Search

Drop shape techniques are extensively used for surface tension measurement. The accuracy of surface tension measurements using a pendant drop setup depends mainly on the drop shape. As the pendant drop shape becomes close to spherical, the performance of drop shape techniques deteriorates. It is important to consider how to optimize the pendant drop design and how to quantify the accuracy of the surface tension measurements. In this paper, the design and accuracy of pendant drop methods is considered in detail. Dimensional analysis is used to describe similarity in pendant drop shapes and to express the problem using appropriate dimensionless groups. A quantitative criterion called shape parameter is then used to express quantitatively the difference in shape between a given experimental drop and a spherical shape. The shape parameter is found to depend only on two dimensionless groups: the dimensionless volume and the Bond number. A critical shape parameter (minimum value of the shape parameter that guarantees a specified accuracy) is shown to depend only on Bond number. A set of experiments were performed with pure liquids to illustrate the change of the critical shape parameter with the Bond number. Results show that high accuracy is indeed reachable independent of the algorithms used.

This paper presents a continuation of the development of a drop shape method for film studies, ADSA-CSD (Axisymmetric Drop Shape Analysis-Constrained Sessile Drop). ADSA-CSD has certain advantages over conventional methods. The development presented here allows complete exchange of the subphase of a spread or adsorbed film. This feature allows certain studies relevant to lung surfactant research that cannot be readily performed by other means. The key feature of the design is a second capillary into the bulk of the drop to facilitate addition or removal of a secondary liquid. The development will be illustrated through studies concerning lung surfactant inhibition. After forming a sessile drop of a basic lung surfactant preparation, the bulk phase can be removed and exchanged for one containing different inhibitors. Such studies mimic the leakage of plasma and blood proteins into the alveolar spaces altering the surface activity of lung surfactant in a phenomenon called surfactant inhibition. The resistance of the lung surfactant to specific inhibitors can be readily evaluated using the method. The new method is also useful for surfactant reversal studies, i.e. the ability to restore the normal surface activity of an inhibited lung surfactant film by using special additives. Results show a distinctive difference between the inhibition when an inhibitor is mixed with and when it is injected under a preformed surfactant film. None of the inhibitors studied (serum, albumin, fibrinogen, and cholesterol) were able to penetrate a preexisting film formed by the basic preparation (BLES and protasan), while all of them can alter the surface activity of such preparation when mixed with the preparation. Preliminary results show that reversal of serum inhibition can be easily achieved and evaluated using the modified methodology.

The film adsorption of bovine lipid extract surfactant (BLES) onto the air–liquid interface was examined using axisymmetric drop shape analysis. In combination with a pendant drop constellation, BLES concentrations as high as 10 mg/ml were studied, i.e. concentrations far higher than those accessible with the captive bubble set-ups. ‘Adsorption clicks’, i.e. dynamic processes in which the interfacial tension of surfactant films decreases quickly in a stepwise fashion, were studied at concentrations below 1 mg/ml. Adsorption clicks with high magnitudes up to approximately 35 mJ/m2 (within 0.2 s) were observed. The rate of adsorption was investigated as a function of surfactant concentration. At concentrations below 1 mg/ml, the rate of adsorption is highly concentration dependent. Surfactant films formed on 1 mg/ml BLES solutions reached a surface tension of about 25 mJ/m2 in approximately 10 s, while 0.1 mg/ml BLES required more than 100 s to reach a similar value.

Molecular interactions between small molecules and proteins, such as binding of lipids to proteins, are of fundamental importance in various biological processes. A recently-developed method based on dynamic surface tension measurement is efficient and versatile in detecting such molecular interactions: Axisymmetric Drop Shape Analysis (ADSA) provides a tool for measuring the surface tension (γ) response to surface area changes. Through the analysis of the γ response pattern, surface competitive adsorption between small organic molecules and protein molecules can be detected. Surface squeeze-out of small molecules by proteins can also be observed. Molecular binding of lipids to proteins manifests itself in a modification of the γ response which is not compatible with a simple superposition of the two individual patterns. The specific binding can be studied in terms of dose effects and specificity.

Helicobacter pylori is a recently recognized bacteria which colonizes the stomach and causes gastritis, gastric cancers and peptic ulceration. However gastric and duodenal ulcers occur in only a minority of H. pylori -infected individuals. Surface hydrophobicity is an important virulence factor of many pathogenic bacteria since it is known to be involved in mediating mucosal adherence. In this study, surface hydrophobicity of H. pylori strains originally isolated from children and adolescents was correlated with gastroduodenal pathology including both endoscopic evidence of duodenal ulceration and histologic evidence of duodenitis. Contact angle measurements on 11 H. pylori isolates were used to characterize surface hydrophobicity. Axisymetric drop-shape analysis revealed contact angles ranging from 7.5 to 15.3°. Contact angles of the strains isolated from patients with duodenal ulcers (11.9±2.6°) were not different from values in isolates obtained from children with gastritis alone (8.9±5.2°; P =0.16). Contact angles of H. pylori isolated from patients with and without duodenitis were also comparable (10.9±2.8° and 10.0±9.6°; P =0.69). These findings indicate that additional virulence properties likely are required for H. pylori to induce peptic ulceration in infected humans.

The surface tension of aqueous solutions of Polymer JR400, the chloride salt of a trimethylammonium derivative of hydroxyethyl cellulose, has been measured as a function of polymer concentration. Axisymmetric drop shape analysis profile has been employed to record the dynamic surface tension response of the solutions to a sawtooth-shaped variation in the surface area. Dilute solutions of the polymer (concentration

Samples of pulmonary surfactant were obtained from rabbits by repeated lung lavage. Surface tension measurements of the samples were made in such a way as to avoid the possibility of film leakage. It was confirmed that the relative magnitudes of the surface tension values are strongly correlated to the amount of phospholipids in the samples. Furthermore, the surface tension measurements are more sensitive means to detect the difference between the lavage samples than the total phospholipid content.

Of the many methodologies used in surface thermodynamic experimentation, Axisymmetric Drop Shape Analysis (ADSA) has proven to be one of the most accurate and versatile. Until recently, however, ADSA has had two main limitations: (1) the inability to analyse drops with near-zero curvature at the apex; and (2) problems with drop edge detection when the contrast between the drop and the background is not adequately sharp. In this paper, ‘second-generation’ ADSA algorithms and new image analysis schemes are introduced as solutions to these shortcomings. To illustrate these improvements, an inverted sessile air bubble in a pulmonary surfactant suspension is used. This ‘captive bubble’ geometry was chosen to overcome the problems of film leakage at low surface tensions. The modified experimental apparatus, called ADSA-captive bubble (CB), in conjunction with the new software, proves to be a powerful technique to investigate these types of systems. ADSA-CB is fully automated, requiring less user intervention, enabling the collection of large amounts of data, and thereby providing intricate details of surface behaviour. The so-called ‘squeeze-out’ and film collapse phenomena are examined closely. To demonstrate the problems of film leakage, pendant (hanging) drop experimental results are also presented.

Axisymmetric Drop Shape Analysis has been employed to measure the dynamic surface tension response to a sawtooth-shaped variation in the surface area. Four systems were studied: (a) bovine serum albumin (BSA) aqueous solution; (b) dimethyl sulfoxide (DMSO) added to the BSA aqueous solution; (c) ethyl alcohol (ethanol) added to the BSA aqueous solution; and (d) hepoxilin methyl ester, a lipidic hydroxy epoxide derived from arachidonic acid, dissolved in DMSO and then added to the BSA aqueous solution. In the mixtures of the solutions, competitive adsorption to the surface is revealed between the smaller molecules and the protein molecules, through the change of the pattern in the dynamic surface tension response to the surface area change. In the initial stage of the adsorption, the smaller organic molecules (DMSO, ethanol) diffuse to the surface at a faster rate than the protein molecules, impeding protein adsorption. However, in the cases of DMSO/BSA and ethanol/BSA solutions, the experiment indicates that the small organic molecules are squeezed out of the surface because of the compression/dilation process. After steady state is established, the surface tension pattern shows a skewed shape suggesting a conformational change in the protein. When hepoxilin, a naturally occurring biologically active C20 fatty acid containing a hydroxyl epoxide functionality, is present in the BSA solution, a rather symmetric shape is found in the dynamic surface tension changes, different from the skewed shape found in that of the pure BSA solution. This suggests that hepoxilin molecules interact with the protein, leading to a stabilization of one conformation of the protein molecules at the surface.

Axisymmetric drop shape analysis-profile (ADSA-P) was employed to obtain accurate measurements of the concentration dependence of the film pressure of human serum albumin at a water/decane interface. Fifteen aqueous human albumin solutions ranging in concentration from 0.0001 to 5 mg ml−1 were used. Three domains were identified in the effect of the bulk protein concentration, C, on the interfacial tension, γ: (a) a slow change of γ at low bulk concentrations (C 10−2 mg ml−1). At the two lowest bulk albumin concentrations, 0.0001 and 0.001 mg ml−1, negative interfacial pressures were observed that may be attributed to repulsive electrostatic interactions and the formation of a surface depletion layer. At all other concentrations, a positive surface pressure was measured. Moreover, based on our data on the variation of γ with C, Gibbs' adsorption equation was used to provide an account of the dependence of the surface concentration on C. The possibility of the applicability of the Gibbs' adsorption equation exists for low bulk protein concentrations (C < 10−3 mg ml−1).

The adsorption kinetics of human albumin (HA) at a 0.02 mg ml−1 aqueous solution/decane interface and the temperature dependence of the interfacial tension from 20 to 60°C were studied using Axisymmetric Drop Shape Analysis. This is a non-perturbation method which allows us to study the interfacial tension for a long period of time, without a significant change in the interfacial area. Two criteria were applied to obtain “experimental equilibrium” values of the interfacial tension and interfacial pressure. The rate of change of interfacial pressure with temperature was found to be about 0.1 mJm−2°C−1 from 20 to 45°C. The interfacial pressures were found to be approximately 10 mJ m−2 larger than those reported elsewhere for the HA solution/air interface at room temperature. For the concentration used, a plateau in the interfacial pressure was obtained and can be interpreted on the basis of saturation at temperatures above 45°C.

A pendant drop arrangement is employed in conjunction with the axisymmetric drop shape analysis-profile (ADSA-P) technique to study the effect of concentration and pH on the surface tension of insulin layers adsorbed at the solution-air interface. Three different insulin preparations are used: human zinc-free insulin and two of its analogs, B9AspB27Glu and B10Asp, at three concentrations, 0.01, 0.1 and 1.0 mg ml −1 , and two pH levels, 3 and 7.5. ADSA-P is shown to permit high resolution between the insulin preparations which differ only by one or two amino acids in their primary structures. Furthermore, the effects of pH and concentration on the surface tensions of insulin solutions are examined.

Contact angle data, measured by using a sessile drop arrangement in conjunction with Axisymmetric Drop Shape Analysis-contact Diameter (ADSA-CD), were used to quantify the effects of ammonia gas plasma treatment on the surface properties of previously untreated polystyrene surfaces. The surface tension of treated polystyrene samples is considerably higher than that of untreated samples. The increase in surface tension following plasma treatment is attributed to the addition of amine groups to the surface.

A micropipette aspiration technique was used to study the extent of adhesion of live and glutaraldehyde-fixed endothelial cells to four solid fibres with different surface tensions, γSV: glass (γSV > 72mj m−2); nylon (γSV = 41 mJ m−2); polyester (γSV = 30 mJ m−2); FC721-coated glass (γSV = 13 mJ m−2). The cells and the fibres were immersed in phosphatebuffered saline (PBS), distilled water (for fixed cells), and complete culture medium. The contact angles of PBS on layers of cells were measured by the Axisymmetric Drop Shape Analysis-Contact Diameter (ADSA-CD) technique. Subsequently, the contact angle data were used in conjunction with the equation of state approach to determine the cell surface tensions, γCV. For a live cell layer, the contact angle was 3.1 ° corresponding to a γCV value of approximately 72 mJ m−2, and for fixed cells, an average contact angle of 36.6° was measured corresponding to a γCV of approximately 60 mJ m−2. A small positive pressure of 1 Pa was applied to deposit the cells onto the fibres before the initiation of aspiration. The force of detachment is reported as a function of the duration of pressure application for the four test fibres in each medium. The results are interpreted by using a thermodynamic model for cell adhesion based on the relevant interfacial tensions. For both live and fixed cells, the forces of detachment increase with increasing duration of pressure application. The measured forces ranged from (3 ± 0) × 10−11N (live cells in culture medium on glass) to (3709 ± 374) × 10−11 N (fixed cells in PBS on FC721), where the errors are the 95% confidence limits. For fixed cells with PBS as a medium, the measured forces of detachment increased with decreasing solid surface tension. This corresponds to a trend predicted by the thermodynamic adhesion model for the case of surface tension of the liquid γLV > γCV and confirmed by the contact angle measurements. In water, although the magnitude of the forces is smaller than that in PBS, the trend is the same as that in PBS and is again predicted by the model. In complete culture medium, the extent of adhesion decreases greatly and the forces of detachment for all test surfaces are indistinguishable from one another. This can be attributed to the prior adsorption of proteins from the medium onto the fibres, resulting in an alteration of their surface properties. For live endothelial cells in PBS, the trend is opposite to that for the fixed cells. According to the thermodynamic model, this trend corresponds to the case of γLV

The adsorption kinetics of human albumin (HA) at the aqueous solution/air interface and the surface relaxation behaviour of HA adsorption layers after transient area disturbances are studied using an axisymmetric drop shape technique (ADSA). Transient relaxation measurements were performed with the ADSA technique by changing the volume of the pendent drop by definite increments. In the concentration interval studied, the relaxation behaviour of HA is not diffusion controlled. Relaxation phenomena observed during the induction time show the presence of structured adsorption layers. The relaxation behaviour of a quasi-equilibrium HA adsorption layer is independent of the HA concentration. The calculated apparent diffusion coefficients represent the order of magnitude of the rate constant of the real mechanism. A comparison of results calculated from adsorption kinetics and relaxation data shows good agreement with an apparent diffusion coefficient in the order of 0.001 cm2s−1.

The surface tensions of human albumin, immunoglobulin G, and fibrinogen were determined by using two different techniques: (1) the freezing front technique and (2) the pendant drop arrangement in conjunction with axisymmetric drop shape analysis-profile (ADSA-P). The changes in surface tension, resulting from the adsorption of proteins from solution, at various concentrations, onto polymer particles are reported by using the freezing front technique. Furthermore, the adsorption of proteins onto the solution—air interface is also reported by using the pendant drop arrangement. The large differences between the two types of measurements are discussed in terms of the different extents of conformational changes in response to different interfacial tensions driving these changes. A further qualitative difference between the two methods as used here is that the ADSA surface tension measurement registers only the properties of the top few angstroms of the layer whereas the solidification front technique registers the presence of much thicker layers.

Shortage or malfunction of pulmonary surfactant in alveolar space leads to a critical condition termed respiratory distress syndrome (RDS). Surfactant replacement therapy, the major method to treat RDS, is an expensive treatment. In this paper, the effect of poly(ethylene glycol) (PEG) to improve dynamic surface activity of a bovine lipid extract surfactant (BLES) was studied by axisymmetric drop shape analysis (ADSA) and a captive bubble method. The activity of BLES + PEG mixtures was compared to that of a natural surfactant containing surfactant proteins A and D. When PEG was added into BLES mixtures, the surface tension hysteresis of BLES films was minimized when the films were compressed by more than 50%. PEG also helps to quickly restore surfactant films after film collapse. Thus, as far as surface tension effects go, the findings suggest that PEG might be used as a substitute for surfactant-associated protein SP-A in therapeutic surfactant products, and might also be used to reduce the amount of BLES required in clinical applications.

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 3,350; 8,000; 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/m(2). 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 8,000 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.

The sedimentation volume technique was employed to characterize the surface properties of proteincoated polymers. The sedimentation volume, V sed , of polymer particles suspended in aqueous solutions at 23°C was determined. The suspending binary liquid mixtures were chosen such that the surface tension,γ PV , of the protein-coated polymers fell between the surface tensions,γ LV , of the two liquids. A constant mass of a given polymer sample was suspended in constant volumes of the liquid mixtures of different concentrations. It was found that the sedimentation volume changed with varying compositions of the suspending liquid mixtures, as did the liquid surface tension. A maximum in V sed occurred when the surface tension of the suspending liquid was equal to that of the particles, i.e., whenγ LV = γ PV . The position of the V sed maxima, and hence the surface tension,γ PV , of the particles, was found to depend on polymer surface tension as well as on the type and bulk concentration of the coating protein solution. At high bulk concentrations the nature of the underlying substrate materials are entirely masked. Under these conditions V sed maxima, and hence the particle surface tensions, are independent of the nature of the substrate surface properties and are reflective only of the adsorbed protein. At low bulk concentrations a range in V sed maxima are observed, suggestive of substrate-induced conformational changes in the adsorbed protein molecules.

Thermodynamic model considerations suggest that the adhesion of biological cells to polymeric surfaces depends on the relative magnitude of the surface tension γSV of the substrates, the surface tension γCV of the cells, and the surface tension γLV of the suspending liquid medium. Glutaraldehyde-fixed erythrocytes are identified as suitable model particles for such studies. Experimental results of the extent of erythrocyte adhesion from suspension of mixtures of buffer and varying amounts of dimethyl sulfoxide (DMSO) confirm qualitatively and in certain aspects quantitatively the thermodynamic predictions. Specifically, the prediction that the extent of cell adhesion should become independent of substrate surface tension when γLV = γCV is confirmed by experiment. On the other hand for this case the free energy of adhesion predicting zero cell adhesion is not experimentally confirmed completely. However, performing these experiments with distilled water rather than buffer reduces erythrocyte adhesion to a negligible level. It emerges that van der Waals interactions are, by a large margin, the most important forces involved in cell adhesion to polymer surfaces. Nevertheless, not only divalent cations (through bridging effects), but also monovalent cations play a certain, though limited, role.