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Bulk Solution (bulk + solution)
Selected AbstractsProtein crystal nucleation: Recent notionsCRYSTAL RESEARCH AND TECHNOLOGY, Issue 1 2007Christo N. Nanev Abstract The nucleation of protein crystals is reconsidered taking into account the specificity of the protein molecules. In contrast to the homogeneous surface properties of small molecules, the protein molecule surface is highly inhomogeneous. Over their surfaces proteins exhibit high anisotropic distribution of patches, which are able to form crystalline bonds, the crystallization patch representing only a small fraction of the total surface of the protein molecule. Therefore, an appropriate spatial orientation of the colliding protein molecules is required in order to create a crystalline cluster. This scenario decreases considerably the success ratio of the attempt frequency for crystal nucleation. On the other hand a heterogeneous nucleation of (protein) crystals may be accelerated due to the arrival on some support of under-critical clusters that are formed in bulk solution; when arriving there they may acquire the property of critical nuclei. Thus, a plausible explanation of important peculiarities of protein crystal nucleation, as inferred from the experimental data, is suggested. (© 2007 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) [source] Adsorptive Stripping Analysis of Riboflavin at Electrically Heated Graphite Cylindrical ElectrodesELECTROANALYSIS, Issue 21 2007Shao-Hua Wu Abstract Electrically heated graphite cylindrical electrodes (HGCEs) made from ground pencil leads have been used to perform adsorptive stripping square wave voltammetry (SWV) measurements of trace riboflavin (RF). The SWV stripping peak current was significantly enhanced with increasing the electrode temperature only during preconcentration step. This enhancement was due to the forced thermal convection induced by heating the electrode rather than the bulk solution. It is the thermal convection that has the ability to improve mass transfer and facilitate adsorption thus enhance stripping responses. It was found that the detection limit of 5×10,9,M (S/N=3) could be obtained at an electrode temperature of 72,°C during 5,min accumulation, more than one magnitude lower than that at 22,°C (room temperature), the sensitivity could be enhanced ca. eight or four folds for two different RF concentration ranges. So it is possible to develop a new highly sensitive method to determine riboflavin at HGCEs. Such HGCEs were also successfully used to determine RF in multivitamin tablets. [source] A New Optically Reflective Thin Layer Electrode (ORTLE) Window: Gold on a Thin Porous Alumina Film Used to Observe the Onset of Water ReductionELECTROANALYSIS, Issue 1-2 2004Abstract The fabrication and unique characteristics of a new type of thin layer electrode, an optically reflective thin layer electrode (ORTLE), are described. The electrode was fabricated by the anodization of a thin layer of aluminum sputtered onto a plain glass microscope slide to create a 750,nm-thick porous alumina film. A thin film of gold was then sputtered atop the porous and transparent alumina film. The gold layer remained porous to allow solution into the pores but was optically thick and reflective. Reflectance measurements made through the microscope slide did not interrogate the bulk solution, but show spectral features that shift with the optical properties of the material filling the pores of the alumina film. A simple series of experiments, in which the potential of the ORTLE was stepped negatively to various values in an aqueous sodium sulfate solution, shows that interference fringes shift measurably in the ORTLE spectrum at potentials several hundred millivolts positive of the potential at which gas evolution was visible to the naked eye. [source] Electrolytic Deposition of Hydroxyapatite Coating on CoNiCrMo SubstratesADVANCED ENGINEERING MATERIALS, Issue 1-2 2010Dong-Yang Lin Hydroxyapatite (HA) coating was fabricated on CoNiCrMo alloy by electrolytic deposition (ELD). Different kinds of uncharged substrates were placed close to the cathode separately during the ELD process. Both CoNiCrMo and uncharged substrates were covered with uniform HA coatings composed of hexagonal prism crystals after 60,min deposition. The pH value of the bulk solution changed hardly while the local pH had a sharp increase after ELD. The results demonstrate the local pH plays a crucial role in the ELD process. [source] Substrate Channelling in a Creatine Kinase System of Rat Skeletal Muscle Under Various pH ConditionsEXPERIMENTAL PHYSIOLOGY, Issue 1 2003M. Gregor The aim of this study was to evaluate myofibrillar creatine kinase (CK) activity and to quantify the substrate channelling of ATP between CK and myosin ATPase under different pH conditions within the integrity of myofibrils. A pure myofibrillar fraction was prepared using differential centrifugation. The homogeneity of the preparation and the purity of the fraction were confirmed microscopically and by enzymatic assays for contaminant enzyme activities. The specific activity of myofibrillar CK reached 584 ± 33 nmol PCr min,1 mg,1 at pH 6.75. Two methods were used to detect CK activity: (1) measurement of direct ATP production, and (2) measurement of PCr consumption. This method of evaluation has been tested in experiments with isolated creatine kinase. No discrepancy in CK activity between the methods was observed in the pH range tested (6.0-7.5). However, the same procedures resulted in a significant discrepancy between the amounts of reacted PCr and produced ATP within the pure myofibrillar fraction. This discrepancy represents the portion of ATP produced by the CK reaction, which is preferentially channelled to the myosin ATPase before diffusing into the bulk solution. The maximum evaluated difference reached 42.3 % at pH 6.95. The substrate channelling between myofibrillar-bound CK and myosin ATPase was evaluated under various pH levels within the physiological range and it reached a maximum value in a slightly acidic environment. These results suggest that ATP/ADP flux control by the CK system is more important at lower pH, corresponding to the physiological state of muscle fatigue. [source] Self-assembled pearling structure of long duplex DNA with histone H1FEBS JOURNAL, Issue 9 2001Yuko Yoshikawa We report that complexes of giant DNA molecules with histone H1 proteins form a pearl necklace-like structure when the complexes are prepared by natural dilution from a high-salt solution (2 m NaCl) to a low-salt solution (0.2 m and 50 mm NaCl). We performed real-time observations on the conformational changes of individual T4 phage DNA (166 kb) molecules in bulk solution by fluorescence microscopy. To identify H1-binding regions on individual DNA molecules, we also performed immunofluorescence microscopic observations on the DNA,H1 complex spread on a glass surface. It was found that histone H1 binds DNA in a highly co-operative manner and is accompanied by local folding of the DNA. On the basis of the experimental observations and a theoretical simulation, we propose a self-assembling mechanism for the pearling structure. [source] Chemical modification of pyroclastic rock by hot water: an experimental investigation of mass transport at the fluid,solid interfaceGEOFLUIDS (ELECTRONIC), Issue 1 2009J. HARA Abstract Hydrothermal water,(pyroclastic) rock interactions were examined using flow-through experiments to deduce the effect of mass transport phenomena on the reaction process. A series of experiments were conducted over the temperature range 75,250°C, with a constant temperature for each experiment, and at saturated vapour pressure, to estimate the apparent rate constants as a function of temperature. Based on the chemistry of analysed solutions, the water,rock interaction in the experiments was controlled by diffusion from the reaction surface and by the existence of a surface layer at the rock,fluid interface, which regulated the chemical reaction rate. The reaction progress depended to a high degree on flow velocity and temperature conditions, with element abundances in the fluid significantly affected by these factors. Mass transport coefficients for diffusion from the rock surface to the bulk solution have been estimated. Ca is selectively depleted under lower temperature conditions (T < 150°C), whereas Na is greatly depleted under higher temperature conditions (T > 150°C), and K reaction rates are increased when flow velocity increases. Using these conditions, specific alkali and alkali earth cations were selectively leached from mineral surfaces. The ,surface layer' comprised a 0.5,1.8 mm boundary film on the solution side (the thickness of this layer has no dependence on chemical character) and a reaction layer. The reaction layer was composed of a Si, Al-rich cation-leached layer, whose thickness was dependent on temperature, flow velocity and reaction length. The reaction layer varied in thickness from about 10,4 to 10,7 mm under high temperature/low fluid velocity and low temperature/high fluid velocity conditions, respectively. [source] Bioactivity of novel carboxymethyl chitosan scaffold incorporating MTA in a tooth modelINTERNATIONAL ENDODONTIC JOURNAL, Issue 10 2010R. Budiraharjo Budiraharjo R, Neoh KG, Kang ET, Kishen A. Bioactivity of novel carboxymethyl chitosan scaffold incorporating MTA in a tooth model. International Endodontic Journal, 43, 930,939, 2010. Abstract Aim, To characterise the bioactivity of a novel carboxymethyl chitosan (CMCS) scaffold with and without incorporating mineral trioxide aggregate (MTA) in a tooth model. Methodology, Cross-linked CMCS scaffold (CaC) and MTA-coated CaC (CaMT) scaffold were prepared by freeze-drying. The bioactivity of the scaffolds was tested in vitro in four different mineralisation solutions (bulk system) and ex vivo in simulated body fluid (SBF) in the tooth model. After mineralisation, the mineral deposits on the scaffolds were analysed using scanning electron microscopy, energy dispersive X-ray, and inductively coupled plasma mass spectroscopy. All data were statistically analysed using the two-sample t -test (P < 0.05). Results, Hydroxyapatite (HAP) deposition was observed on CaC and CaMT scaffolds after 1 week of mineralisation in the tooth model and in the bulk system. The deposition was significantly higher (P < 0.05) on CaMT scaffold than that on CaC scaffold. The amount of HAP formed in the tooth model was significantly lower (P < 0.05) than that in the bulk solution. Conclusions, The CMCS scaffolds are bioactive and capable of biomineralisation by forming HAP within a tooth model ex vivo. The bioactivity of the CMCS scaffold can be enhanced by incorporating MTA. [source] Solvent effects on glycine II.JOURNAL OF COMPUTATIONAL CHEMISTRY, Issue 5 2004Water-assisted tautomerization Abstract The water-assisted tautomerization of glycine has been investigated at the B3LYP/6-31+G** level using supermolecules containing up to six water molecules as well as considering a 1:1 glycine,water complex embedded in a continuum. The conformations of the tautomers in this mechanism do not display an intramolecular H bond, instead the functional groups are bridged by a water molecule. The replacement of the intramolecular H bond by the bridging water reduces the polarity of the NH bond in the zwitterion and increases that of the OH bond in the neutral, stabilizing the zwitterion. Both the charge transfer effects and electrostatic interactions stabilize the nonintramolecularly H-bonded zwitterion conformer over the intramolecularly hydrogen bonded one. The nonintramolecularly H-bonded neutral is favored only by charge transfer effects. Although there is no strong evidence whether the intramolecularly hydrogen bonded or non hydrogen bonded structures are favored in the bulk solution represented as a dielectric continuum, it is likely that the latter species are more stable. The free energy of activation of the water-assisted mechanism is higher than the intramolecular proton transfer channel. However, when the presumably higher conformational energy of the zwitterion reacting in the intramolecular mechanism is taken into account, both mechanisms are observed to compete. The various conformers of the neutral glycine may form via multiple proton transfer reactions through several water molecules instead of a conformational rearrangement. © 2004 Wiley Periodicals, Inc. J Comput Chem 25: 690,703, 2004 [source] Direct monitoring of molecular recognition processes using fluorescence enhancement at colloid-coated microplatesJOURNAL OF MOLECULAR RECOGNITION, Issue 4 2001Ch. Lobmaier Abstract Direct monitoring of recognition processes at the molecular level is a valuable tool for studying reaction kinetics to assess affinity constants (e.g. drugs to receptors) and for designing rapid single step immunoassays. Methods currently used to gain information about binding processes predominantly depend on surface plasmon resonance. These systems use excitation with coherent light in attenuated total reflection geometry to obtain discrimination between surface-bound and free molecules in solution. Therefore labeling of the compounds is not necessary, but due to the complexity of the measuring setup the method is rather costly. In this contribution we present a simple method for performing kinetic single step biorecognition assays with fluorophore labeled compounds using the fluorescence enhancement properties of surface bound silver colloids. Silver colloids are bound to standard microplates via silanization of the plastic surface. Fluorophores close to this colloid coated surface show a significant gain in fluorescence compared to fluorophores farther away in the bulk solution. Therefore discrimination between surface bound and free fluorophores is possible and the binding of, for example, fluorophore labeled antibodies to antigens immobilized on the colloid surface results in increasing fluorescence intensity. Utilization of standard microplates makes this method fully compatible with conventional microplate processing and reading devices. Neither excitation with coherent laser light nor ATR geometry is required, the measurement is performed in a standard fluorescence microplate reader in front face geometry with a xenon flash lamp as excitation source. Methods for the preparation of colloid-coated microplates and fluorescence-enhanced biorecognition assays are presented. Additionally the dependence of the system performance on the structure and properties of the metal colloid coated surface is described. A two-component biorecognition model system shows a detection limit in the subnanomolar range. The ease of colloid-surface preparation and the high sensitivity makes fluorescence enhancement at colloid-coated microplates a valuable tool for studying reaction kinetics and performing rapid single-step immunoassays. Copyright © 2001 John Wiley & Sons, Ltd. [source] Analysis of unstirred water layer in in vitro permeability experimentsJOURNAL OF PHARMACEUTICAL SCIENCES, Issue 12 2009Timo Korjamo Abstract In vitro permeability experiments are used widely in drug discovery and other areas of pharmaceutical research. Much effort has been expended in developing novel epithelial models but generally much less attention has been paid to the hydrodynamic barrier in the actual experiments. The restricted liquid flow in the vicinity of solid surfaces leads to a zone where the diffusional movement of molecules exceeds the convection. This leads to formation of a concentration gradient between the bulk solution and the surface. The formed unstirred water layer (UWL) reduces the apparent permeability (Papp) of compounds that rapidly pass through the actual epithelial layer. This lowers the resolution of Papp versus fraction-absorbed assay, complicates the structure-permeability analysis and skews apparent kinetic parameters of transporter substrates. This review describes the techniques that can be used to determine the UWL thickness in permeability experiments and apparatuses described in the literature to control the in vitro hydrodynamics. © 2009 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 98:4469,4479, 2009 [source] (203) Manufacturing Process for a Highly Purified, Stable Liquid Formulation of Botulinum Toxin Type B (MyoblocÔ)PAIN MEDICINE, Issue 3 2001Andrew Grethlein Botulinum toxin type B (BoNT-B; MyoblocÔ) is a new botulinum toxin with demonstrated efficacy in patients with cervical dystonia, and has been found to decrease neck pain associated with this disorder. Developmental work has led to the commercial-scale production of a uniform, highly purified toxin complex in a liquid formulation. Production of BoNT-B involves fermentation, recovery and purification from Clostridium botulinum. The reliability and robustness of the process were tested by altering critical process parameters (eg, pH, temperature) and showing that a high-quality product resulted even in conditions detrimental to C botulinum fermentation. Consistency and key quality attributes (purity, complex integrity, percent nicking, specific activity) of the toxin were assessed using a series of biochemical tests, which were validated as precise and accurate and are now routinely used in quality control analysis. Results confirmed production of an intact, uniform toxin complex with consistent purity, percentage nicking (a measure of toxin activation) of over 70%, and specific activity over 90 U/ng in three manufacturing runs. BoNT-B is manufactured as a slightly acidic liquid formulation that maintains complex integrity, reducing the potential for generating neutralizing antibodies. The potency of drug substance, dilute bulk solution, and finished product was shown to be reliable using the validated mouse intraperitoneal LD50 potency assay. The liquid formulation of BoNT-B was found to be stable for at least 9 months at 25°C and at least 3 years at 2,8°C. BoNT-B has a long shelf-life and may be produced on a commercial scale reliably and reproducibly, making it readily available and convenient to store and use. Support of Elan Pharmaceuticals is gratefully acknowledged. [source] Protein crystallization in hydrogel beadsACTA CRYSTALLOGRAPHICA SECTION D, Issue 9 2005Ronnie Willaert The use of hydrogel beads for the crystallization of proteins is explored in this contribution. The dynamic behaviour of the internal precipitant, protein concentration and relative supersaturation in a gel bead upon submerging the bead in a precipitant solution is characterized theoretically using a transient diffusion model. Agarose and calcium alginate beads have been used for the crystallization of a low-molecular-weight (14.4,kDa, hen egg-white lysozyme) and a high-molecular-weight (636.0,kDa, alcohol oxidase) protein. Entrapment of the protein in the agarose-gel matrix was accomplished using two methods. In the first method, a protein solution is mixed with the agarose sol solution. Gel beads are produced by immersing drops of the protein,agarose sol mixture in a cold paraffin solution. In the second method (which was used to produce calcium alginate and agarose beads), empty gel beads are first produced and subsequently filled with protein by diffusion from a bulk solution into the bead. This latter method has the advantage that a supplementary purification step is introduced (for protein aggregates and large impurities) owing to the diffusion process in the gel matrix. Increasing the precipitant, gel concentration and protein loading resulted in a larger number of crystals of smaller size. Consequently, agarose as well as alginate gels act as nucleation promoters. The supersaturation in a gel bead can be dynamically controlled by changing the precipitant and/or the protein concentration in the bulk solution. Manipulation of the supersaturation allowed the nucleation rate to be varied and led to the production of large crystals which were homogeneously distributed in the gel bead. [source] Boronic Acid Functionalized Core,Satellite Composite Nanoparticles for Advanced Enrichment of Glycopeptides and GlycoproteinsCHEMISTRY - A EUROPEAN JOURNAL, Issue 39 2009Lijuan Zhang Abstract A core,satellite-structured composite material has been successfully synthesized for capturing glycosylated peptides or proteins. This novel hybrid material is composed of a silica-coated ferrite "core" and numerous "satellites" of gold nanoparticles with lots of "anchors". The anchor, 3-aminophenylboronic acid, designed for capturing target molecules, is highly specific toward glycosylated species. The long organic chains bridging the gold surface and the anchors could reduce the steric hindrance among the bound molecules and suppress nonspecific bindings. Due to the excellent structure of the current material, the trap-and-release enrichment of glycosylated samples is quite simple, specific, and effective. Indeed, the composite nanoparticles could be used for enriching glycosylated peptides and proteins with very low concentrations, and the enriched samples can be easily separated from bulk solution by a magnet. By using this strategy, the recovery of glycopeptides and glycoproteins after enrichment were found to be 85.9 and 71.6,% separately, whereas the adsorption capacity of the composite nanoparticles was proven to be more than 79,mg of glycoproteins per gram of the material. Moreover, the new composite nanoparticles were applied to enrich glycosylated proteins from human colorectal cancer tissues for identification of N-glycosylation sites. In all, 194 unique glycosylation sites mapped to 155 different glycoproteins have been identified, of which 165 sites (85.1,%) were newly identified. [source] Inlaid Multi-Walled Carbon Nanotube Nanoelectrode Arrays for ElectroanalysisELECTROANALYSIS, Issue 1 2005Jun Li Abstract The rapid development in nanomaterials and nanotechnologies has provided many new opportunities for electroanalysis. We review our recent results on the fabrication and electroanalytical applications of nanoelectrode arrays based on vertically aligned multi-walled carbon nanotubes (MWCNTs). A bottom-up approach is demonstrated, which is compatible with Si microfabrication processes. MWCNTs are encapsulated in SiO2 matrix leaving only the very end exposed to form inlaid nanoelectrode arrays. The electrical and electrochemical properties have been characterized, showing well-defined quasireversible nanoelectrode behavior. Ultrasensitive detection of small redox molecules in bulk solutions as well as immobilized at the MWCNT ends is demonstrated. A label-free affinity-based DNA sensor has shown extremely high sensitivity approaching that of fluorescence techniques. This platform can be integrated with microelectronics and microfluidics for fully automated microchips. [source] Reversible Hydrogen Storage in Hydrogel Clathrate HydratesADVANCED MATERIALS, Issue 23 2009Fabing Su The use of inexpensive hydrogels as supports to significantly improve H2 enclathration kinetics and capacities in THF,H2O clathrate hydrate with respect to bulk solutions is demonstrated. Polymer hydrogels give rise to significant rate and capacity enhancements for hydrogen clathrate formation with respect to unmixed bulk systems, suggesting potential for accelerated gas-storage kinetics in clathrate-based technologies. [source] Study on bulk polymerization of methyl methacrylate initiated by low intensity ultrasonic irradiationJOURNAL OF APPLIED POLYMER SCIENCE, Issue 6 2010Sude Ma Abstract Methyl methacrylate (MMA) was polymerized in bulk solutions using low intensity ultrasonic radiation of 0.25 W/cm2. The polymerization occurred after 1 h of irradiation time was applied. The polymerization rate was greatly accelerated either by increasing the amount of poly (methyl methacrylate) (PMMA) granular added into the system or by elongating the irradiation time. However, it was found that the reaction rate increased with the decreasing of the ultrasonic frequencies when the exposure time of the polymerization under the irradiation was less than 3 h. Experimental results verified that the polymerization was initiated by free radicals, which were mainly generated from the degradation of PMMA macromolecular chains, the friction between the polymer macromolecular chains and the solvent monomer. These findings were obviously different from those obtained when high intensity ultrasonic irradiation was used. The polymers fabricated in this study by using ultrasound irradiation have a narrower molecular weight distribution compared to those obtained from the polymerizations induced by the conventional initiators. © 2010 Wiley Periodicals, Inc. J Appl Polym Sci, 2010 [source] IgG binding kinetics to oligo B protein A domains on lipid layers immobilized on a 27,MHz quartz-crystal microbalanceJOURNAL OF MOLECULAR RECOGNITION, Issue 2 2007Hideyuki Mitomo Abstract Although molecular recognitions between membrane receptors and their soluble ligands have been analyzed using their soluble proteins in bulk solutions, molecular recognitions of membrane receptors should be studied on lipid membranes considering their orientation and dynamics on membrane surfaces. We employed Staphylococcal Protein A (SpA) oligo B domains with long trialkyl-tags from E. coli (LppBx, x,=,1, 2, and 5) and immobilized LppBx on lipid layers using hydrophobic interactions from the trialkyl-tag, while maintaining the orientation of B domain-chains on a 27,MHz quartz-crystal microbalance (QCM; AT-cut shear mode). The binding of IgG Fc regions to LppBx on lipid layers was detected by frequency decreases (mass increases) on the QCM. The maximum amount bound (,mmax), association constants (Ka), association and dissociation rate constants (k1 and k,1, respectively) were obtained. Binding kinetics of IgG to LppB2 and LppB5 were quite similar, showing a simple 1:1 binding of the IgG Fc region to the B domain, when the surface coverage of LppB2 and LppB5 on the lipid surface is low (1.4%). When LppB5 was immobilized at the high surface coverage of 3.5%, the complex bindings of IgG such as one IgG bound to one or two LppB5 on the membrane could be observed. IgG-LppB1 binding was largely restricted because of steric hindrance on lipid surfaces. This gives a suggestion why Protein A has five IgG binding domains. Copyright © 2006 John Wiley & Sons, Ltd. [source] | ||||||||||||||||