A Preliminary Study of Cell Membrane Mediated Immobilization of a Recombinant Acyl-homoserine Lactonase AidH

The aim of this work was to inhibit biofilm formation by taking advantages of bacterial surface display technology in combination with cell membrane chromatography.A recombinant protein INPAidH was constructed by fusing a quorum signal hydrolase AidH to the C-terminus of the ice nucleation protein(INP).Expression of INP-AidH was achieved on E.coli cell surface at an expression level of 30%of total membrane proteins.Activity of INP-AidH on cell membranes was confirmed in degrading the quorum signal C6-HSL as well as inhibiting bacterial biofilm.Immobilization of INP-AidH anchored cell membranes on silica gel particles was facilitated by taking advantages of cell membrane chromatography.The functionalized silica gel particles also exhibit activities in degrading C6-HSL and inhibiting bacterial biofilm.This article presents a new approach to prevent biofilm formation of silica-based materials.

Facile purification and immobilization of organophosphorus hydrolase on protein-inorganic hybrid phosphate nanosheets

Oriented immobilization of enzymes helps to maintain their native structure and proper orientation for high-performance engineering to meet extensive biocatalysis demands.However,the supporting materials used for orientated immobilization are usually costly or complicated in preparation,affecting their practical applications.In this work,a facile purification and immobilization method was proposed for enzyme immobilization based on organic-inorganic hybrid calcium phosphate nanocrystal(Ca Ps)induced by Cu^(2+) modified bovine serum albumin(BSA-Cu).Then,the as-prepared hybrid calcium phosphate nanosheet,BSA-Cu@Ca Ps,was utilized for one-pot purification and immobilization of His-tagged organophosphorus hydrolase(OPH)by metal-affinity binding to the incorporated BSA.BSA-Cu@Ca PsOPH exhibited enhanced p H stability and thermal stability compared to the free enzyme.Moreover,BSA-Cu@Ca Ps-OPH could retain more than 75%and 56%of initial activity after reuse 5 and 10 times,respectively.The results demonstrated that this facile strategy was promising for the effective biodegradation of organophosphorus pesticides with the immobilized enzyme.

Immobilization of Oxyanions on the Reconstructed Heterostructure Evolved from a Bimetallic Oxysulfide for the Promotion of Oxygen Evolution Reaction

Efficient and durable oxygen evolution reaction(OER)requires the electrocatalyst to bear abundant active sites,optimized electronic structure as well as robust component and mechanical stability.Herein,a bimetallic lanthanum-nickel oxysulfide with rich oxygen vacancies based on the La_(2)O_(2)S prototype is fabricated as a binder-free precatalyst for alkaline OER.The combination of advanced in situ and ex situ characterizations with theoretical calculation uncovers the synergistic effect among La,Ni,O,and S species during OER,which assures the adsorption and stabilization of the oxyanion SO_(4)^(2-)onto the surface of the deeply reconstructed porous heterostructure composed of confining Ni OOH nanodomains by La(OH)_3 barrier.Such coupling,confinement,porosity and immobilization enable notable improvement in active site accessibility,phase stability,mass diffusion capability and the intrinsic Gibbs free energy of oxygen-containing intermediates.The optimized electrocatalyst delivers exceptional alkaline OER activity and durability,outperforming most of the Ni-based benchmark OER electrocatalysts.

Effect of Hydrated Calcium Aluminate Cement on the Chloride Immobilization of Portland Cement Paste

To improve the efficiency and stability of chloride immobilization of portland cement paste,hydrated calcium aluminate cement(HCAC)prepared by wet grinding of CAC was added into portland cement paste as an additive.The immobilized chloride ratio(ICR)was evaluated,and the mechanism of chloride immobilization was researched by XRD,DTG,NMR,and MIP tests.The analysis results demonstrated that HCAC could improve the chloride immobilization capacity of portland cement paste.The mechanism was attributed to the following aspects:chemical binding capacity was enhanced via producing more Kuzel’s salt;physical adsorption capacity was reduced by decreasing the C-S-H gel;migration resistance was enhanced through refining the pore structure.

Formation process of extension knee joint contracture following external immobilization in rats

BACKGROUND Current research lacks a model of knee extension contracture in rats.AIM To elucidate the formation process of knee extension contracture.METHODS We developed a rat model using an aluminum external fixator.Sixty male Sprague-Dawley rats with mature bones were divided into the control group(n=6)and groups that had the left knee immobilized with an aluminum external fixator for 1,2,and 3 d,and 1,2,3,4,6,and 8 wk(n=6 in each group).The passive extension range of motion,histology,and expression of fibrosis-related proteins were compared between the control group and the immobilization groups.RESULTS Myogenic contracture progressed very quickly during the initial 2 wk of immobilization.After 2 wk,the contracture gradually changed from myogenic to arthrogenic.The arthrogenic contracture progressed slowly during the 1^(st) week,rapidly progressed until the 3^(rd) week,and then showed a steady progression until the 4^(rd) week.Histological analyses confirmed that the anterior joint capsule of the extended fixed knee became increasingly thicker over time.Correspondingly,the level of transforming growth factor beta 1(TGF-β1)and phosphorylated mothers against decapentaplegic homolog 2(p-Smad2)in the anterior joint capsule also increased with the immobilization time.Over time,the cross-sectional area of muscle fibers gradually decreased,while the amount of intermuscular collagen and TGF-β1,p-Smad2,and p-Smad3 was increased.Unexpectedly,the amount of intermuscular collagen and TGF-β1,p-Smad2,and p-Smad3 was decreased during the late stage of immobilization(6-8 wk).The myogenic contracture was stabilized after 2 wk of immobilization,whereas the arthrogenic contracture was stabilized after 3 wk of immobilization and completely stable in 4 wk.CONCLUSION This rat model may be a useful tool to study the etiology of joint contracture and establish therapeutic approaches.

Immobilization of activated sludge using improved polyvinyl alcohol (PVA) gel

The microbial immobilization method using polyvinyl alcohol （PVA） gel as an immobilizing material was improved and used for entrapment of activated sludge. The oxygen uptake rate （OUR） was used to characterize the biological activity of immobilized activated sludge. Three kinds of PVA-immobilized particles of activated sludge, that is, PVA-boric acid beads, PVA-sodium nitrate beads and PVA-orthophosphate beads were prepared, and their biological activity was compared by measuring the OUR value. The bioactivity of both autotrophic and heterotrophic microorganisms of activated sludge was determined using different synthetic wastewater media （containing 250 mg/L COD and 25 mg/L NH4^＋ -N）. The experimental results showed that the bioactivity and stability of the three kinds of immobilized activated sludge was greatly improved after activation. With respect of the bioactivity and the mechanical stability, the PVA-orthophosphate method may be a promising and economical technique for microbial immobilization.

Immobilization study of biosorption of heavy metal ions onto activated sludge

Activated sludge was immobilized into Ca-alginate beads via entrapment, and biosorption of three heavy metal ions, copper(Ⅱ), zinc(Ⅱ), and chromimum(Ⅱ), from aqueous solution in the concentration range of 10\_100 mg/L was studied by using both entrapped activated sludge and inactivated free biomass at pH≤5. A biphasic metal adsorption pattern was observed in all immobilized biomass experiments. The biosorption of metal ions by the biosorbents increased with the initial concentration increased in the medium. The adsorption rate of immobilized pre-treated activated sludge(PAS) was much lower than that of free PAS due to the increase in mass transfer resistance resulting from the polymeric matrix. Biosorption equilibrium of beads was established in about 20 h and the adsorbed heavy metal ions did not change further with time. No significant effect of temperature was observed in the test for free biomass while immobilized PAS appeared to be strong temperature dependent in the test range of 10 and 40℃. Besides, the content of activated sludge in the calcium alginate bead has an influence on the uptake of heavy metals. The sorption equilibrium was well modeled by Langmuir isotherm, implying monomolecular adsorption mechanism. Carboxyl group in cell wall played an important role in surface adsorption of heavy metal ions on PAS.

Inhibition of acid mine drainage and immobilization of heavy metals from copper flotation tailings using a marble cutting waste

Acid mine drainage （AMD） with high concentrations of sulfates and metals is generated by the oxidation of sulfide beating wastes. CaCO3-rich marble cutting waste is a residual material produced by the cutting and polishing of marble stone. In this study, the feasibility of using the marble cutting waste as an acid-neutralizing agent to inhibit AMD and immobilize heavy metals from copper flotation tailings （sul- fide-beating wastes） was investigated. Continuous-stirring shake-flask tests were conducted for 40 d, and the pH value, sulfate content, and dissolved metal content of the leachate were analyzed every 10 d to determine the effectiveness of the marble cutting waste as an acid neu- tralizer. For comparison, CaCO3 was also used as a neutralizing agent. The average pH value of the leachate was 2.1 at the beginning of the experiment （t = 0）. In the experiment employing the marble cutting waste, the pH value of the leachate changed from 6.5 to 7.8, and the sul- fate and iron concentrations decreased from 4558 to 838 mg/L and from 536 to 0.01 mg/L, respectively, after 40 d. The marble cutting waste also removed more than 80wt% of heavy metals （Cd, Cr, Cu, Ni, Pb, and Zn） from AMD generated by copper flotation tailings.

Neuropeptide Y and leptin receptor expression in the hypothalamus of rats with chronic immobilization stress

In this study, Sprague-Dawley rats were immobilized to a frame for 3 hours a day for 21 days to establish a model of chronic immobilization stress. The body weight and food intake of rats subjected to chronic immobilization stress were significantly decreased compared with the control group. Dual-labeling immunofluorescence revealed that the expression of leptin receptor and the co-localization coeffient in these leptic receptor neurons in the arcuate nucleus of the hypothalamus were both upregulated, while the number of neuropeptide Y neurons was decreased. Chronic immobilization stress induced high expression of leptin receptor in the arcuate nucleus and suppressed the synthesis and secretion of neuropeptide Y, thereby disrupting the pathways in the arcuate nucleus that regulate feeding behavior, resulting in diminished food intake and reduced body weight.

Immobilization of organophosphorus hydrolase enzyme by covalent attachment on modified cellulose microfibers using different chemical activation strategies:Characterization and stability studies

The plant cellulose powder was activated by two different methods using 1,4-butanediol diglycidyl ether(BTDE)and 1,1′-Carbonyldiimidazole(CDI) as the chemical coupling agents.Organophosphorus hydrolase(OPH) from Flavobacterium ATCC 27551 was immobilized on any of activated support through covalent bonding.The optimal conditions of affecting parameters on enzyme immobilization in both methods were found, and it was demonstrated that the highest activity yields of immobilized OPH onto epoxy and CDI treated cellulose were 68.32%and 73.51%, respectively.The surface treatment of cellulose via covalent coupling with BTDE and CDI agents was proved by FTIR analysis.The kinetic constants of the free and immobilized enzymes were determined, and it was showed that both immobilization techniques moderately increased the Kmvalue of the free OPH.The improvements in storage and thermal stability were investigated and depicted that the half-life of immobilized OPH over the surface of epoxy modified cellulose had a better growth compared to the free and immobilized enzymes onto CDI treated support.Also, the pH stability of the immobilized preparations was enhanced relative to the free counterpart and revealed that all enzyme samples would have the same optimum pH value for stability at 9.0.Additionally, the immobilized OPH onto epoxy and CDI activated cellulose retained about 59% and 68% of their initial activity after ten turns of batch operation, respectively.The results demonstrated the high performance of OPH enzyme in immobilized state onto an inexpensive support with the potential of industrial applications.

Human sperm immobilization effect of Carica papaya seed extracts: an in vitro study

Aim: To examine if the seed extracts of Carica papaya, which showed anfispermatogenic/sperm immobilizationproperties in animal models, could cause human sperm immobilization in vitro. Methods: Chloroform extract, ben-zene chromatographic fraction of the chloroform extract, its methanol and ethyl acetate sub-fractions and the isolatedcompounds from the sub-fractions i. e., ECP 1 & 2 and MCP 1 & 2, of the seeds of Carica papaya were used at con-centrations of 0.1%, 0.5%, 1% and 2%. Sperm motility was assessed immediately after addition of extracts and ev-ery 5 minutes thereafter for 30 minutes. Results: There were dose-dependent spermicidal effects showing an instantfall in the sperm motility to less than 20% at 2% concentration. Isolated compounds ECP 1 & 2 were more effective in-ducing a motility of less than 10%. Many of the spermatozoa became vibratory on the spot. Total inhibition of motilitywas observed within 20-25 rain at all concentrations of all products. Scanning and transmission electron microscopyrevealed deleterious changes in the plasma membrane of the head and mid-piece of spermatozoa. Sperm viability testand the number of abnormal spermatozoa after completion of incubation suggested that the spermatozoa were infertile.The effects were spermicidal but not spermiostatic as revealed by the sperm revival test. Conclusion: The results re-veal spermicidal activity in vitro of the seed extracts of Carica papaya.

Preparation of Magnetic Chitosan Nanoparticles and Immobilization of Laccase

The magnetic chitosan nanoparticles were prepared by reversed-phase suspension method using Span-80 as an emulsifier, glutaraldehyde as cross-linking reagent. And the nanoparticles were characterized by TEM, FT-IR and hysteresis loop. The results show that the nanoparticles are spherical and almost superparamagnetic. The laccase was immobilized on nanoparticles by adsorption and subsequently by cross-linking with glutaraldehyde. The immobilization conditions and charac-terizations of the immobilized laccase were investigated. The optimal immobilization conditions were as follows： 10 mL of phosphate buffer （0.1 M, pH 7.0） containing 50 mg of magnetic chitosan nanoparticles, 1.0 mg·mL-1 of laccase and 1% （v/v） glutaraldehyde, immobilization temperature of 4 ℃ and immobilization time of 4 h. The immobilized laccase exhibited an appreciable catalytic capability （480 units·g-1 support） and had good storage stability and operation stability. The Km of immobilized and free laccase for ABTS were 140.6 and 31.1 μM in phosphate buffer （0.1 M, pH 3.0） at 37 ℃, respectively. The immobilized laccase is a good candidate for the research and development of biosensors based on laccase catalysis.

Sperm immobilization activity of Allium sativum L. and other plant extracts

<abstract>Aim: To identify possible spermicidal agents through screening a number of edible medicinal plants with antimicrobial activity. Methods: Initial screening was made on the basis of ram cauda epididymal sperm immobilization immediately after addition of extracts. The most potent extract was selected and was evaluated on both ram and human spermatozoa. To unravel its mode of action several sperm functional tests were carried out, namely viability of cells, hypo-osmotic swelling test for membrane integrity and assays of membrane-bound enzyme 5'-nucleotidase and acrosomal marker enzyme acrosin. Results: The crude aqueous extract of the bulb of Allium sativum L. showed the most promising results by instant immobilization of the ram epididymal sperm at 0.25 g/mL and human ejaculated sperm at 0.5 g/mL. Sperm immobilizing effects were irreversible and the factor of the extract responsible for immobilization was thermostable up to 90 癈. On boiling at 100 癈 for 10 minutes, this activity was markedly reduced. Moreover, this extract was able to cause aggregation of ram sperms into small clusters after 30 minutes of incubation at 37 癈. However this property was not found in human spermatozoa. More than 50 % reduction in sperm viability and hypo-osmotic swelling occurred in treated sperm as compared with the controls, indicating the possibility of plasma membrane disintegration which was further supported by the significant reduction in the activity of membrane bound 5'-nucleotidase and acrosomal acrosin. Conclusion: The crude aqueous extract of A. sativum bulb possesses spermicidal activity in vitro.

Synthesis and Heavy Metal Immobilization Behaviors of Fly Ash based Gepolymer

Two aspects of studies were carried out： 1） synthesis of geopolymer by using fly ash and metakaolin; 2） Immobilization behaviors of fly ash based geopolymer in a presence of Pb and Cu ions. As for the synthesis of fly ash based geopolymer, 4 different fly ash content （10%, 30%, 50%, 70%） and 3 types of curing regimes （standard curing, steam curing and autoclave curing） were investigated to obtain the optimum synthesis condition based on the compressive and flexural strength. The experimental results show that geopolymer, containing 30% fly ash and synthesized at steam curing （80 ℃ for 8 h）, exhibits higher mechanical strengths. The compressive and flexural strengths of fly ash based geopolymer reach 32.2 MPa and 7.15 MPa, respectively. Additionally, Infrared （IR） and X-ray diffraction （XRD） techniques were used to characterize the microstructure of the fly ash geopolymer. IR spectra shows that the absorptive band at 1086 cm^-1 shifts to lower wave number around 1033 cm^-1, and the 6-coordinated Al transforms into 4-coordination during the syn-thesis of fly ash based geopolymer. The resulting geopolymeric products were X-ray amorphous materials. As for immobilization of heavy metals, the leaching tests were employed to investigate the immobilization behaviors of the fly ash based geopolymer synthesized under the above optimum condition. The leaching tests showed that fly ash based geopolymer can effectively immobilize Cu and Pb heavy metal ions, and the immobilization efficiency reached 90% greater when heavy metals were incorporated in the fly ash geopolymer in the range of 0.1% to 0.3%. The Pb exhibits better immobilization efficiency than the Cu, especially in the case of large dosages of heavy metals.

Immobilization of carbonic anhydrase for facilitated CO2 capture and separation

Carbonic anhydrase(CA)as a typical metalloenzyme in biological system can accelerate the hydration/dehydration of carbon dioxide(CO2,the major components of greenhouse gases),which performer with high selectivity,environmental friendliness and superior efficiency.However,the free form of CA is quite expensive(~RMB 3000/100 mg),unstable,and non-reusable as the free form of CA is not easy for recovery from the reaction environment,which severely limits its large-scale industrial applications.The immobilization may solve these problems at the same time.In this context,many efforts have been devoted to improving the chemical and thermal stabilities of CA through immobilization strategy.Very recently,a wide range of available inorganic,organic and hybrid compounds have been explored as carrier materials for CA immobilization,which could not only improve the tolerance of CA in hazardous environments,but also improve the efficiency and recovery to reduce the cost of large-scale application of CA.Several excellent reviews about immobilization methods and application potential of CA have been published.By contrast,in our review,we stressed on the way to better retain the biocatalytic activity of immobilized CA system based on different carrier materials and to solve the problems facing in practical operations well.The concluding remarks are presented with a perspective on constructing efficient CO2 conversion systems through rational combining CA and advanced carrier materials.

Immobilization of microorganisms using carrageenan gels coated with chitosan and application to biodegradation of 4-chlorophenol

A new cell immobilization method based on the replacement of KCl by KCl+chitosan as the gelling agent was developed. The experimental results showed that through addition of chitosan into gelling agent, the mechanical strength and the thermal stability of the carrageenan gel were greatly improved. The new immobilization method was used to entrap a chlorophenol degrading microorganism. The immobilized microbial cells were applied for chlorophenol biodegradation. The experiments demonstrated that immobilized cells exhibit a higher bioactivity in the degradation of chlorophenol than free cells.

Radiotherapy-customized head immobilization masks:from modeling and analysis to 3D printing

Immobilization devices may be a valuable aid to ensure the improved effectiveness of radiotherapy treatments where constraining the movements of specific anatomical segments is crucial. This need is also present in other situations, specifically when the superposition of various medical images is required for fine identification and characterization of some pathologies. Because of their structural characteristics, existing head immobilization systems may be claustrophobic and very uncomfortable for patients, during both the modeling and usage stages. Because of this, it is important to minimize all the discomforts related to the mask to alleviate patients’ distress and to simultaneously guarantee and maximize the restraint effectiveness of the mask. In the present work, various head immobilization mask models are proposed based on geometrical information extracted from computerized tomography images and from 3D laser scanning point clouds. These models also consider the corresponding connection to a radiotherapy table, as this connection is easily altered to accommodate various manufacturers’ solutions. A set of materials used in the radiotherapy field is considered to allow the assessment of the stiffness and strength of the masks when submitted to typical loadings.

Immobilization of Bovine Serum Albumin on Poly(ether ether ketone) for Surface Biocompatibility Improvement

UV-induced graft polymerization of acrylic acid（AA） on poly（ether ether ketone）（PEEK） films was carried out to introduce ―COOH for the subsequent immobilization of bovine serum albumin（BSA）.BSA was introduced on PEEK surface based on the condensation reaction between ―NH 2 and ―COOH.The modified surface（PEEK-BSA） was characterized by energy-disperse spectrometry（EDS）,X-ray photoelectron spectroscopy（XPS）,water contact angle measurement and UV spectrum analysis.The contact angle was found to decrease from 104° for the virgin PEEK films to 63° for the BSA-immobilized PEEK films,demonstrating a significant improvement of surface hydrophilicity.Moreover,the appearance of nitrogen on PEEK film confirmed by XPS and EDS indicates the immobilization of BSA on PEEK surface.

Fabrication and characterization of epoxylated zwitterionic copolymergrafted silica nanoparticle as a new support for lipase immobilization

Our previous work proved that the thermal stability of Candida rugosa lipase(CRL)immobilized on zwitterionic polymer(poly(carboxybetaine methacrylate))grafted silica nanoparticle(SNP)was much higher than that on poly(glycidyl methecrylate)(pGMA)grafted SNP,while the latter showed significantly increased activity.Inspired by the research,we have herein proposed to synthesize copolymers of zwitterionic sulfobetaine methacrylate(SBMA)and GMA for CRL immobilization.The copolymers were grafted onto SNP surface at three GMA/SBMA(G/S)molar ratios(G100/S0,G50/S50,G10/S90),followed by the covalent coupling of CRL to the surface copolymers.The immobilized CRLs on the corresponding supports were denoted as p(G100-S0)-CRL,p(G50-S50)-CRL and p(G10-S90)-CRL.The enzyme loading increased with the increase of GMA content in the copolymer,while the activity varied with the grafted copolymer composition.Kinetic study proved the improvement of enzyme-substrate affinity after immobilization.In comparison to p(G100-S0)-CRL,p(G50-S50)-CRL and p(G10-S90)-CRL presented remarkably enhanced thermal stability and pH tolerance,and p(G10-S90)-CRL showed the highest stability.These results suggest that the copolymer design is promising for development as a versatile platform for enzyme immobilization.

Surface Modification of Polypropylene Microporous Membrane by Atmospheric-Pressure Plasma Immobilization of N,N-dimethylamino Ethyl Methacrylate

Surface modification of polypropylene microporous membrane （PPMM） was performed by atmospheric pressure dielectric barrier discharge plasma immobilization of N,Ndimethylamino ethyl methacrylate （DMAEMA）. Structural and morphological changes on the membrane surface were characterized by attenuated total reflection-Fourier transform infrared spectroscopy （FT-IR/ATR）, X-ray photoelectron spectroscope （XPS） and field emission scanning electron microscopy （FE-SEM）. Water contact angles of the membrane surfaces were also measured by the sessile drop method. Results reveal that both the plasma-treating conditions and the adsorbed DMAEMA amount have remarkable effects on the immobilization degree of DMAEMA. Peroxide determination by 1,1-diphenyl-2-picrvlhydrazyl （DPPH） method verifies the exsistence of radicals induced by plasma, which activize the immobilization reaction. Pure water contact angle on the membrane surface decreased with the increase of DMAEMA immobilization degree, which indicates an enhanced hydrophilicity for the modified membranes. The effects of immobilization degrees on pure water fluxes were also measured. It is shown that pure water fluxes first increased with immobilization degree and then decreased. Finally, permeation of bovine serum albumin （BSA） and lysozyme solution were measured to evaluate the antifouling property of the DMAEMA-modified membranes, from which it is shown that both hydrophilicity and electrostatic repulsion are beneficial for membrane antifouling.