Robert R. Stromberg

The conformation of adsorbed blood proteins by infrared bound fraction measurements

Abstract The likelihood that surface-induced blood coagulation results from specific protein-material interactions has led to a study of the conformation of adsorbed blood proteins. Infrared difference spectroscopy was used to determine the bound fraction, i.e., the fraction of carbonyl groups of an adsorbed molecule directly interacting with the surface, of serum albumin, prothrombin, and fibrinogen in situ. Measurements were carried out on individual proteins as a function of the amount adsorbed, time of absorption, pD, and ionic strength using a silica surface. The results obtained for serum albumin and prothrombin indicate that the internal bonding of these globular proteins is sufficient to prevent changes in the structure while adsorbed, even at low surface population. The bound fraction of fibrinogen increases with increasing adsorbance, suggesting possible interfacial aggregation. The conformation of all three proteins was found to be independent of the time of adsorption, although major differences in the rates of adsorption were observed. Studies of cross-linked and denatured serum albumin have provided information on the conformational changes concomitant with adsorption of the native protein. Qualitatively, such changes, if they occur, are small. This conclusion is supported by computer simulation studies of lysozyme adsorption. Studies of the effect of pD and ionic strength on the adsorbance and bound fraction of serum albumin show that caution must be exercised when identifying the plateau adsorbance of a protein isotherm with a close-packed monolayer.

Ellipsometric investigation of the effect of potential on blood protein conformation and adsorbance

Abstract The possible relationship between the surface charge of an implant material and intravascular thrombosis has long been explored. In order to characterize the effect of potential on protein-surface interactions, ellipsometric measurements have been carried out in situ to determine the conformation and adsorbance of fibrinogen, γ-globulin, and serum albumin adsorbed on platinum and germanium for a range of imposed potentials. On platinum at pH 7.4, fibrinogen, γ-globulin, and serum albumin all exhibited a reproducible “onset” potential, at which enhanced adsorption occurred. The adsorbance for all three proteins was unchanged from the rest potential value as the applied potential became more anodic until the onset potential was reached. Desorption, as a result of changes in applied potential, was not observed for any protein studied. The adsorbed conformation of all three proteins changed reversibly at moderately cathodic potentials as indicated, for example, by a sharp increase in extension at −0.2 V/SCE. The extension decreased as the imposed potential became progressively anodic. Germanium was found to be unsuitable for these ellipsometric studies due to etching of the surface by the 0.15 M NaCl immersion medium.

Adsorption and desorption rates of polystyrene on flat surfaces

The rates of adsorption and desorption of polystyrene have been measured in cyclohexane and benzene solutions by radiotracer techniques. Relatively long times were required to attain maximum adsorbance values for adsorption of both low and high molecular weight polymer on a solid chrome surface under theta conditions. This was interpreted as indicating a molecular reorientation during the adsorption process. Under these conditions it was observed that the maximum adsorbance values increased with increasing molecular weight, but the amounts adsorbed were independent of molecular weight at very short adsorbance times. This was interpreted as indicating two separate rates of adsorption : an initial rate dependent on the rate of arrival and attachment at the surface and the rate of re-arrangement on the surface. More rapid attainment of maximum adsorbance values on a mercury surface was attributed to participation of the liquid mercury surface in accommodating the changing polymer conformation. The rates of desorption were very slow for both chrome and mercury and were dependent on the surface populations as determined by the adsorbance values.

Measurement of Optical Constants: Optical Constants of Liquid Mercury at 5461 Å*

The optical constants of liquid mercury have been measured by ellipsometry at a wavelength of 5461 A. The measured results are n = 1.485 and κ = 3.061. Discrepancies among values from the literature are discussed and the magnitude of the important sources of error are evaluated. The effects on the measured index of several different types of surface films between the mercury and several contact media have been calculated and compared. Errors in alignment are shown to cause large changes in the measured refractive index as well as an apparent dependence upon the angle of incidence.

Ellipsometry techniques and their application to polymer adsorption

The application of ellipsometry to the study of the adsorption of polymers at interfaces is discussed. The correction of certain instrumental errors such as imperfect components and the use of solutions containing optically active materials are described. Results for the measurement of the extension of polystyrene, poly(ethylene o-phthalate), and certain proteins present in blood adsorbed on surfaces are given and interpreted in terms of changes in molecular conformation that occur during the adsorption process. The differences observed are ascribed to the interaction energies between the different polymers and surfaces.

The Conformation of Adsorbed Polystyrene

The properties of synthetic polymers at the solid-solution interface are relevant to the fields of adhesion, composites, coatings, and flocculation. The description of both the microscopic and macroscopic systems requires the determination of both the conformation and changes in the conformation of the polymer molecule upon interaction with a surface. A number of theoretical and experimental techniques have been employed to characterize adsorbed polystyrene and to test the theoretical models describing the system. These include investigations of: 1) the extension of the adsorbed molecule normal to the surface by ellipsometry and viscosity studies, 2) the distribution of segments of the adsorbed molecule normal to the surface by attenuated total reflection, 3) the fraction of segments of the polymer chain attached to the surface using infrared difference spectroscopy, and 4) the rates of polymer adsorption and desorption using radiotracer techniques. In general, the conformation of adsorbed polystyrene is dependent upon competition among polymer molecules for the available surface sites. At low surface concentration, the adsorbed polymer molecules show a relatively low extension and flat conformation. As the adsorption time continues, or the surface concentration increases, the molecular conformation becomes more extended. Rates of adsorption and desorption provide additional information on the adsorbed molecular conformation. The rate of adsorption is dependent on the molecular weight, solvent, and substrate. For a given molecular weight, the rate of desorption is dependent on the initial adsorbance.

Determination of distributions in inhomogeneous films

Abstract The distribution of material in an inhomogeneous film is an important parameter in many problems. By representing the inhomogeneous film as a stack of thin homogeneous films, we have developed a method for utilizing concomitant ellipsometric and attenuated total reflection measurements to determine the distribution of films whose thickness can be modified stepwise. We have compared our method to that of Beckman and Harrick, using their data for SiH impurity within an Si0 2 film. For two distributions, an exponential and a Gaussian, our method produced good fits with an accuracy limited only by the incremental thickness used for the calculations. Application of the method of Beckmann and Harrick yielded less accurate results which can also show artificial minima in the distribution.

Polywater—A search for alternative explanations

Abstract The case for the existence of “polywater” has been re-examined. Studies were made of the method of preparation of this material. It was determined that surface creep is a factor in the transport of “polywater” into the capillary tubes used for its preparation. It was found possible to reproduce approximately the infrared spectrum previously attributed to “polywater” by synthetic mixtures of salts of carboxylic acids, particularly acetic acid. The inclusion of lactates and pyruvates was also considered. It was established both by infrared and microprobe analyses of “polywater”, that neither silicon nor silicon compounds were present in significant quantities. Several impurity elements were detected by microprobe analysis, but the very thin films did not permit quantitative interpretation of the data. Carbon was not detected in the microprobe analysis. Although several mass spectrographic analyses of “polywater” preparations were made no peaks characteristic of multiple water units were ever observed. Attempts to reproduce the Raman spectrum previously reported have been unsuccessful. It was concluded that there is reason to seriously question the concept that water exists in a stable polymeric condition and that contaminants, both inorganic and organic, may account for a number of the physical properties and other phenomena associated with “polywater”.