Compartment modeling.
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Administration, Cutaneous↗
Biomedical subjects
M Holz
Publications and source records attributed to M Holz.
Electrical transport in a disordered medium: NMR measurement of diffusivity and electrical mobility of ionic charge carriers.
Electrical transport in porous media plays an important role in many fields of pure and applied science. The basic microscopic processes of the charge transport have attracted considerable theoretical interest for a long time. However, on a microscopic level there was up to now no experimental access to this problem. In the present paper we demonstrate, by using a suited porous system, that two combined NMR methods can offer such a first experimental access. We apply common PFG NMR methods and the special electrophoretic NMR (ENMR) technique for the measurement of self-diffusion coefficient D+ and electric mobility u+ of a cation ((C4H9)+4) in a disordered gel-like medium (Sephadex LH-20) filled with electrolyte solution. We find a, qualitatively expected, observation time-dependence of D+, but for the first time such a time-dependence is also observed for u+, which means the detection of the phenomenon of "anomalous field assisted diffusion" or "anomalous mobility." For the measurement of the short-time behavior of the mobility a new pulse sequence is presented. The time-dependent mobilities were measured at three different external electrical fields E. From the long-time behavior of D+, u+, and DH2O three independent values for the tortuosity T of the porous system could be derived. We find equality of the tortuosities T(D+) and T(u+), which represents a first experimental proof of the validity of the Einstein relation (D+ approximately u+) in a disordered medium. Finally, we discuss advantages of the possible use of "anomalous field assisted diffusion" over the commonly used "anomalous diffusion" in morphology studies by dynamic imaging in porous media.
Dextrans↗
Liposomal formulations of cyclosporin A: influence of lipid type and dose on pharmacokinetics.
PURPOSE: Liposomal formulations of Cyclosporin A (CyA)3 have been described in more than 30 publications to substitute Cremophor EL (CrEL), a triricinoleate ester of ethoxylated glycerol, as drug carrier. However, conflicting reports did not allow to draw consistent conclusions about the influence of liposomes on CyA pharmacokinetics (PK) and pharmacodynamics. METHODS: A series of liposomal CyA-formulations with varying liposome composition and lipid dose but constant CyA dose was compared in rats. Data were analysed with a PK-model taking into account the varying volume of distribution with the varying lipid concentration in blood. RESULTS: Surface properties and lipid type of liposomes are not important PK predictors of liposomal CyA, at least for small dosages of liposomes. Rather, the absolute lipid amount and the lipophilicity of cyclosporins are critical factors influencing the PK of liposomal CyA. The higher the concentration of lipid in blood and the greater the lipophilicity of cyclosporin is, the higher are the concentrations of CyA in blood. CONCLUSIONS: These relations may explain the inconsistent literature results. Together with earlier observations from our group the above findings indicate, that CyA is not caged in the liposomal membranes. Reports in literature, which claim lower clearance and a lower volume of distribution of CyA in obese rats compared to lean rats, support our assumption about the involved mechanisms. A semi-quantitative model of CyA distribution is presented, which points to the variable free fraction of CyA in plasma as the crucial factor for all previously reported phenomena in liposomal CyA formulations.
Animals↗
Substitution of amino acids Asp-85, Asp-212, and Arg-82 in bacteriorhodopsin affects the proton release phase of the pump and the pK of the Schiff base.
Photocycle and flash-induced proton release and uptake were investigated for bacteriorhodopsin mutants in which Asp-85 was replaced by Ala, Asn, or Glu; Asp-212 was replaced by Asn or Glu; Asp-115 was replaced by Ala, Asn, or Glu; Asp-96 was replaced by Ala, Asn, or Glu; and Arg-82 was replaced by Ala or Gln in dimyristoylphosphatidylcholine/3-[(3-cholamidopropyl)dimethylammonio]-1- propanesulfonate micelles at pH 7.3. In the Asp-85----Ala and Asp-85----Asn mutants, the absence of the charged carboxyl group leads to a blue chromophore at 600 and 595 nm, respectively, and lowers the pK of the Schiff base deprotonation to 8.2 and 7, respectively, suggesting a role for Asp-85 as counterion to the Schiff base. The early part of the photocycles of the Asp-85----Ala and Asp-85----Asn mutants is strongly perturbed; the formation of a weak M-like intermediate is slowed down about 100-fold over wild type. In both mutants, proton release is also slower but clearly precedes the rise of M. The amplitude of the early (less than 0.2 microseconds) reversed photovoltage component in the Asp-85----Asn mutant is very large, and the net charge displacement is close to zero, indicating proton release and uptake on the cytoplasmic side of the membrane. The data suggest an obligatory role for Asp-85 in the efficient deprotonation of the Schiff base and in the proton release phase, probably as proton acceptor. In the Asp-212----Asn mutant, the rise of the absorbance change at 410 nm is slowed down to 220 microsecond, its amplitude is small, and the release of protons is delayed to 1.9 ms. The absorbance changes at 650 nm indicate perturbations in the early time range with a slow K intermediate. Thus Asp-212 also participates in the early events of charge translocation and deprotonation of the Schiff base. In the Arg-82----Gln mutant, no net transient proton release was observed, whereas, in the Arg-82----Ala mutant, uptake and release were reversed. The pK shift of the purple-to-blue transition in the Asp-85----Glu, Arg-82----Ala, and Arg-82----Gln mutants and the similarity in the photocycle and photoelectrical signals of the Asp-85----Ala, Asp-85----Asn, and Asp-212----Asn mutants suggest the interaction between Asp-85, Arg-82, Asp-212, and the Schiff base as essential for proton release.
Arginine↗
Aspartic acid-96 is the internal proton donor in the reprotonation of the Schiff base of bacteriorhodopsin.
Above pH 8 the decay of the photocycle intermediate M of bacteriorhodopsin splits into two components: the usual millisecond pH-independent component and an additional slower component with a rate constant proportional to the molar concentration of H+, [H+]. In parallel, the charge translocation signal associated with the reprotonation of the Schiff base develops a similar slow component. These observations are explained by a two-step reprotonation mechanism. An internal donor first reprotonates the Schiff base in the decay of M to N and is then reprotonated from the cytoplasm in the N----O transition. The decay rate of N is proportional to [H+]. By postulating a back reaction from N to M, the M decay splits up into two components, with the slower one having the same pH dependence as the decay of N. Photocycle, photovoltage, and pH-indicator experiments with mutants in which aspartic acid-96 is replaced by asparagine or alanine, which we call D96N and D96A, suggest that Asp-96 is the internal proton donor involved in the re-uptake pathway. In both mutants the stoichiometry of proton pumping is the same as in wild type. However, the M decay is monophasic, with the logarithm of the decay time [log (tau)] linearly dependent on pH, suggesting that the internal donor is absent and that the Schiff base is directly reprotonated from the cytoplasm. Like H+, azide increases the M decay rate in D96N. The rate constant is proportional to the azide concentration and can become greater than 100 times greater than in wild type. Thus, azide functions as a mobile proton donor directly reprotonating the Schiff base in a bimolecular reaction. Both the proton and azide effects, which are absent in wild type, indicate that the internal donor is removed and that the reprotonation pathway is different from wild type in these mutants.
Aspartic Acid↗
Replacement of aspartic acid-96 by asparagine in bacteriorhodopsin slows both the decay of the M intermediate and the associated proton movement.
The photocycle, electrical charge translocation, and release and uptake of protons from the aqueous phase and release and uptake of protons from the aqueous phase were investigated for bacteriorhodopsin mutants with aspartic acid-96 replaced by asparagine or glutamic acid. At neutral pH the main effect of the Asp-96----Asn mutation is to slow by 2 orders of magnitude the decay of the M intermediate and the concomitant charge displacement associated with the reprotonation of the Schiff base from the cytoplasmic side of the membrane. The proton uptake measured with the indicator dye pyranine is likewise slowed without affecting the stoichiometry of proton pumping. The corresponding results for the Asp-96----Glu mutant, on the other hand, are very close to those for the wild-type protein. These results provide a kinetic explanation for the fact that at pH 7 and saturating light intensities the steady-state proton pumping is almost abolished in the Asp-96----Asn mutant but is close to normal in the Asp-96----Glu mutant. Thus, the pump is simply turning over much more slowly in the Asp-96----Asn mutant. The time constants of the decay of M and the associated charge translocation increase strongly with increasing pH for the Asp-96----Asn mutant but are virtually pH-independent for the Asp-96----Glu mutant and wild-type bacteriorhodopsin. At pH 5 the M decay of the Asp-96----Asn mutant is as fast as for wild type. These results suggest that Asp-96 serves as an internal proton donor in the proton-uptake pathway from the cytoplasm to the Schiff base.
Asparagine↗
NMR tomography for the visualization of the diffusion of Gd-DTPA in agar-gel and in brain tissue--in vitro studies.
Serial magnetic resonance images of a diffusion model and of brain specimens have been performed to study the diffusion behavior of a paramagnetic complex (Gd-DTPA). The model consisted of solidified agar gel with layered Gd-DTPA solution on top of the gel. Corresponding studies have been performed with the brain specimen to visualize the penetration of the marker through the extracellular space into the brain tissue. Diffusion coefficients for the agar gel and the brain tissue model have been calculated. Magnetic resonance tomography proves to be a promising method for the study of complicated diffusion processes of paramagnetic complexes in biological tissue.
Agar↗
Influence of NMR measuring sequences and gradient variation on signals from flowing systems. Magnetic resonance tomography on experimental models.
The influence of flow on MR tomography images and signal intensities has been studied using experimental model tubes and an aqueous NiCl2 solution having the same relaxation time as human blood. We applied the inversion recovery (IR) and the spin-echo (SE) sequence on a .15T MR tomography system. The influence of RF pulse distance (tau) in the IR and SE experiment as well as the influence of magnetic z-gradient strength on the flow images has been investigated. IR images revealed that signals from flowing systems recover more rapidly due to influx of non-inverted longitudinal magnetization into the scan slice. SE images in presence of flow are characterized by signal intensity loss with increasing time caused by the outflow and dephasing of transverse magnetization. With increasing strength of the z-gradient, the MR signals of flowing fluids decrease drastically. Thus for detection of flow, all the above mentioned parameters are of importance.
Blood↗
Spatio-temporal structure of migrating chemotactic band of Escherichia coli. I. Traveling band profile.
We developed a rapid-scanning, light-scattering densitometer by which extensive measurements of band migration speeds and band profiles of chemotactic bands of Escherichia coli in motility buffer both with and without serine have been made. The purpose is to test the applicability of the phenomenological model proposed by Keller and Segel (J. Theor. Biol. 1971. 30:235) and to determine the motility (mu) and chemotactic (delta) coefficients of the bacteria. We extend the previous analytical solution of the simplified Keller-Segel model by taking into account the substrate diffusion which turns out to be significant in the case of oxygen. We demonstrate that unique sets of values of mu and delta can be obtained for various samples at different stages of migration by comparing the numerical solution of the model equation and the experimental data. The rapid-scanning technique also reveals a hitherto unobserved time-dependent fine structure in the bacterial band. We give a qualitative argument to show that the fine structure is an example of the dissipative structure that arises from a nonlinear coupling between the bacterial density and the oxygen concentration gradient. Implications for a further study of the dissipative structure in testing the Keller-Segel model of chemotaxis are briefly discussed.
Chemotaxis↗
Quasi-elastic light scattering from migrating chemotactic bands of Escherichia coli.
We report the observation of migrating chemotactic bands of Escherichia coli in a buffer solution. The temporal development of the bacterial density profile is observed by the scattered light intensity as the band migrates through a stationary laser beam. We have made a preliminary analysis of the observed band profile with help of the Keller-Segel theory. The model accounts for only some aspects of the observed time evolution of the density profile. The microscopic motility characteristics of the E. coli in the band are simultaneously studied by photon correlation. The measured correlation functions are analyzed to obtain the spatial dependence of the half-width within the band. A simple analytical model is proposed to account for the contribution of the twiddle motion to the correlation function. By analyzing the correlation function as a superposition of straight-line and twiddle motions, we obtain a satisfactory agreement between the theory and the measured angular dependence of the line shape. As a consequence we are able to extract a parameter beta, which measures the average fraction of twiddling bacteria in the center of the band at a given time.
Cell Movement↗
Medical application of photon correlation spectroscopy.
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Bacterial Physiological Phenomena↗
Raman studies of the conformation of the basic pancreatic trypsin inhibitor.
The vibrational Raman spectra of the basic pancreatic trypsin inhibitor in aqueous solution, as lyophilized powder and in a single crystal and presented. The thermal stability of this protein is demonstrated by the fact that minor alterations in the spectrum, mainly in the amide III band near 1260 cm-1, occur in the solution spectrum only at temperatures above 75 degrees C. No significant spectral changes appear when the pH value of the solution is varied in the range from 1.5 to 8.7. The distinct differences of the powder spectrum compared to that of the solution, show that lyophilization causes appreciable conformational changes both in the main-chain and in the side-chains. A difference in main chain conformation of the basic pancreatic trypsin inhibitor in single crystal and in solution is suggested by different amide III frequencies.
Amino Acids↗
Raman studies on native, reduced, and modified basic pancreatic trypsin inhibitor.
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Disulfides↗
Does consumption of beer, alcohol, and bitter substances affect bitterness perception?
The relation between consumption of beer, alcohol, and bitter substances was investigated by comparing 20 each of male adults, low users ( < 720 ml/week) and high users ( > 3.61/week) of beer for sensitivity to and like/dislike for the bitterness of isohumulones. Intake of beer, alcohol, and bitter substances was measured with a food frequency questionnaire and a 14-day recall journal. Intake of alcohol (including alcohol from sources other than beer) and bitter substances was higher for high users of beer than for low users. The average bitterness units (BUs) in the beers consumed by the two groups did not differ, indicating that high users do not consume more bitter beers than low users. Detection thresholds for isohumulones in water did not differ between the two groups, but high users had higher difference thresholds for isohumulones in beer (p < 0.10). In time-intensity measurements of bitterness in beer, low users of beer recorded significantly higher maximum intensities (p < 0.001) and shorter total durations (p < 0.001) than high users, with no difference between the two groups for time to maximum intensity and area under the TI curve. There was no significant difference between the two groups for BUs of beers produced by ad lib mixing of low- and high-bitterness beers, or for mean hedonic ratings of various concentrations of isohumulones in beer. These results suggest that intakes of beer, alcohol, and bitter substances are not major determinants of taste responses to the bitterness of isohumulones.
Adult↗
Electrophoretic NMR studies of electrical transport in fluid-filled porous systems.
An NMR technique is described which allows the observation of ionic charge carriers moving in the electric field within a porous system saturated with electrolyte solution. This method, which was recently developed in our laboratory, gives experimental access to the study of electric transport in disordered media on a microscopic level and offers new potential for morphology studies. We performed 1H NMR PFG self-diffusion measurements on ions combined with ionic drift velocity measurements by electrophoretic NMR (ENMR), each as a function of observation time Delta. In this way we obtained time-dependent self-diffusion coefficients D(+/-) (Delta) and time-dependent electric mobilities mu(+/-) (Delta) of polyatomic cations and anions in porous media. The porous media used were gels and glass bead packs. From the behaviour of D(+/-) (Delta) and mu(+/-) (Delta) at long observation times the tortuosities T(p) (D(+/-)) and T(p) (mu(+/-)) are derived, allowing a direct experimental check of the validity of the Einstein relation (D(+/-) is proportional to mu(+/-)) in a disordered medium. The tortuosities obtained via the diffusivity of ions are compared with those obtained via the diffusivity of water molecules. We also make a first attempt to derive the specific surface S/V(p) from the time-dependence of the ionic mobility at short observation times and discuss possible advantages of those measurements in morphology studies of porous media.
Ion Transport↗