Dielectric relaxation of ice in a partially crystallized poly(N-isopropylacrylamide)microgel suspension compared to other partially crystalized polymer-water mixtures.

A broadband dielectric spectroscopy study was conducted on a partially crystallized 10 wt% poly(N-isopropylacrylamide) [PNIPAM] microgel aqueous suspension to investigate the dielectric relaxation of ice in microgel suspensions. The measurements covered a frequency range of 10 mHz to 10 MHz and at temperatures ranging from 123 K to 273 K. Two distinct relaxation processes were observed at specific frequencies below the melting temperature. One is associated with the combination of the local chain motion of PNIPAM and interfacial polarization in the uncrystallized phase, while another is associated with ice. To understand the temperature-dependent behaviour of the ice relaxation process, the relaxation time of ice was compared with those observed in other frozen polymer water mixtures, including gelatin, poly-vinylpyrrolidone (PVP), and bovine serum albumin (BSA). For concentrations ≥ 10 wt%, the temperature dependence of the relaxation time of ice was found to be independent. Therefore, the study primarily focused on the 10 wt% data for easier comprehension of the ice relaxation process. It was found that the microgel and globular protein BSA had no significant effect on ice crystallization, while gelatin slowed down the crystallization process, and PVP accelerated it. To discuss the mechanism of the dielectric relaxation of ice, the trap-controlled proton transport model developed by Khamzin et al. [Chem. Phys., 2021, 541, 111040.] was employed. The model was used to discuss the dynamic heterogeneity of ice observed in this investigation, distinguishing it from the spatial heterogeneity of ice commonly discussed.

Dielectric relaxations of ice and uncrystallized water in partially crystallized bovine serum albumin-water mixtures.

To investigate the dielectric relaxations of ice in low-concentration protein-water mixtures, broadband dielectric spectroscopy measurements were performed on partially crystallized bovine serum albumin (BSA)-water mixtures with BSA concentrations of 1-10 wt% at temperatures in the range of 123-298 K. The temperature dependence of the relaxation time of ice observed in all these mixtures changes twice at TC1 (∼240 K) and TC2 (200-160 K) (TC1 > TC2), i.e., at which the apparent activation energy, Ea, changes. Below 200 K, the relaxation of ice separates as 3-4 relaxations with different TC2 and Ea values. The presence of the multiple ice relaxations is the same as that observed for the gelatin-water mixtures (T. Yasuda, K. Sasaki, R. Kita, N. Shinyashiki and S. Yagihara, J. Phys. Chem. B, 2017, 121, 2896), but the concentration dependences of TC1 and TC2 are different. The relaxation interpreted to be due to uncrystallized water in 20 wt% and 40 wt% BSA-water mixtures reported (N. Shinyashiki, W. Yamamoto, A. Yokoyama, T. Yoshinari, S. Yagihara, R. Kita, K. L. Ngai and S. Capaccioli, J. Phys. Chem. B, 2009, 113, 14448) was re-examined and concluded to be due to one of the multiple relaxations of ice.

The Clonal Population of Trichophyton equinum from Dermatophytoses of Japanese Racehorses.

Trichophyton equinum is a zoophilic dermatophyte that is frequently isolated from horse dermatophytosis and rare infections in humans. In the present study, molecular and physiological testing were performed on T. equinum isolates from dermatophytoses of Japanese racehorses to assess genotype and phenotype patterns of these strains. Comparative nucleotide sequence analysis showed that internal transcribed spacer (ITS) region sequences amplified from all Japanese isolates were 99.5% identical to T. equinum reference strains. ITS sequences amplified among the isolates were 100% (BT2) showed that isolates were 100% identical and harbored a "T" single nucleotide polymorphism at position 18. The sequences of ß-tubulin (BT2) showed that isolates were 100% identical to T. equinum reference strains. The MAT1-2 allele was detected by PCR in all seven isolates, whereas none of the isolates contained the MAT1-1 allele. All isolates grew only on Trichophyton Agar 5 and did not grow on Trichophyton Agar 1 and 4, indicating nicotinic acid requirement. These results suggest that Japanese T. equinum isolates are derived from a clonal population.

Antifungal susceptibility of dermatophytes from racehorses in Japan.

BACKGROUND: Luliconazole (LCZ) is an imidazole antifungal medication that exhibits excellent activity against dermatophytes. As a topical cream and lotion (approved for human use), LCZ has demonstrated a broad spectrum of activity against human dermatophytoses. OBJECTIVES: This is the first study to investigate the in vitro susceptibility of clinical isolates from horse dermatophytoses to LCZ. ANIMALS: No animals were used in this study. METHODS AND MATERIALS: In the present study, the in vitro susceptibilities of clinical isolates of dermatophytes to LCZ, clotrimazole (CTZ), miconazole (MCZ) and terbinafine (TRF) were investigated using the Clinical & Laboratory Standards Institute M38-A2 test. RESULTS: The minimum inhibitory concentrations (MICs) for all 16 clinical isolates of Trichophyton equinum, Microsporum equinum/canis and M. gypseum for LCZ were <0.03 mg/L. The MICs of all isolates were <0.03-0.5 mg/L for CTZ, 0.03-16 mg/L for MCZ and <0.03-1 mg/L for TRF. CONCLUSIONS: LCZ demonstrated a broad spectrum of activity against clinical isolates from horse dermatophytoses. We consider that LCZ will become the primary antifungal agent for treating horse dermatophytosis.

Electric-field penetration depth and dielectric spectroscopy observations of human skin.

BACKGROUND: The dynamic behavior of water molecules remains an important subject for understanding human skin. The change in the dynamics of water molecules from those in bulk water can be effectively observed by dielectric spectroscopy. To study water in the human skin in vivo, non-invasive and non-destructive measurements are essential. Since many unknowns remain from previous research, in this report we employ a two-layer dielectric model to evaluate the penetration depth of the electric field and use the results in measurements on human skin. MATERIALS AND METHODS: We used open-ended coaxial probes with different diameters to perform time-domain reflectometry (TDR) measurements for an acetone-Teflon double-layer model and for human skin from various parts of the body. RESULTS: The electric-field penetration depth obtained from model measurements increases with the increasing outer diameter of open-ended coaxial electrodes. For skin measurements, the relaxation strength corresponding to the water content shows a clear dependence on the epidermal thickness of the measured body parts. CONCLUSION: We determined the depth distribution of the water content of skin from results of dielectric measurements obtained using electrodes with various electric-field penetration depths. We found exponential decays with the thickness of the epidermis of each body part for several examinees. This study suggests an effective method for detailed evaluations of human skin.

Physical Meanings of Fractal Behaviors of Water in Aqueous and Biological Systems with Open-Ended Coaxial Electrodes.

The dynamics of a hydrogen bonding network (HBN) relating to macroscopic properties of hydrogen bonding liquids were observed as a significant relaxation process by dielectric spectroscopy measurements. In the cases of water and water rich mixtures including biological systems, a GHz frequency relaxation process appearing at around 20 GHz with the relaxation time of 8.2 ps is generally observed at 25 °C. The GHz frequency process can be explained as a rate process of exchanges in hydrogen bond (HB) and the rate becomes higher with increasing HB density. In the present work, this study analyzed the GHz frequency process observed by suitable open-ended coaxial electrodes, and physical meanings of the fractal nature of water structures were clarified in various aqueous systems. Dynamic behaviors of HBN were characterized by a combination of the average relaxation time and the distribution of the relaxation time. This fractal analysis offered an available approach to both solution and dispersion systems with characterization of the aggregation or dispersion state of water molecules. In the case of polymer-water mixtures, the HBN and polymer networks penetrate each other, however, the HBN were segmented and isolated more by dispersed and aggregated particles in the case of dispersion systems. These HBN fragments were characterized by smaller values of the fractal dimension obtained from the fractal analysis. Some examples of actual usages suggest that the fractal analysis is now one of the most effective tools to understand the molecular mechanism of HBN in aqueous complex materials including biological systems.

Phase Behavior of Co-Nonsolvent Systems: Poly( N-isopropylacrylamide) in Mixed Solvents of Water and Methanol.

Cloud points of poly( N-isopropylacrylamide) in aqueous mixed solvents, with methanol as the cosolvent, are experimentally measured for polymer concentrations varied up to as high as the weight fraction 0.25. They are shown to form closed loops on the ternary phase plane in the temperature region between 5 and 30 °C, and hence co-nonsolvency is complete. Miscibility loops shrink by cooling, or equivalently, they exhibit lower critical solution temperature behavior. For a fixed polymer concentration, there is a composition of the mixed solvent at which the cloud-point temperature takes the lowest value. This minimum cloud-point temperature composition of the mixed solvent turned out to be almost independent of the polymer concentration, at least within the measured dilute region below the weight fraction 0.25. On the basis of the assumption that the phase separation is closely related to the preferential adsorption of the solvents by hydrogen bonding, we employ a model solution of Flory-Huggins type, augmented with direct and cooperative polymer-solvent hydrogen bonds, to construct the ternary phase diagrams. Theoretical calculation of the spinodal curves is performed, and the results are compared with the obtained experimental cloud-point data. The effect of molecular volume of the cosolvent is also studied within the same theoretical framework. Possibility for a upper critical solution temperature co-nonsolvency to appear for cosolvents with larger molecular volume is discussed.

Ludwig-Soret effect of aqueous solutions of ethylene glycol oligomers, crown ethers, and glycerol: Temperature, molecular weight, and hydrogen bond effect.

The thermal diffusion, also called the Ludwig-Soret effect, of aqueous solutions of ethylene glycol oligomers, crown ethers, and glycerol is investigated as a function of temperature by thermal diffusion forced Rayleigh scattering. The Soret coefficient, ST, and the thermal diffusion coefficient, DT, show a linear temperature dependence for all studied compounds in the investigated temperature range. The magnitudes and the slopes of ST and DT vary with the chemical structure of the solute molecules. All studied molecules contain ether and/or hydroxyl groups, which can act as acceptor or donor to form hydrogen bonds, respectively. By introducing the number of donor and acceptor sites of each solute molecule, we can express their hydrogen bond capability. ST and DT can be described by an empirical equation depending on the difference of donor minus acceptor sites and the molecular weight of the solute molecule.

How does thermodiffusion of aqueous solutions depend on concentration and hydrophobicity?

The thermal diffusion of aqueous solutions of mono-, di-ethylene glycols, poly(ethylene glycol), methanol, and glycerol is investigated systematically as a function of concentration using the Thermal Diffusion Forced Rayleigh Scattering (TDFRS). For all investigated binary mixtures, the Soret coefficient, S(T), decays with increasing concentration of the non-aqueous component showing two regions. For aqueous solution of ethylene glycol, at a very low solute content the decay is steep, while it becomes less steep for higher solute concentration. All mixtures show a sign change of S(T) with concentration. The sign change concentration is discussed with respect to chemical structures of solute molecules and the partition coefficient, log p. It turns out that the number of hydroxyl groups plays an important role. For the investigated aqueous mixtures, we find empirical linear relations between the sign change concentration and the ratio of the number of hydroxyl groups to the number of carbon atoms as well as the partition coefficient, log p.

Glass transition of partially crystallized gelatin-water mixtures studied by broadband dielectric spectroscopy.

The glass transition of partially crystallized gelatin-water mixtures was investigated for gelatin concentrations of 40 and 20 wt. % by broadband dielectric spectroscopy (BDS) in wide frequency (10 mHz-50 GHz) and temperature (113-298 K) ranges. Three dielectric relaxation processes were clearly observed. The origin of each relaxation process was the same as that observed for partially crystallized bovine serum albumin (BSA)-water mixtures [N. Shinyashiki et al., J. Phys. Chem. B 113, 14448 (2009)]. The relaxation process at the highest frequency is originated from uncrystallized water (UCW) in the hydration shell of gelatin. Its relaxation time is almost the same as that of water in uncrystallized system; water in various binary aqueous mixtures and confined water in nanoscale region. The relaxation process at the intermediate frequency is originated from ice, and its relaxation time and strength were similar to those for the relaxation of pure ice, particularly above 240 K. The glass transition temperature Tg, is defined by BDS measurement as the temperature at which dielectric relaxation time τ, is 100-1000 s. The relaxation process at the lowest frequency, Tg is approximately 200 K, is originated from the cooperative motion of water and gelatin. This relaxation is strong and has a similar relaxation strength to that of hydrated BSA. At Tg for the relaxation process involving the cooperative motion of gelatin and water, the temperature dependence of the relaxation process of UCW crosses over from Vogel-Fulcher behavior to Arrhenius behavior with decreasing temperature. A characteristic property of the gelatin-water mixture is a change in the temperature dependence of the relaxation time of the relaxation processes of hydrated gelatin at approximately 260 K.

Dielectric Study on Supramolecular Gels by Fiber Structure Formation from Low-Molecular-Weight Gelator/Water Mixtures.

There are various types of gel materials used in a wide range of fields, and their gelation mechanisms are extremely diverse. Furthermore, in the case of hydrogels, there exist some difficulties in understanding complicated molecular mechanisms especially with water molecules interacting through hydrogen bonding as solvents. In the present work, the molecular mechanism of the structural formation of fibrous super-molecular gel by the low molecular weight gelator, N-oleyl lactobionamide/water mixture was elucidated using the broadband dielectric spectroscopy (BDS) method. The dynamic behaviors observed for the solute and water molecules indicated hierarchical structure formation processes in various time scales. The relaxation curves obtained at various temperatures in the cooling and heating processes showed relaxation processes respectively reflecting the dynamic behaviors of water molecules in the 10 GHz frequency region, solute molecules interacting with water in MHz region, and ion-reflecting structures of the sample and electrode in kHz region. These relaxation processes, characterized by the relaxation parameters, showed remarkable changes around the sol-gel transition temperature, 37.8 °C, determined by the falling ball method and over the temperature range, around 53 °C. The latter change suggested a structure formation of rod micelles appearing as precursors before cross-linking into the three-dimensional network of the supramolecular gels. These results clearly demonstrate how effective relaxation parameter analysis is for understanding the gelation mechanism in detail.

UV-Crosslinked Poly(N-isopropylacrylamide) Interpenetrated into Chitosan Structure with Enhancement of Mechanical Properties Implemented as Anti-Fouling Materials.

High-performance properties of interpenetration polymer network (IPN) hydrogels, based on physically crosslinked chitosan (CS) and chemically crosslinked poly(N-isopropylacrylamide) (PNiPAM), were successfully developed. The IPN of CS/PNiPAM is proposed to overcome the limited mechanical properties of the single CS network. In this study, the viscoelastic behaviors of prepared materials in both solution and gel states were extensively examined, considering the UV exposure time and crosslinker concentration as key factors. The effect of these factors on gel formation, hydrogel structures, thermal stabilities of networks, and HeLa cell adhesion were studied sequentially. The sol-gel transition was effectively demonstrated through the scaling law, which agrees well with Winter and Chambon's theory. By subjecting the CS hydrogel to the process operation in an ethanol solution, its properties can be significantly enhanced with increased crosslinker concentration, including the shear modulus, crosslinking degree, gel strength, and thermal stability in its swollen state. The IPN samples exhibit a smooth and dense surface with irregular pores, allowing for much water absorption. The HeLa cells were adhered to and killed using the CS surface cationic charges and then released through hydrolysis by utilizing the hydrophilic/hydrophobic switchable property or thermo-reversible gelation of the PNiPAM network. The results demonstrated that IPN is a highly attractive candidate for anti-fouling materials.

Universal Behavior of Fractal Water Structures Observed in Various Gelation Mechanisms of Polymer Gels, Supramolecular Gels, and Cement Gels.

So far, it has been difficult to directly compare diverse characteristic gelation mechanisms over different length and time scales. This paper presents a universal water structure analysis of several gels with different structures and gelation mechanisms including polymer gels, supramolecular gels composed of surfactant micelles, and cement gels. The spatial distribution of water molecules was analyzed at molecular level from a diagram of the relaxation times and their distribution parameters (τ-ß diagrams) with our database of the 10 GHz process for a variety of aqueous systems. Polymer gels with volume phase transition showed a small decrease in the fractal dimension of the hydrogen bond network (HBN) with gelation. In supramolecular gels with rod micelle precursor with amphipathic molecules, both the elongation of the micelles and their cross-linking caused a reduction in the fractal dimension. Such a reduction was also found in cement gels. These results suggest that the HBN inevitably breaks at each length scale with relative increase in steric hindrance due to cross-linking, resulting in the fragmentation of collective structures of water molecules. The universal analysis using τ-ß diagrams presented here has broad applicability as a method to characterize diverse gel structures and evaluate gelation processes.

Aberrant Water Structure Dynamics in B16 Melanoma-Bearing Mice by Time Domain Refractometry Analysis.

Living bodies comprise approximately 55-75% water to maintain homeostasis. However, little is known about the comprehensive differences in in vivo water molecule dynamics (water structure dynamics; WSD) between physiological and pathophysiological statuses. Here, we examined the WSD of ex vivo tumor tissues and organs from tumor-bearing mice with engrafted mouse malignant melanoma cells (B16-F10) in the right flanks to compare with those in healthy mice, using time domain reflectometry of dielectric spectroscopy at days 9, 11, and 14 after engrafting. The relaxation parameters of relaxation time (τ), relaxation time distribution parameter (ß), and relaxation strength (∆ε) were measured on tumor tissues and lung, liver, kidney, and skin tissues. Immediately afterward, the water contents (%) in the tumor and the other organs were calculated by measuring their weights before and after freeze-drying. Each parameter of the tumor was compared to that of pooled values of other organs in tumor-bearing (TO) and healthy mice (HO). The tumor water content temporarily increased compared to that of HO at day 11; the tumor volume was also prone to increase. In contrast, tumor tissues exhibited significantly higher values of ß close to 1 of ultrapure water and ∆ε compared to TO and HO at all times. Moreover, ß in the viscera of TO was prone to increase compared to that of HO with significantly higher levels at day 11. Conclusively, tumor-bearing mice exhibited systemically aberrant WSD, unlike healthy mice. Thus, dielectric spectroscopy in terms of WSD may provide novel pathophysiological perspectives in tumor-bearing living bodies.

First events in the coil-to-globule transition of PVME in water: An ultrafast temperature jump - time-resolved elastic light scattering study.

HYPOTHESIS: The coil-to-globule transition is an essential phenomenon in protein and polymer solutions. Late stages of such transitions, >1 µs, have been thoroughly studied. Yet, the initial ones are a matter of speculations. Here, we present the first observation of a sub-nanosecond stage of the coil-to-globule transition of poly (vinyl methyl ether), PVME, in water. EXPERIMENTS: The detection of an early stage of the coil-to-globule transition has been possible thanks to a novel experimental approach - time-resolved elastic light scattering study, following an ultrafast temperature jump. We identified a molecular process active in the observed stage of the transition with use of broadband dielectric spectroscopy. FINDINGS: In the experiment's time window, from a few ps to around 600 ps, we observed an increase in the light scattering intensity 300-400 ps after the temperature jump that heated the sample above its lower critical solution temperature (LCST). The observed time coincides with the time of segmental relaxation of PVME, determined by broadband dielectric spectroscopy in the temperature range of the LCST of the PVME/water mixture. This coincidence strongly suggests that the observed herein stage of coil-to-globule transition is the rapid formation of local nuclei along the polymer chain. Those nuclei may grow and aggregate in later stages of the process, which are out of our experimental time window.

Thermoreversible gelation of isotactic-rich poly(N-isopropylacrylamide) in water.

We report the experimentally determined phase diagram for an aqueous solution of isotactic-rich poly(N-isopropylacrylamide) (PNiPAM) composed of the sol-gel transition curve and the cloud-point curve. The meso diad content of isotactic-rich PNiPAM is 64%, and it is soluble in water at low temperatures, but undergoes a sol-to-gel transition with increasing temperature in the investigated concentration range of 1.8 wt. %-6.0 wt. %. With a further increase in temperature, the system becomes turbid. The gel formation and clouding behavior are thermally reversible. This is the first observation of thermoreversible gelation under the cloud-point temperature for an aqueous solution of PNiPAM. On the basis of the determined phase diagram, we carried out light scattering experiments to characterize the sol-gel transition behavior as a function of temperature.

Interfacial polarization of in vivo rat sciatic nerve with crush injury studied via broadband dielectric spectroscopy.

Electrical stimulation is one of the candidates for elongation-driven regeneration of damaged peripheral nerves. Different organs and tissues have an inherent cell structure and size. This leads to variation in the tissue-specific electrical properties of the frequency of interfacial polarization. Although nervous tissues have a membrane potential, the electrical reaction inside these tissues following electrical stimulation from outside remains unexplored. Furthermore, the pathophysiological reaction of an injured nerve is unclear. Here, we investigated the electrical reaction of injured and non-injured rat sciatic nerves via broadband dielectric spectroscopy. Crush injured and non-injured sciatic nerves of six 12-week-old male Lewis rats were used, 6 days after infliction of the injury. Both sides of the nerves (with and without injury) were exposed, and impedance measurements were performed at room temperature (approximately 25°C) at frequencies ranging from 100 mHz to 5.5 MHz and electric potential ranging from 0.100 to 1.00 V. The measured interfacial polarization potentially originated from the polarization by ion transport around nerve membranes at frequencies between 3.2 kHz and 1.6 MHz. The polarization strength of the injured nerves was smaller than that of non-injured nerves. However, the difference in polarization between injured and non-injured nerves might be caused by inflammation and edema. The suitable frequency range of the interfacial polarization can be expected to be critical for electrical stimulation of injured peripheral nerves.

First isolation of Trichophyton bullosum from a horse with dermatophytosis in Japan.

Trichophyton bullosum is a zoophilic dermatophyte that has been rarely isolated from horses and humans in Africa and Europe. This is the first reported isolation of T. bullosum from a horse with dermatophytosis in Japan. The isolate from a skin lesion formed a cream-colored and waxy colony that was slightly elevated in the center. Sequencing of the internal transcribe spacer region of the isolate revealed that it was 100% identical to that of T. bullosum.

Dynamics of Uncrystallized Water, Ice, and Hydrated Polymer in Partially Crystallized Poly(vinylpyrrolidone)-Water Mixtures.

In this study, we investigated the cooperative molecular dynamics of poly(vinylpyrrolidone) (PVP), ice, and uncrystallized water (UCW) in partially crystallized PVP-water mixtures by means of broadband dielectric spectroscopy. Three relaxation processes, denoted I, II, and III, were observed at temperatures ranging from immediately below the crystallization temperature (Tc) to approximately 200 K. At temperatures of 173-193 K, processes I and II cannot be distinguished. Below 168 K, process II separates into two processes: process IV at higher frequencies and process V at lower frequencies. Process I contributes to process V. In partially crystallized mixtures, process I originates from UCW in an uncrystallized phase with PVP. Process II is attributed to ice in the mixture, with a relaxation time that is 2 orders of magnitude smaller than that of pure ice. The concentration dependence of the strength of process II and the relaxation time relative to that of ice in bovine serum albumin (BSA)-water and gelatin-water mixtures strongly support this conclusion. Observation of processes IV and V indicates the presence of multiple ice relaxation processes. Process III is attributed to the α process of PVP in the uncrystallized phase in 40 and 50 wt % PVP mixtures. For mixtures with 30 wt % PVP or less, process III is attributed not only to the α process of PVP but also to interfacial polarization.

Phase transition and abnormal behavior of a nematic liquid crystal in benzene.

Differential scanning calorimetry and broadband dielectric spectroscopy measurements were performed to investigate the phase transitions in nematic liquid crystal (LC)/benzene mixtures. Noticeable changes of the crystal-isotropic phase transition and the phase transition enthalpy were observed. We also estimated the number of unfreezable benzene molecules from the LC/benzene mixtures. The crystal-isotropic phase transition temperature was described very accurately from the temperature-dependent relaxation strength, the relaxation time, and the symmetric shape parameter obtained from the fitting procedure. Surprisingly, the abnormal behavior of the isotropic phase was observed in LC/benzene mixtures that suggested the presence of local structure in the mixture, which set off the dipole moments along the long axis. The interaction between the LC molecules was also discussed from the tau-beta diagram.