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Study on the concentration effects in size exclusion chromatography VII. A quantitative verification for the model theory of concentration and molecular mass dependences of hydrodynamic volumes for polydisperse polymers.

A model theory of concentration effects for polydisperse polymers was proposed in 1988. It is successful in relating the concentration of the injected solution to the effective hydrodynamic volumes of peak, the retention volumes of peak and the polydispersity index (Dc = (Vhcw)/(Vhcn) of hydro-dynamic volume distribution for polydisperse polymers at a given concentration. The dependence of the concentration of injected polymer solution on the effective hydrodynamic volumes, the retention volumes of peak and the polydispersity index of hydrodynamic volume distribution for narrow disperse and polydisperse polystyrene, poly(dodecyl methacrylate), poly(tridecyl methacrylate) and poly(methyl methacrylate) in tetrahydrofuran solvent were studied. The proposed theory was verified by these experimental data. Results show that the proposed theory can predict the concentration effects in GPC for polydisperse polymers quantitatively and can provide a theoretical foundation for the two methods of calibrating the universal calibration curves with polydisperse polymers and of determining the second virial coefficients (A2) of polymers. It is found that the determined values of A2 for narrow disperse and polydisperse polymers by the proposed method are in agreement with those obtained by the LALLS method, and the two universal calibration curves with narrow disperse and polydisperse polymers are in excellent agreement.

Calibration↗

Hydrodynamic instability and coalescence in trains of emulsion drops or gas bubbles moving through a narrow capillary.

We investigate the effect of surfactant on the hydrodynamic stability of a thin liquid film formed between two emulsion drops or gas bubbles, which are moving along a narrow capillary. A ganglion (deformed drop or bubble in a pore) is covered by an adsorption monolayer of surfactant. Due to the hydrodynamic viscous friction, the surfactant is dragged from the front part of a moving ganglion toward its rear part. Consequently, the front and rear parts are, respectively, depleted and enriched in adsorbed surfactant. When such two ganglia move one after another, surfactant molecules desorb from the rear part of the first ganglion and are transferred by diffusion, across the intermediate liquid film, to the front part of the second ganglion. This leads to the appearance of a diffusion-driven hydrodynamic instability, which may cause coalescence of the two neighboring drops or bubbles. The coalescence occurs through a dimple-like perturbation in the film thickness, which is due to a local lowering in the pressure caused by a faster circulation of the liquid inside the film, which in turn is engendered by the accelerated surfactant diffusion across the thinner parts of the film. The developed theory predicts the critical distance between the two ganglia, which corresponds to the onset of coalescence, and its dependence on the radius of the capillary channel, velocity of motion, surfactant concentration and type of the operative surface forces. The results can be useful for a better understanding and quantitative description of the processes accompanying the flow of emulsions and foams though porous media.

Journal Article↗

Hydrodynamic and biochemical effects of isolated hypothermic renal perfusion depending on the pump model and perfusion solution.

OBJECTIVES: To evaluate hydrodynamic, biochemical, and histological consequences of hypothermic isolated renal perfusion using a new computerized perfusion system. MATERIALS AND METHODS: The device that allowed us to obtain on renal hydrodynamics during perfusion included multiple parts. The organ was perfused at 4 degrees C with a constant flow either using a classic roller pump or a pump designed in our laboratory to employ vacuum or atmospheric pressure sequentially to achieve a truly pulsatile wave (vacuum-powered tubular pump). The study included 16 minipigs with Eurocollins or Belzer perfusion solutions sampled at predefined interval and histological studies of the organs performed. RESULTS: There was a significant difference in weight increase between the two types of pumps; those perfused with Eurocollins showed greater values than those with Belzer solution. Onset of nitric oxide (NO) in the perfusion solution increased inversely with the renal vascular resistance. The highest NO levels were observed with the Belzer solution and vacuum pump. CONCLUSIONS: Changes in renal hydrodynamics, as induced by perfusion wave form and solution type, may be recorded in real time using a computerized system. A vacuum pump with the Belzer solution achieved the best experimental results.

Animals↗

Hydrodynamic delivery.

Hydrodynamic delivery has emerged as a near-perfect method for intracellular DNA delivery in vivo. For gene delivery to parenchymal cells, only essential DNA sequences need to be injected via a selected blood vessel, eliminating safety concerns associated with current viral and synthetic vectors. When injected into the bloodstream, DNA is capable of reaching cells in the different tissues accessible to the blood. Hydrodynamic delivery employs the force generated by the rapid injection of a large volume of solution into the incompressible blood in the circulation to overcome the physical barriers of endothelium and cell membranes that prevent large and membrane-impermeable compounds from entering parenchymal cells. In addition to the delivery of DNA, this method is useful for the efficient intracellular delivery of RNA, proteins, and other small compounds in vivo. This review discusses the development, current application, and clinical potential of hydrodynamic delivery.

Animals↗

Hydrodynamic models and computational methods for NMR relaxation.

Interpretation of NMR relaxation data of macromolecules is based on the analysis of their dynamic behavior in solution. For quasirigid molecules, in addition to a minor, separable contribution from local mobility, the main contribution corresponds to the overall rotational diffusion of the complete molecule. Therefore, theoretical descriptions and computational methodologies for hydrodynamic calculations, which yield the full, anisotropic rotational diffusion tensor of rigid molecules, are extremely helpful in the analysis of NMR relaxation. Recent approaches allow realistic predictions of the rotational diffusion tensor from structures at atomic detail. This enables measured relaxation rates and structural models to be compared. Such a comparison (1) provides an independent test of the structural model, (2) provides a framework for the interpretation of local motion, even for highly anisotropic systems, (3) provides a simple method for the detection of additional sources of relaxation, such as chemical exchange, and (4) provides a sensitive method for the detection of nonspecific aggregation or oligomer formation. Although hydrodynamic calculations usually assume a rigid structure, Brownian dynamics simulations extend their range of applications to flexible multidomain structures. Hydrodynamic applications are not restricted to globular proteins. Small DNA fragments, which could be otherwise considered cylindrical objects, can also be treated with atomic detail using the same methodology used for proteins.

Computational Biology↗

Value of phase contrast magnetic resonance imaging for investigation of cerebral hydrodynamics.

OBJECTIVE: Phase Contrast Magnetic Resonance Imaging (PCMRI) is a noninvasive technique that can be used to quantify variations of flow during the cardiac cycle. PCMRI allows investigations of blood flow dynamics in the main arteries and veins of the brain but also the dynamics of cerebrospinal fluid. These cerebral flow investigations provide a description of the regulation mechanisms of intracranial pressure during the cardiac cycle. The objective of this paper is to describe the contribution of this technique in diseases related to disorders of cerebral hydrodynamics in the light of 5 clinical cases. METHOD: Flow measurements were performed using PCMRI sequences on a 1.5 Tesla MR imager in 4 patients with symptomatic ventricular dilation and 1 patient with a syringomyelic cavity. RESULTS: Flow quantification in these 5 patients, representative of the diseases mainly concerned by cerebral hydrodynamics, is useful to guide the indication for ventricular shunting in patients with hydrocephalus, to demonstrate obstruction of the cerebral aqueduct, to demonstrate recirculation of ventricular CSF after ventriculostomy and to characterize the dynamic features of CSF inside a spinal cavity. CONCLUSION: PCMRI, now available to neurosurgeons, is complementary to morphological MR and provides quantitative information on cerebral hydrodynamics. This information is mainly used to confirm alteration of CSF flow in the cerebral and spinal compartments. PCMRI is also a functional tool to better understand the pathophysiology of hydrocephalus and syringomyelia.

Adult↗

The role of inducible nitric oxide synthase in a murine acute hepatitis B virus (HBV) infection model induced by hydrodynamics-based in vivo transfection of HBV-DNA.

BACKGROUND/AIMS: Inducible nitric oxide synthase (iNOS) is found to have antiviral activity. Its role is evaluated using a murine acute hepatitis B virus (HBV) infection model. METHODS: pHBV3.6 plasmid containing HBV genome was injected into mice by hydrodynamics-based in vivo transfection. HBV antigenemia and serum HBV-DNA were detected by enzyme-linked immunosorbent assay (ELISA) and polymerase chain reaction, respectively. HBV replication in liver was analyzed by Northern and Southern blot. Intrahepatic leukocytes were isolated and analyzed with flow cytometry. RESULTS: After hydrodynamics injection of pHBV3.6, HBV genome was synthesized in the liver and HBV-DNA, as well as hepatitis B surface antigen and hepatitis B e antigen were secreted into the blood. Anti-HBV antibody responses developed afterward. A murine acute HBV infection model was established with hydrodynamics injection of non-transponase based HBV-DNA. Using this protocol in iNOS deficient or wild type B6 mice, the level of HBV transcript, replicative intermediate, and antigenemia were higher in iNOS(-/-) than in B6 mice. The intrahepatic leukocytes in iNOS(-/-) mice were also affected after transfection. CONCLUSIONS: Our data suggests that the iNOS expression not only affects the HBV clearance, but also modulates the infiltrating leukocytes response to HBV antigens.

Acute Disease↗

Hydration from hydrodynamics. General considerations and applications of bead modelling to globular proteins.

The effect of hydration on hydrodynamic properties of globular proteins can be expressed in terms of two quantities: the delta (g/g) parameter and the thickness of the hydration layer. The two paradigms on hydration that originate these alternative measures are described and compared. For the numerical calculation of hydrodynamic properties, from which estimates of hydration can be made, we employ the bead modelling with atomic resolution implemented in programs HYDROPRO and HYDRONMR. As typical, average values, we find 0.3 g/g and a thickness of only approximately 1.2 A. However, noticeable differences in this parameter are found from one protein to another. We have made a numerical analysis, which leaves apart marginal influences of modelling imperfections by simulating properties of a spherical protein. This analysis confirms that the errors that one can attribute to the experimental quantities suffice to explain the observed fluctuations in the hydration parameters. However, for the main purpose of predicting protein solution properties, the above mentioned typical values may be safely used. Particularly for atomic bead modelling, a hydrodynamic radius of approximately 3.2 A yields predictions in very good agreement with experiments.

Models, Chemical↗

Effects of hydrodynamic chromatography on colloid-facilitated migration of radionuclides in the fractured rock.

The performance assessment of high level radioactive waste disposal has emphasized the role of colloids in the migration of radionuclides in the geosphere. Previous literature [Nagasaki S, Tanaka S, Suzuki A. Fast transport of colloidal particles through quartz-packed columns. J. Nucl. Sci. Technol. 1975;30(11):1136] indicates that owing to hydrodynamic chromatography the colloid velocity may not be equal to that of groundwater. Using hydrodynamic chromatography, this work investigates the effects of the size of colloidal particles on the radionuclide migration facilitated by colloids in a single fractured porous rock. Also, a methodology is proposed to develop a predictive model to assess transport within the fracture rock as well as various other phenomenological coefficients, particularly the size of colloidal particles. In addition, a fully developed concentration profile for non-reactive colloids in the fracture is developed to elucidate hydrodynamic chromatography of colloids in geological media. The external forces acting on colloidal particles hypothesized in the model proposed herein include inertial force, van der Waals attractive force, double layer force as well as gravitational force. The dispersion coefficient of colloids and the distribution coefficient for radionuclides with colloids are also considered as they pertain to the size of the colloid. In addition, the size distributions of colloids are utilized to investigate the effects of polydispersed colloids.

Chromatography↗

Hydrodynamic function of tilting disc prostheses and bileaflet valves in double valve replacement.

OBJECTIVE: To determine the energy loss attributable to prosthetic valve size and design in double valve replacement, energy consumption of mitral valves (size #25 to #29), of two different designs (Bjork Shiley tilting disc and Carbomedics bileaflet valves), in combination with a small (#21) and large sized (#27) aortic prosthesis, were analyzed in a flow simulator. METHODS: A pulsatile flow simulator was used to reproduce physiological ventricular dynamics and to match the input and output impedances of the human left ventricle. Hydrodynamic performance was determined as stroke work, closing work and leak work for each combination of valves at low flow (5 liters per minute) and high flow (9 liters per minute) conditions. RESULTS: At low flow no decrease of energy requirement was found with the use of a mitral valve larger than #25. At high flow the #27 and #29 mitral valves required less energy compared to a #25 mitral valve, in combination with a large aortic prosthesis. The #29 mitral prosthesis revealed similar results as the #27. With the use of a large aortic prosthesis a remarkable reduction of total flow work was shown. These results were found in both designs. In comparison of the two designs, tilting disc valves required more energy for stroke and closure of the valve, although less energy for leakage. In total, energy requirements were higher for tilting disc valves. COMMENTS: A hydrodynamic advantage for the use of a mitral valve larger than #25 was found only with the combination of a large aortic prosthesis and high flow. Hydrodynamic data favor the use of bileaflet prosthesis especially for a patient who is expected to exercise.

Aortic Valve↗

Hydrodynamic thrombectomy of hemodialysis fistulas: first clinical results.

PURPOSE: A hydrodynamic thrombectomy system was used for the treatment of recent dialysis shunt thrombosis. PATIENTS AND METHODS: Sixteen shunt thromboses in 14 patients were included in the study. There were seven polytetrafluoroethylene grafts and nine native arteriovenous fistulas. Occlusion time ranged from 6 to 48 hours, and thrombus length ranged from 4 to 40 cm. RESULTS: Thrombectomy was technically successful in 15 of 16 instances. No significant residual thrombus was found in 15 cases. In one case, half of the thrombus remained in the vessel and the procedure failed technically. One embolus to the radial artery occurred after balloon dilation following hydrodynamic thrombectomy and was removed percutaneously. Early rethrombosis within 24 hours occurred in five shunts; four more rethrombosed within 2 weeks to 3 months. Eleven shunts were available for follow-up. Cumulative patency was 41% after 6 months. CONCLUSION: Hydrodynamic thrombectomy is a promising concept for declotting of both hemodialysis grafts and native shunts and may offer an alternative to thrombolysis and surgical thrombectomy.

Adult↗

Hydrodynamic relaxation and sample concentration in field-flow fractionation using permeable wall elements.

The advantages of hydrodynamic relaxation in field-flow fractionation, in which an injected sample is driven rapidly toward its equilibrium distribution by flow, are described relative to conventional field-driven relaxation. A new concept for achieving hydrodynamic relaxation, based on the use of permeable wall elements (or frit elements) embedded in the channel walls, is introduced. Here an auxiliary substream of carrier fluid, permeating uniformly into the FFF channel near the inlet, drives the sample, entrained in its own substream, close to its equilibrium configuration. Such frit elements can also be used to enrich the sample at the outlet. Equations are derived and plots are provided for the position of the splitting plane dividing the two substreams; this position defines the strength of the hydrodynamic relaxation. Variations in shear through these frit-modified end regions are also formulated and plotted. The effects of frit elements on band broadening are discussed. It is concluded that permeable wall elements in many configurations may be broadly applicable to FFF and related methods for improved sample introduction, increased separation speed, reduced risk of sample adhesion to the wall, improved flow stability, and sample enrichment.

Chemistry Techniques, Analytical↗

Kinetic and hydrodynamic analysis of blood clotting factor V-membrane binding.

The kinetics and hydrodynamic properties of factor V-membrane interaction were characterized. Factor V bound to membranes containing acidic phospholipids with a high collisional efficiency. For membranes of 20% phosphatidyl-serine-80% phosphatidylcholine, an association rate constant of (1.13 +/- 0.10) X 10(8) M-1 s-1 was obtained. These membranes contained about 20 factor V binding sites per vesicle of 3.6 X 10(6) daltons. This association rate represented about a 30% collisional efficiency. Dissociation of factor V was measured by a fluorescence energy transfer method with a dissociation rate constant of 0.0055 s-1 at 10 degrees C. The equilibrium dissociation constant for binding to these membranes at 10 degrees C and 0.14 M ionic strength was 5 X 10(-11) M. Ionic strength, pH, calcium, and charge density in the membrane had large effects on the rate of factor V-membrane dissociation, indicating a strongly ionic interaction between protein and membrane. In contrast, the association rate was nearly insensitive to ionic strength. The membrane-binding properties were relatively unchanged after thrombin digestion of factor V or after long-term protein storage which resulted in loss of procoagulant activity. Other proteins of the prothrombinase reaction greatly decreased the rate of factor Va-membrane dissociation. At protein saturation, factor V increased the hydrodynamic radius of phospholipid vesicles by 11.4 nm. In contrast, factor Va increased the hydrodynamic vesicle radius by only about 5 nm. The mass of membrane-bound protein was comparable for both proteins.

Animals↗

Hydrodynamic diameters of RNA tumor viruses. Studies by laser beat frequency light scattering spectroscopy of avian myeloblastosis and Rauscher murine leukemia viruses.

The diffusion constants of avian myeloblastosis virus (AMV) and murine leukemia virus (MuLV) (Rauscher) suspensions in buffer and in 30% sucrose were determined by laser beat frequency light scattering spectroscopy at a series of temperatures ranging rom 5 to 25 degrees. By the use of the Stokes-Einstein equation, the following hydrodynamic diameters are calculated at 20 degrees: MuLV, 154 plus or minus 3 nm in sucrose and 145 plus or minus 7 nm in buffer; AMV, 144 plus or minus 3 nm in sucrose and 138 plus or minus 4 nm in buffer. While the diameters measured in buffer were temperature independent, the diameters measured in sucrose decreased by about 20% as the temperature was raised from 5 to 25 degrees. The concentration of virus particles in the suspensions ranged from 10 7 to 10 9 particles/ml. The absolute particle concentrations are estimated within plus or minus 30% by determining the dilution needed to reach a concentration sufficiently low that the particle number fluctuation contribution was comparable to that of the interference scattering. Particle weights of 3.9 x 10 8 daltons for MuLV and 4.0 x 10 8 daltons for AMV were calculated from the diffusion constants and from our own experimentally determined sedimentation coefficients. From these particle weights and the hydrodynamic diameters of the viruses, we calculated the per cent of the hydrodynamic volume of the viruses which could be freely penetrated by water, viz., 57% for AMV and 69% for MuLV.

Avian Leukosis Virus↗

Hydrodynamic analysis of the human progesterone receptor A-isoform reveals that self-association occurs in the micromolar range.

Human progesterone receptors exist as two functionally distinct isoforms, an 83 kDa A-receptor (PR-A) and a 99 kDa B-receptor (PR-B). The isoforms are identical except that PR-B has an additional 164 amino acids at its N-terminus. We have previously characterized the hydrodynamics and solution assembly energetics of PR-B [Heneghan, A. F., et al. (2005) Biochemistry 44, 9528-9537], and here we present an analysis of PR-A. At micromolar concentrations of the receptor, sedimentation velocity studies demonstrate that PR-A undergoes a concentration-dependent change in its sedimentation coefficient distribution, indicative of a self-associating system. Global analysis of data sets collected at multiple PR-A concentrations supports the presence of a hydrodynamically homogeneous 3.50 S monomer species in equilibrium with a 7.15 S dimer species. Sedimentation equilibrium analysis demonstrates that self-association can be rigorously described by a monomer-dimer assembly reaction and a dimerization free energy of -7.6 +/- 0.6 kcal/mol. Both the PR-A monomer and dimer are structurally asymmetric, although the extent of asymmetry is significantly decreased for the dimer, indicative of quaternary-induced hydrodynamic compaction. Limited proteolysis studies suggest that PR-A asymmetry arises from an ensemble of partially folded conformations within the N-terminal half of the molecule. Finally, comparison to our previous work on PR-B self-association energetics demonstrates that it dimerizes, under identical solution conditions, with an affinity at least 8-fold weaker than that of PR-A. Thus, residues unique to the B-isoform destabilize receptor assembly energetics. Importantly, the physical and chemical driving forces underlying isoform-specific dimerization suggest that B-unique amino acids modulate the energetics through an allosteric mechanism.

Dimerization↗

Hydrodynamic radii of native and denatured proteins measured by pulse field gradient NMR techniques.

Pulse field gradient NMR methods have been used to determine the effective hydrodynamic radii of a range of native and nonnative protein conformations. From these experimental data, empirical relationships between the measured hydrodynamic radius (R(h)) and the number of residues in the polypeptide chain (N) have been established; for native folded proteins R(h) = 4.75N (0.29)A and for highly denatured states R(h) = 2.21N (0.57)A. Predictions from these equations agree well with experimental data from dynamic light scattering and small-angle X-ray or neutron scattering studies reported in the literature for proteins ranging in size from 58 to 760 amino acid residues. The predicted values of the hydrodynamic radii provide a framework that can be used to analyze the conformational properties of a range of nonnative states of proteins. Several examples are given here to illustrate this approach including data for partially structured molten globule states and for proteins that are unfolded but biologically active under physiological conditions. These reveal evidence for significant coupling between local and global features of the conformational ensembles adopted in such states. In particular, the effective dimensions of the polypeptide chain are found to depend significantly on the level of persistence of regions of secondary structure or features such as hydrophobic clusters within a conformational ensemble.

Adhesins, Bacterial↗

Hydrodynamic dimensions, electrophoretic mobility, and stability of hydrophilic quantum dots.

Luminescent semiconductor quantum dots (QDs) have great potential for use in biological assays and imaging. These nanocrystals are capped with surface ligands (bifunctional molecules, amphiphilic polymers, phospholipids, etc.) that render them hydrophilic and provide them with functional properties. These coatings alters their hydrodynamic radii and surface charge, which can drastically affect properties such as diffusion within the cell cytoplasm. Heavy atom techniques such as transmission electron microscopy and X-ray scattering probe the inorganic core and do not take into account the ligand coating. Herein we use dynamic light scattering to characterize the hydrodynamic radius (R(H)) of CdSe-ZnS QDs capped with various hydrophilic surface coatings (including dihydrolipoic acid and amphiphilic polymers) and self-assembled QD-protein bioconjugates. Experiments were complemented with measurements of the geometric size and zeta potential using agarose gel electrophoresis and laser Doppler velocimetry. We find that the effects of surface ligands on the hydrodynamic radius and on the nanoparticle mobility are complex and strongly depend on a combination of the inorganic core size and nature and lateral extension of the hydrophilic surface coating. These properties are critical for the design of QD-based biosensing assays as well as QD bioconjugate diffusion in live cells.

Biosensing Techniques↗

Application of binary polymer system in drug release rate modulation. 2. Influence of formulation variables and hydrodynamic conditions on release kinetics.

The significance of factors such as drug solubility, polymer molecular weight, drug loading dose, compression force, and hydrodynamic conditions on drug release from a swellable hydrophilic delivery system was investigated. Hydroxypropyl methylcellulose (HPMC) and pectin were major polymeric constituents of the delivery system. Nifedipine, prednisolone, theophylline anhydrous, and diltiazem hydrochloride with solubilities of < 0.001%, <0.1%, <1%, and >50%, respectively, were used as drug models. Results show that changes in pectin:HPMC ratios, HPMC molecular weight, and hydrodynamic conditions exert notable influences on release rate and release duration from the designed system. In the case of prednisolone, drug loading up to 30% (w/w) of the matrix composition (pectin:HPMC K4M; 3:6) had no effect on zero-order release kinetics, and the delivery system was insensitive to changes in compression force (2000 to 5000 lb). For nifedipine, theophylline, and diltiazem, determination of mean dissolution time (MDT) for 50 and 80% drug release provided accurate information on release behavior. The dominating effect of matrix composition over variations in drug solubilities in controlling drug release from the delivery system was evident from similarities in dissolution profiles. It is further shown that hydrodynamic stress and intensity of fluid flow causes greater attrition at the swollen periphery and is responsible for dramatic increases in release rates. This latter observation confirms that the mechanism of drug release from this swellable system is erosion dependent. Influence of polymer molecular weight and drug solubility on release kinetics and the potential of the delivery system is discussed.

Delayed-Action Preparations↗