I. Ermolina

Time domain dielectric spectroscopy study of biological systems

The main principles of time domain dielectric spectroscopy, its application to conductive systems and possible methods of electrode polarization corrections in time domain are introduced. A comprehensive theoretical and experimental study of static and dynamic dielectric properties of different biological systems including globular, and membrane proteins, hydrate water, human erythrocytes, and normal and malignant blood cells of different types is presented in the paper.

Time domain dielectric spectroscopy: An advanced measuring system

The new time domain measurement system for dielectric measurements is described. The current model is comprised of an IBM PC‐AT/486, ‘‘TDM‐2,’’ a new time domain measurement system, a set of thermostabilized sample holders, and operation and analysis software. This system is designed for use in the measurement of dielectric parameters of liquid and solid materials over the frequency range 100 kHz–10 GHz. Software consists of programs of registration, accumulation and data collection, Fourier analysis, time domain treatment, analysis software: fast and reliable nonlinear curve fitting programs to determine spectroscopic parameters and correlation analysis in time domain. The system utilizes the difference method of measurement with the registration of primary signals with multiwindow nonuniform sampling. Such a system permits the overlap of a frequency range of five orders in a single measurement.

Time domain dielectric spectroscopy with nonuniform signal sampling

The hardware and software designs for fast measurement of dielectric properties of different substances over a wide range of characteristic times (10−5–10−10 s) by means of time domain dielectric spectroscopy (TDS) are presented. The characteristic feature of the described installation is the nonuniform time sampling of measured signals, which enables one to cover a wide range of characteristic times with a single measurement. Comparative results for an aqueous solution of lysozyme obtained on the traditional apparatus and on the described one are presented.

NMR and dielectric spectroscopy investigation of protein dynamical structure.

Abstract The general approach to globular proteins dynamical structure investigation by NMR and time domain dielectric spectroscopy (TDDS) is presented. The information on macromolecular dynamical behavior in the case of these two methods is obtained in terms of correlation function and its parameters of atom motions. The interpretation of experimental results in the present work will be carried out in the framework of model-free approach which is common both for magnetic and dielectric relaxation. The lysozyme pH-dependence investigation is presented as an example.

Structure and dynamic behavior of protein molecules in solution

The results of the investigation of structure and dynamic behavior of protein in solution by time-domain dielectric spectroscopy are presented. The dipole correlation functions for myoglobin, lysozyme and RNase A solutions at different concentration, pH and temperature were obtained. The processes of the intramolecular interactions of myoglobin molecules, the pH-induced dimerization of lysozyme and thermal denaturation of RNase A are discussed.

A THz spectroscopy method for quantifying the degree of crystallinity in freeze-dried gelatin/amino acid mixtures: An application for the development of rapidly disintegrating tablets

OBJECTIVE In rapidly disintegrating tablets (RDTs) manufactured by freeze-drying processes, the % crystallinity of the freeze-dried matrix underpins the physical properties such as mechanical strength and dissolution profile. This study examines the feasibility of using terahertz pulsed spectroscopy (TPS) as an off-line tool (in the first instance) for assessing the degree of crystallinity in co-freeze dried amino-acid/gelatine mixtures. MATERIALS AND METHODS Three amino acids (l-alanine, serine and proline) were studied by TPS (in the wave number range of 3-100cm(-1)) both in the pure crystalline form and in the form of a co-freeze-dried matrix with gelatin (in weight fractions of 10:90, 30:70, 50:50, and 70:30). RESULTS In co-freeze dried proline/gelatin matrix no crystallinity was observed using TPS, whereas ~62±1% (n=3) and 72±0.5% (n=3) crystallinity was observed in the co-freeze dried alanine/gelatin matrix (50% w/w and 70% w/w, respectively). Similarly, ~41±0.5% (n=3) and ~80±0.5% (n=3) crystallinity was observed in the serine/gelatin matrix (30% w/w and 50% w/w respectively). CONCLUSION The potential for using TPS as a quantitative in-line tool for determining the degree of crystallinity in a range of complex systems such as RDTs, conventional tablets, sprayed dried micro particles, is suggested from these results.

Effect of acidic and basic pH on Thioflavin T absorbance and fluorescence.

Thioflavin T (ThT) is a fluorescent dye able to enhance significantly its fluorescence quantum yield upon binding to protein amyloids. ThT assay is widely used to detect and quantify amyloids in a variety of conditions, including solutions with different pH levels. In the present work, the effect of acidic and basic pH on the conformation of the ThT molecule and its absorption and fluorescence properties was studied. The results show that both acidic and basic pH decrease significantly the intensity of ThT absorption in the visible region and fluorescence emission intensity. Low pHs induce an immediate “all-or-nothing” decrease in the ThT signal, while in alkaline solutions the ThT signal decreases gradually over time. pH-induced signal quenching is less in the presence of glycerol or protein aggregates. Two different mechanisms are responsible for the ThT signal quenching—the ThT hydroxylation at basic pH and protonation of the nitrogen atom of the dimethylamino group at acidic pH. ThT assays should be carefully carried out at basic or acidic pH as strong pH dependence of ThT could be responsible for misinterpretation and false positive/negative experimental results. The potential unsuitability of ThT as a probe in solutions with high pH (>9) has been shown.

Dielectric spectra broadening as a signature for dipole-matrix interaction. IV. Water in amino acids solutions

In this paper, the fourth one of our series on the dielectric spectrum symmetrical broadening of water, we consider amino acid (AA) aqueous solutions. The developed 3D-trajectory is applied here to the variety of zwitterion amino acids representing both the hydrophobic and hydrophilic nature of their residues. The dipole moment of amino acids due to their zwitterion determines their interaction with the solvent and reflects mostly the dipole-matrix interactions described in our Paper I [E. Levy et al., J. Chem. Phys. 136, 114502 (2012)]. It is also shown that in the case of charged AAs at high concentrations, the shape of the 3D trajectory transforms to the pattern typical of the dipole-charge interactions that were described in our Paper III [A. Puzenko et al., J. Chem. Phys. 137, 194502 (2012)].

Dielectric Relaxation, Molecular Motion and Interprotein Interactions in Myoglobin Solution

The results of the investigation of protein molecule dynamic in solution by Time Domain Dielectric Spectroscopy are presented. The horse myoglobin solutions in wide range of concentration from 0.6% to 54% at 20 degrees C have been investigated. The result of analysis produced in the term of dipole correlation function has shown that the obtained correlation function of macromolecule motion may be presented as sum of three components corresponding to three kinds of protein motions: anisotropic intramolecular motion, anisotropic Brownian tumbling and isotropic slow motion. We suppose that the cause of protein tumbling anisotropy and the possibility to keep slow motion is the interprotein electrostatic interactions. The characteristic time of slow motion depends on the concentration of protein and perhaps is controlled by translational diffusion. The dipole moment of myoglobin calculated by the Onsager-Oncley equation is 200D for solutions less than 10% protein concentration. It is in a good agreement with the theoretical value.

Quantification of residual crystallinity in ball milled commercially sourced lactose monohydrate by thermo-analytical techniques and terahertz spectroscopy.

The quantification of crystallinity is necessary in order to be able to control the milling process. The use of thermal analysis for this assessment presents certain challenges, particularly in the case of crystal hydrates. In this study, the residual crystallinity on ball milling of lactose monohydrate (LMH), for periods up to 90min, was evaluated by thermo-analytical techniques (TGA, DSC) and terahertz spectroscopy (THz). In general, the results from one of the DSC analysis and the THz measurements agree showing a monotonous decrease in relative residual crystallinity with milling time (∼80% reduction after 60min milling) and a slight increase at the 90min time point. However, the estimates from TGA and two other methods of analyzing DSC curve do not agree with the former techniques and show variability with significantly higher estimates for crystallinity. It was concluded that, the thermal techniques require more complex treatment of the data in the evaluation of changes in crystallinity of a milled material (in particular to account for the de-vitrification and mutarotation of the material that inevitably occurs during the measurement cycle) while the analysis of THz data is more straightforward, with the measurement having no impact on the native state of the material.