Viscosity at low shear and circular dichroism studies of heparin

The viscosity of heparin solution was investigated under conditions of low shear stress between 0.0193 and 0.222 dyne cm^?2, in water, in the presence of various cations (Na⁺, K⁺, Cs⁺, Mg²⁺, Ba²⁺, Cu²⁺) and at several pH's. The viscosity was found to decrease with in creasing shear stress. Shear dependence was greatest in the absence of added salts, and decreased as the ionic strength increased. Differences in viscosity in the presence of various cations appear to be related to the binding affinity of these cations to heparin. Viscosity studies of the periodate oxidation of heparin confirmed that heparin contains vicinal hydroxyl groups in its primary structure. Circular dichroism spectra of the same heparin solutions were also studied. The binding process between Cu²⁺ and heparin appears to be different from that of other divalent ions. A reduction in the pitch of the helix would qualitatively explain the conformational changes that occur on binding Cu²⁺ to heparin. These changes are reversible on removal of Cu²⁺ and replacement with Na⁺. The circular dichroism spectrum was virtually lost for periodateoxidized heparin.     

Theoretical calculation of the dielectric constant of a bilayer membrane.

The dielectric constant (epsilon) and refractive index (n) of a bilayer lipid membrane is determined from the known values of the polarizabilities of the carbon-carbon and carbon-hydrogen bonds. It is assumed that the hydrocarbon chains are hexagonally arranged in an all-trans conformation perpendicular to the plane of the membrane. The only variable in the calculation is the average separation between the chains and the theory relates epsilon to this separation. The calculation and results differ significantly from those presented in a 1968 publication by Ohki. It is shown that a thin membrane is not homogeneously polarized by the applied field. This effect is analysed and the dependence of epsilon on the membrane thickness is determined. The theoretical results are in good quantitative agreement with experimental measurements on bulk paraffins and on oriented multilayers of saturated fatty acids. The most important conclusion is that the dielectric constant for an applied field perpendicular to the membrane (which is the appropriate value for capacitance measurements) differs by only a few percent from the value for the macroscopic (bulk) liquid hydrocarbon. Thus the dielectric constant of a bilayer membrane can be approximated by the value for the appropriate bulk hydrocarbon.     

Hydration properties of DNA-lysine gels by microwave dielectric measurements as a function of temperature

Dielectric measurements by a cavity perturbation method at 10 GHz in the temperature range from-20°C to +45°C are reported for aqueous gels of herring sperm DNA in the presence of 1 or 3 lysine molecules per nucleotide. Measurements for lysine-water and DNA-water systems are also reported. The experimental results can be accounted for by the presence of interfacial water, with dielectric properties different from those of bulk water, and are analyzed in terms of a three component equation (solute molecules, interfacial water and bulk water) to calculate hydration parameters of the systems. The lysine molecule is found to coordinate a particular number of water molecules, in agreement with the literature. The specific hydration of DNA is reduced by the presence of lysine, indicating a direct interaction between the polyion and the aminoacid: a decrease to about 50% was observed at a ratio of one molecule of lysine per nucleotide. A suggestion is made that the interaction is mainly electrostatic in nature.     

Experimental evaluation of the effective dielectric constant of proteins

Chemical modifications that alter the net charge of residues in reduction-oxidation proteins influence the redox potential of the protein by changing the electrostatic potential at the redox center. If the locations of the modified charges are known, the shift in redox potential may be used to determine the effective dielectric constant for the interactions between the redox center and modified residues. From the shift in redox potential upon charge neutralization of specific lysines in the hemoprotein cytochrome c, an effective dielectric constant of approximately 50 is calculated for the electrostatic interaction between the modified lysines and heme iron in the native protein.     

A theoretical study of the dielectric constant of protein

The dielectric properties of a protein molecule were investigated by calculating a 'local dielectric constant' with the aid of normal mode analysis. This local dielectric constant was calculated from the electronic polarization of atoms and the orientational polarization of local dipoles. The former was obtained from atomic polarizations of the whole atoms in a protein molecule. The latter was determined from the fluctuation-dissipation theorem. The degree of dipole fluctuation was calculated from the positional fluctuation of each atom obtained by the normal mode analysis. Assuming a minimum volume for a continuum model, the resulting local dielectric constants ranged from 1 to 20 inside the protein.     

Circadian changes in anticoagulant effect of heparin infused at a constant rate.

Six patients with venous thromboembolism were treated with heparin, administered intravenously by a constant infusion pump. The initial daily dose of heparin was adjusted to keep the activated partial thromboplastin time, sampled at 0800, between 1.5 and 2.5 times the control level. Once that level was obtained, this dose was kept constant. Anticoagulation was thereafter measured, every four hours for 48 hours, by activated partial thromboplastin time, thrombin time, and coagulation factor Xa inhibition assay. The results of all three coagulation tests showed a circadian variation in the six patients. Maximum values were achieved at night and minimum values in the morning. These circadian variations were reproduced for two consecutive days. Differences between night and morning values reached almost 50% for activated partial thromboplastin time, 60% for thrombin time, and 40% for factor Xa inhibition assay. This circadian variation resulted from two rhythms, a circadian rhythm lasting 24 hours and an ultradian rhythm lasting 12 hours, which were detected by cosinor analysis for each coagulation test (p less than 0.01). A circadian rhythm was detected individually in most of the patients for each coagulation test (p less than 0.05). All patients had a nocturnal peak in activated partial thromboplastin time on both days. In four patients this peak exceeded the upper desired limit of activated partial thromboplastin time. These rhythms should be taken into account when evaluating the dosage of heparin to be administered.     

Modification of the erythrocyte membrane dielectric constant by alcohols

Summary Aliphatic alcohols are found to stimulate the transmembrane fluxes of a hydrophobic cation (tetraphenylarsonium, TPA) and anion (AN-12) 5–20 times in red blood cells. The results are analyzed using the Born-Parsegian equation (Parsegian, A., 1969,Nature (London) 221:844–846), together with the Clausius-Mossotti equation to calculate membrane dielectric energy barriers. Using established literature values of membrane thickness, native membrane dielectric constant, TPA ionic radius, and alcohol properties (partition coefficient, molar volume, dielectric constant), the TPA permeability data is predicted remarkably well by theory. If the radius of AN-12 is taken as 1.9 Å, its permeability in the presence of butanol is also described by our analysis. Further, the theory quantitatively accounts for the data of Gutknecht and Tosteson (Gutknecht, J., Tosteson, D.C., 1970,J. Gen. Physiol. 55:359–374) covering alcohol-induced conductivity changes of 3 orders of magnitude in artificial bilayers. Other explanations including perturbations of membrane fluidity, surface charge, membrane thickness, and dipole potential are discussed. However, the large magnitude of the stimulation, the more pronounced effect on smaller ions, and the acceleration of both anions and cations suggest membrane dielectric constant change as the primary basis of alcohol effects.     

Study of helix-coil transition of DNA by dielectric constant measurement

The thermal helix–coil transition of DNA was studied by means of dielectric constant measurements. The dielectric dispersion of native helical DNA is characterized by a large dielectric increment and a large relaxation time, whereas that of denatured coil DNA is characterized by a small dielectric increment and a small relaxation time. The dielectric dispersion of partially denatured DNA is of particular interest. At the intermediate stage of the helix–coil transition, dispersion curves which are different from either that of helix DNA or that of coil DNA appear. This is particularly pronounced for large DNA. This indicates the presence of an intermediate form of DNA. Flow birefringence measurements were carried out simultaneously. The negative birefringence of helical DNA diminishes as the helix–coil transition proceeds. However, the extinction angle remains constant, as long as it can be measured. These results indicate the absence of intermediate forms during the helix–coil transition. The discrepancy between dielectric and birefringence measurements can be resolved by assuming that the intermediate forms are not birefringent. The size distribution of native DNA and of the indicated intermediate form of DNA was studied. It is found that a logarithmic normal distribution function explains the distribution of size of DNA reasonably well.     

Effect of dielectric constant of medium on protonation equilibria of ethylenediamine

Abstract

The effect of dielectric constant of medium on protonation equilibria has been studied by determining protonation constants of ethylenediamine pH metrically in various concentrations (0–60%v/v) of acetoni-trile– and ethylene glycol–water mixtures, at an ionic strength of 0.16mol L^?1 and at 303.0 K. MINIQUAD75 computer program has been used for the calculation of protonation constants. Linear and non-linear variations of step-wise protonation constants with reciprocal of dielectric constant of the solvent mixtures have been attributed to the dominance of the electrostatic and non-electrostatic forces, respectively. The trend is explained on the basis of solute–solute and solute–solvent interactions, solvation, proton transfer processes and dielectric constants of the media.     

Lipid Bilayer Domain Fluctuations as a Probe of Membrane Viscosity

We argue that membrane viscosity, η_m, plays a prominent role in the thermal fluctuation dynamics of micron-scale lipid domains. A theoretical expression is presented for the timescales of domain shape relaxation, which reduces to the well-known η_m = 0 result of Stone and McConnell in the limit of large domain sizes. Experimental measurements of domain dynamics on the surface of ternary phospholipid and cholesterol vesicles confirm the theoretical results and suggest domain flicker spectroscopy as a convenient means to simultaneously measure both the line tension, σ, and the membrane viscosity, η_m, governing the behavior of individual lipid domains.     

Temperature dependence of dielectric constant at 10 GHz of Na-DNA gels

Permittivity ε′ and dielectric loss ε″ of aqueous Na-DNA gels have been measured at 10 GHz in the temperature interval ?15 to + 45°C. The experimental results are analyzed in terms of a three-component equation (Na-DNA, interfacial water, bulk water) and yield a value of 35 water molecules/nucleotide interacting with DNA. According to theoretical and experimental data the presence of strongly bonded and weakly bonded water is considered. The modified water exhibits a mean dielectric relaxation time two-or threefold greater than that of bulk water.     

The use of SAX-HPLC-CD as a heparin screening strategy

Heparin, a heterogeneous polysaccharide, has been widely used as an anticoagulant for decades. Recently, however, international events involving the sudden onset of allergic-type reactions following heparin administration led to numerous fatalities, and demanded the use of multiple laborious, time consuming techniques to identify an economically motivated adulterant. Using these methods cooperatively, the semi-synthetic molecule known as oversulfated chondroitin sulfate (OSCS), was found to be present at significant concentrations. Since the discovery of this adulterant, several analytical methods have been put forth or updated to advance the process of screening pharmaceutical heparins; of these, strong anion exchange high performance liquid chromatography (SAX-HPLC) methods have now become routine. In this preliminary work, we report the use of circular dichroism (CD) detection in conjunction with existing SAX-HPLC methods to quantitate various sulfated polysaccharides. The proposed strategy exploits the selectivity associated with CD detection of heparin and heparin-like polysaccharides, while taking advantage of the method's insensitivity to the use of mobile phase additives and programmed gradients. The limit of detection of heparin by CD was found to be ～0.22 mg/mL, whereas traditional UV/Vis detection yielded a detection limit of ～1.09 mg/mL. The success of CD detection varied for other polymers, however no significant modifications were made to the separations method to capitalize on the advantages of CD detection.     

The correlation of the complex dielectric constant and blood glucose at low frequency

A new needle-type sample cell was designed and produced to investigate the correlation between blood glucose and electrical parameters using an impedance analyzer. The characteristics of the measurement cells were optimized to give high sensitivity. High sensitivity complex dielectric constant measurements were obtained by calibration with several known fluids. It was observed that the values of the real (epsilon') and the imaginary (epsilon") dielectric constant increase with decreasing glucose contents in the water/glucose system, and that the value of epsilon' in hamster tail changes according to the variation in blood glucose. It is likely that there is a correlation between blood glucose and the value of epsilon', the electrical parameter.