Influence of the dissolution rate on the collapse and shedding behavior of monostearin/monopalmitin-rich coated microbubbles

A low cost food grade emulsifier (a mixture of monoglycerides, diglycerides, and sodium stearoyl lactylate) in combination with polyethylene glycol-40 stearate (PEG-40S) was used as an alternative to pure saturated phospholipids to form the thin shell of a microbubble. To investigate the stability of these microbubbles in a water system over time, their dissolution behavior was studied at various degas factors and at two percentages of PEG-40S. It was found that the favored shell collapse/shedding mechanism switched, as the dissolution rate increased (degas factor decreased), from folding with a smooth surface contour to buckling accompanied by surface folding/expulsion with a cyclic buckled-smooth surface contour. The compositional change that we made played a more minor role, mainly controlling the resistance to mass transfer of the microbubble shell and again modifying the mechanism-determinant dissolution rate. The shell resistance behavior for these microbubbles varied from that of previous lipid/PEG-40S-coated microbubbles by the presence of a maximum in shell resistance during dissolution. We hypothesize that the dominance of one collapse mechanism over another for these compositions is related to the time scales of two competing processes, fold nucleation and area compression. For these mixtures at room temperature, we estimate that the maximum area compression rate for folding as the major collapse mechanism is approximately 0.2 s (-1), a rate unattainable in a traditional Langmuir trough but achievable by the use of a dissolving microbubble.     

Stability and rheological behavior of concentrated monodisperse food emulsifier coated microbubble suspensions

Nearly monodispersed populations of microbubbles were produced using flow focusing with a food grade emulsifier. The microbubbles produced by this technique have diameters in the range of 120-200 microm. The flow focusing device uses metered streams of air and liquid to produce a jet that periodically pinches to make individual microbubbles. The size of the microbubbles can be controlled by changing the relative flow rates of the gas and the liquid. The emulsifier consists of a mixture of monoglycerides, diglycerides and sodium stearoyl lactylate in combination with polyethylene glycol (PEG)-40 stearate. The emulsifier forms a thin shell that stabilizes the microbubbles. The microbubbles are stable over time with their sizes remaining roughly constant over 2 h. Such stability allows suspensions of microbubbles to be formed and their rheological properties tested. The sizes of the microbubbles are also monitored off-line while testing, examining the effect of shearing on the bubble sizes, as well as their stability over time. These results show that the microbubble suspensions are viscoelastic and exhibit power law behavior. The relationship between the air fraction of the suspension and fluid viscosity is determined.     

Surface phase behavior and microstructure of lipid/PEG-emulsifier monolayer-coated microbubbles

Langmuir trough methods and fluorescence microscopy were combined to investigate the phase behavior and microstructure of monolayer shells coating micron-scale bubbles (microbubbles) typically used in biomedical applications. The monolayer shell consisted of a homologous series of saturated acyl chain phospholipids and an emulsifier containing a single hydrophobic stearate chain and polyethylene glycol (PEG) head group. PEG-emulsifier was fully miscible with expanded phase lipids and phase separated from condensed phase lipids. Phase coexistence was observed in the form of dark condensed phase lipid domains surrounded by a sea of bright, emulsifier-rich expanded phase. A rich assortment of condensed phase area fractions and domain morphologies, including networks and other novel structures, were observed in each batch of microbubbles. Network domains were reproduced in Langmuir monolayers under conditions of heating–cooling followed by compression–expansion, as well as in microbubble shells that underwent surface flow with slight compression. Domain size decreased with increased cooling rate through the phase transition temperature, and domain branching increased with lipid acyl chain length at high cooling rates. Squeeze-out of the emulsifier at a surface pressure near 35 mN/m was indicated by a plateau in Langmuir isotherms and directly visualized with fluorescence microscopy, although collapse of the solid lipid domains occurred at much higher surface pressures. Compression of the monolayer past the PEG-emulsifier squeeze-out surface pressure resulted in a dark shell composed entirely of lipid. Under certain conditions, the PEG-emulsifier was reincorporated upon subsequent expansion. Factors that affect shell formation and evolution, as well as implications for the rational design of microbubbles in medical applications, are discussed.     

DNA and polylysine adsorption and multilayer construction onto cationic lipid-coated microbubbles

We report on a novel application of the layer-by-layer (LbL) assembly technique to attach multiple layers of DNA and poly-l-lysine (PLL) onto preformed lipid-coated microbubbles to increase the DNA loading capacity. We first measured the effects of the cationic lipid fraction and salt concentration on the microbubble stability. Microbubble production and stability were robust up to a cationic lipid fraction of 40 mol % in 10 mM NaCl. DNA adsorption was heterogeneous over the microbubble shell and occurred primarily on the condensed phase domains. The amount of adsorbed DNA, and subsequently adsorbed PLL, increased linearly with the fraction of cationic lipid in the shell. DNA loading was further enhanced by the LbL assembly method to construct polyelectrolyte multilayers (PEMs) of DNA and PLL. PEM buildup was demonstrated by experimental results from zeta potential analysis, fluorescence microscopy, UV spectroscopy, and flow cytometry. The PEMs exhibited two growth stages and were heterogeneously distributed over the microbubble surface. The DNA loading capacity onto the microbubbles was enhanced by over 10-fold by using five paired layers. However, the PEM shell did not prevent oscillation or destruction during ultrasound insonification. These results suggest that the surface can be compartmentalized to make multifunctional, high-payload ultrasound contrast agents for targeted gene therapy.     

Miscibilities of methanol and ethanol with dodecylammonium chloride in the adsorbed film and micelle

The surface tension of the aqueous solutions of methanol– dodecylammonium chloride (DAC) and ethanol–DAC mixtures has been measured as a function of the total molality of the mixture m^ and the mole fraction of DAC X^₂ in the mixture at 298.15?K under atmospheric pressure. The compositions of the adsorbed film at 50, 40, and 30?mN?m^-1, and that of the micelle at the critical micelle concentration (CMC) have been evaluated by applying the thermodynamic equations derived previously and shown in the form of phase diagrams of adsorption and micelle formation. It has been found that (i) methanol is hardly miscible in M28.8nthe adsorbed film and micelle, and (ii) ethanol molecules are very slightly incorporated into adsorbed film of DAC at high-surface tension and into the micelle, although hardly miscible in the adsorbed film at low-surface tension. By comparing the corresponding phase diagrams of ethanol-2- (octylsulfinyl)ethanol (OSE) mixture, furthermore, it has been shown that there exists a difference in the miscibility of ethanol between DAC and OSE.     

Aqueous Suspensions of Poly(ethylene Glycol)/Pyrocarbon/Fumed Silica

Aqueous suspensions of fumed silica and pyrocarbon/silica (CS) in the presence of dissolved poly(ethylene glycol) (PEG) were studied using (1)H NMR spectroscopy with freezing-out of bulk water and quantum chemical computations of the chemical shifts. The freezing effect for PEG/water is akin to that for low-molecular organics, as formation of solid phases of water (ice) and PEG occurs, and their mixture forms at the eutectic temperature. In the aqueous suspensions of fumed silica or CS, PEG molecules are localized at the solid-liquid interfaces and do not form the bulk solution even at large concentrations; however, the amount of bulk undisturbed water rises due to formation of the immobilized PEG layer. For such suspensions of silica or CS at a low amount of pyrocarbon (C(C)=4 wt%), there is a portion of the graph of the surface free energy (gamma(S)) increasing nearly linearly with the PEG concentration (C(PEG)); however, in the case of large C(C)=40 wt% in CS, a similar effect is not observed, as gamma(S) is maximal at low C(PEG)=0.1 wt%. Copyright 2001 Academic Press.     

Fabrication of hollow melamine-formaldehyde microcapsules from microbubble templates

A fabrication method for hollow melamine-formaldehyde microcapsules from microbubble templates is presented. This method is based on the direct encapsulation of microbubbles, and thus does not require a liquid- or solid-core decomposition process. This study determined the conditions for controlling the surface morphology, shell thickness, and diameter distribution of hollow microcapsules. Results showed that the surface morphology of these hollow microcapsules depended on the reaction time, glycine concentration (pH of aqueous continuous phase) and pre-polymer concentration. The capsule shell thickness could be controlled by adjusting the concentration of aniline that had adsorbed on the microbubble surface and reacted with pre-polymer. The capsule diameter depended on the dissolution rate of gases, and the diameter of the hollow microcapsules fabricated from air microbubble templates ranged from 5 to 200 microm.     

Poly(ethylene glycol) enhances the surface activity of a pulmonary surfactant

The primary role of lung surfactant is to reduce surface tension at the air–liquid interface of alveoli during respiration. Axisymmetric drop shape analysis (ADSA) was used to study the effect of poly(ethylene glycol) (PEG) on the rate of surface film formation of a bovine lipid extract surfactant (BLES), a therapeutic lung surfactant preparation. PEG of molecular weights 3350; 8000; 10,000; 35,000; and 300,000 in combination with a BLES mixture of 0.5 mg/mL was studied. The adsorption rate of BLES alone at 0.5 mg/mL was much slower than that of a natural lung surfactant at the same concentration; more than 200 s are required to reach the equilibrium surface tension of 25 mJ/m². PEG, while not surface active itself, enhances the adsorption of BLES to an extent depending on its concentration and molecular weight. These findings suggest that depletion attraction induced by higher molecular weight PEG (in the range of 8000 to 35,000) may be responsible for increasing the adsorption rate of BLES at low concentration. The results provide a basis for using PEG as an additive to BLES to reduce its required concentration in clinical treatment, thus reducing the cost for surfactant replacement therapy.     

Use of Nitric Oxide Donor-Loaded Microbubble Destruction by Ultrasound in Thrombus Treatment

In the presence of a vascular thrombus, the recovery of blood flow and vascular recanalization are very important to prevent tissue damage. An alternative procedure to thrombolysis is required for patients who are unable to receive surgery or thrombolytic drugs due to other physical conditions. Recently, the performance of thrombolysis combined with microbubbles has become an attractive and effective therapeutic procedure. Indeed, in a recent study, we demonstrated that, upon exposure to ultrasound, liposomes loaded with nitric oxide release agonists conjugated to microbubbles; therefore, there is potential to release the agonist in a controlled manner into specific tissues. This means that the effect of the agonist is potentiated, decreasing interactions with other tissues, and reducing the dose required to induce nitric-oxide-dependent vasodilation. In the present study, we hypothesized that a liposome microbubble delivery system can be used as a hydrophilic agonist carrier for the nitric oxide donor spermine NONOate, to elicit femoral vasodilation and clot degradation. Therefore, we used spermine-NONOate-loaded microbubbles to evaluate the effect of ultrasound-mediated microbubble disruption (UMMD) on thromboembolic femoral artery recanalization. We prepared spermine NONOate-loaded microbubbles and tested their effect on ex vivo preparations, hypothesizing that ultrasound-induced microbubble disruption is associated with the vasorelaxation of aortic rings. Thrombolysis was demonstrated in aorta blood-flow recovery after disruption by spermine NONOate-loaded microbubbles via ultrasound application in the region where the thrombus is located. Our study provides an option for the clinical translation of NO donors to therapeutic applications.     

Effect of solution viscosity on dynamic surface tension detection

A dynamic surface tension detector (DSTD) was used to examine the molecular diffusion and surface adsorption characteristics of surface-active analytes as a function of solution viscosity. Dynamic surface tension is determined by measuring the differential pressure across the air/liquid interface of repeatedly growing and detaching drops. Continuous surface tension measurement throughout the entire drop growth is achieved for each eluting drop (at a rate of 30 drops/min for 2 μl drops), providing insight into the kinetic behavior of molecular diffusion and orientation processes at the air/liquid interface. Three-dimensional data are obtained through a calibration procedure previously developed, but extended herein for viscous solutions, with surface tension first converted to surface pressure, which is plotted as a function of elution time axis versus drop time axis. Thus, an analyte that lowers the surface tension results in an increase in surface pressure. The calibration procedure derived for the pressure-based DSTD was successfully extended and implemented in this report to experimentally determine standard surface pressures in solutions of varied viscosity. Analysis of analytes in viscous solution was performed at low analyte concentration, where the observed analyte surface activity indicates that the surface concentration is at or near equilibrium when in a water mobile phase (viscosity of 1.0 Cp). Two surface-active analytes, sodium dodecyl sulfate (SDS) and polyethylene glycol (MW 1470 g/mol, PEG 1470), were analyzed in solutions ranging from 0 to 60% (v/v) glycerol in water, corresponding to a viscosity range of 1.0-15.0 Cp. Finally, the diffusion-limited surface activity of SDS and PEG 1470 were observed in viscous solution, whereby an increase in viscosity resulted in a decreased surface pressure early in drop growth. The dynamic surface pressure results reported for SDS and PEG 1470 are found to correlate with solution viscosity and analyte diffusion coefficient via the Stokes-Einstein equation.     

Ultrasound-responsive polymer-coated microbubbles that bind and protect DNA

Ultrasound in combination with microbubbles has recently been considered by gene delivery scientists to be an interesting approach to enhance gene transfer into cells. Its low toxicity and simplicity to apply in vivo without major complications make this technology (sonoporation) especially attractive. Sonoporation of DNA has been evaluated in vivo by the injection of free plasmid DNA (pDNA) together with microbubbles (as used in diagnostic imaging) in the bloodstream. However, the in vivo gene-transfer efficiency in these experiments remained rather low. Both the enzymatic degradation of the injected pDNA as well as the low pDNA concentration in the neighborhood of sonoporated cell membranes may explain this low efficiency. Therefore, we developed polymer-coated microbubbles that can bind and protect the pDNA. Coating albumin-shelled microbubbles with poly(allylamine hydrochloride) (PAH) makes the surface charge of the microbubbles positive without drastically affecting the size distribution of the microbubbles, thereby not affecting the ultrasound responsiveness and injectability. The cationic coating allowed both to bind up to 0.1 pg of DNA per microbubble as well as to protect the bound DNA against nucleases. Finally, the PAH coating significantly increased the lifetime of the microbubbles (half-life approximately 7 h), making them more convenient for in vivo applications because more microbubbles are expected to reach the target organ. Binding and nuclease protection of DNA by polymer-coated diagnostic microbubbles has, to our knowledge, never been demonstrated. We conclude that these LbL-coated microbubbles might be significant in the further development of ultrasound-mediated gene delivery.     

丙烯酰胺型阴离子表面活性单体的化学结构与其胶束化行为

对两种丙烯酰胺型阴离子表面活性单体(2-丙烯酰胺基十四烷磺酸钠, NaAMC₁₄S; 2-丙烯酰胺基十二烷磺酸钠, NaAMC₁₂S)的化学结构与胶束化行为的关系进行了较深入的研究. 使用紫外分光光度法测定了NaAMC₁₄S, NaAMC₁₂S及十二烷基磺酸钠(SDS)在水中的溶解度, 同时采用表面张力法(环法)测定了它们在不同温度下的临界胶束浓度CMC; 采用稳态荧光探针法测定了不同浓度的胶束聚集数与本征胶束聚集数. 实验结果表明, 与普通表面活性剂相比, 由于丙烯酰胺型阴离子表面活性单体分子中具有两个亲水头基, 在水中的溶解性能较强, 故具有较低的Krafft温度; 在溶液表面的饱和吸附量低, 故降低水表面张力的能力较差, 即表面活性差; 疏水缔合的胶团较为疏松, 故聚集数很小; 胶束内分子间的疏水相互作用较弱, 故临界胶束浓度CMC较高.     

Novel calibration of a dynamic surface tension detector: flow injection analysis of kinetically-hindered surface active analytes

First, a novel technique for calibration of a dynamic surface tension detector (DSTD) is described. The DSTD measures the differential pressure as a function of time across the liquid-air interface of growing drops that repeatedly form and detach at the end of a capillary tip. The calibration technique utilizes the ratio of pressure signals acquired from the drop growth of two separate solutions, i.e. a standard solution and a corresponding mobile phase, such as water, both of which have a known surface tension. Once calibrated, the dynamic surface tension of an analyte is obtained from the ratio of the pressure signals from the analyte solution to that of the mobile phase solution. Thus, this calibration technique eliminates the need to optically image the radius of the expanding drop of liquid. Accurate dynamic surface tension determinations were achieved for aqueous sodium dodecyl sulfate (SDS) solutions over a concentration range of 0.5-5.4 mM. The measured surface tensions for these SDS solutions range from 70.3 to 46.8 dyne/cm and were in excellent agreement with the literature. A precision of 0.2 dyne/cm (1 S.D.) was routinely obtained. Second, the DSTD with this calibration technique was combined with flow injection analysis (FIA) for the study of model protein solutions and polymer solutions. The kinetic surface tension behavior of aqueous bovine serum albumin (BSA) solutions as a function of concentration and flow rate is presented. Evaluation of the dynamic surface tension data illustrates that a protein such as BSA initially exhibits kinetically-hindered surface tension lowering, i.e. a time dependence, as BSA interacts with the liquid-air interface of an expanding drop. FIA/DSTD is then shown to be an effective tool for the rapid study of kinetically-hindered surfactant mixtures. It was found that mixtures of SDS and the polymeric surfactant Brij(R)-35 (lauryl polyoxyethylene ether with an average molecular weight of 1200 g/mol) result in essentially an additive lowering of the surface tension. Mixtures of polyethylene glycol (PEG), with an average molecular weight of 1470 g/mol, and Brij(R)-35, however, result in a competitive (non-additive) surface tension with the Brij(R)-35 dominating the response.     

Synthesis and interfacial properties of sophorolipid derivatives

Biosurfactants made by fermentation from renewable resources provide “environmental friendly” processes and products. A natural sophorolipid mixture was produced by the yeast Candida bombicola when cultured on glucose and oleic acid. The sophorolipid mixture was chemically modified to form the corresponding sophorolipid alkyl (methyl, ethyl, propyl, and butyl) esters by reaction with the corresponding sodium alkoxides. Interfacial properties of these surfactants, such as surface tension reduction, aggregation, and adsorption, were systematically studied. It was found that the critical micelle concentration of sophorolipid esters decreases to about 1/2 per additional one CH₂ group to the alkyl ester moiety. Interestingly, these surfactants were found to adsorb strongly on alumina but weakly on silica. They have properties that make them attractive candidates for uses in detergents, cosmetics, soil remediation, and enhanced oil recovery.     

Miscibility of sodium chloride and sodium dodecyl sulfate in the adsorbed film and aggregate

The adsorption, micelle formation, and salting out of sodium dodecyl sulfate in the presence of sodium chloride were studied from the viewpoint of their mixed adsorption and aggregate formation. The surface tension of aqueous solutions of a sodium chloride–sodium dodecyl sulfate mixture was measured as a function of the total molality and composition of the mixture. Phase diagrams of adsorption and aggregate formation were obtained by applying thermodynamic equations to the surface tension. Judging from the phase diagrams, sodium chloride and sodium dodecyl sulfate are miscible in the adsorbed film at very large composition of sodium chloride and in the salted-out crystalline particle, while they are immiscible in the micelle. The miscibilities in the adsorbed film, micelle, and crystalline particle increase in the following order: particle > adsorbed film > micelle. The difference in miscibility among the oriented states was ascribed to the difference in geometry between the adsorbed film and micelle and to the interaction between bilayer surfaces in the particle.     

Microbubble formation using asymmetric Shirasu porous glass (SPG) membranes and porous ceramic membranes—A comparative study

We have recently proposed a new method for generating uniformly sized microbubbles from Shirasu porous glass (SPG) membranes with a narrow pore size distribution. In this study, to obtain a high gas permeation rate through SPG membranes in microbubble formation process, asymmetric SPG membranes were used. At the transmembrane/bubble point pressure ratio of less than 1.50, uniformly sized microbubbles with a bubble/pore diameter ratio of approximately 9 were generated from an asymmetric SPG membrane with a mean pore diameter of 1.58 μm and a skin-layer thickness of 12 ± 2 μm at a gaseous-phase flux of 2.1–24.6 m³ m⁻² h⁻¹, which was much higher than that through a symmetric SPG membrane with the same pore diameter. This is mainly due to the much smaller membrane resistance of the asymmetric SPG membrane. Only 0.27–0.43% of the pores of the asymmetric SPG membrane was active under the same conditions. The proportion of active pores increased with a decrease in the thickness of skin layer. In contrast to the microbubble formation from asymmetric SPG membranes, polydispersed larger bubbles were generated from asymmetric porous ceramic membranes used in this study, due to the surface defects on the skin layer. The surface defects were observed by the scanning electron microscopy and detected by the bubble point method.     

Prediction of the Properties of a Binary Surfactant Mixture of Arlacel‐165 and Myrj‐59 in Aqueous System

Foamability and foam stability, emulsifying power, surface tension, and interfacial tension were investigated for different ratios of binary surfactant system of Arlacel‐165 (glyceryl stearate (and) PEG‐100 stearate) and Myrj‐59 (polyoxyethylene 100 stearate). Among all the ratios tested for their foaming power and foamabilty, the ratios 8:2, 5:5, 4:6, 2:8, and 1:9 of Arlacel‐165 and Myrj‐59 showed the best results. At these ratios, the foaming power and foamability was found to be 100%. The surfactants having foam stability more than 50% can be considered as metastable and those less than 50% are considered as low‐stability foams. In case of surface tension and interfacial tension property measurements, 8:2 and 9:1 showed the best results. At 8:2 and 9:1 of Arlacel‐165 and Myrj‐59, the surface tension was found to be 37.7 dynes/cm and 1.33 dynes/cm respectively at 30°C ambient temperature. Also, 7:3 of this binary mixture was found to exhibit the best emulsifying power among all the ratios tested. At 30°C, the emulsifying property of the binary mixture was 6 hours.     

疏水改性水溶性聚合物/表面活性剂溶液性质

表面张力;疏水改性水溶性聚合物/表面活性剂溶液性质     

Tunable diacetylene polymerized shell microbubbles as ultrasound contrast agents

Monodisperse gas microbubbles, encapsulated with a shell of photopolymerizable diacetylene lipids and phospholipids, were produced by microfluidic flow focusing, for use as ultrasound contrast agents. The stability of the polymerized shell microbubbles against both aggregation and gas dissolution under physiological conditions was studied. Polyethylene glycol (PEG) 5000, which was attached to the diacetylene lipids, was predicted by molecular theory to provide more steric hindrance against aggregation than PEG 2000, and this was confirmed experimentally. The polymerized shell microbubbles were found to have higher shell-resistance than nonpolymerizable shell microbubbles and commercially available microbubbles (Vevo MicroMarker). The acoustic stability under 7.5 MHz ultrasound insonation was significantly greater than that for the two comparison microbubbles. The acoustic stability was tunable by varying the amount of diacetylene lipid. Thus, our polymerized shell microbubbles are a promising platform for ultrasound contrast agents.