Application of experimental design to the formulation of glucose oxidase encapsulation by liposomes

Fractional factorial screening design and response surface methodology were applied to optimize the entrapment of glucose oxidase in liposomes by the dehydration–rehydration vesicle (DRV) method. Phosphatidylcholine from different sources, cholesterol:phosphatidylcholine (Ch:PC) and enzyme:lipid (E:L) ratios, buffer pH, sonication frequency and trehalose concentration were the parameters selected for this study. The type of phosphatidylcholine was found to be the most important factor followed by the trehalose concentration, Ch:PC ratio, sonication frequency and E:L ratio. The pH did not play an important role in the response. By treating liposomes with trehalose, as cryoprotectant, the activity of entrapped enzyme decreased by 16%. Two of the factors (cholesterol:phosphatidylcholine and enzyme:lipid ratios) were further studied in a 3² central composite design. The optimized liposomal formulation with an entrapment efficiency of 24% was obtained for egg yolk PC with Ch:PC and E:L ratios of 0.95 and 14.69, respectively, at pH 6 and applying a sonication frequency of 150 W. Copyright © 2004 Society of Chemical Industry     

Elaboration of Lutein-Loaded Nanoliposomes Using Supercritical CO2

A batch process for producing lutein-loaded liposomes using supercritical CO₂ is studied. The effects of the variation of pressure (10 and 15 MPa), temperature (308, 313, and 318 K), and lutein to lipid ratio (0.5 and 1 wt%) on the liposome average size and size distribution are investigated, as well as on the encapsulation efficiency (EE) of lutein. This process is worked in a repeatable manner and is allowed the production of nanoliposomes with mean diameters (MDs) ranging from 65 ± 33 to 77 ± 40 nm, obtaining lutein EEs ranging from 82.1 ± 3.7% to 91.9 ± 2.9%. Temperature, pressure, and lutein to lipid ratio seem to have no impact on size, size distribution, and EE on formed liposomes. The use of low temperatures and low pressures allows the obtainment of liposomes with diameters less than 100 nm and limits the process energy cost. Moreover, the supercritical CO₂-assisted batch process effectively encapsulates lutein into liposome, an antioxidant molecule used for the prevention of retinal damage. Liposomes formed by this supercritical process have the desired characteristics for human target delivery. Practical applications: This work on the optimization of a process for developing liposomes in a supercritical environment has applications in medicine. Indeed, the liposomes formed with this process are nanoliposomes with a size of less than 80 nm. In addition, excellent lutein EEs (hydrophobic molecules) show that the liposomes formed constitute excellent coating matrices for the protection of active ingredients. These reasons make these liposome matrices applicable in nanomedicine (injection of sensitive drugs requiring protection before injection). The elaboration process also makes it possible to form liposomes with desired properties by changing pressure, temperature, or lecithin concentration. Therefore, this work focuses on the properties of liposomes as a function of the operating conditions.     

The sonication effect on CNT‐epoxy composites finally clarified

Carbon nanotube‐epoxy composites were prepared using amino‐functionalized CNTs and sonication as a mixing process. Different times and sonication powers were used for preparing composites in order to study how the sonication process may influence the curing reaction of both systems: neat epoxy resin and amino‐CNT/epoxy composite.The curing reaction was investigated with differential scanning calorimetry and the results were associated with analysis of gel permeation chromatography. The results showed that the effect of CNTs on the cure behavior of the epoxy resin depends on the sonication power. The sonication of neat resin with a 150 W powered device led to a molar mass reduction of the resin and an increase in the cure enthalpy. The CNT addition to this system reduced the cure enthalpy. However, when neat epoxy resin was sonicated with a 200 W powered device, the molar mass did not decrease (i.e., it was increased or was not changed) and the cure enthalpy did not increase (essentially it decreased or did not change). The CNT addition to such solutions did not reduce (i.e., it was increased or did not change) the cure enthalpy, which is a contrary result from that obtained with a 150 W powered device. POLYM. COMPOS., 38:1964–1973, 2017. © 2015 Society of Plastics Engineers     

In situ polycondensation of amino acid modified liposomes and their properties

Three amphiphilic amino acids based on glutamic acid, i.e. S-[1-carboxy-2-([N-bistetradecyl- -glutamate]carbon-yl)ethyl]cysteine (1), S-[1-carboxy-2-([N-bishexadecyl- -glutamate]carbonyl)ethyl]cysteine (2), S-[1-carboxy-2-([N-bis-octadecyl- -glutamate]carbonyl)ethyl]cysteine (3), were synthesized. The aggregation behavior of them in water or buffer solution was studied. It was found that upon hydration and sonication in water, they could form stable liposomes. This kind of amino acid modified liposome was then polycondensed locally on the liposome surface to form a polypeptide-surfaced liposome and the peptide formation was detected by Fr-IR, GPC, etc. The effect of polycondensation of amino acid on the properties of liposomes were studied by detecting the phase transition temperatures with DSC or measuring the leakage of the encapsulated fluorescent probe from the liposomes. It was observed that the phase transition temperatures of the peptide liposomes moved down and the polycondensation of amino acid moieties obviously increased the leakage of the encapsulated molecules.     

Functional Properties of Soy Protein Isolates Produced from Ultrasonicated Defatted Soy Flakes

This study aimed to determine the effect of pretreating defatted soy flakes with ultrasound on soy protein isolate (SPI) yield and functionality. Defatted soy flakes dispersed into water (16%, w/w) were sonicated for 30, 60 and 120 s at ultrasonic amplitudes of 21 and 84 μm_pp (peak to peak amplitude in μm), representing low and high power, respectively. The power densities were 0.30 and 2.56 W mL⁻¹, respectively. The SPI yield increased by 13 and 34%, after sonication for 120 s at low and high power, respectively. The sonication of defatted soy flakes for 120 s at the higher power level improved the SPI solubility by 34% at pH 7.0, while decreasing emulsification and foaming capacities by 12 and 26%, respectively, when compared to control SPI. Rheological behavior of the SPI was also modified with significant loss in consistency coefficient due to sonication. Some of these results could be explained by the loss of the protein native state with increased sonication time and power.     

Direct and indirect power ultrasound assisted pre-osmotic treatments in convective drying of guava slices

Application of ultrasound to osmotic dehydration of guava slices via indirect sonication using an ultrasonic bath system and direct sonication using an ultrasonic probe system was studied. Pre-treatments were designed in three osmotic solution concentrations of 0, 35, and 70 °Brix at indirect ultrasonic bath power from 0 to 2.5 kW for immersion times ranging for 20–60 min and direct ultrasonic probe amplitudes from 0 to 35% for immersion times of 6–20 min. The calculated ultrasound intensities from calorimetric ultrasound power dissipated indicated that direct sonication was more intensive than indirect sonication. The general linear model (GLM) showed that ultrasound input (power and amplitude), osmotic solution concentrations, and immersion time increased the water loss, solid gain, and total colour change of guava slices significantly with P < 0.0005. Indirect sonication in osmotic solutions contributed to high water loss and solid gain with acceptable total colour change than direct sonication. Applying ultrasound pre-osmotic treatment in 70 °Brix prior to hot-air drying reduced the drying time by 33%, increased the effective diffusivity by 35%, and decreased the total colour change by 38%. A remarkable decrease of hardness to 4.2 N obtained was also comparable to the fresh guava at 4.8 N.     

超声在陶瓷膜处理乳化含油废水中的作用研究

Ultrasonic field was applied in the treatment of oil emulsification wastewater by ZrO2 ceramic mem-brane. The permeate flux, rejection ratio in the filtration process and recovery ratio of flux in the membrane cleaning process were measured. Great improvement in the permeate flux and rejection ratio have been observed for the membrane process enhanced by the ultrasonic field. The permeate flux of water through the membrane was about 210L.m^-2.h^-1 and the oil rejection ratio was over 99.9% under the optimal ultrasonic treatment conditions, which were 8W of ultrasonic power, 7cm of ultrasonic probe length introduced into the membrane channel and the same ultrasonic radiation direction as the wastewater flow. The resistance of the membrane process was compared between the cases with and without ultrasound, and the total resistance was reduced 68% by the use of ultrasound, Four methods including water cleaning, water cleaning under sonication, chemical cleaning and chemical cleaning under sonication were used to recover membrane flux. It was found that the flux recovery ratio increased with the increase of ultrasonic cleaning power. In addition, the use of chemical agents combining with ultrasonic irradiation showed a synergistic effect, which resulted in the highest cleaning efficiency and the shorter cleaning time.     

Investigation of the cell disruption methods for maximizing the extraction of arginase from mutant <Emphasis Type="Italic">Bacillus licheniformis</Emphasis> (M09) using statistical approach

Arginase, an intracellular enzyme produced by Bacillus licheniformis (NRS-1264) is effectively used as a drug in the treatment of arginine-dependent cancers, and it is essential for controlling acute neurological disorders. We investigated the effect of various cell disruption methods for maximizing the extraction of intracellular arginase from mutant Bacillus licheniformis (M09), followed by comparing optimization methods, one factor at a time (OFAT), evolutionary operation (EVOP) and response surface method (RSM). Ultrasonication for 2-5 min having a suspension volume in the range of 12-20 mL at a radio frequency power between 30–70 W appeared to be the most effective extraction technique for arginase. The arginase yield decreased in the range of 50–70 W of RF power/16-20 mL suspension volume and 4-5 min sonication time. EVOP predicted a maximum arginase extraction of 3,910 IU·L^-1 at 2 min sonication having 16 mL suspension volume at 30W RF power. However, response surface optimization suggested an optimized condition of 3 min sonication having 14.5 mL suspension volume at 35W RF power in which the maximum arginase activity in the medium was 3,600 IU·L^-1.     

Effect of Ultrasonically Induced Nucleation on the Drying Kinetics and Physical Properties of Freeze-Dried Strawberry

Power ultrasound has proved to be very useful in controlling crystallization processes since sonication can enhance both the nucleation rate and crystal growth rate by producing fresh and/or more nucleation sites. Therefore, in this study, power ultrasound was applied to control the freezing step of freeze-dried strawberry. The results showed that when power ultrasound was applied at different temperatures, it increased the nucleation temperature and shortened the characteristic freezing time. The application of power ultrasound in the freezing step increased the drying time in subsequently freeze-dried strawberry samples. This longer drying time was found to be due to increased resistance to moisture diffusion due to the formation of a network of small pores caused by sublimation of small ice crystals induced by the power ultrasound. Scanning electron microscopic images revealed that the freeze-dried sample frozen by ultrasound-assisted freezing (UAFD) had finer cellular structure compared to those frozen in other freezing conditions. UAFD samples had better textural hardness, while the rehydration capacity was lower compared to those of NRFD and NIFD samples.     

Antisolvent Crystallization of Roxithromycin and the Effect of Ultrasound

Antisolvent crystallization was performed to precipitate roxithromycin particles from organic solutions. Roxithromycin was dissolved in acetone at different concentrations and each solution was injected into an aqueous antisolvent leading to prompt particle formation. The effects of various experimental variables (solution injection rate, solution concentration, and temperature) on the particle size of roxithromycin were investigated. In addition to these variables, the effect of ultrasound on the resulting particle size was investigated by changing process parameters such as wave intensity (power output), sonication time, and the moment of ultrasonic application. When the drug solution was rapidly injected into the antisolvent, smaller crystals were obtained. Smaller crystals were obtained when solutions with high drug concentrations were used and also when the crystallization took place at lower temperatures. The particle size decreased with the increasing power output of ultrasound and with the increasing sonication time. It was also found that the ultrasonic wave induced the reduction of the particle size only when the ultrasound was applied to the solution at the initial stage of crystallization.     

Optimization and sensitivity analysis of rheological properties of high concentration γ-alumina/water suspension

In this study, a high concentration γ-alumina/water suspension containing 75% (wt) γ-alumina nanoparticle was prepared by using a 3D-mixer. TEM imaging, Dynamic Light Scattering (DLS), and X-Ray Diffraction (XRD) analyses were used to investigate the stability of nanoparticles in the base fluid, purity, and crystallinity as well as chemical structure of nanoparticles, respectively. Then, the variation of dynamic viscosity and rheological behavior of the suspension and their dependency on pH, sonication durations, dispersing agent concentrations, and ratios of ceramic balls to nanoparticles weight was studied. The results showed that the rheological behavior of the suspension was similar to non-Newtonian fluids, and a power-law model with yield stress was able to justify this behavior. Moreover a correlation, including pH, sonication time, dispersing agent concentration, and the ratio of ceramic balls to nanoparticles weight as independent parameters, was developed to predict the power of the power-law model. The model showed a low deviation, about 5%, from the experimental values and revealed the significant effect of pH, sonication time, and the ratio of mixing ceramic balls to nanoparticles on the rheological behavior of the suspension. More importantly, by implementing the method used in this work, a very low dynamic viscosity of 11.4 mPa.s was achieved.     

Ultrasound‐induced nucleation in microcellular polymers

In this study, microcellular Acrylonitrile–Butadiene–Styrene foams with high cell density and expansion ratio has been manufactured using ultrasound‐induced nucleation technique in solid‐state batch foaming process. Influence of sonication time, sonication frequency, and ultrasound power were found very crucial in designing of cellular morphology. The initial 10 s of ultrasound exposure was found to influence the foam morphology critically. Longer periods of ultrasound exposure developed foams with lower average cell size as compared to foams developed with lesser ultrasound exposure time. Higher sonication power resulted in foams with uniform morphology and higher cell densities as compared to foams developed with lower sonication intensities. Finally, the ultrasonic frequency was also found to influence the morphology intensely. Low frequency sonication resulted in foams with uniform cell distribution, whereas high frequency sonication developed bimodal microcellular type of microstructure. The results coherently demonstrate that with the advent of ultrasonic waves, the energy barrier for cell nucleation swiftly decreases which enhances the cell density in the final foamed product. © 2014 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2014 , 131, 40742.     

Bioimprinted Immobilization of <Emphasis Type="Italic">Candida antarctica</Emphasis> Lipase A for Concentration of Omega-3 Polyunsaturated Fatty Acids

Bioimprinting and immobilization of Candida antarctica lipase A (CALA) was optimized by response surface methodology to improve its selectivity for concentration of omega-3 polyunsaturated fatty acids (PUFA) by lipase-catalyzed ethanolysis of salmon oil obtained from salmon by-products. Enzyme-to-support ratio, oleic acid concentration, and immobilization time were the factors investigated. Total omega-3 PUFA lost to the ethyl esters (EE) fraction was monitored in relation to the varying levels of the factors. Results showed that the second-order polynomial model generated adequately fitted the experimental data. The most critical factor was the enzyme-to-support ratio, which led to decreased recovery of omega-3 PUFA in the glycerides fraction with increasing levels of enzyme amount. Optimum conditions were selected as follows: enzyme-to-support ratio, 0.2 (w/w); oleic acid concentration, 50 mM; immobilization time, 85 min. The total omega-3 PUFA lost to the EE fraction through ethanolysis of salmon oil decreased from 3.76 to 1.47 (mol%) under the optimum conditions.     

Studies on ultrasonic disintegration kinetics of SEBS in PP melts and the influencing factors

The ultrasonic disintegration behavior of polystyrene‐block‐poly(ethylene‐co‐butylenes)‐block‐polystyrene (SEBS) in polypropylene (PP) melts was studied by means of scanning electron microscopy (SEM) and dynamic rheological measurements. A disintegration kinetics model of the dispersed phase in polymer matrix was proposed to explain the ultrasonic disintegration kinetics, and it is in accordance with the equation: . The experimental results showed that the average particle diameter of SEBS decreased with sonication time and approached a limiting value (the equilibrium diameter), below which no further disintegration occurred. The effects of ultrasonic power, ultrasonic frequency, sonication distances, and heating temperatures on the morphology of PP/SEBS blends were also investigated. SEM results demonstrated that the increase of ultrasonic power could enhance the dispersion of SEBS. And SEBS particles were more easily disintegrated at 20 kHz. Moreover, the disintegration extent decreased with the increase of sonication distance or melt temperature. The dispersion state of SEBS was indirectly evaluated by the dynamic rheological properties. It was found that there was an increase in the complex viscosity, storage, and loss moduli as the ultrasonic intensity was in an appropriate scope. © 2016 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2017 , 134, 44386.     

Effect of ultrasonication on the dilution crystallization of naproxen

This work investigates the use of ultrasonication for the dilution crystallization of naproxen. Naproxen solution was prepared in acetone, while water was used as the anti-solvent. The main objective of this study was to determine the effect of sonication on the thermodynamic properties (solubility, metastable zone width, and induction time) and particle size of naproxen crystals. The use of sonication was compared to conventional stirred crystallization methods. When the solvent ratio (V_acetone/V_water) is low, no significant difference exists between the naproxen solubility in either the sonicated or stirred solutions. However, at higher solvent ratios (V_a/V_w = 9) and at low temperatures (0°C), ultrasonication increases the solubility of naproxen from 0.171 to 0.193 g/mL. Compared to conventional stirred crystallization techniques (the metastable zone width is 0.024 g/mL), ultrasonic crystallization is characterized by a narrower metastable zone (the width is 0.010 g/mL), and with ultrasonic crystallization the induction time decreases by about 50 seconds. During ultrasonic crystallization, the average naproxen particle size is reduced from 22.6 to 10.5 μm by increasing the power and time of ultrasonication. Particle sizes and crystal morphologies obtained using ultrasonication were significantly improved compared to those produced with stirring.     

Enhanced localization of anticancer drug in tumor tissue using polyethylenimine-conjugated cationic liposomes

Liposome-based drug delivery systems hold great potential for cancer therapy. However, to enhance the localization of payloads, an efficient method of systemic delivery of liposomes to tumor tissues is required. In this study, we developed cationic liposomes composed of polyethylenimine (PEI)-conjugated distearoylglycerophosphoethanolamine (DSPE) as an enhanced local drug delivery system. The particle size of DSPE-PEI liposomes was 130 ± 10 nm and the zeta potential of liposomes was increased from -25 to 30 mV by the incorporation of cationic PEI onto the liposomal membrane. Intracellular uptake of DSPE-PEI liposomes by tumor cells was 14-fold higher than that of DSPE liposomes. After intratumoral injection of liposomes into tumor-bearing mice, DSPE-PEI liposomes showed higher and sustained localization in tumor tissue compared to DSPE liposomes. Taken together, our findings suggest that DSPE-PEI liposomes have the potential to be used as effective drug carriers for enhanced intracellular uptake and localization of anticancer drugs in tumor tissue through intratumoral injection.     

Development and characterization of lactoferrin nanoliposome: cellular uptake and stability

Lactoferrin was purported in consumer literature to enhance and support the immune system response through their antioxidant, antibacterial, and anticarcinogenic properties. To improve the effectiveness of lactoferrin, liposomes were used as a carrier in this study. The main purpose of this study was to compare three different methods to prepare the lactoferrin nanoliposomes based on the encapsulation efficiency and size distribution and evaluate the stability and cellular uptake of lactoferrin nanoliposomes. Encapsulation efficiency and size distribution indicated the reverse-phase evaporation method was fit for preparing the lactoferrin nanoliposomes. The stabilities of lactoferrin nanoliposomes in simulated gastrointestinal juice, sonication treatment time and lipoperoxidation extent of storage time were evaluated. The lactoferrin nanoliposomes showed an acceptable stability in simulated gastrointestinal juice at 37°C for 4 h and short treatment times were required to achieve nano-scaled liposomes. Furthermore, the viability of cells was decreased by increasing the concentration of the various lactoferrin nanoliposomes. The methyl thiazolyl tetrazolium results demonstrated that Lf nanoliposomes and Lf activated in the cells in a manner of dose-effect relation and Lf nanoliposomes had a statistically significantly different (p<0.01) between the concentration 5 and 10 mg/mL. According to the results, nanoliposomes may be fit for the oral administration of lactoferrin and could be useful approach for lactoferrin availability in tumor cells.     

The effects of sonication time and frequencies on degradation,crystallization behavior,and mechanical properties of polypropylene

The effects of ultrasound with different frequencies on the degradation, crystallization behavior, and mechanical properties of polypropylene were investigated by means of intrinsic viscosity, gel permeation chromatography, dynamic rheological measurements, X‐ray diffraction, differential scanning calorimetry, polarized optical microscopy, and tensile and impact measurements. The results demonstrated that the degradation extent of polypropylene (PP) increased with increasing sonication time, especially for the samples treated in more than 5 min. Meanwhile, PP treated at 20 KHz was more easily degraded than other two frequencies. On the other hand, β‐form PP could be induced by ultrasonic irradiation, its amount increased as the sonication time increased to 3 min and then decreased as the time further increased due to the degradation of PP. Likewise, β‐phase was more easily formed when PP was treated at 20 KHz. The results of mechanical properties showed that the toughness of PP could be enhanced by the formation of β?phase, while the mechanical properties would be weakened by the substantial degradation under ultrasonic irradiation with higher intensity. Therefore, it was believed that the effects of both sonication time and frequency should account for the balance between toughening and degradation of PP. POLYM. ENG. SCI., 55:2566–2575, 2015. © 2015 Society of Plastics Engineers     

Explosion Performance of High‐Temperature Degraded Emulsion Explosives

Experiments were conducted to study the underwater explosion performance of emulsion explosives (EE) after hot water bath. Spherical charges of EE with different sensitizers and hot water bath were prepared and tested. As for as‐prepared charges, the detonation velocity experiments and underwater explosion experiments were carried out and the crystallization ratio was measured and calculated by the dissolution and neutralization method. The results showed an inverse relationship between explosion parameters (pressure peak, specific impulse, detonation velocity and specific total energy) and heating time. It also revealed that the effective explosive weight of EE was reduced with long time of hot water bath. Moreover, the crystallization ratio and the decreasing rate of explosion parameters of EE sensitized by NaNO₂ were apparently higher than EE containing physical sensitizers (glass microballoon and perlite), which was attributed to the different destruction mechanism of EE. After 6‐hour hot water bath, the EE containing physical sensitizers still retained detonator sensitivity and more than 80 % of specific total energy. Meanwhile, the crystallization ratio was less than 20 %. Whereas, the EE sensitized by NaNO₂ lost the detonator sensitivity and the crystallization ratio of EE was also above 40 %.     

Target-selective drug delivery through liposomes labeled with oligobranched neurotensin peptides

The structure and the in vitro behavior of liposomes filled with the cytotoxic drug doxorubicin (Doxo) and functionalized on the external surface with a branched moiety containing four copies of the 8-13 neurotensin (NT) peptide is reported. The new functionalized liposomes, DOPC-NT?Lys(C??)?, are obtained by co-aggregation of the DOPC phospholipid with a new synthetic amphiphilic molecule, NT? Lys(C??)?, which contains a lysine scaffold derivatized with a lipophilic moiety and a tetrabranched hydrophilic peptide, NT8-13, a neurotensin peptide fragment well known for its ability to mimic the neurotensin peptide in receptor binding ability. Dynamic light scattering measurements indicate a value for the hydrodynamic radius (RH) of 88.3±4.4?nm. The selective internalization and cytotoxicity of DOPC-NT? Lys(C??)? liposomes containing Doxo, as compared to pure DOPC liposomes, were tested in HT29 human colon adenocarcinoma and TE671 human rhabdomyosarcoma cells, both of which express neurotensin receptors. Peptide-functionalized liposomes show a clear advantage in comparison to pure DOPC liposomes with regard to drug internalization in both HT29 and TE671 tumor cells: FACS analysis indicates an increase in fluorescence signal of the NT?-liposomes, compared to the DOPC pure analogues, in both cell lines; cytotoxicity of DOPC-NT? Lys(C??)?-Doxo liposomes is increased four-fold with respect to DOPC-Doxo liposomes in both HT29 and TE671 cell lines. These effects could to be ascribed to the higher rate of internalization for DOPC-NT? Lys(C??)?-Doxo liposomes, due to stronger binding driven by a lower dissociation constant of the NT?-liposomes that bind the membrane onto a specific protein, in contrast to DOPC liposomes, which approach the plasma membrane unselectively.