Immobilization of penicillin acylase on copolymer of butyl acrylate and ethylene glycol dimethacrylate

The effects of glutaraldehyde, enzyme concentrations and reactants volumes, ionic strength, pH value and carrier particle diameter on immobilization of penicillin acylase onto acrylic carriers were studied. The activity of immobilized enzyme preparations was also studied over a range of pH values and temperatures and thermal and pH stabilities were determined. The use of the immobilized preparation for penicillin G hydrolysis in a batch reactor was investigated. The immobilized enzyme gave a significant reduction in hydrolysis time compared to hydrolysis by the native enzyme.     

Behavior of immobilized glucose oxidase on membranes from polyacrylonitrile and copolymer of methylmethacrylate‐dichlorophenylmaleimide

Two new ultrafiltration membranes were obtained from a polymer mixture, containing 60% polyacrylonitrile (PAN) and 40% copolymer of methylmethacrylate‐dichlorophenylmaleimide (MMA‐DCPMI). Membrane 1 (MB1) contains 40% DCPMI of the copolymer, and membrane 2 (MB2) contains 15% of the copolymer. The pore size, the specific surface, the water content, the water flux, and the selectivity were determined for the two membranes. The presence of dichlorophenylmaleimide in the copolymer ensures the preparation of membranes suitable for direct covalent enzyme immobilization without further modifications. These membranes were used for immobilization of glucose oxidase (GOD). High amount of bound protein was found on each of the membranes. High relative activities of the immobilized GOD were achieved, 72% for MB1 and 68% for MB2. The properties of the immobilized enzyme (GOD) were determined: optimum pH and temperature and pH, thermal, and storage stability, and then compared with the properties of the native enzyme. The kinetic parameters of the enzyme reaction, Michaelis constant (K_m) and maximum reaction rate (V_max), were also investigated. The results obtained showed that the ultrafiltration membranes prepared from the mixture of PAN and the copolymer MMA‐DCPMI were suitable for use as carriers for the immobilization of GOD. © 2006 Wiley Periodicals, Inc. J Appl Polym Sci 101: 4334–4340, 2006     

功能化磁性纳米颗粒固定化脂肪酶研究

用葡萄糖酸对Fe3O4磁性纳米颗粒表面进行修饰,然后用水溶性碳化二亚胺（EDC）作偶联剂,对脂肪酶进行固定化。考察了偶联剂浓度、给酶量和反应时间对脂肪酶固定化过程的影响。结果表明,制备功能化磁性颗粒固定化酶的最佳条件为：偶联剂浓度为12．5mg／mL磷酸缓冲液（PBS）,给酶量为2．5mg／mLPBS,反应时间为24h。固定化脂肪酶表现出优异的热稳定性,60℃时酶活为游离酶的6倍。重复使用10次后,酶促活力依然保持80％以上。     

Development of superparamagnetic functional carriers and application for affinity separation of subtilisin Carlsberg

Superparamagnetic functional carriers to obtain high selectivity were reported in this study. Magnetic carriers with epoxy groups were synthesized by spraying suspension polymerization (SSP). The measurement of scanning electron microscopy (SEM) and vibrating sample magnetometer (VSM) showed that the magnetic carriers have a narrow size distribution and displayed superparamagnetic characteristics. The magnetic carriers with epoxy groups were modified by various affinity ligands to magnetic functional carriers such as copper-IDA carriers, benzamidine carriers and phenylboronic acid carriers. The effects of medium pH and subtilisin Carlsberg concentration on adsorption capacity were investigated. It was found that phenylboronic acid and benzamidine immobilized magnetic carriers were effective magnetic carriers for affinity adsorption of subtilisin Carlsberg, and the maximum adsorption capacity (about 65 mg/g) was obtained at pH 9.5. The adsorbed subtilisin Carlsberg was also desorbed successfully by using dissociation agents, and the recovery of the enzyme activity was still around 85%.     

融合蛋白CR2-GDH固定化及不对称还原制备(S)-4-氯-3-羟基丁酸乙酯

比较介孔分子筛材料SBA-15、MCM-41、海藻酸钙、改性二氧化硅4种载体固定化融合蛋白CR2-GDH其酶固载量和酶活回收率,选择SBA-15为固定化载体。研究固定化条件对固定化融合酶量的影响以及固定化酶的稳定性,固定化酶在双相体系催化不对称还原反应。结果表明,在pH值为5.5、酶浓度为1.4mg/mL、反应1h条件下,固定化酶量为27.7mg/g。加入25mmol/L的Ca²⁺,固定化酶的酶活回收率由58.6%提高到78.1%。与游离酶相比,固定化酶的热稳定性显著提高,40℃条件下酶活回收率提高19.1%。固定化酶水相中反复使用7批次后,剩余活性仍超过30%,具有较好的操作稳定性。与游离酶相比,固定化酶更耐受烷烃类有机溶剂。在水/有机溶剂双相反应体系中,Ca²⁺/SBA-15固定化酶和游离酶催化相比,产物得率提高23.8%。     

An Synthesepulp immobilisierte Enzyme,I.

Synthetic pulp—a polyethylene fibrid which contains poly(vinyl alcohol)—was reacted with 2-(3-aminophenyl)-1,3-dioxolane. The amino derivative was transferred into the reactive diazonium compound. The content of reactive diazonium groups was determined by reaction with tyrosine. The diazonium derivatives of synthetic pulp were used for the immobilization of the hydrolases trypsin, chymotrypsin, papain, aminoacrylase, esterase and urease of the oxidoreductases glucose oxidase, catalase, peroxidase, and glucose-6-phosphate-dehydrogenase, and of the transferase hexokinase. Furthermore soybean trypsin inhibitor was immobilized on synthetic pulp via azo coupling. The protein binding ability of the reactive carriers and the enzymatic properties of the immobilization products were investigated. The thermostability of immobilized trypsin and immobilized urease, as well as the dependence of the immobilized tryptic activity on the temperature were studied. The activities of immobilized trypsin were assayed with low molecular weight substrates as well as with high molecular weight substates. The pH-optima of immobilized chymotrypsin, papain and urease were studied and the influence of buffers on the pH-activity profiles was investigated. Hexokinase and glucose-6-phosphatedehydrogenase were co-immobilized on synthetic pulp as an example for a two enzyme system and the properties of the immobilization products were investigate.     

Liposomes in wool dyeing — the stability of dye-liposome systems and their application to untreated wool fibres

The use of two different types of liposome suspensions (multilamellar vesicles, MLV, and large unilamellar vesicles, LUV) as carriers in the commercial dyeing of untreated wool with a milling acid dye is described. Liposomes prepared with egg phosphatidylcholine and containing the dye CI Acid Blue 90 were used. The physico-chemical stability of liposomes was studied by measuring the mean particle size distribution of phospholipidic vesicles during dyeing. The possible hydrolysis of phospholipid molecules was also determined. Kinetic aspects involving dye adsorption and bonding were investigated. Dye exhaustion on untreated wool fibres was inhibited and dye bonding was improved. The lipid concentration and type of liposomes were important factors in this process.     

用于酶固定化的高分子载体材料研究进展

固定化酶是一种高效、高选择性和反应条件温和的生物催化剂。近年来,高分子材料作为酶固定化载体的研究越来越受到重视,相关的研究报道很多。对近十多年来用于固定化酶的高分子载体材料以及它们的优缺点进行了综述,并对用于酶固定化高分子载体材料的发展前景作了展望。     

Ion mobility time-of-flight measurements: isolating the mobility of charge carriers during an epoxy–amine reaction

Ion mobility time-of-flight (ITOF) measurements can isolate the mobility of the charge carriers during a chemical reaction. The ITOF technique complements conductivity measurements. Together they more accurately monitor changes in the chemical and physical properties such as viscosity of a material as it polymerizes. The use of conductivity due to migration of charges to monitor the advancement of reaction has been widely reported. It is commonly assumed that a correlation should exist between the decrease in conductivity, and the increasing viscosity and/or extent of reaction. This is based on the assumption that the number of charge carriers in the polymer-forming system is constant. Unfortunately, this assumption is not correct in an epoxy/amine system where the number of OH groups and NH groups changes as the reaction advances.The epoxy–amine reaction (diglycidyl ether of bisphenol-A (DGEBA)/amine 4,4′-methylene bis[3-chloro 2,6-diethylaniline] (MCDEA)) was investigated by ITOF and dielectric measurement. Together these two techniques were used to separately monitor the changes in the ion mobility and the number of charge carriers. Parameters, such as voltage, pulse length and sensor geometry, were determined and optimized to obtain the repeatable results. The results were compared to dynamic mechanical measurements and the reaction advancement. A most interesting finding is the significant changes in the number of charge carriers during the reaction at the three different temperatures measured. An explanation of the observed changes in ion mobility and the number of charge carriers is offered in terms of the inter-relationship of the changing chemical structure and the density of an epoxy during cure.     

Stabilization of entrapped catalase using photo‐crosslinked resin gel for use in wastewater containing hydrogen peroxide

Catalase immobilized using photo‐crosslinkable resin was applied to treatment of wastewater containing a low concentration of hydrogen peroxide. The structure of photo‐crosslinked resin gel was stable even in the acceleration test of concentrated hydrogen peroxide for a long period. Accordingly, the most important subject of this process was maintaining long term catalase activity. The stabilization of immobilized catalase was investigated by modifying gel carriers with some prepolymers and functional monomers, and adding some stabilizing materials without modification of catalase itself. But these gel carriers did not improve the stability beyond 20%. Addition of Tween 80, polyoxyethylene (5) sorbitan monooleate (hydrophilic/lipophilic balance, HLB: 10.0), to catalase was most effective for maintenance of the activity when the enzyme was immobilized in hydrophilic photo‐crosslinked resin, resulting in 80% of its initial activity. It was assumed that this surfactant did not influence the permeation of hydrogen peroxide into gel, but acted as a stabilizer for catalase in the gel carrier. © 2000 Society of Chemical Industry     

Associations between apolipoprotein E genotype and circulating F<Subscript>2</Subscript>-isoprostane levels in humans

Apolipoprotein E (apoE), an important determinant of plasma lipoprotein metabolism, has three common alleles (ε2, ε3, and ε4). Population studies have shown that the risk of diseases characterized by oxidative damage, such as coronary heart disease and Alzheimer’s disease, is significantly higher in ɛ4 carriers. We evaluated the association between apoE genotypes and plasma F₂-isoprostane levels, an index of lipid peroxidation, in humans. Two hundred seventy-four healthy subjects (104 males, 170 females; 46.9±13.0 yr; 200 whites, 74 blacks; 81 nonsmokers, 64 passive smokers, and 129 active smokers) recruited for a randomized clinical antioxidant intervention trial were included in this analysis. ApoE genotype was determined by PCR and restriction enzyme digestion. Free plasma F₂-isoprostane was measured by GC-MS. Genotype groups were compared using multiple regression analysis with adjustment for sex, age, race, smoking status, body mass index, plasma ascorbic acid, and β-carotene. Subjects with ε3/ε4 and ε4/ε4 genotype (ε4-carriers) and with ε2/ε3 and ε3/ε3 (non-ε4-carriers) were pooled for analysis. In subjects with high cholesterol levels (total cholesterol above 200 mg/dl), plasma F₂-isoprostane levels were 29% higher in ε4 carriers than in non-ε4-carriers (P=0.0056). High-cholesterol subjects that are ε4 carriers have significantly higher levels of lipid peroxidation as assessed by circulating F₂-isoprostane levels.     

Low‐temperature carrier dyeing of poly(vinyl chloride) fibers with disperse dyes

The effects of various carriers on the color yield of poly(vinyl chloride) (PVC) filaments in a disperse dyeing system were investigated. The dyeability of disperse dyes on PVC fiber was improved using appropriate carriers. The enhancement of dye uptake imparted by carriers coincided with the reduction in glass‐transition temperature, which in turn coincided approximately with the similarity of the inorganic/organic ratio values between PVC fibers and carriers. The results clearly demonstrate that hydrophobic carriers function as diluents of PVC fiber, facilitating dye diffusion by increasing segmental mobility of the fiber. The dyed PVC filaments with disperse dyes showed reasonable levels of build‐up and color fastness properties in the carrier‐assisted dyeing. © 2003 Wiley Periodicals, Inc. J Appl Polym Sci 90: 3896–3904, 2003     

Antibacterial polymer composites based on low‐density polyethylene and essential oils immobilized on various solid carriers

In this study, we investigated the antibacterial modification of polymers with biologically active substances in essential oils [EOs; linalool, 4‐allylanisole (ALY), and trans‐anethole]. These compounds were thermoplastically incorporated into a low‐density polyethylene matrix via solid inert carriers [wood flour (WF) and talc and molecular sieves] with physically immobilized EOs. The concentrations of the antibacterial modifiers on the carriers and in the resulting composites were determined with three chromatographic techniques (gas chromatography with mass spectrometry, pyrolysis and gas chromatography with mass spectrometry, and high‐performance liquid chromatography). The effects of such modifications to the mechanical properties of the prepared composites were studied by stress–strain analysis. Interactions on the polymer matrix carriers were observed by scanning electron microscopy. The prepared composites were also tested for antibacterial activity against both Gram‐negative and Gram‐positive bacterial strains. The highest efficiency of isothermal immobilization was found for systems consisting of ALY and WF. This finding was in accordance with microbiological studies. The phase of immobilizing the EOs did not influence the mechanical properties of the studied composites. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015 , 132, 42816.     

Immobilization of α-amylase on polymeric carriers having different structures

α-Amylase was immobilized onto several polymeric carriers having carboxyl groups by the Woodward's reagent K better than by other methods. The amount of immobilized α-amylase mainly depended on the surface area of carriers, while the enzymic activity depended on the texture of carrier surface. Flat surface was favorable for making the enzyme exhibit high activity and for keeping it active in wide pH range. The durability of immobilized enzyme was excellent at 20°C and pH 5.5.     

Magnetic and electrical properties of polypyrrole nanocomposites with iron nanoparticles attached to 2‐acrylamido‐2‐methyl‐1‐propanesulfonic acid

Polypyrrole (PPy) nanocomposites with iron nanoparticles attached to 2‐acrylamido‐2‐methyl‐1‐propanesulfonic acid (AMP) were chemically prepared by varying the concentration of AMP using ammonium persulfate as an oxidant. The synthesized composites were characterized by using Fourier transform infrared (FTIR) spectroscopy, and their surface morphology and amended crystallinity were determined by using the transmission electron microscopy (TEM) and X‐ray diffraction (XRD), respectively. The electrical and magnetic properties of PPy–Fe–AMP composites were investigated at room temperature. The increase of AMP has brought out a significant increase of DC conductivity and magnetic saturation moment of the composites, which might have been caused by the increased mobility of charge carriers due to the hydrophilic AMP components. POLYM. COMPOS., 35:364–371, 2014. © 2013 Society of Plastics Engineers     

Pluronic嵌段共聚物磁性纳米颗粒的制备及其对漆酶的分离纯化

选择合适的双亲性嵌段共聚物Pluronic产品接枝到Fe3O4粒子的表面,合成了粒径均一、亲水性好、非特异性吸附低的纳米级磁性分离载体(PMNPs). 通过对漆酶发酵液的分离纯化研究了Pluronic对排除蛋白非特异性吸附的作用. 结果表明,PMNPs对漆酶的最大吸附量可达0.535 mg/mg,在5~40℃内基本不受温度变化影响,对漆酶发酵液的分离纯化因子为3.4,酶活收率达62.9%,可实现一步分离提纯.     

Development of a new two‐enzyme biosensor based on poly(pyrrole‐co‐3,4‐ethylenedioxythiophene) for lactose determination in milk

A new lactose biosensor was developed by preparing a suitable copolymer of polypyrrole and poly(3,4‐ethylenedioxythiophene) synthesized using the electropolymerization method. Pyrrole and 3,4‐ethylenedioxythiophene monomers were deposited in the presence of sodium dodecylbenzene sulphonic acid on a platinum disc electrode, which was used as the working electrode. The sensor is based on the serial reactions of β‐galactosidase and galactose oxidase immobilized on a copolymer‐modified platinum disc electrode. Successful synthesis of the enzyme‐immobilized copolymer was confirmed by FT‐IR spectrometry, SEM, and electrochemical analysis. The response of the enzyme electrode to lactose was determined by cyclic voltammetry at + 0.40 V. The response time of the biosensor was found to be from 8 to 10 s, and the upper limit of the linear working portion was found to be at a lactose concentration of 2.30 mM with a detection limit of 1.4 × 10^?5 M. The apparent Michaelis–Menten constant was found to be 0.65 mM of lactose. The effects of interferents were also investigated. Lactose concentrations determined by the biosensor were in good agreement with those measured by the reference methods. Our results show that the developed biosensor has a significant potential to the determination of lactose concentration in milk. POLYM. ENG. SCI., 58:839–848, 2018. © 2017 Society of Plastics Engineers     

Effects of reaction conditions on laccase‐catalyzed α‐naphthol polymerization

Enzymatic oxidative polymerization of α‐naphthol was carried out batch‐wise with the laccase enzyme, produced by Trametes versicolor (ATCC 200801). The polymerization reaction was conducted in a closed, temperature controlled system containing acetone (solvent) and sodium acetate buffer for pH control. The effects of the organic solvent (acetone) composition, monomer (α‐naphthol) and enzyme concentrations, buffer pH and temperature on the polymerization rate were investigated with respect to initial reaction conditions and depletion rate of dissolved oxygen. The optimum acetone composition, pH, monomer, dissolved oxygen and enzyme concentrations were determined as 50% (v/v), 5, 3409 gm⁻³, 20.3 gm⁻³ and 0.173 U cm⁻³, respectively; these values provided the most desirable conditions for initial reaction rate. Temperature rise supported the rate increase up to 37 °C, after which the rate tended to be stable due to a drop in dissolved oxygen concentration. The product polymer, poly(α‐naphthol), with an average molecular weight of 4920 Da was soluble in common organic solvents. © 2000 Society of Chemical Industry     

氨基功能载体固定化酶研究进展

综述了甲壳素、壳聚糖天然氨基功能载体和氨基化硅胶化学合成载体的制备方法,并介绍了利用戊二醛直接固定、载体活化和酶活化固定化酶的方法,最后对氨基功能载体固定化酶的发展趋势加以评述:在利用氨基功能载体固定化酶过程中,有必要有针对性地合成一些新的氨基功能载体,使其反应条件更温和、酶的固载量更大、酶活力回收率更高、稳定性更强。或者针对特定的载体和酶,通过结合配体、添加稳定剂、固定前修饰、固定后修饰和后固定化技术处理等方法,进一步改善固定化酶的性能。     

Reaktive träger zur immobilisierung von enzymen auf der grundlage von polyvinylalkohol

On the basis of poly(vinyl alcohol) various reactive carriers for the immobilization of enzymes were synthesized. Starting as well from hydrolyzed beads of crosslinked poly(vinylacetate), as from tubes of poly(vinyl acetate-co-ethylene) which have been coated with poly(vinyl alcohol) resp. from poly(vinyl alcohol) containing synthetic pulp reactive carriers were synthesized by reaction with 2-(3-aminophenyl)-1,3-dioxolane followed by diazotization. Furthermore reactive isothiocyanato groups as well as mixed polymeric disulfides with 2-thiopyridine groups were introduced into gels of poly(vinyl alcohol) which were crosslinked with terephthalaldehyde. The immobilization of enzymes was carried out with papain, trypsin, glucose oxidase, and catalase. The properties of the immobilized enzymes were investigated.