Inactivation of Enterobacter aerogenes in reconstituted skim milk by high- and low-frequency ultrasound

The inactivation of Enterobacter aerogenes in skim milk using low-frequency (20 kHz) and high-frequency (850 kHz) ultrasonication was investigated. It was found that low-frequency acoustic cavitation resulted in lethal damage to E. aerogenes. The bacteria were more sensitive to ultrasound in water than in reconstituted skim milk having different protein concentrations. However, high-frequency ultrasound was not able to inactivate E. aerogenes in milk even when powers as high as 50 W for 60 min were used. This study also showed that high-frequency ultrasonication had no influence on the viscosity and particle size of skim milk, whereas low-frequency ultrasonication resulted in the decrease in viscosity and particle size of milk. The decrease in particle size is believed to be due to the breakup of the fat globules, and possibly to the cleavage of the κ-casein present at the surface of the casein micelles. Whey proteins were also found to be slightly affected by low-frequency ultrasound, with the amounts of α-lactalbumin and β-lactoglobulin slightly decreasing.     

Effect of sonication frequency on the disruption of algae

In this study, the efficiency of ultrasonic disruption of Chaetoceros gracilis, Chaetoceros calcitrans, and Nannochloropsis sp. was investigated by applying ultrasonic waves of 0.02, 0.4, 1.0, 2.2, 3.3, and 4.3 MHz to algal suspensions. The results showed that reduction in the number of algae was frequency dependent and that the highest efficiency was achieved at 2.2, 3.3, and 4.3 MHz for C. gracilis, C. calcitrans, and Nannochloropsis sp., respectively. A review of the literature suggested that cavitation, rather than direct effects of ultrasonication, are required for ultrasonic algae disruption, and that chemical effects are likely not the main mechanism for algal cell disruption. The mechanical resonance frequencies estimated by a shell model, taking into account elastic properties, demonstrated that suitable disruption frequencies for each alga were associated with the cell’s mechanical properties. Taken together, we consider here that physical effects of ultrasonication were responsible for algae disruption.     

Ultrasound-induced inactivation of gram-negative and gram-positive bacteria in secondary treated municipal wastewater

The effect of 24 kHz, high energy ultrasound in the presence and absence of titanium dioxide particles on the destruction of different bacteria groups was studied. Applying a total of 1500 W/L for 60 min (this corresponds to 5400 kJ/L specific nominal energy), the mean destruction of gram-negative bacteria such as total coliforms, faecal coliforms and Pseudomonas spp. was 99.5%, 99.2% and 99.7%, respectively. More recalcitrant to sonolytic inactivation were the gram-positive bacteria Clostridium perfringens and faecal streptococci with a mean removal of 66% and 84%, respectively. The presence of 5 g/L TiO₂ generally enhanced the destruction of gram-negative bacteria, yielding three to five logs reduction. On the other hand, the relatively weak sonochemical inactivation of gram-positive bacteria was only slightly affected by the presence of solid particles. Inactivation was found to follow first-order kinetics regarding bacteria population and was not affected significantly by the wastewater quality. Ultrasound irradiation at 4000 kJ/L specific nominal energy and in the presence of 5 g/L TiO₂ achieved less than 10³ CFU/100 mL total coliforms, thus meeting USEPA quality standards for wastewater reuse.     

Combination of therapeutic ultrasound with antibiotics interfere with the growth of bacterial culture that colonizes skin ulcers: An in-vitro study

Staphylococcus aureus and Escherichia coli are among the major bacterial species that colonize skin ulcers. Therapeutic ultrasound (TUS) produces biophysical effects that are relevant to wound healing; however, its application over a contaminated injury is not evidence-based. The objective of this research was to analyze the effect of TUS on in vitro-isolated S. aureus and E. coli, including the combination of ultrasound and antibiotics, in order to assess their antibiotic action on bacterial susceptibility. For the experiments, the bacterial strains were suspended in saline, then diluted (10⁴ CFU/mL) for irradiation (at 1 and 3 MHz, 0.5 and 0.8 W/cm² for 0 and 15 min) and the combination treatment of ultrasonication and antibiotics was administered by adding nalidixic acid (S. aureus) and tetracycline (E. coli) at concentrations equivalent to 50% of the minimum inhibitory concentration (MIC). The experiments were carried out in duplicate with six repetitions. The suspensions were inoculated on to Petri plates and incubated at 37 °C and the colony forming units (CFUs) were counted after 24 h. The results were subjected to the Shapiro–Wilk normality test, followed by parametric ANOVA and Tukey’s post hoc test at a significance level of 1%. The results demonstrated that the action of TUS at 1 MHz inhibited bacterial growth while at 3 MHz, bacterial growth was observed in both species. However, the synergistic combination of ultrasound and antibiotics was able to inhibit the growth of both bacteria completely after 15 min of ultrasonication. The results suggest that the action of ultrasound on S. aureus and E. coli are dependent on the oscillation frequency as well as the intensity and time of application. The combination of ultrasound with antibiotics was able to inhibit bacterial growth fully at all frequencies and doses in both species.     

Effect of ultrasound on the extraction of total anthocyanins from Purple Majesty potato

This study examined anthocyanin extraction using the application of ultrasound to raw freeze dried, microwaved and raw sliced Purple Majesty potato, a new pigmented potato variety rich in anthocyanins. A 20 kHz probe was used for the sonication at 3 different amplitudes (30%, 50% and 70%) and ethanol in water at different ratios (50:50 and 70:30 v/v) was used for the extraction. Anthocyanin extraction from raw freeze dried purple potato was optimal at an ethanol:water ratio (70:30; v/v) after 5 min of ultrasonication, while the least amount of anthocyanins was extracted from raw sliced potatoes. The application of microwaves (as a pre-treatment) before the UAE resulted in an increase in the amount of anthocyanins extracted and a decrease in the amount of solvent used. Analysis of variance showed that potato form, ultrasonication time, ultrasonication amplitude and solvent ratio as well as two and three way interactions between some of these factors had a very significant effect (p < 0.000) on the amount of anthocyanins extracted.     

Investigation of the growth and local stoichiometric point group symmetry of titania nanotubes during potentiostatic anodization of titanium in phosphate electrolytes

Potentiostatic anodization of commercially pure, 50 µm-thick titanium (Ti) foil was performed in aqueous, phosphate electrolytes at increasing experimental timeframes at a fixed applied potential for the synthesis of titania nanotube arrays (TNAs). High resolution scanning electron microscopy images, combined with energy dispersive spectroscopy and x-ray diffraction spectra reveal that anodization of the Ti foil in a 1 M NaF+0.5 M H₃PO₄ electrolyte for 4 h yields a titanate surface with pore diameters ranging between 100 and 500 nm. The presence of rods on the Ti foil surface with lengths exceeding 20 µm and containing high concentrations of phosphor on the exterior was also detected at these conditions, along with micro-sized coral reef-like titanate balls. We propose that the formation of these structures play a major role during the anodization process and impedes nanotube growth during the anodization process. High spatially resolved scanning transmission electron microscopy-electron energy loss spectroscopy (STEM-EELS) performed along the length of a single anodized TiO₂ nanotube reveals a gradual evolution of the nanotube crystallinity, from a rutile-rich bottom to a predominantly anatase TiO₂ structure along its length.     

Green synthesis of nanocomposites consisting of silver and protease alpha chymotrypsin

The synergy of ultrasonication and the exposure to light radiation was found to be necessary in the formation of nanocomposites of silver and a protease alpha chymotrypsin. The reaction was carried out in aqueous medium and the process took just less than 35 min. Ultrasonication alone formed very negligible number of nanoparticles of <100 nm size whereas light alone produced enough number but the size of the particles was >100 nm.The effects of pH (in the range of 3–5, 9–10), ultrasonication time periods (0–30 min), ultrasonication intensity (33–83 W cm^?2), energy of light radiation (short UV, long UV and Fluorescent light) and time period of exposure (5–60 min) to different light radiations were studied.The formation of nanocomposites under these effects was followed by surface plasmon resonance (SPR) spectra, dynamic light scattering (DLS), transmission electron microscopy (TEM). Ag–chymotrypsin nanocomposites of sizes ranging from 13 to 72 nm were formed using the synergy of ultrasonication and exposure to short UV radiation. Results show that ultrasonication promoted nuclei formation, growth and reduction of polydispersity by Ostwald ripening.     

Inactivation of Escherichia coli and Streptococcus mutans by ultrasound at 500 kHz

This paper shows a systematic study of the 500 kHz frequency ultrasound efficiency on the microbial inactivation as a function of ultrasonic power delivered into the bacterial suspension. The inactivation of Escherichia coli IAM 12058, a Gram-negative bacterium and Streptococcus mutans JCM 5175, a Gram-positive bacterium is enhanced by increasing the ultrasonic power in the range of 1.7–12.4 W and the logarithm of survival ratio decreases linearly with irradiation time, except for E. coli sonicated with the highest power level. The rate constants were estimated in the linear region of the plots representing survival ratio logarithm vs. sonication time. A better understanding of the inactivation process at 500 kHz could be gained by suppressing the chemical effects with a radical scavenger. We find out that the rate constants increase with the ultrasonic power delivered into the solution and dramatically decrease by the addition of t-butanol as a radical scavenger to the bacterial suspension. For comparison, experiments were carried out at a low frequency level of 20 kHz. It was found out that for the same ultrasonic power delivered into the bacterial suspension, the inactivation was slightly enhanced at 500 kHz frequency. The examinations of bacterium performed with a TEM revealed lethal damages arising from the interaction of bacterial cells with the cavitational bubbles. A significant amount of empty cell envelopes as well as their cytoplasmatic content was detected. Thus, based on these new data, the mechanism of bacterial inactivation by ultrasounds at high frequency is discussed here.     

Ultrasound stimulated production of a fibrinolytic enzyme

The present study is aimed at enhanced production of a fibrinolytic enzyme from Bacillus sphaericus MTCC 3672 under ultrasonic stimulation. Various process parameters viz; irradiation at different growth phases, ultrasonication power, irradiation duration, duty cycle and multiple irradiation were studied for enhancement of fibrinolytic enzyme productivity. The optimum conditions were found as follows, irradiation of ultrasonic waves to fermentation broth at 12 h of growth phase with 25 kHz frequency, 160 W ultrasound power, 20% duty cycle for 5 min. The productivity of fibrinolytic enzyme was increased 1.82-fold from 110 to 201 U/mL compared with the non sonicated control fermentation. Drop in glucose concentration from 0.41% to 0.12% w/v in ultrasonicated batch implies that, ultrasonication increases the cell permeability, improves substrate intake and progresses metabolism of microbial cell. Microscopic images before and after ultrasonic stimulation clearly signifies the impact of duty cycle on decreasing biomass concentration. However, environmental scanning electron micrograph does not show any cell lysis at optimum ultrasonic irradiation. Offshoots of our results will contribute to fulfill the demand of enhancement of microbial therapeutic enzyme productivity, through ultrasonication stimulation.     

Laser micro-machined semi-slinky like MEMS structures: Novel interface coolers

Laser micro-machining has recently been considered a precision and reproducible manufacturing technique in MEMS fabrication because of the superior characteristics of a focused laser beam. It is not only a unique tool but also an invisible optical drill. The aim of the present paper is two-fold: to manufacture novel miniaturized titanium 3D MEMS surface structures in order to increase the cooling performance. Second is to find the behaviors of the operational parameters which controlling the laser-material interaction mechanisms and also suggest the best adjustments in order to achieve this novel semi-slinky like spiral MEMS surface structures with using a 20 W ytterbium fiber laser. Pure titanium micro-MEMS product which has novel interface coolers was manufactured using a ytterbium fiber laser (λ=1060 nm) with 40 ns pulse duration. Best adjustments were, respectively, the pulse duration: 40 ns, the pulse energy: 0.4 mJ, the laser scanning speed: 336.1 mm/s, the peak power density: 17.46 ? 10⁸ W/cm².     

Inactivation of microorganisms by low-frequency high-power ultrasound: 1. Effect of growth phase and capsule properties of the bacteria

The aim of this study was to determine the effects of high-intensity low-frequency (20 kHz) ultrasound treatment on the viability of bacteria suspension. More specifically, we have investigated the relationship between the deactivation efficiency and the physical (size, hydrophobicity) and biological (gram-status, growth phase) properties of the microbes. Enterobacter aerogenes, Bacillus subtilis, Staphylococcus epidermidis, S. epidermidis SK and Staphylococcus pseudintermedius were chosen for this study owing to their varying physical and biological properties. The survival ratio of the bacteria suspension was measured as a function of the ultrasound power (up to 13 W) for a constant sonication time of 20 min. Transmission electron microscopy was used to evaluate the ultrasound-induced damages to the microbes. Ultrasound treatment resulted in lethal damage to E. aerogenes and B. subtilis (up to 4.5-log reduction), whereas Staphylococcus spp. were not affected noticeably. Further, E. aerogenes suspensions were more sensitive to ultrasonication in exponential growth phase than when they were in stationary phase. The results of this study demonstrate that the main reason for bacterial resistance to ultrasonic deactivation is due to the properties of the bacterial capsule. Microbes with a thicker and “soft” capsule are highly resistant to ultrasonic deactivation process.     

Surface treatment of iron by electrochemical oxidation and subsequent annealing for the improvement of anti-corrosive properties

Corrosion resistance of iron oxides on iron foils prepared by anodization, annealing or a combination of both was characterized by electrochemical methods. Even though iron oxide film with a thickness of more than 2 μm could be prepared by single anodization, corrosion resistance deteriorated because the oxide film was in the amorphous phase and contained many defects. Corrosion resistance of iron oxides was also not enhanced by single annealing. Conversely, combination of anodization and subsequent annealing led to a positive shift of the corrosion potential in the Tafel plot, indicating that corrosion resistance was improved. Formation of thicker oxide during anodization was associated with a more positive shift in corrosion potential after annealing. Electrochemical impedance spectroscopy showed that the slowest charge transfer was observed in oxide films grown by a combination of anodization and annealing. We found that the optimum annealing temperature of anodic films in terms of the most positive shift of E_corr was 500 °C.     

Ultrasound assisted three phase partitioning of a fibrinolytic enzyme

The present investigation is aimed at ultrasound assisted three phase partitioning (UATPP) of a fibrinolytic enzyme from Bacillus sphaericus MTCC 3672. Three phase partitioning integrates the concentration and partial purification step of downstream processing of a biomolecule. Three phase system is formed with simultaneous addition of ammonium sulfate to crude broth and followed by t-butanol. UATPP of a fibrinolytic enzyme was studied by varying different process parameters such as ammonium sulfate saturation concentration, pH, broth to t-butanol ratio, temperature, ultrasound frequency, ultrasonication power, and duty cycle. The optimized parameters yielding maximum purity of 16.15-fold of fibrinolytic enzyme with 65% recovery comprised of 80% ammonium sulfate saturation, pH 9, temperature 30 °C, broth to t-butanol ratio 0.5 (v/v), at 25 kHz frequency and 150 W ultrasonication power with 40% duty cycle for 5 min irradiation time. SDS PAGE analysis of partitioned enzyme shows partial purification with a molecular weight in the range of 55–70 kDa. Enhanced mass transfer of UATPP resulted in higher fold purity of fibrinolytic enzyme with reduced time of operation from 1 h to 5 min as compared to conventional TPP. Outcome of our findings highlighted the use of UATPP as an efficient biosepartion technique.     

Ultrasonic energy input influence οn the production of sub-micron o/w emulsions containing whey protein and common stabilizers

Ultrasonication may be a cost-effective emulsion formation technique, but its impact on emulsion final structure and droplet size needs to be further investigated. Olive oil emulsions (20 wt%) were formulated (pH ～ 7) using whey protein (3 wt%), three kinds of hydrocolloids (0.1–0.5 wt%) and two different emulsification energy inputs (single- and two-stage, methods A and B, respectively). Formula and energy input effects on emulsion performance are discussed. Emulsions stability was evaluated over a 10-day storage period at 5 °C recording the turbidity profiles of the emulsions. Optical micrographs, droplet size and viscosity values were also obtained. A differential scanning calorimetric (DSC) multiple cool–heat cyclic method (40 to ?40 °C) was performed to examine stability via crystallization phenomena of the dispersed phase.Ultrasonication energy input duplication from 11 kJ to 25 kJ (method B) resulted in stable emulsions production (reduction of back scattering values, dBS ～ 1% after 10 days of storage) at 0.5 wt% concentration of any of the stabilizers used. At lower gum amount samples became unstable due to depletion flocculation phenomena, regardless of emulsification energy input used. High energy input during ultrasonic emulsification also resulted in sub-micron oil-droplets emulsions (D₅₀ = 0.615 μm compared to D₅₀ = 1.3 μm using method A) with narrower particle size distribution and in viscosity reduction.DSC experiments revealed no presence of bulk oil formation, suggesting stability for XG 0.5 wt% emulsions prepared by both methods. Reduced enthalpy values found when method B was applied suggesting structural modifications produced by extensive ultrasonication. Change of ultrasonication conditions results in significant changes of oil droplet size and stability of the produced emulsions.     

Sonochemical degradation of textile dyes in aqueous solution using sulphate radicals activated by immobilized cobalt ions

Decolorisation of dye solutions by cobalt activated persulphate and ultrasonication has been investigated. Rhodamine B, Methylene Blue dye (basic dyes) and Acid orange II, Acid scarlet red 3R (acid dyes) were used as model compounds in this study. Immobilized cobalt ions, activated the persulphate to form highly reactive sulphate radicals. The degradation studies were conducted with only persulphate (PS), cobalt activated persulphate (PS + Co), persulphate + ultrasonication (PS + US) and cobalt activated persulphate + ultrasonication (PS + US + Co). The decolorisation efficiency were in the order of PS < PS + Co < PS + US < PS + US + Co for all the four dye solutions. The effect of pH, dosage of persulphate as well as catalyst and contact time was investigated. Under the optimum condition, the decolorisation obeyed first-order kinetics. Nearly 90–97% of decolorisation was achieved with COD and TOC removal of about 65–73% and 53–62%, respectively, were achieved within an hour.     

Low frequency ultrasound-assisted leaching of sewage sludge for toxic metal removal,dewatering and fertilizing properties preservation

The aim of this research was to evaluate the efficiency of an ultrasonication process in combination with METIX-AC technology, a chemical process, for metal removal and dewatering ability, with preservation of fertilizing properties of sewage sludge. Waste activated sludge samples having a total solids concentration of 4 and 20 g L^?1, were enriched with Cu and Zn in order to exceed the limiting values recommended by Québec regulation for sludge valorization. Ultrasonication was applied at low frequency (22 kHz) at specific energies ranging from 180 to 66,000 kJ kg^?1 of dry sludge. Ultrasound-assisted leaching rates and yields were similar to chemical leaching alone for Zn, whereas solubilisation was reduced for Cu, regardless the specific energy. Fertilizing properties preservation and dewatering ability were similar in ultrasound-assisted leaching, compared to chemical leaching alone. These trends were mainly attributed to the changes of metal speciation, particle size and morphology during ultrasonication, in addition to untreated sludge properties.     

Preparation of resveratrol-enriched grape juice from ultrasonication treated grape fruits

Grape (Vitis spp.) is a major source of resveratrol that can be eaten directly or after making jam, jelly, wine and juice. Resveratrol (3,5,4′-trihydroxystilbene) has a profound positive influence on human health, including anti-carcinogenic, anti-cancer, anti-inflammatory, and anti-ageing effects and the ability to lower blood sugar. During industrial production of grape juice, resveratrol is lost because of the use of clarifying agents and filtration; therefore, commercial grape juice contains very low amounts of resveratrol. In this study, we investigated the accumulation of resveratrol in grape juice prepared from three varieties of grape, viz. Campbell Early, Muscat Bailey A (MBA) and Kyoho, following post-harvest ultrasonication cleaning for 5 min and 6 h of incubation in the dark at 25 °C. This process resulted in the amounts of resveratrol increasing by 1.53, 1.15 and 1.24 times in juice prepared from Campbell Early, MBA and Kyoho, respectively, without changing the amounts of total soluble solids. Overall, our results indicate that ultrasonication treatment of post-harvested grape fruits can be an effective method for producing resveratrol-enriched grape juice as well as cleaning grapes thoroughly.     

Ultrasonic-assisted synthesis and structural characterization of two new nano-structured Hg(II) coordination polymers

Two new Hg(II) coordination polymers containing N,N′-Bis-pyridin-3-ylmethylene-naphtalene-1,5-diamine ligand were synthesized by conventional and sonochemical methods, characterized by spectroscopic techniques (FT-IR and elemental analysis), and their X-ray crystallographic structures were determined. The crystal packing and supramolecular features of these coordination polymers were studied using geometrical analysis and Hirshfeld surface analysis. The crystal structure analysis revealed that H⋯H contacts, C–H⋯π and C–H⋯X (X = Cl for 1 and X = Br for 2) hydrogen bonding interactions are strong enough to govern the supramolecular architecture. The BFDH analysis helps us to compare the predicted morphology to that obtained under ultrasonication. This study may provide further insight into discovering the role of weak intermolecular interactions in the context of nano-supramolecular assembly.     

Ultrasonic emulsification of eucalyptus oil nanoemulsion: Antibacterial activity against Staphylococcus aureus and wound healing activity in Wistar rats

The plant derived essential oil nanoemulsion was prepared using a mixture of components containing eucalyptus oil as organic phase, water as continuous phase, and non ionic surfactant, Tween 80, as emulsifier at a particular proportion of 1:1 v/v%. The ultrasonication was applied for varied processing time from 0 to 30 min to study the effect of time on the formation of nanoemulsion and physical stability of formulation by this method. The transparency and stability of emulsion was enhanced when the sonication time was increased compared to hand blender emulsion. The most stable nanoemulsion was obtained in 30 min sonication having the mean droplet diameter of 3.8 nm. The antibacterial studies of nanoemulsion against Staphylococcus aureus by time kill analysis showed complete loss of viability within 15 min of interaction. Observations from scanning electron microscopy of treated bacterial cells confirmed the membrane damage compared to control bacteria. Furthermore, the wound healing potential and skin irritation activity of the formulated nanoemulsion in Wistar rats, suggested non-irritant and higher wound contraction rate with respect to control and neomycin treated rats. These results proposed that the formulated system could be favourable for topical application in pharmaceutical industries.     

Experimental study on the influence of low-frequency and low-intensity ultrasound on the permeability of the Mycobacterium smegmatis cytoderm and potentiation with levofloxacin

Tuberculosis is an infectious disease caused by the bacterium M. tuberculosis. The aim of this study was to investigate the bactericidal effect and underlying mechanisms of low-frequency and low-intensity ultrasound combined with levofloxacin treatment against M. smegmatis (a surrogate of M. tuberculosis). As part of this study, M. smegmatis was continuously irradiated with low frequency ultrasound (42 kHz) using several different doses whereby both intensity (0.138, 0.190 and 0.329 W/cm²) and exposure time (5, 15 and 20 min) were varied. Flow cytometric analyses revealed that the permeability of M. smegmatis increased following ultrasound exposure. The survival rate, structure and morphology of bacteria in the lower-dose (I_SATA = 0.138 W/cm² for 5 min) ultrasound group displayed no significant differences upon comparison with the untreated group. However, the survival rate of bacteria was significantly reduced and the bacterial structure was damaged in the higher-dose (I_SATA = 0.329 W/cm² for 20 min) ultrasound group. Ultrasound irradiation (0.138 W/cm²) was subsequently applied to M. smegmatis in combination with levofloxacin treatment for 5 min. The results demonstrated that the bactericidal effect of ultrasonic irradiation combined with levofloxacin is higher compared to ultrasound alone or levofloxacin alone.