Ultrasound responsive block copolymer micelle of poly(ethylene glycol)–poly(propylene glycol) obtained through click reaction

The well-defined amphiphilic poly(ethylene glycol)-block-poly(propylene glycol) copolymer containing 1, 2, 3-triazole moiety and multiple ester bonds (PEG-click-PPG) was prepared by click reaction strategy. The PEG-click-PPG copolymer can self-assemble into spherical micelles in aqueous solution. It is found that high intensity focused ultrasound (HIFU) can open the copolymer PEG-click-PPG micelles and trigger the release of the payload in the micelle. The multiple ester bonds introduced in the junction point of the copolymer chain through click reactions were cleaved under HIFU, and leads to the disruption of the copolymer micelle and fast release of loaded cargo. The click reaction provides a convenient way to construct ultrasound responsive copolymer micelles with weak bonds.     

Biodegradable self-assembled PEG-PCL-PEG micelles for hydrophobic drug delivery,part 2: in vitro and in vivo toxicity evaluation

Polymeric micelles, prepared by self-assembly of biodegradable poly(ethylene glycol)-poly(ε-caprolactone)-poly(ethylene glycol) (PEG–PCL–PEG, PECE) copolymer in aqueous solution, were proved to be a potential carrier for hydrophobic drug honokiol in our previous contribution. In this study, the safety of blank PECE micelles was evaluated in vitro and in vivo before its further application in biomedical field. The average particle size of obtained micelle was 83.47 ± 0.44 nm, and polydisperse index was 0.27 ± 0.01. Also, the zeta potential of prepared micelles was about −0.41 ± 0.02 mV. Otherwise, cytotoxicity of PECE micelles was evaluated by cell viability assay using L929 cells, and in vitro hemolytic test was also performed. In vivo acute toxicity evaluation and histopathological study of PECE micelles were conducted in BALB/c mice by intravenous administration. Furthermore, serum chemistry profile and complete blood count test were performed. In acute toxicity test, the mice were observed continuously for 7 days. For histopathological study, samples including heart, liver, spleen, lung, and kidneys were histochemical prepared and stained with hematoxylin-eosin (H&E). No mortality or significant signs of acute toxicity was observed during the whole observation period, and there is no significant lesion to be shown in histopathological study of major organs. The maximal tolerance dose of PECE micelles (100 mg/mL) by intravenous administration was calculated to be higher than 10 g/kg body weight (b.w.). The results indicated that the obtained PECE micelles was non-toxic after intravenous administration, and could be a safe candidate for hydrophobic drug delivery system.     

Polymeric nano-micelles as drug carrier using polyethylene glycol and polytrimethylene carbonate linear and star-shaped block copolymer

The polymerization of trimethylene carbonate (TMC) in the presence of HCl · Et₂O via activated monomer mechanism was performed to synthesize linear and star-shaped block copolymers composed of polyethyleneglycol (PEG) and poly(trimethylene carbonate) (PTMC). The obtained PTMCs had molecular weights close to the theoretical values calculated from the TMC to PEG molar ratios and exibited monomodal GPC curves. We successfully prepared PEG and PTMC linear and star-shaped block copolymers by activated monomer mechanism. The characterization for formation of micelle of block copolymers in an aqueous phase was carried out by using NMR, dynamic light scattering (DLS), AFM and fluorescence techniques (FL). The block copolymers gave micelles with a critical micelle concentration (CMC) ranging 1.88 × 10⁻²–3.09 × 10⁻³ mg/mL depended on the molecular shape of block copolymers. The diameters of micelles, measured by DLS, were 100–250 nm. Most micelles exhibited a spherical shape in AFM.     

Thioflavin T Displays Enhanced Fluorescence Selectively Inside Anionic Micelles and Mammalian Cells

Thioflavin T (ThT) has been widely employed to detect amyloid fibrils in tissues and recently in presence of SDS micelles. However, the contribution of membranes or micelles to ThT fluorescence has never been investigated. In this paper, we show for the first time that the anionic micellar microenvironment of SDS has a profound impact on the absorption and fluorescence spectra of ThT in sharp contrast to cationic (CTAB) and neutral micelles (Triton X-100 & Tween 20). Unlike CTAB or Triton X-100 or Tween 20 micelles, formation of SDS micelles shifts the λ_max for ThT absorption from 412 nm in buffer to 428 nm inside the micelle, with a 28% increase in the peak molar absorptivity and a ∼13 fold increase in ThT fluorescence (λ_max = 489 nm). Extending these observations to cell plasma membranes, we show that ThT can quickly enter and appear selectively fluorescent inside mammalian cells like BHK21 and HT29, against a dark background owing to negligible fluorescence from free ThT in aqueous medium. The above results suggest that ThT can be a useful probe for live cell imaging and for selectively labeling micelles on the basis of the charge in the polar headgroup. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

The effects of power ultrasound (24 kHz) on the electrochemical reduction of CO2 on polycrystalline copper electrodes

The electrochemical CO₂ reduction reaction (CO2RR) on polycrystalline copper (Cu) electrode was performed in a CO₂-saturated 0.10 M Na₂CO₃ aqueous solution at 278 K in the absence and presence of low-frequency high-power ultrasound (f = 24 kHz, P_T ~ 1.23 kW/dm³) in a specially and well-characterized sonoelectrochemical reactor. It was found that in the presence of ultrasound, the cathodic current (I_c) for CO₂ reduction increased significantly when compared to that in the absence of ultrasound (silent conditions). It was observed that ultrasound increased the faradaic efficiency of carbon monoxide (CO), methane (CH₄) and ethylene (C₂H₄) formation and decreased the faradaic efficiency of molecular hydrogen (H₂). Under ultrasonication, a ca. 40% increase in faradaic efficiency was obtained for methane formation through the CO2RR. In addition, and interestingly, water-soluble CO₂ reduction products such as formic acid and ethanol were found under ultrasonic conditions whereas under silent conditions, these expected electrochemical CO2RR products were absent. It was also found that power ultrasound increases the formation of smaller hydrocarbons through the CO2RR and may initiate new chemical reaction pathways through the sonolytic di-hydrogen splitting yielding other products, and simultaneously reducing the overall molecular hydrogen gas formation.     

Ultrasonic stimulation of the brain to enhance the release of dopamine – A potential novel treatment for Parkinson’s disease

Parkinson’s disease (PD) is characterized by the decrease of dopamine (DA) production and release in the substantia nigra and striatum regions of the brain. Transcranial ultrasound has been exploited recently for neuromodulation of the brain in a number of fields. We have stimulated DA release in PC12 cells using low-intensity continuous ultrasound (0.1 W/cm² − 0.3 W/cm², 1 MHz), 12 h after exposure at 0.2 W/cm², 40 s, the amount of DA content eventually increased 78.5% (p = 0.004). After 10-day ultrasonic treatment (0.3 W/cm², 5 min/d), the DA content in the striatum of PD mice model restored to 81.07% of the control (vs 43.42% in the untreated PD mice model). In addition to this the locomotion activity was restored to the normal level after treatment. We suggest that the low intensity ultrasound-induced DA release can be attributed to a combination of neuron regeneration and improved membrane permeability produced by the mechanical force of ultrasound. Our study indicates that the application of transcranial ultrasound applied below FDA limits, could provide a candidate for relatively safe and noninvasive PD therapy through an amplification of DA levels and the stimulation of dopaminergic neuron regeneration without contrast agents.     

Effect of thermosonication pre-treatment on mung bean (Vigna radiata) and white kidney bean (Phaseolus vulgaris) proteins: Enzymatic hydrolysis,cholesterol lowering activity and structural characterization

Protein hydrolysates have attained great attention due to a good nutritive food ingredient and higher biological activities. In this study, thermosonication, ultrasound and heat were used as a pre-treatment to obtain (<3KDa) hydrolysate from mung bean and white kidney bean to understand the mechanism of cholesterol absorption into micelle and inhibition of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG-CoA) activity. Size exclusion high performance liquid chromatography (SE-HPLC) results of mung bean showed that the concentration of peptides (0.5KDa-1KDa and 1-3KDa) in the hydrolysate were significantly (p < 0.05) increased after thermosonication while, the peptides concentration (1-3KDa) in white kidney bean was significantly (p < 0.05) decreased. Thermosonication of mung bean hydrolysate exhibited higher inhibition of cholesterol solubilization, hydrophobicity and antioxidant activities. In addition, there was no difference observed in HMG-CoA activity and hydrophobicity between ultrasound alone and ultrasound combined with heat i.e. thermosonication treated hydrolysate of white kidney bean. Changes in secondary and tertiary structures were also analyzed under different processing conditions with maximum change due to thermosonication. Results indicated that mung bean hydrolysate had a great potential for inhibition of cholesterol synthesis and its solubility in the micelle, antioxidant activity and also convinced for its application in food and nutraceutical industries.     

Enhancement of artificial promoter activity by ultrasound-induced oxidative stress

We previously developed artificial promoters that were activated in response to X-ray irradiation. Sonication with 1.0 MHz ultrasound that causes intracellular oxidative stress was found to activate some of these promoters though to lesser degrees. The most sensitive one among these promoters showed intensity- and duration-dependent activations by sonication. In addition, its activation by sonication was attenuated when N-acetyl cysteine was present, suggesting the involvement of intracellular oxidative stress in the activation mechanism. Improved promoters for sensitivity to X-ray irradiation were also found more sensitive to sonication. The most improved one showed 6.0 fold enhancement after sonication with 1.0 MHz ultrasound at 1.0 W/cm² for 60 s. This enhancement was also attenuated with the presence of N-acetyl cysteine. When stably transfected HeLa cells with the most sensitive promoter were transplanted on to mice and sonicated, luciferase activity by the promoter increased to 1.35 fold in average though it was not statistically significant compared to control. Although gene regulation in vivo by sonication was not clear, this is the first report on artificially constructed promoters responsive to ultrasound.     

Measurement of microstructural changes promoted by ultrasound application on plant materials with different porosity

This research investigated the effects of ultrasound application (192 ± 6 W/L) on the microstructure of vegetables/fruits with different porosities, cell sizes and patterns (eggplants, beetroots, and apples), submitted to an immersion treatment in different liquids: distilled water, citric acid (1% w/v), and the vegetable/fruit juice, at 25 °C during 5 min. The ultrasound application did not significantly (p > 0.05) affect the size of the cells of the most porous material (eggplant) compared to the samples immersed without ultrasound assistance. The apple samples (with a middle-high porosity and the largest cells) were the most affected by ultrasound application. The median cell areas of samples treated with ultrasound in water and apple juice were 26 and 20% larger than those of samples treated without ultrasound, mainly because of cell wall disruption which caused the cells to merge into bigger clusters, but no effect was observed with the citric acid. Ultrasound application significantly (p < 0.05) increased the median cell area of the less porous raw matter (beetroot) only when the treatment was carried out in the vegetable juice (cells were 26% larger after treatment assisted with ultrasound than without it). Thus, the effects of ultrasound differ in materials with initially different characteristics.     

Ultrasound-assisted transesterification of soybean oil using low power and high frequency and no external heating source

In this work, high frequency and low power ultrasound without external heating source and mechanical stirring in biodiesel production were studied. Transesterification of soybean oil with methanol and catalyzed by KOH was investigated using ultrasound equipment and ultrasonic transducer. The effect of ultrasonic output power (3 W–9 W), ultrasonic frequency (1 MHz and 3 MHz), and alcohol to oil molar ratio (6:1 and 8:1) have been investigated. The increase in ultrasonic power provided higher conversion rates. In addition, higher conversion rates were obtained by increasing the ultrasonic frequency from 1 MHz to 3 MHz (48.7% to 79.5%) for the same reaction time. Results also indicate that the speed of sound can be used to evaluate the produced biodiesel qualitatively. Further, the ultrasound system presented electric consumption (46.2 W∙h) four times lower than achieved using the conventional method (211.7 W∙h and 212.3 W∙h). Thus, biodiesel production using low power ultrasound in the MHz frequency range is a promising technology that could contribute to biodiesel production processes.     

Structural Transition in the Micellar Assembly: A Fluorescence Study

Structural transition can be induced in charged micelles by increasing the ionic strength of the medium. Thus, spherical micelles of sodium dodecyl sulfate (SDS) that exist in water at concentrations higher than the critical micelle concentration assume an elongated rod-like structure in the presence of an increased electrolyte concentration. This is known as sphere-to-rod transition. In this paper, we characterize the change in organization and dynamics that is accompanied by the salt-induced sphere-to-rod transition in SDS micelles using wavelength-selective fluorescence and other steady-state and time-resolved fluorescence parameters. Since the change in micelle organization during such structural transition may not be limited to one region of the micelle, we have selectively introduced fluorophores in two distinct regions of the micelle. We used two probes, N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)-1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine (NBD-PE) and 25-[N-[(7-nitrobenz-2-oxa-1,3-diazol-4-yl)-methyl]amino]-27-norcholesterol (NBD-cholesterol), for monitoring the two regions of the micelle. NBD-PE monitors the interfacial region of the micellar assembly, while NBD-cholesterol acts as a reporter for the deeper regions of the micellar interior. Our results show that wavelength-selective fluorescence, in combination with other fluorescence parameters, offers a powerful way to monitor structural transitions induced in charged micelles. These results could be significant to changes in membrane morphology observed under certain physiological conditions.     

Sonocatalytic injury of cancer cells attached on the surface of a nickel–titanium dioxide alloy plate

The present study demonstrates ultrasound-induced cell injury using a nickel–titanium dioxide (Ni–TiO₂) alloy plate as a sonocatalyst and a cell culture surface. Ultrasound irradiation of cell-free Ni–TiO₂ alloy plates with 1 MHz ultrasound at 0.5 W/cm² for 30 s led to an increased generation of hydroxyl (OH) radicals compared to nickel–titanium (Ni–Ti) control alloy plates with and without ultrasound irradiation. When human breast cancer cells (MCF-7 cells) cultured on the Ni–TiO₂ alloy plates were irradiated with 1 MHz ultrasound at 0.5 W/cm² for 30 s and then incubated for 48 h, cell density on the alloy plate was reduced to approximately 50% of the controls on the Ni–Ti alloy plates with and without ultrasound irradiation. These results indicate the injury of MCF-7 cells following sonocatalytic OH radical generation by Ni–TiO₂. Further experiments demonstrated cell shrinkage and chromatin condensation after ultrasound irradiation of MCF-7 cells attached on the Ni–TiO₂ alloy plates, indicating induction of apoptosis.     

Xenon prevents cellular damage in differentiated PC-12 cells exposed to hypoxia

Background  

The neuroprotective effect of xenon has been demonstrated for glutamatergic neurons. In the present study it is investigated if dopaminergic neurons, i.e. nerve-growth-factor differentiated PC-12 cells, are protected as well against hypoxia-induced cell damage in the presence of xenon.     

Sol-gel transition in worm-like micelles

Summary The statical and dynamical properties of lecithin/H₂O/cyclohexane cylindrical reverse micelles are investigated as a function of lecithin volume fraction, ϕ, and temperature at fixed water/lecithin ratio,w ₀. The viscosity data are well fitted by the Cates model when the breathing mode of micelles is taken into consideration, overlapping with the breaking and reforming mechanisms. We present some results from Brillouin-scattering experiment, performed across the sol-gel transition. In order to explain the experimentally observed ϕ-dependence of the hyperacoustic parameters, a mechanical model was developed from which the ϕ-dependence of the micelle size distribution was obtained. From a comparison with the viscosity data the entanglement length was estimated. Furthermore some new results from an incoherent quasi-elastic neutron scattering experiment are presented. The whole body of the experimental results suggests for a sol-gel transition triggered by topologic phenomena. When the lecithin volume fraction increases, the kinetic equilibrium between the breaking and reforming mechanisms of the micelles shifts the mean micelle length towards higher values and the entanglement of the micelles becomes highly favourable. The obtained results are discussed and compared with other findings in literature. Paper presented at the I International Conference on Scaling Concepts and Complex Fluids, Copanello, Italy, July 4–8, 1994.     

Positive effects and mechanism of ultrasound on chitin preparation from shrimp shells by co-fermentation

The objective of this study is to explore the effect and mechanism of ultrasound on chitin extraction from shrimp shells powder (SSP) by the co-fermentation of Bacillus subtilis and Acetobacter pasteurianus. After pre-treating the SSP with high-intensity ultrasound (HIU) at 800 W, the protease activity in the fermentation solution reached 96.9 U/mL on day 3, which was significantly higher than for SSP that had not been pre-treated with ultrasound (81.8 U/mL). The fermentation time of the chitin extraction process was 5.0 d without ultrasound pre-treatment, while it was shortened to 4.5 d when using ultrasound at 800 W to treat SSP. However, there were no obvious differences when we applied ultrasound at low power (200 W, 400 W). Furthermore, chitin purified from shrimp shells pre-treated with HIU at 800 W exhibited lower molecular weight (11.2 kDa), higher chitin purity (89.8%), and a higher degree of deacetylation (21.1%) compared to SSP with no ultrasound pre-treatment (13.5 kDa, 86.6%, 18.5%). Results indicate that HIU peels off the protein/CaCO₃ matrix that covers the SSP surface. About 9.1% of protein and 4.7% of Ca²⁺ were released from SSP pre-treated with HIU at 800 W. These figures were both higher than with no ultrasound pre-treatment (4.5%, 3.2%). Additionally, the amount of soluble protein extracted from SSP through HIU at 800 W was 50% higher than for the control sample. SDS-PAGE analysis indicated that the soluble protein was degraded to the micromolecule. It also revealed that HIU (600, 800 W) induced the secondary and tertiary structure destruction of protein extracted from SSP. In conclusion, HIU-induced degradation and structural damage of protein enhances the protein/CaCO₃ matrix to be peeled off from SSP. Also, in the co-fermentation process, an increase of protease activity further accelerates deproteinization.     

Nonradiative energy transfer in carbocyanine dye compositions inside surfactant micelles

Utilizing forced concentration of carbocyanine dye molecules DiI and DiD within a surfactant micelle, we have detected nonradiative transfer of electronic excitation energy for low initial dye concentrations in the aqueous micellar solution. Luminescence and fluorescence anisotropy decay studies allowed us to establish that in the micelles, the hydrophilic head of the dye is in contact with the water while the C18H37 groups are located in the hydrocarbon core of the micelle. We show that due to the structural features of the DiD acceptor molecule, rotational diffusion of the acceptor in the micelle is more free than for the donor DiI, the motion of which is constrained in the micelle. __________ Translated from Zhurnal Prikladnoi Spektroskopii, Vol. 73, No. 2, pp. 152–157, March–April, 2006.     

Synergetic decolorization of azo dyes using ultrasounds,photocatalysis and photo-fenton reaction

In the present work, ultrasound irradiation, photocatalysis with TiO₂, Fenton/Photo-Fenton reaction, and the combination of those techniques were investigated for the decolorization of industrial dyes in order to study their synergy. Three azo dyes were selected from the weaving industry. Their degradation was examined via UV illumination, Fenton and Photo-Fenton reaction as well as ultrasound irradiation at low (20 kHz) and high frequencies (860 kHz). In these experiments, we investigated the simultaneous action of the ultrasound and UV irradiation by varying parameters like the duration of photocatalysis and ultrasound irradiation frequency. At the same time, US power, temperature, amount of TiO₂ photocatalyst and amount of Fenton reagent remained constant. Due to their diverse structure, each azo dye showed different degradation levels using different combinations of the above-mentioned Advanced Oxidation Processes (AOPs). The Photo-Fenton reagent is more effective with US 20 kHz and US 860 kHz for the azo dyes originated from the weaving industry at pH = 3 as compared to pH = 6.8. The combination of the Photo-Fenton reaction with 860 kHz ultrasound irradiation for the same dye gave an 80% conversion at the same time. Experiments have shown a high activity during the first two hours. After that threshold, the reaction rate is decreased. FT-IR and TOC measurements prove the decolorization due to the destruction of the chromophore groups but not complete mineralization of the dyes.     

Ultrasound-assisted micellar cleanup coupled with large-volume-injection enrichment for the analysis of polar drugs in blood and zebrafish samples

A novel ultrasound-assisted micellar cleanup strategy (UAMC) coupled with large volume injection (LVI) high performance liquid chromatography (HPLC) method was proposed and successfully applied to the analysis of cefathiamidine in complex biological samples such as whole blood, plasma, serum and even zebrafish, a challenging positive real sample. Based on the micelle-biomacromolecule interaction, the phase-separation feature of surfactant micelles and ultrasound cavitation, UAMC possessed an impressive matrix cleanup capability and could rapidly reach distribution equilibrium (approximately 2 min), which enabled simultaneous sample cleanup and analyte extraction within 8 min. Due to the high cleanup efficiency of UAMC, large volume of pretreated samples could be injected for analysis without peak broadening, impurity interference and column degradation. Thus, online analyte enrichment could be automatically performed to significantly improve method sensitivity by the column-switching LVI-HPLC system, a commercial HPLC system with small modifications. The UAMC-LVI-HPLC method creatively integrated sample cleanup, analyte extraction and on-column enrichment into simple operation. In addition, the UAMC-LVI-HPLC method enabled non-matrix-matched analysis of cefathiamidine in complex biological samples. This feature was helpful to address the problems caused by conventional matrix-matched or internal standard calibration methods, such as matrix bias, increased workload, limited availability of suitable blank matrices and the use of expensive internal standards. The method had low limits of detections (e.g., 0.0051 mg/L and 0.038 μg/g), wide linear ranges (0.030–100 mg/L and 0.15–489 μg/g), good linear correlation (R² = 0.9999), satisfactory accuracy (97.6–109.7%) and excellent intra- and interday precision (0.5–4.9%). Thus, UAMC-LVI-HPLC is expected to be a promising candidate for bioanalysis in therapeutic drug monitoring or pharmacokinetic and toxicology studies in the future.     

表面活性剂SDS/TX-100混合体系的NMR研究

用NMR测量了不同比例的SDS/TX-100混合溶液中质子化学位移,结合表面活性剂溶液的两态交换模型,分析了质子化学位移随浓度的变化趋势, 求出了不同比例混合溶液中两种表面活性剂各自的临界胶束浓度及混合胶束的临界胶束浓度. 依据理想混合溶液理论,预测了混合胶束的临界胶束浓度,计算了溶液中SDS与TX-100之间的相互作用参数和SDS在混合胶束中的摩尔分数. 根据所得参数讨论了混合胶束的形成过程. 利用文中和文献中混合体系的实验数据验证了协同作用理论改进前后的适用性,表明改进后的协同作用理论完善一些.     

Axonal outgrowth is associated with increased ERK 1/2 activation but decreased caspase 3 linked cell death in Schwann cells after immediate nerve repair in rats

Background  

Extracellular-signal regulated kinase (ERK1/2) is activated by nerve damage and its activation precedes survival and proliferation of Schwann cells. In contrast, activation of caspase 3, a cysteine protease, is considered as a marker for apoptosis in Schwann cells. In the present study, axonal outgrowth, activation of ERK1/2 by phosphorylation (p-ERK 1/2) and immunoreactivity of cleaved caspase 3 were examined after immediate, delayed, or no repair of transected rat sciatic nerves.