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The wetting behavior of aqueous two-phase polymer test systems on dextran coated glass surfaces: Effect of molecular weight
The Wetting Behavior of Aqueous Two-Phase Polymer Test Systems on Dextran Coated Glass Surfaces: Effect of Molecular Weight J O H N F. B O Y C E , * B R U C E A. H O V A N E S , t J. M I L T O N H A R R I S , t J A M E S M. V A N A L S T I N E , $ AND D O N A L D E. B R O O K S ~'* * Departments of Pathology and Chemistry, University of British Columbia, Vancouver, British Columbia V6T 2B5, Canada; t Department of Chemistry, University of Alabama in Huntsville, Huntsville, Alabama 35899; and ~Department of Biological Sciences, University of Alabama in Huntsville, Huntsville, Alabama 35899
Received August 21, 1989; accepted November 6, 1991 The wetting behavior of phase-separated aqueous solutions of dextran and poly (ethylene glycol) on quartz substrates coated with covalently bound dextran fractions was examined as a function of coating molecular weight. The effect of dissolved polymer size was also investigated by using two-phase test systems containing high and low molecular weight dextrans. Sharply cut dextran fractions of Mw from 27,900 to 205,000 were attached to quartz activated with 3-aminopropyldimethylethoxysilane through reductive amination of the dextran's reducing end group in the presence of sodium cyanoborohydride. X-ray photoelectron spectroscopy and contact angle measurements in the two-phase polymer solutions were used to assess the quantity and quality of the surface coatings. Our results demonstrate that wetting by the dextran-rich phase improves, as evidenced by a declining contact angle, with increasing molecular weight of the bound dextran, decreasing molecular weight of the free dextran in solution and increasing time of exposure to the polymer phases. The usual relationship of increasing contact angle with increasing interracial tension is not observed in these phase-separated polymer mixtures. © 1992AcademicPress,Inc. INTRODUCTION W h e n two incompatible, neutral polymers are mixed in aqueous solution a critical concentration is reached above which the system separates into two phases, Each phase is enriched in one o f the polymers a n d depleted in the other. Since these systems can be buffered and m a d e isotonic, they have proven useful, both in the laboratory and on an industrial scale, for separating by partition a variety o f biological materials ranging from proteins and nucleic acids to m e m b r a n e fragments and whole cells ( 1, 2). O u r interest in the wetting behavior o f these mixtures stems f r o m two sources. First, partition o f particulates such as biological cells generally occurs between one bulk phase and the liquid/liquid interface. T h e contact angle To whom correspondence should be addressed at: Department of Pathology, 2211 Wesbrook Mall, University of British Columbia, Vancouver, Canada V6T 2B5.
o f the particle/phase b o u n d a r y contact line is a measure o f the difference in free energy for a cell located at the interface or i m m e r s e d in one bulk phase and hence is a determinant o f the cell partition coefficient. Since cell m e m branes bear a coat o f a n c h o r e d glycoprotein and glycolipid, the dependence o f the contact angle on the molecular weight ( M W ) o f surface-attached and dissolved p o l y m e r chains is o f interest as a partition determinant. The second motivation for our interest in wetting derives f r o m work on the effects of gravity on cell partitioning ( 3 - 5 ) . Partitioning is initiated by shaking the phase system, containing cells, into a fine emulsion that rapidly begins to coalesce once shaking has ceased. The coalescing drops develop into structured regions which begin to stream past each other as the phases demix and slowly reach the equilibrium location determined by their density. The cells are carried along in the process, either adsorbed to the liquid-liquid interfaces or en-
153 0021-9797/92 $3.00 Journal of Colloid and Interface Science, Voh 149, No. I, March L 1992
Copyright © 1992 by Academic Press, Inc. All rights of reproduction in any form reserved.
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BOYCE ET AL.
closed within the individual bulk phases. Sep- to reverse the inner and outer phases and posaration yields can be excellent, but theory sibly alter the rate of demixing. based on thermodynamic measurements predicts that separation efficiency should be much MATERIALS AND METHODS higher (6). This analysis indicates that the Dextrans used in this project were obphase partitioning process is not dominated by thermal energies as one might expect, but tained from Pharmacia (Uppsala, Sweden) rather by other, unidentified randomizing fac- and were custom prepared, sharply cut fractions originally kindly supplied by Dr. K. tors (6). Careful observation of the demixing process Granath with the following polydispersities: has suggested that the convective flow intro- (Mw/Mn): (27,900/23,000), (52,000/41,700), duced during the coalescence of phase com- (107,000/73,000), (147,000/94,300), and ponents is one potential source of randomizing (205,000/143,000). The fractions are referred energy in the system. The fluid shear forces to as dextran (Mw × 10-3). Polyethylene glycreated during this stage may randomly dis- col, nominal MW 8,000 (PEG 8,000), was lodge cells adsorbed at the liquid-liquid in- from Union Carbide. The surface used was terface, thus giving separations of poorer quartz, cut into 1-cm 2 × 1-mm thick slides quality than indicated by thermodynamic (Quartz Scientific). analysis. This density-driven, convective streaming can be expected to be absent in the Dextran-Derivatized Quartz diminished gravity of space. On earth, in a strong gravitational field, Dextrans were covalently coupled to the polymer systems separate within a few minutes slides by first aminating the surface with an into an upper and a lower phase. Experiments aminopropyl silane and then reductively amiwith isopycnic phase systems and on board nating the dextran reducing terminus (only the space shuttle have shown slower demixing one is available per molecule (9)) with sodium that terminates in an inner sphere of one phase cyanoborohydride. Macroscopically uniform surrounded by the second phase (much like coatings (as revealed by XPS and contact anthe yolk and white of an egg) (5, 7). Mea- gles) were only achieved when extreme care surement of contact angles has shown that the was taken to insure cleanliness of all glassware outer phase in these experiments is the one and substrates to be coated. Glassware was which preferentially wets the container wall. soaked with chromic acid or "Nochromix" (a (For typical aqueous phase systems composed cleaning solution from Godax) and rinsed of dextran and poly (ethylene glycol) (PEG) thoroughly with distilled, deionized, charcoal it is the PEG-rich phase that wets the glass or filtered water. Quartz slides were treated with poly (methyl methacrylate) containers (8).) cleaning solution, rinsed with water, and fiOne goal of the present research is to deter- nally submitted to cleaning in a Tegal Plasmod mine if wall coatings can be applied to control 02 plasma cleaning device. wall wetting and in turn affect both the dySilanization with 3-aminopropyldimethylnamics of phase demixing and the ultimate ethoxysilane (Petrarch Systems, Bristol, PA) disposition of the phases. was performed by, first, covering the slide with In this paper, we present a study of the wet- a 1% solution ofsilane in mesitylene, degassing ting of dextran-coated glass by dextran-PEG by applying a vacuum or sonication and rephase systems as a function of dextran MW fluxing at 160°C overnight. The liquid was both in the coating and in the phase system. then drained from the slides, and they were lfthe dextran-rich phase, rather than the PEG- rinsed and sonicated with several washes of rich phase, can be made to be the wetting phase ethanol. The liquid was again drained from in the space experiments, it should be possible the quartz and the slide cured at 110°C for 4 Journal of Colloid and Interface Science, Vol. 149, No. 1, March 1, 1992
WETTING BEHAVIOR h. The quartz finally was rinsed and sonicated in 50% aqueous ethanol. Reductive amination was performed by adding a 10% aqueous solution of dextran in 10% NaC1, 0.05% sodium azide to cover the silanated slide and incubating in an 80°C bath. Small amounts of cyanoborohydride (about 0.3 g per gram of quartz) were added daily for 2 days (no improvement in coating was found if further additions were made). The quartz was then washed thoroughly with distilled water.
155
eV ( 11 ), but as there were no other components near the carbon region this did not interfere with identification of the C-C, C-O, and O - C - O peaks. The atomic percentages of the surface constituents identified were calculated using photoemission cross sections relative to fluorine of: carbon 0.296, oxygen 0.71, nitrogen 0.477, and silicon 0.339, These cross sections were supplied by Perkin-Elmer for the omnifocus lens system used in the spectrometer.
Phase Systems XPS Analysis of Coupled Dextran X-ray photoelectron spectra were obtained on a Perkin-Elmer 5400 spectrometer on 1cm 2 slides using a beam diameter of approximately 1100 t~m and an AIKo~ source. A typical spectrum of the carbon region is illustrated in Fig. 1, expressed as electron counts in arbitrary units, as a function of binding energy. The complex peak in the C ls region was deconvoluted into three peaks associated with C-C, C-O, and one other constituent, which we assume to be due to O - C - O (10). The quartz slides showed evidence of varying degrees of electrostatic charging, the source of the difference between the location of the C C peak in Fig. 4 and the accepted value of 285
296
294
292
290
Binding Energy (eV)
288
286
FIG. 1. XPS spectrum of quartz slide derivatizedwith dextran 147.5 showingdeconvolutedpeaks in the carbon region of the spectrum. Curves are identified as (A), observed experimental curve; (B), sum of deconvoluted curves; (C), deconvolutedC-C peak; (D), deconvoluted C-O peak; (E) deconvolutedO-C-O peak.
To investigate the effect of free dextran M W on the interaction with bound dextran, two phase separated d e x t r a n ] P E G test systems were used, one containing a low M W dextran 27.9 fraction and one a high MW dextran 205. Since polymer M W has a strong effect on phase behavior, phase diagrams for each dextran were constructed in order to identify two test systems with similar phase properties (i.e., two systems with the same tie line length and hence the same mean difference in mass concentration between the phases). These diagrams appear in Fig. 2. Polymer concentrations were determined using polarimetry and refractometry by methods described elsewhere (12). The two systems chosen from these phase diagrams and used throughout the experiments were: 5% w / w dextran 27.9-8.4% w / w PEG 8,000, and 5% w / w dextran 2055.25% w / w PEG 8,000, each in 150 m M NaCI. Interfacial tensions were determined from sessile drop measurements using Rotenberg's axisymmetric drop program (13). These and other relevant physical data for the test systems are summarized in Table I. Over 24 h of exposure to coated and uncoated slides the interracial tension varied by at most 7%.
Contact Angle Measurements After rinsing thoroughly in distilled water the coated slides were placed in a 1-cm deep × 2.5-cm high X 10-cm long glass-sided chamber and immersed in the upper PEG-rich phase of the test system. A layer ofhexadecane Journal of Colloid and Interface Science, Vol. 149, No. 1, March I, 1992
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B O Y C E ET AL.
light source projected through the rear of the chamber. To give some measure of the effect of time on the polymeric interaction with the coated surface, the same droplets were photographed again 24 h later. All photographs were taken with a pipette of known diameter showing in the frame to serve as a scale for deriving a magnification factor. Contact angles were calculated from the digitized profiles of enlarged drops using Rotenberg's axisymmetric drop program (13).
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floated on the surface of the PEG-rich phase prevented evaporation of the aqueous solution. The results obtained were independent of the time the hexadecane was present before the drops were placed on the slide. Droplets of the more dense dextran-rich bottom phase were placed randomly over the sample surface and allowed to equilibrate for 1 h. At this time photographs of the drops were taken through a horizontally mounted Zeiss Tessovar microscope equipped with an optical extension tube for increased magnification. Illumination of the chamber was achieved with a 150-W fiber
The coating technique used here was optimized on porous glass chromatography beads with commercially available dextran because the large surface area available per unit mass permitted straightforward, quantitative determination of the amount of dextran per unit area by wet chemical means (data not shown). However, there are potential restrictions on molecular packing within the pores of the beads, and it is unlikely that the available surfaces of the beads and quartz slides are chemically identical. Hence, extrapolation from the porous bead results to derive accurate values for the amount ofdextran bound per unit area to the slides was not warranted. The amount of dextran on the quartz slides is too small to be readily measured by wet chemical techniques. The quality and extent of dextran coatings was analyzed in two ways. Uniformity of coatings was indicated by the consistency of contact angles measured for dextran phase droplets randomly distributed on the surfaces and by the excellent agreement among XPS spectra
TABLE I Test Systems a n d Physical D a t a
System
Tie line length (%)
Densitydifference (kg m-3)
Interfacialtension (naN m-j)
5% W/W D e x t r a n 27.9/8.4% w / w P E G 8,000 5% w / w D e x t r a n 205/5.25% w / w P E G 8,000
14.7 14.7
58.1 48.0
0.0351 0.0228
Journal oj'Colloid and IntetJbce Science, Vol. 149, No. l, March I, 1992
157
W E T T I N G B EH A V IO R
obtained at five different locations on each slide. The relative amount of dextran on the surfaces as a function of MW was estimated using XPS to measure the fraction of the derivatized surface occupied by C-O carbon from which photoelectrons could be collected. Results of the XPS analysis are given in Table II expressed as the atomic percentage of the surface constituents accessible to XPS. The CC, C-O, and O-C-O carbon fractions were obtained from computer deconvolution of overlapping peaks, as illustrated in Fig. 1.
Contact Angle Measurements The results are summarized in Table III. They indicate a consistent decline in the contact angle as the MW of the coating increases. This effect was more pronounced for the lower MW dextran 27.9 two-phase system than for the system containing dextran 205. A decrease in the contact angle over 24 h is apparent for both test systems on all coated surfaces. This observation implies that there is a time dependency associated with the interaction between the free polymers in solution and the polymer bound to the glass surface, particularly for the higher MW coatings. DISCUSSION
XPS is a surface spectroscopic technique which provides analytical information on material located within about 50 A of the external boundary of polymeric samples (11 ). As a
high vacuum is required for the collection of photoelectrons, no solvent will be present. In our samples the dextran detected therefore will be collapsed into a more compact configuration than would be present in solution or when the molecule was bound to the surface by its reducing end in an aqueous environment. XPS spectra of carefully cleaned quartz surfaces always show a hydrocarbon (C-C) peak (11 ), presumably due to impurities in the environments to which the surfaces are exposed. Covalent attachment of a polymer having carbon-oxygen bonds (such as dextran) should show an overlapping, companion carbon peak due to C-O bonds of the carbohydrate. Figure 1 shows this and a companion, less intense peak at a higher binding energy in the deconvoluted spectrum. The higher energy peak presumably represents the O-C-O signal, since diethers, esters, and carboxylic acids all show peaks in this region (10). The C-C peak accounts for a relatively constant fraction of the total carbon detected as the MW of bound dextran is varied (Table II). A portion of this constituent likely derives from the silane coupling reagent but, considering the level of nitrogen detected is about tenfold lower, most of this material must be from other sources. Since dextran is the only source of C-O and the coupling reagent the only nitrogen-containing compound in the sample, the C-O and N signals must represent the levels of these two materials detectable. Evidently there is a great deal of silane on the surface which is not
TABLE lI Atomic Percentages of Constituents of Dextran-Derivatized Quartz Surfaces Detected by XPS for the Dextran Fractions Indicated Dextran
% C-C
% C-O
% O-C-O
%N
% Si
%O
% Other
Control a 52 107 147.5 205
12.2 b 7.9 7.7 6.8 11.1
-2.0 3.7 8.9 11.9
-0.2 0.5 0.8 1.2
2.1 0.7 0.5 0.7 0.7
29.8 21.5 20.5 18.2 14.7
55.9 67.1 66.1 63.9 57.2
-0.8 1.5 1.5 4.3
Amin ated quartz. b Uncertainty typically +0.15% for all constituents. a
Journal of Colloid and Interface Science, Vol. 149,No. 1, March 1, 1992
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BOYCE ET AL. TABLE III
Contact Angles Measured (Degrees + SEM a) for Dextran-Derivatized Quartz Slides in Dextran/PEG Two-Phase Systems Containing Dextran 27.9 or Dextran 205 at the Times Indicated Dx 27.9 (5, 8.4) Coating (molec wt)
No Dx Dx Dx Dx Dx
coating 27.9 52 107 147.5 205
Dx 205 (5, 5.25)
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168.3 148.5 141.6 125.3 118.5 105.4
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170.2 133.9 132.3 118.3 113.0 84.0
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176.1 161.4 158.0 134.0 145.7 132.5
24 h
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164.5 157.9 156.0 131.3 127.0 113.8
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a Standard error of the mean for 5-7 measurements.
coupled to dextran. If all the coupling compound was derivatized with dextran, taking dextran 52 as an example there would be 1,900 C - O groups per N, compared to the observed value of 2.9. However, the surface activation procedure used here gives an amine surface concentration of about one per 70 A2 (14). A fully hydrated molecule ofdextran 52 in solution has a cross sectional area of approximately 1.2 × 10 4 fik 2, calculated from its radius of gyration (15). This implies a ratio of one dextran per 170 nitrogens or a C-O:N ratio of about 11 if the polymers were close packed, each with the same cross-sectional area as a molecule in solution. The density of surface-attached polymer may be greater than this, however. Measurement of the amount of a commercial dextran l0 fraction bound to porous glass beads of known specific area gave a value of 1,400 ~2 per molecule (data not shown), half the value calculated from the solution radius of gyration, which translates to a C - O : N ratio of 20. This number may be somewhat underestimated due to the method of specific area measurement (gas adsorption) for the porous glass beads, and it ignores the breadth of the molecular weight distribution of the dextran. Nonetheless these calculations support the conclusion that most of the surface amines must be underivatized. As is seen in Fig. 3 (data taken from Table IIL the fractions of the detected surface material consisting of C - O and O - C - O increase Journal of Colloid and Interface Srience, Vol. 149, No. 1, March 1, 1992
approximately linearly with dextran MW, concomitant with a roughly linear decrease in the Si signal. The C - O and O - C - O linearities imply that the mass ofdextran present within the XPS beam sampling volume is greater the higher the MW. If the depth of the sampling volume was such that all the C - O and O - C - O groups of each M W contributed to the signal, the linearities would indicate that the number of molecules coupled per unit area was approximately constant as a function of MW, since the number of C - O and O - C - O groups per molecule are directly proportional to this parameter. However, the decrease in the Si signal as M W increases implies that the larger molecules are covering some of the Si detectable at lower MW. Evidently with higher MWs
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WETTING
some portion of the surface is occupied by regions of collapsed polymer chains which are thicker than the sampling depth of the spectrometer. Calculation of the diameter of spheres with the same mass and density as unhydrated dextran molecules is consistent with this idea, the values ranging from 47 to 73 for the MWs in Table II. The reduction with MW is not seen in the N percentage, presumably because uncertainty in the measurement masks any regular variation in the very small signal values observed. From the XPS results we conclude, therefore, that dextran is bound to the surfaces and that an increasing mass of dextran is bound per unit area as the dextran M W increases. In this work the contact angle is defined as the interior angle between the solid substrate and the surface of the dextran-rich drop. Hence, decreasing contact angles correspond to progressively greater wetting tendency by the dextran-rich phase. It is apparent from Table III that the contact angle decreases with: (a) increasing M W of the bound dextran, (b) decreasing M W of the free dextran in the twophase system, and (c) time of exposure. To attempt to understand these effects we consider the free energy differences which determine the contact angle, 0, using Young's equation appropriate to the present situation ( 16): cos 0 - (%~ - "rsb) _ 2X~ "Ytb
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where 3'tb is the interfacial tension of the interface between the top, PEG-rich phase and the bottom, dextran-rich phase, and 7~t and 7sb are the interfacial tensions between the coated quartz surface and the PEG-rich and dextran-rich phases, respectively; A7 is the interracial free energy difference. Thus for a given phase system, where 7tb is constant, observed changes in cos 0 may be interpreted as being due to differences in the interfacial tensions between the coated surface and the two phases. The underivatized quartz surface exhibits a contact angle of nearly 180 °, implying a much lower energy interaction with the PEG-rich
BEHAVIOR
159
phase than with the dextran-rich one. This may be attributable to a stronger adsorption of PEG than dextran. However, we have not measured adsorption in this study so this explanation must remain a conjecture. Derivatization with any of the dextran fractions reduces 0 by making the surface more compatible with dextran and less so with PEG, presumably by the same molecular mechanisms that drive phase separation in these mixtures, namely an unfavorable interaction free energy between the monomers of the two polymers ( 17 ). Since we have been able neither to measure the absolute amounts ofdextran bound to the quartz nor to measure independently the average thickness of these layers, it is not possible to quote values for the concentration of polymer chains in the surface-attached layer. We therefore have not attempted to interpret any of the A3, data quantitatively using current theories of polymer mixtures ( 18, 19). However, the trends in changes in 0 may be rationalized qualitatively as follows. For a given phase system, 0 decreases with increasing MW ofdextran on the surface. As discussed above, the higher the MW, the greater the mass of polymer bound per unit area. Moreover, because of the larger molecular size, the bound layer would be expected to extend farther out into solution for higher M W fractions. Hence, since PEG is incompatible with dextran it might be expected to be rejected more readily by the more extensive high M W coatings than by lower molecular analogues, increasing 7~t and producing the effects observed. The second obvious trend in the data is the greater decrease in 0 observed for a given surface with the phase system containing the lower M W dextran fraction. These data illustrate an important difference between immiscible polymer solutions and simple two component mixtures as far as contact angle studies are concerned. The latter systems show a predictable decline in 0 as the surface or interfacial tension decreases (20, 21 ), while this may or may not be true for aqueous phase mixtures. The behavior of polymer systems is more Journal of Colloid and Interface Science, Vol. 149, No. I, March 1, 1992
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BOYCE ET AL.
complex as 0 can decline or, as with the systems described here, increase with a falling interfacial tension. In the study at hand, the dextran 27.9 phase mixture yielded the lower contact angles of the two systems tested. When the values of AT are calculated and plotted as a function of coating MW, Fig. 4, the low MW dextran line falls below that of the dextran 205, except at the highest bound polymer MW. A MW effect is not unexpected, since a similar result is seen when macromolecules are partitioned in these systems where it is universally observed that if the MW of one of the phase-forming polymers is reduced, partition into the phase in which that polymer predominates is increased (22). That is, the interaction between the "substrate" and the phase becomes more favorable as the MW of the polymer in that phase is lowered. The likely reason for both effects is that the entropy of mixing of the "substrate" (either the macromolecule being partitioned or the surface-attached chains on the surface) with the phase polymer is greater per unit mass of polymer for lower MW chains (17). Hence, the free energy of mixing between the phase polymer and the substrate will be lower the lower the MW of the former. Lowering %b lowers 0, as observed. For the highest MW coating this effect is not observed, however, and the trend in the data suggests other ~
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factors may be involved, possibly polymer adsorption. It might be noted that the dependence on dissolved polymer molecular weight for all but the highest MW coatings is opposite to that which would be predicted if dextran adsorption to an uncoated surface were occurring and determining the relative surface compatibility. Polymer adsorption isotherms generally saturate at higher values for higher MWs in the range used here (23), so pure adsorption would be expected to have the opposite MW dependence, with respect to the phase polymers, to that which was observed. The present results do not imply that adsorption does not occur, however, since as discussed above PEG binding may well predominate on bare quartz and the PEG MW was held constant throughout. Considering the time dependencies observed, it is seen that with the phase system containing the lower MW dextran there are significant decreases in 0 between l and 24 h exposure for all molecular coatings. With the higher dextran MW system, however, significant time effects are only seen with the two highest MW coatings. This could reflect differences in transport rates of the two dextran fractions through the coatings following initial exposure of the surface to the phase systems. It would be expected that the diffusion ofdextran would be significantly hindered by layers whose thicknesses were of the order of their own dimension or greater. Thus, all surfaceattached layers would be expected to affect the time taken for the lower molecular weight dextran fraction to reach its equilibrium profile, while the higher MW polymer should be little impeded by the relatively low MW coatings, as observed. ACKNOWLEDGMENTS
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FIG. 4. Interfacial tension difference, 4% vs molecular weight of dextran coating.
Journa/of Colloid and Interface Srience, Vol. 149, No. 1, March 1, i 9 9 2
The authors gratefully acknowledge the financial support of this work by the National Aeronautics and Space Administration, the National Institutes of Health, and the Medical Research Council of Canada (Grant MT 5759 to DEB).
WETTING BEHAVIOR REFERENCES 1. Albertsson, P. A., "Partition of Cell Particles and Macromolecules," 3rd ed. Wiley-Interscience, New York, 1986. 2. Magnussen, K.-E., and Stendahl, O., in "Partitioning in Aqueous Two Phase Systems" (H. Walter, D. E. Brooks, and D. Fisher, Eds.), p. 415. Academic Press, New York, 1985. 3. Brooks, D. E., Bamberger, S. B., Harris, J. M., and Van Alstine, J. M., Eur. Space Agency Publ. SP222, 315 (1984). 4. Van Alstine, J. M., Harris, J. M., Snyder, R. S., Curreri, P. A., Bamberger, S. B., and Brooks, D. E., Eur. Space Agency Publ. SP-222, 309 (1984). 5. Brooks, D. E., Bamberger, S. B., Harris, J. M., Van Alstine, J. M., and Snyder, R. S., Eur. Space Agency Publ. SP-256, 131 (1978). 6. Brooks, D. E., in "Frontiers in Bioprocessing (M. Bier, S. Sikdar, and P. Todd, Eds.), p. 259. CRC Press, Cleveland, 1989. 7. Bamberger, S. B., Van Alstine, J. M., Baird, J. M., Harris, J. M., and Brooks, D. E., Sep. Sci. Technol. 23, 17 (1987). 8. Harris, J. M., Brooks, D. E., Boyce, J. F., Snyder, R. S., and Van AIstine, J. M., in "Dynamic Aspects of Polymer Surfaces" (J. D. Andrade, Ed.), p. 111. Plenum Press, New York, 1988. 9. Harris, J. M., and Yalpani, M., in "Partitioning in Aqueous Two Phase Systems" (H. Waiter, D. E. Brooks, and D. Fisher, Eds.), p. 589. Academic Press, New York, 1985. 0. Gelius, U., Hed6n, P. F., Hedman, J., Lindberg, B. J., Manne, R., Nordberg, R., Nordling, C., and Siegbahn, K., Phys. Scr. 2, 70 (1980).
161
11. Ratner, B. D., and McElroy, B. J., in "Spectroscopy in the Biomedical Sciences" (R. M. Gendreau, Ed.), p. 107. CRC Press, Boca Raton, Florida, 1986. 12. Bamberger, S. B., Brooks, D. E., Sharp, K. A., Van Alstine, J. M., and Webber, T. J., in "Partitioning in Aqueous Two Phase Systems" (H. Walter, D. E. Brooks and D. Fisher, Eds.), p. 104. Academic Press, New York, 1985. 13. Rotenberg, Y., Boruvka, L., and Neumann, A. W., J. Colloid Interface Sci. 93, 169 (1983). 14. Herren, B. J., Shafer, S. G., Van Alstine, J. M., and Snyder, R. S., J. Colloid Interface Sci. 115, 46 (1987). 15. Granath, K. A., J. ColloidSci. 13, 308 (1958). 16. Young, T., Philos. Trans. R. Sac. London 95, 65 (1805). 17. Brooks, D. E., Sharp, K. A., and Fisher, D., in "Partitioning in Aqueous Two Phase Systems" (H. Walter, D. E. Brooks, and D. Fisher, Eds.), p. 11. Academic Press, New York, 1985. 18. de Gennes, P.-G., "Scaling Concepts in Polymer Physics." Cornell Univ. Press, Ithaca, New York, 1979. 19. Solc, K., Macromolecules 19, 1166 (1986). 20. Fox, H. W., and Zisman, W. A., J. ColloidSci. 7, 428 (1952). 21. Good, R. J., and Girifalco, L. A., J. Phys. Chem. 64, 561 (1960). 22. Albertsson, P. A., Cajarville, A., Brooks, D. E., and Tjerneld, F., Biochim. Biophys. Acta 926, 87 (1987). 23. Cohen Stuart, M. A., Scheutjens, J. M. H. M., and Fleer, G. J., J. Polym. Sci. 18, 559 (1980).
Journal of Colloid and lnterface Science, Vol. 149, No. I, March 1, 1992
Received August 21, 1989; accepted November 6, 1991 The wetting behavior of phase-separated aqueous solutions of dextran and poly (ethylene glycol) on quartz substrates coated with covalently bound dextran fractions was examined as a function of coating molecular weight. The effect of dissolved polymer size was also investigated by using two-phase test systems containing high and low molecular weight dextrans. Sharply cut dextran fractions of Mw from 27,900 to 205,000 were attached to quartz activated with 3-aminopropyldimethylethoxysilane through reductive amination of the dextran's reducing end group in the presence of sodium cyanoborohydride. X-ray photoelectron spectroscopy and contact angle measurements in the two-phase polymer solutions were used to assess the quantity and quality of the surface coatings. Our results demonstrate that wetting by the dextran-rich phase improves, as evidenced by a declining contact angle, with increasing molecular weight of the bound dextran, decreasing molecular weight of the free dextran in solution and increasing time of exposure to the polymer phases. The usual relationship of increasing contact angle with increasing interracial tension is not observed in these phase-separated polymer mixtures. © 1992AcademicPress,Inc. INTRODUCTION W h e n two incompatible, neutral polymers are mixed in aqueous solution a critical concentration is reached above which the system separates into two phases, Each phase is enriched in one o f the polymers a n d depleted in the other. Since these systems can be buffered and m a d e isotonic, they have proven useful, both in the laboratory and on an industrial scale, for separating by partition a variety o f biological materials ranging from proteins and nucleic acids to m e m b r a n e fragments and whole cells ( 1, 2). O u r interest in the wetting behavior o f these mixtures stems f r o m two sources. First, partition o f particulates such as biological cells generally occurs between one bulk phase and the liquid/liquid interface. T h e contact angle To whom correspondence should be addressed at: Department of Pathology, 2211 Wesbrook Mall, University of British Columbia, Vancouver, Canada V6T 2B5.
o f the particle/phase b o u n d a r y contact line is a measure o f the difference in free energy for a cell located at the interface or i m m e r s e d in one bulk phase and hence is a determinant o f the cell partition coefficient. Since cell m e m branes bear a coat o f a n c h o r e d glycoprotein and glycolipid, the dependence o f the contact angle on the molecular weight ( M W ) o f surface-attached and dissolved p o l y m e r chains is o f interest as a partition determinant. The second motivation for our interest in wetting derives f r o m work on the effects of gravity on cell partitioning ( 3 - 5 ) . Partitioning is initiated by shaking the phase system, containing cells, into a fine emulsion that rapidly begins to coalesce once shaking has ceased. The coalescing drops develop into structured regions which begin to stream past each other as the phases demix and slowly reach the equilibrium location determined by their density. The cells are carried along in the process, either adsorbed to the liquid-liquid interfaces or en-
153 0021-9797/92 $3.00 Journal of Colloid and Interface Science, Voh 149, No. I, March L 1992
Copyright © 1992 by Academic Press, Inc. All rights of reproduction in any form reserved.
154
BOYCE ET AL.
closed within the individual bulk phases. Sep- to reverse the inner and outer phases and posaration yields can be excellent, but theory sibly alter the rate of demixing. based on thermodynamic measurements predicts that separation efficiency should be much MATERIALS AND METHODS higher (6). This analysis indicates that the Dextrans used in this project were obphase partitioning process is not dominated by thermal energies as one might expect, but tained from Pharmacia (Uppsala, Sweden) rather by other, unidentified randomizing fac- and were custom prepared, sharply cut fractions originally kindly supplied by Dr. K. tors (6). Careful observation of the demixing process Granath with the following polydispersities: has suggested that the convective flow intro- (Mw/Mn): (27,900/23,000), (52,000/41,700), duced during the coalescence of phase com- (107,000/73,000), (147,000/94,300), and ponents is one potential source of randomizing (205,000/143,000). The fractions are referred energy in the system. The fluid shear forces to as dextran (Mw × 10-3). Polyethylene glycreated during this stage may randomly dis- col, nominal MW 8,000 (PEG 8,000), was lodge cells adsorbed at the liquid-liquid in- from Union Carbide. The surface used was terface, thus giving separations of poorer quartz, cut into 1-cm 2 × 1-mm thick slides quality than indicated by thermodynamic (Quartz Scientific). analysis. This density-driven, convective streaming can be expected to be absent in the Dextran-Derivatized Quartz diminished gravity of space. On earth, in a strong gravitational field, Dextrans were covalently coupled to the polymer systems separate within a few minutes slides by first aminating the surface with an into an upper and a lower phase. Experiments aminopropyl silane and then reductively amiwith isopycnic phase systems and on board nating the dextran reducing terminus (only the space shuttle have shown slower demixing one is available per molecule (9)) with sodium that terminates in an inner sphere of one phase cyanoborohydride. Macroscopically uniform surrounded by the second phase (much like coatings (as revealed by XPS and contact anthe yolk and white of an egg) (5, 7). Mea- gles) were only achieved when extreme care surement of contact angles has shown that the was taken to insure cleanliness of all glassware outer phase in these experiments is the one and substrates to be coated. Glassware was which preferentially wets the container wall. soaked with chromic acid or "Nochromix" (a (For typical aqueous phase systems composed cleaning solution from Godax) and rinsed of dextran and poly (ethylene glycol) (PEG) thoroughly with distilled, deionized, charcoal it is the PEG-rich phase that wets the glass or filtered water. Quartz slides were treated with poly (methyl methacrylate) containers (8).) cleaning solution, rinsed with water, and fiOne goal of the present research is to deter- nally submitted to cleaning in a Tegal Plasmod mine if wall coatings can be applied to control 02 plasma cleaning device. wall wetting and in turn affect both the dySilanization with 3-aminopropyldimethylnamics of phase demixing and the ultimate ethoxysilane (Petrarch Systems, Bristol, PA) disposition of the phases. was performed by, first, covering the slide with In this paper, we present a study of the wet- a 1% solution ofsilane in mesitylene, degassing ting of dextran-coated glass by dextran-PEG by applying a vacuum or sonication and rephase systems as a function of dextran MW fluxing at 160°C overnight. The liquid was both in the coating and in the phase system. then drained from the slides, and they were lfthe dextran-rich phase, rather than the PEG- rinsed and sonicated with several washes of rich phase, can be made to be the wetting phase ethanol. The liquid was again drained from in the space experiments, it should be possible the quartz and the slide cured at 110°C for 4 Journal of Colloid and Interface Science, Vol. 149, No. 1, March 1, 1992
WETTING BEHAVIOR h. The quartz finally was rinsed and sonicated in 50% aqueous ethanol. Reductive amination was performed by adding a 10% aqueous solution of dextran in 10% NaC1, 0.05% sodium azide to cover the silanated slide and incubating in an 80°C bath. Small amounts of cyanoborohydride (about 0.3 g per gram of quartz) were added daily for 2 days (no improvement in coating was found if further additions were made). The quartz was then washed thoroughly with distilled water.
155
eV ( 11 ), but as there were no other components near the carbon region this did not interfere with identification of the C-C, C-O, and O - C - O peaks. The atomic percentages of the surface constituents identified were calculated using photoemission cross sections relative to fluorine of: carbon 0.296, oxygen 0.71, nitrogen 0.477, and silicon 0.339, These cross sections were supplied by Perkin-Elmer for the omnifocus lens system used in the spectrometer.
Phase Systems XPS Analysis of Coupled Dextran X-ray photoelectron spectra were obtained on a Perkin-Elmer 5400 spectrometer on 1cm 2 slides using a beam diameter of approximately 1100 t~m and an AIKo~ source. A typical spectrum of the carbon region is illustrated in Fig. 1, expressed as electron counts in arbitrary units, as a function of binding energy. The complex peak in the C ls region was deconvoluted into three peaks associated with C-C, C-O, and one other constituent, which we assume to be due to O - C - O (10). The quartz slides showed evidence of varying degrees of electrostatic charging, the source of the difference between the location of the C C peak in Fig. 4 and the accepted value of 285
296
294
292
290
Binding Energy (eV)
288
286
FIG. 1. XPS spectrum of quartz slide derivatizedwith dextran 147.5 showingdeconvolutedpeaks in the carbon region of the spectrum. Curves are identified as (A), observed experimental curve; (B), sum of deconvoluted curves; (C), deconvolutedC-C peak; (D), deconvoluted C-O peak; (E) deconvolutedO-C-O peak.
To investigate the effect of free dextran M W on the interaction with bound dextran, two phase separated d e x t r a n ] P E G test systems were used, one containing a low M W dextran 27.9 fraction and one a high MW dextran 205. Since polymer M W has a strong effect on phase behavior, phase diagrams for each dextran were constructed in order to identify two test systems with similar phase properties (i.e., two systems with the same tie line length and hence the same mean difference in mass concentration between the phases). These diagrams appear in Fig. 2. Polymer concentrations were determined using polarimetry and refractometry by methods described elsewhere (12). The two systems chosen from these phase diagrams and used throughout the experiments were: 5% w / w dextran 27.9-8.4% w / w PEG 8,000, and 5% w / w dextran 2055.25% w / w PEG 8,000, each in 150 m M NaCI. Interfacial tensions were determined from sessile drop measurements using Rotenberg's axisymmetric drop program (13). These and other relevant physical data for the test systems are summarized in Table I. Over 24 h of exposure to coated and uncoated slides the interracial tension varied by at most 7%.
Contact Angle Measurements After rinsing thoroughly in distilled water the coated slides were placed in a 1-cm deep × 2.5-cm high X 10-cm long glass-sided chamber and immersed in the upper PEG-rich phase of the test system. A layer ofhexadecane Journal of Colloid and Interface Science, Vol. 149, No. 1, March I, 1992
156
B O Y C E ET AL.
light source projected through the rear of the chamber. To give some measure of the effect of time on the polymeric interaction with the coated surface, the same droplets were photographed again 24 h later. All photographs were taken with a pipette of known diameter showing in the frame to serve as a scale for deriving a magnification factor. Contact angles were calculated from the digitized profiles of enlarged drops using Rotenberg's axisymmetric drop program (13).
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floated on the surface of the PEG-rich phase prevented evaporation of the aqueous solution. The results obtained were independent of the time the hexadecane was present before the drops were placed on the slide. Droplets of the more dense dextran-rich bottom phase were placed randomly over the sample surface and allowed to equilibrate for 1 h. At this time photographs of the drops were taken through a horizontally mounted Zeiss Tessovar microscope equipped with an optical extension tube for increased magnification. Illumination of the chamber was achieved with a 150-W fiber
The coating technique used here was optimized on porous glass chromatography beads with commercially available dextran because the large surface area available per unit mass permitted straightforward, quantitative determination of the amount of dextran per unit area by wet chemical means (data not shown). However, there are potential restrictions on molecular packing within the pores of the beads, and it is unlikely that the available surfaces of the beads and quartz slides are chemically identical. Hence, extrapolation from the porous bead results to derive accurate values for the amount ofdextran bound per unit area to the slides was not warranted. The amount of dextran on the quartz slides is too small to be readily measured by wet chemical techniques. The quality and extent of dextran coatings was analyzed in two ways. Uniformity of coatings was indicated by the consistency of contact angles measured for dextran phase droplets randomly distributed on the surfaces and by the excellent agreement among XPS spectra
TABLE I Test Systems a n d Physical D a t a
System
Tie line length (%)
Densitydifference (kg m-3)
Interfacialtension (naN m-j)
5% W/W D e x t r a n 27.9/8.4% w / w P E G 8,000 5% w / w D e x t r a n 205/5.25% w / w P E G 8,000
14.7 14.7
58.1 48.0
0.0351 0.0228
Journal oj'Colloid and IntetJbce Science, Vol. 149, No. l, March I, 1992
157
W E T T I N G B EH A V IO R
obtained at five different locations on each slide. The relative amount of dextran on the surfaces as a function of MW was estimated using XPS to measure the fraction of the derivatized surface occupied by C-O carbon from which photoelectrons could be collected. Results of the XPS analysis are given in Table II expressed as the atomic percentage of the surface constituents accessible to XPS. The CC, C-O, and O-C-O carbon fractions were obtained from computer deconvolution of overlapping peaks, as illustrated in Fig. 1.
Contact Angle Measurements The results are summarized in Table III. They indicate a consistent decline in the contact angle as the MW of the coating increases. This effect was more pronounced for the lower MW dextran 27.9 two-phase system than for the system containing dextran 205. A decrease in the contact angle over 24 h is apparent for both test systems on all coated surfaces. This observation implies that there is a time dependency associated with the interaction between the free polymers in solution and the polymer bound to the glass surface, particularly for the higher MW coatings. DISCUSSION
XPS is a surface spectroscopic technique which provides analytical information on material located within about 50 A of the external boundary of polymeric samples (11 ). As a
high vacuum is required for the collection of photoelectrons, no solvent will be present. In our samples the dextran detected therefore will be collapsed into a more compact configuration than would be present in solution or when the molecule was bound to the surface by its reducing end in an aqueous environment. XPS spectra of carefully cleaned quartz surfaces always show a hydrocarbon (C-C) peak (11 ), presumably due to impurities in the environments to which the surfaces are exposed. Covalent attachment of a polymer having carbon-oxygen bonds (such as dextran) should show an overlapping, companion carbon peak due to C-O bonds of the carbohydrate. Figure 1 shows this and a companion, less intense peak at a higher binding energy in the deconvoluted spectrum. The higher energy peak presumably represents the O-C-O signal, since diethers, esters, and carboxylic acids all show peaks in this region (10). The C-C peak accounts for a relatively constant fraction of the total carbon detected as the MW of bound dextran is varied (Table II). A portion of this constituent likely derives from the silane coupling reagent but, considering the level of nitrogen detected is about tenfold lower, most of this material must be from other sources. Since dextran is the only source of C-O and the coupling reagent the only nitrogen-containing compound in the sample, the C-O and N signals must represent the levels of these two materials detectable. Evidently there is a great deal of silane on the surface which is not
TABLE lI Atomic Percentages of Constituents of Dextran-Derivatized Quartz Surfaces Detected by XPS for the Dextran Fractions Indicated Dextran
% C-C
% C-O
% O-C-O
%N
% Si
%O
% Other
Control a 52 107 147.5 205
12.2 b 7.9 7.7 6.8 11.1
-2.0 3.7 8.9 11.9
-0.2 0.5 0.8 1.2
2.1 0.7 0.5 0.7 0.7
29.8 21.5 20.5 18.2 14.7
55.9 67.1 66.1 63.9 57.2
-0.8 1.5 1.5 4.3
Amin ated quartz. b Uncertainty typically +0.15% for all constituents. a
Journal of Colloid and Interface Science, Vol. 149,No. 1, March 1, 1992
158
BOYCE ET AL. TABLE III
Contact Angles Measured (Degrees + SEM a) for Dextran-Derivatized Quartz Slides in Dextran/PEG Two-Phase Systems Containing Dextran 27.9 or Dextran 205 at the Times Indicated Dx 27.9 (5, 8.4) Coating (molec wt)
No Dx Dx Dx Dx Dx
coating 27.9 52 107 147.5 205
Dx 205 (5, 5.25)
Ih
168.3 148.5 141.6 125.3 118.5 105.4
_ 3.0 + 0.5 + 1.2 _ 3.0 _+ 3.8 _+ 3.2
24 h
170.2 133.9 132.3 118.3 113.0 84.0
Ih
+ 2.8 _+ 0.9 _ 1.7 _ 4.1 + 0.8 + 1.1
176.1 161.4 158.0 134.0 145.7 132.5
24 h
+_ 2.0 + 2.3 + 1.8 + 0.9 _+ 1.8 _+ 2.3
164.5 157.9 156.0 131.3 127.0 113.8
_+ 2.3 _+ 1.5 -+ 2.6 _ 1.1 _+ 2.3 _+ 2.0
a Standard error of the mean for 5-7 measurements.
coupled to dextran. If all the coupling compound was derivatized with dextran, taking dextran 52 as an example there would be 1,900 C - O groups per N, compared to the observed value of 2.9. However, the surface activation procedure used here gives an amine surface concentration of about one per 70 A2 (14). A fully hydrated molecule ofdextran 52 in solution has a cross sectional area of approximately 1.2 × 10 4 fik 2, calculated from its radius of gyration (15). This implies a ratio of one dextran per 170 nitrogens or a C-O:N ratio of about 11 if the polymers were close packed, each with the same cross-sectional area as a molecule in solution. The density of surface-attached polymer may be greater than this, however. Measurement of the amount of a commercial dextran l0 fraction bound to porous glass beads of known specific area gave a value of 1,400 ~2 per molecule (data not shown), half the value calculated from the solution radius of gyration, which translates to a C - O : N ratio of 20. This number may be somewhat underestimated due to the method of specific area measurement (gas adsorption) for the porous glass beads, and it ignores the breadth of the molecular weight distribution of the dextran. Nonetheless these calculations support the conclusion that most of the surface amines must be underivatized. As is seen in Fig. 3 (data taken from Table IIL the fractions of the detected surface material consisting of C - O and O - C - O increase Journal of Colloid and Interface Srience, Vol. 149, No. 1, March 1, 1992
approximately linearly with dextran MW, concomitant with a roughly linear decrease in the Si signal. The C - O and O - C - O linearities imply that the mass ofdextran present within the XPS beam sampling volume is greater the higher the MW. If the depth of the sampling volume was such that all the C - O and O - C - O groups of each M W contributed to the signal, the linearities would indicate that the number of molecules coupled per unit area was approximately constant as a function of MW, since the number of C - O and O - C - O groups per molecule are directly proportional to this parameter. However, the decrease in the Si signal as M W increases implies that the larger molecules are covering some of the Si detectable at lower MW. Evidently with higher MWs
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WETTING
some portion of the surface is occupied by regions of collapsed polymer chains which are thicker than the sampling depth of the spectrometer. Calculation of the diameter of spheres with the same mass and density as unhydrated dextran molecules is consistent with this idea, the values ranging from 47 to 73 for the MWs in Table II. The reduction with MW is not seen in the N percentage, presumably because uncertainty in the measurement masks any regular variation in the very small signal values observed. From the XPS results we conclude, therefore, that dextran is bound to the surfaces and that an increasing mass of dextran is bound per unit area as the dextran M W increases. In this work the contact angle is defined as the interior angle between the solid substrate and the surface of the dextran-rich drop. Hence, decreasing contact angles correspond to progressively greater wetting tendency by the dextran-rich phase. It is apparent from Table III that the contact angle decreases with: (a) increasing M W of the bound dextran, (b) decreasing M W of the free dextran in the twophase system, and (c) time of exposure. To attempt to understand these effects we consider the free energy differences which determine the contact angle, 0, using Young's equation appropriate to the present situation ( 16): cos 0 - (%~ - "rsb) _ 2X~ "Ytb
[11
"Ytb '
where 3'tb is the interfacial tension of the interface between the top, PEG-rich phase and the bottom, dextran-rich phase, and 7~t and 7sb are the interfacial tensions between the coated quartz surface and the PEG-rich and dextran-rich phases, respectively; A7 is the interracial free energy difference. Thus for a given phase system, where 7tb is constant, observed changes in cos 0 may be interpreted as being due to differences in the interfacial tensions between the coated surface and the two phases. The underivatized quartz surface exhibits a contact angle of nearly 180 °, implying a much lower energy interaction with the PEG-rich
BEHAVIOR
159
phase than with the dextran-rich one. This may be attributable to a stronger adsorption of PEG than dextran. However, we have not measured adsorption in this study so this explanation must remain a conjecture. Derivatization with any of the dextran fractions reduces 0 by making the surface more compatible with dextran and less so with PEG, presumably by the same molecular mechanisms that drive phase separation in these mixtures, namely an unfavorable interaction free energy between the monomers of the two polymers ( 17 ). Since we have been able neither to measure the absolute amounts ofdextran bound to the quartz nor to measure independently the average thickness of these layers, it is not possible to quote values for the concentration of polymer chains in the surface-attached layer. We therefore have not attempted to interpret any of the A3, data quantitatively using current theories of polymer mixtures ( 18, 19). However, the trends in changes in 0 may be rationalized qualitatively as follows. For a given phase system, 0 decreases with increasing MW ofdextran on the surface. As discussed above, the higher the MW, the greater the mass of polymer bound per unit area. Moreover, because of the larger molecular size, the bound layer would be expected to extend farther out into solution for higher M W fractions. Hence, since PEG is incompatible with dextran it might be expected to be rejected more readily by the more extensive high M W coatings than by lower molecular analogues, increasing 7~t and producing the effects observed. The second obvious trend in the data is the greater decrease in 0 observed for a given surface with the phase system containing the lower M W dextran fraction. These data illustrate an important difference between immiscible polymer solutions and simple two component mixtures as far as contact angle studies are concerned. The latter systems show a predictable decline in 0 as the surface or interfacial tension decreases (20, 21 ), while this may or may not be true for aqueous phase mixtures. The behavior of polymer systems is more Journal of Colloid and Interface Science, Vol. 149, No. I, March 1, 1992
160
BOYCE ET AL.
complex as 0 can decline or, as with the systems described here, increase with a falling interfacial tension. In the study at hand, the dextran 27.9 phase mixture yielded the lower contact angles of the two systems tested. When the values of AT are calculated and plotted as a function of coating MW, Fig. 4, the low MW dextran line falls below that of the dextran 205, except at the highest bound polymer MW. A MW effect is not unexpected, since a similar result is seen when macromolecules are partitioned in these systems where it is universally observed that if the MW of one of the phase-forming polymers is reduced, partition into the phase in which that polymer predominates is increased (22). That is, the interaction between the "substrate" and the phase becomes more favorable as the MW of the polymer in that phase is lowered. The likely reason for both effects is that the entropy of mixing of the "substrate" (either the macromolecule being partitioned or the surface-attached chains on the surface) with the phase polymer is greater per unit mass of polymer for lower MW chains (17). Hence, the free energy of mixing between the phase polymer and the substrate will be lower the lower the MW of the former. Lowering %b lowers 0, as observed. For the highest MW coating this effect is not observed, however, and the trend in the data suggests other ~
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factors may be involved, possibly polymer adsorption. It might be noted that the dependence on dissolved polymer molecular weight for all but the highest MW coatings is opposite to that which would be predicted if dextran adsorption to an uncoated surface were occurring and determining the relative surface compatibility. Polymer adsorption isotherms generally saturate at higher values for higher MWs in the range used here (23), so pure adsorption would be expected to have the opposite MW dependence, with respect to the phase polymers, to that which was observed. The present results do not imply that adsorption does not occur, however, since as discussed above PEG binding may well predominate on bare quartz and the PEG MW was held constant throughout. Considering the time dependencies observed, it is seen that with the phase system containing the lower MW dextran there are significant decreases in 0 between l and 24 h exposure for all molecular coatings. With the higher dextran MW system, however, significant time effects are only seen with the two highest MW coatings. This could reflect differences in transport rates of the two dextran fractions through the coatings following initial exposure of the surface to the phase systems. It would be expected that the diffusion ofdextran would be significantly hindered by layers whose thicknesses were of the order of their own dimension or greater. Thus, all surfaceattached layers would be expected to affect the time taken for the lower molecular weight dextran fraction to reach its equilibrium profile, while the higher MW polymer should be little impeded by the relatively low MW coatings, as observed. ACKNOWLEDGMENTS
20
40
60
80
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100
120
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140 Cooflng
160
180
200
220
x l O -3
FIG. 4. Interfacial tension difference, 4% vs molecular weight of dextran coating.
Journa/of Colloid and Interface Srience, Vol. 149, No. 1, March 1, i 9 9 2
The authors gratefully acknowledge the financial support of this work by the National Aeronautics and Space Administration, the National Institutes of Health, and the Medical Research Council of Canada (Grant MT 5759 to DEB).
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Journal of Colloid and lnterface Science, Vol. 149, No. I, March 1, 1992