood, were adopted as model proteins to be separated. The Cell–IDA–Cu membrane had large adsorption capacity for these proteins despite the low degree of modification. The amounts of proteins adsorbed on the Cell–IDA–Cu membrane increased with increasing pH, and BγG was adsorbed more than BSA. High protein recoveries from the Cell–IDA–Cu membrane were obtained. The separation of these proteins was also conducted under the optimum conditions of adsorption and recovery, and BγG was concentrated more than BSA although the initial concentration of BγG was lower than that of BSA. © 1996 John Wiley & Sons, Inc.

Fatty acid acylated peroxidase as a model for the study of interactions of hydrophobically-modified proteins with mammalian cells.

Artificial fatty acylation of proteins has attracted significant attention during the last decade as a method for modification of protein specificity and efficacy of action on mammalian cells (A. V. Kabanov and V. Yu. Alakhov (1994) J. Contr. Release 28, 15-35). Horse radish peroxidase (HRP) is used in this work to study the interaction of a fatty acylated protein with various mammalian cells. The HRP is modified with the chloranhydride of the stearic acid in the reversed micelles of sodium bis-(2-ethylhexyl)sulfosuccinate (Aerosol OT) in octane, a convenient protocol allowing production of protein molecules with a controlled, low modification degree (A.V. Kabanov et al. (1987) Ann. N. Y. Acad. Sci. 501, 63-66). The influence of the hydrophobic group on the binding and internalization of HRP in MDCK, P3-X63-Ag8, CHO, and HepG2 cells is examined. The major results are as follows: (i) the fatty acylation of a protein significantly enhances its binding to all tested mammalian cell lines, with a line-specific efficiency; (ii) the binding efficiency can be modified by changing growth conditions in a defined medium; (iii) along with the enhancement of protein adsorption on the plasma membrane, fatty acylation increases internalization of the protein during incubations at 37 degrees C; (iv) internalized protein was observed in endocytic vesicles; no evidence was obtained for a cytoplasmic distribution. These results are discussed in connection with previously observed effects of the fatty acylated proteins on cell activity.

Biodegradation of soluble redox polymers. 1. (0.017ferrocene)amylose.

The reaction of amylose with NaH and FcCH2NMe3+I- in dimethyl sulfoxide brought about a redox-labeled polymer with a low degree of modification, viz. 1 ferrocene residue per 60 glucose units. The preparation, (0.017ferrocene)amylose, displays one-electron irreversible behavior at a pyrographite electrode in terms of the Delahay formalism, the formal redox potential E degree' being equal to 0.38 V at pH 6 and 40 degrees C versus SCE. Specific to amylose enzymes endo-depolymerases, which carry out random hydrolysis, accept the labeled amylose providing a significant increase in the peak current on cyclic voltammograms. The absence of potential drifts suggests that the effect is due an increase in the diffusion coefficients of the amylose fragments in the course of enzymatic digestion of (0.017ferrocene)amylose. This proposal was confirmed by the simulation of experimental cyclic voltammograms. Several practical applications of the results of this study for electrochemical assaying the amylolytic activity and evaluation of the mechanisms of the enzymatic catalysis by amylases have been demonstrated.