Water Interface As Investigated by X-ray

© Copyright 1999 American Chemical Society

JULY 6, 1999 VOLUME 15, NUMBER 14

Gelatin Adsorption at the Air/Water Interface As Investigated by X-ray Reflectivity K. Abraham Vaynberg,† Norman J. Wagner,† Heiko Ahrens,‡ and Christiane A. Helm*,‡ Center for Molecular and Engineering Thermodynamics, Department of Chemical Engineering, University of Delaware, Newark, Delaware 19716, and Institut fu¨ r Physikalische Chemie, Johannes Gutenberg-Universita¨ t Mainz, Jakob-Welder Weg 11, D-55099 Mainz, Germany Received January 7, 1999. In Final Form: April 21, 1999 The method of X-ray reflectometry with electron density “labeling” of polyampholytes by heavy counterions has been demonstrated for measuring the adsorption profiles of gelatin at both hydrophobic (air/water) and like-charged hydrophilic (arachidic acid) surfaces. The results show that gelatin adsorbs onto a hydrophobic surface with a dense layer corresponding to the chain diameter, whereas this dense layer is absent for adsorption onto a hydrophilic like-charged surface. For both surface types a diffuse and extended adsorption layer is found.

Introduction Gelatin, a polyampholyte, is used in the food, photographic, and pharmaceutical industries as a stabilizer and interfacial property modifier, as well as a modifier of bulk fluid rheology.1 It is an important stabilizer of colloidal particles,2 but only very recently have there been determinations of the molecular level structure on colloidal surfaces.3,4 The importance of understanding protein and polyampholyte adsorption on surfaces for controlling emulsion stability and rheology is pronounced, as discussed in a recent review.5 To date, the molecular structure of adsorbed or anchored polymers at flat surfaces was best investigated by neutron reflectivity (see, for example, refs 6 and 7). The reason is threefold: the contrast between * Corresponding author. E-mail: [email protected]. † University of Delaware. ‡ Johannes Gutenberg-Universita ¨ t Mainz. (1) Ward, A. G.; Courts, A. In The Science and Technology of Gelatin; Academic Press: London, 1977. (2) Howe, A. M.; Clarke, A.; Whitesides, T. H. Langmuir 1997, 13, 2617. (3) Vaynberg, K. A.; Wagner, N. J.; Sharma, R.; Martic, P. J. Colloid Interface Sci. 1998, 205, 131. (4) Cosgrove, T.; Hone, J. H. E.; Howe, A. M.; Heenan, R. K. Langmuir 1998, 14, 5376. (5) Dickinson, E. Curr. Opin. Colloid Interface Sci. 1998, 3 (6), 633. (6) Atkinson, P. J.; Dickinson, E.; Horne, D. S.; Leermakers, F. A. M.; Richardson, R. M. Ber. Bunsen-Ges. Phys. Chem. 1996, 100, 994. (7) Higgins, J. S.; Benoit, H. C. Polymers and Neutron Scattering; Clarendon Press: Oxford, U.K., 1994.

different molecules or molecular groups can be varied by proton/deuterium exchange, neutrons are suitable for studies on a wide range of interfaces (solid/liquid, gaseous/ liquid, gaseous/solid), and the Å-wavelength is appropriate for scattering from structures on the 1-1000 Å length scale. Recent work has investigated the structure of gelatin adsorbed at the polystyrene/water interface, showing it was possible to determine a surface coverage.8 However, molecular resolution was not obtained. Compared to neutrons, X-rays from a standing anode provide more brilliance and a smaller background from incoherent scattering; thus, a higher Q-range is accessible. However, the contrast is proportional to the electron density, and thus, X-ray scattering was considered unsuitable for polymers at interfaces. However, one of us recently reported a successful approach for charged polymers (polyelectrolytes) dissolved in water:9 namely, adding electron-rich counterions. The counterion density F+(r) at a distance r away from a negative point charge on the polyelectrolyte chain is governed by the PoissonBoltzmann equation.10 In the Debye-Hu¨ckel approximation one obtains (8) Turner, S. F.; Clarke, S. M.; Rennie, A. R.; Thirtle, P. N.; Li, Z. X.; Thomas, R. K.; Langridge, S.; Penfold, J. Submitted to Prog. Colloid Polym. Sci. (9) Ahrens, H.; Fo¨rster, S.; Helm, C. A. Phys. Rev. Lett. 1998, 81, 1472. (10) Israelachvili, J. N. Intermolecular and Surface Forces; Academic Press: London, 1991; pp 55 and 215.

10.1021/la990011+ CCC: $18.00 © 1999 American Chemical Society Published on Web 06/05/1999

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F+(r) ) F+∞ exp(-eψ(r)/kT) ) F+∞ exp(-ecψ(0) exp(-κr)/rkT) (1) Here, ψ(r) is the potential of the point charge at a distance r, c is a constant, F+∞ is the bulk density of the positive counterions, and 1/κ is the Debye length. In the case of a 0.1 mol/L monovalent solution, κ ) 1/9.8 Å. This equation demonstrates that the counterion density decreases double-exponentially toward the bulk value. Because of the double-expoenential decay and the short Debye length, most counterions neutralizing the monomer charge are located directly next to charges on the polyelectrolyte chain. (With high ion concentrations, the term “diffuse double layer” is really misleading.) If one species of counterions (in our case the electron-rich Cs+ or Cl-) is offered in excess, these counterions will neutralize the charge and will therefore be embedded in the polyelectrolyte layer. Using exact calculations and other geometries for the charge distribution, one still finds a few angstroms away from the charge the bulk ion density. Thus, “counterion labeling” should allow us to trace the polymer segment density profile. As long as no specific ion binding occurs, the kind of counterion is not important. Since we and others have found that both the gelatin coverage and the free enthalpy of adsorption depend on the hydrophobicity of the surface,3,4 in our work we use this new approach of “counterion labeling” to investigate the electron density profile of gelatin adsorbed onto two different surfaces. The hydrophobic interface is the air/ water interface, whereas the hydrophilic interface is a compressed charged monolayer of arachidic acid.11 Materials and Methods Lime-processed, deionized bone gelatin (IEP ∼ 5.1) was obtained from Eastman Kodak Corp. (number average molecular weight, Mn ) 100K) and is the same material studied previously.3 All other chemicals are from Sigma. The X-ray setup is home built12 with a Cu anode (wavelength λ ) 1.54 Å). In X-ray experiments, the index of refraction n depends linearly on the electron density F and known constants (Thomson radius r0), n ) 1 - r0Fλ2/2π, and deviates only by 10-5 from 1. Therefore, approximations are possible, and the reflectivity can be seen as the Fresnel reflectivity RF of an infinitely sharp interface modulated by interference effects from the thin surface layer.13

|

∫

R 1 ) F′(z)eiQzz dz RF Fsub

|

2

(2)

where Fsub is the electron density of the bulk phase, F′(z) is the gradient of the electron density along the surface normal, and Qz is the wave vector transfer normal to the surface. For a 0.1 M CsCl bulk phase Fsub ) 0.3373 e-/Å3, which is larger than that of clean water (0.333 e-/Å3) due to the electron-rich Cs+ ions. The electron density profiles were calculated first by an indirect Fourier transform of eq 2.14 Then, to quantify the molecular parameters, the exact matrix formalism is used. The surface layer is parametrized as consisting of different slabs (each with an electron density and a thickness, as well as a roughness parameter). In all cases, the simulated reflectivity is convoluted with the angular divergence (0.012°). The electron density profiles obtained in this manner coincide within error with the profiles obtained by indirect Fourier transform, as found previously.9 To (11) Kjaer, K.; Als-Nielsen, J.; Helm, C. A.; Tippman-Krayer, P.; Mo¨wald, H. J. Phys. Chem. 1989, 93, 3200. (12) Baltes, H.; Schwendler, M.; Helm, C. A.; Mo¨hwald, H. J. Colloid Interface Sci. 1996, 176, 135. (13) Pershan, P. S.; Als-Nielsen, J. Phys. Rev. Lett. 1984, 52, 759. (14) Pederson, J. S.; Hamley, I. W. J. Appl. Crystallogr. 1994, 27, 36. Pederson, J. S. J. Appl. Crystallogr. 1992, 25, 129.

recognize and possibly eliminate correlated parameters, an interdependency analysis of the parameters is performed.15 To convert the measured electron densities to gelatin concentrations, a number of assumptions are made. First, we assume that the solvent is replaced in the electron-rich layers by gelatin with stoichiometric amounts of associated cations (cesium) and anions (chloride). In calculating the absolute electron density of this gelatin-ion complex, we consider and report two limiting cases: no hydrated water, and 2 water molecules of hydration per amino acid and 1 water molecule per adsorbed ion. Further, we use the average reported molecular weight of our gelatin (Mn ) 100 K) and the composition and number of ionized amino acids at the measured pH of our experiments.15 For the ions (both hydrated and not), we use radii reported by Israelachvili.10 Finally, using the measured solution density of gelatin (1.3 g/cm3), we can convert the electron densities to volume fractions, where we assume the free water in the swollen, adsorbed layer has its bulk properties. The data used in the caluclation with the resulting values of electron denstities are shown in Table 1. Before the gelatin was introduced in the Langmuir trough, the gelatin/water mixture with the desired ion concentration was allowed to swell for 1 h and then was heated for another hour to 40 °C. Isotherms and X-ray reflectivity were measured at 30 °C. Gelatin is itself a buffer, and so the system was found to have a pH of 7.1. To prepare a hydrophilic surface, arachidic acid was compressed to a surface pressure of 18 mN/m; a concentrated gelatin solution was introduced with Teflon tubing directly beneath the monolayer.

Results Gelatin adsorbs to the water surface,14 and an equilibrium surface pressure of 2.5 mN/m is measured (on a 0.01 wt % solution). The kinetics of the adsorbed surface layer are relatively slow (about 1 h). Several compression/ expansion cycles in a Langmuir trough yield an increase in the equilibrium pressure above that originally obtained for the freshly adsorbed surface layer (cf. Figure 1), suggesting irreversible gelatin rearrangement at the interface. The gelatin isotherm looked the same measured on CsCl and on NaCl solution (both 0.1 mol/L). A hydrophilic surface is prepared by compressing arachidic acid to 18 mN/m (corresponding to one surface charge per 150 Å2) and adsorbing the gelatin onto this surface layer. After the gelatin was introduced, the trough barrier was moved only slightly (cf. Figure 1b) to check stability without lateral manipulation of the adsorbed layer. Representative normalized X-ray reflectivity curves of gelatin adsorbed to a clean water surface are shown in Figure 2a. A typical feature is an apparent sudden increase in the measured reflectivity at the critical angle of the 0.1 M CsCl solution. This jump indicates a surface layer with a higher critical angle and with a thickness similar to or larger than the penetration depth of the X-rays (≈40 Å). Additionally, a small minimum occurs before the global maximum at Qz ≈ 0.09 Å-1. The segment density profile deduced from these curves basically consists of two parts, a thin and concentrated layer at the water surface and a longer, diffuse layer, which gradually disappears into the bulk phase (cf. Figure 3). However, a three-layer model for the electron density is necessary to fit the reflectivity data: the first slab describes the thin surface layer, and the second and third correspond to the long, diffuse part (cf. Table 2). Since this introduces more parameters than can be determined independently, we concentrate on the significant ones that are independent of modeling details. In the first slab, the concentrated gelatin surface layer is 14 ( 2 Å thick. Since 14 Å corresponds to the cross-sectional diameter of a gelatin (15) Asmussen, A.; Riegler, H. J. Chem. Phys. 1996, 104, 8159.

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Table 1. Parameters Used in Estimating Electron Densities of Gelatin-Ion Complexes and the Calculated Results gelatin density (by densitometry) gelatin amino acid Structure Cs+, radius Cs+, hydrated, radius Cl-, radius Cl-, hydrated, radius gelatin electron density gelatin-ion complex electron density gelatin-ion complex, hydrated, electron density

Figure 1. (a) Isotherm of 0.01 wt % gelatin adsorbed to clean water (compression speed 0.14 cm/s, corresponding to 3.85 cm2/ s, 40 °C). (b) Isotherm of arachidic acid on 0.1 M CsCl (dotted line) and with 0.01 wt % gelatin in the bulk phase (30 °C), the latter only for a small pressure range.

chain,16 we deduce that parts of the gelatin chain lay flat at the water surface. Deeper into the bulk, the adsorbed gelatin layer can be distinguished to be less concentrated, extending 108 ( 40 Å into the bulk phase. Within the resolution, the surface coverage is constant and independent of the surface pressure. With increasing compression, the reflectivity curve decays more swiftly for large Qz-values, a feature that is expected if the roughness σair is mainly due to capillary waves17 and increases with the lateral pressure. Yet, no clear interference maximum or minimum is found from the gelatin/water interface, suggesting an extremely high roughness σ23 of the polymer/water interface. Values of 42 ( 13 Å are found (averaged over six different experiments, cf. Table 2). Compressed arachidic acid was chosen as a charged, hydrophilic surface because it is a well-characterized (16) Rose, P. I. The Theory of the Photographic Process; Macmillan Publishing: Riverside, NJ, 1977; pp 52 and 57. (17) Schlossmann, M. L.; Schwartz, D. K.; Pershan, P. S.; Kawamoto, E. H.; Kellog, G. J.; Lee, S. Phys. Rev. Lett. 1991, 66, 1599.

1.3 g/cm3 pH 5.71, 110 (+), 80 (-) (ref 15, p 57) 0.169 nm (ref 10, p 55) 0.33 nm (ref 10, p 55) 0.181 nm (ref 10, p 55) 0.33 nm (ref 10, p 55) 0.408 e-/A3 0.523 e-/A3 0.453 e-/A3

monolayer from which very structured reflectivity curves can be obtained.11 In Figure 2b, reflectivity curves for an arachidic acid monolayer with and without adsorbed gelatin are shown. Beyond the first maximum, the curves are identical. Yet, similar to the observations reported before, with gelatin in the bulk phase one finds a sudden increase of the normalized reflectivity at the critical angle. This indicates a thick layer of adsorbed gelatin. Again, the gelatin/water interface is very diffuse, as observed by the roughness parameter; only a change in slope of the first maximum reflects the structure of the gelatin layer. To quantify the electron density profile, the arachidic acid monolayer was parametrized as described before.11 For the adsorbed gelatin, one slab was added. The thickness and electron density of the gelatin, as well as the gelatin/water roughness, were adjustable parameters (averaged over six different experiments, cf. Table 2). The thickness of the adsorbed gelatin was 47 ( 10 Å with a roughness σ ) 56 ( 10 Å, yet its density was only 0.346 e-/Å3, corresponding to 6 vol %. Thus less gelatin is found beneath the hydrophilic negatively charged monolayer than at the hydrophobic air/water interface. Conversion of the electron densities to gelatin concentrations follows the procedure outlined above. Table 3 summarizes the concentrations in the dense and diffuse layers for both systems. For example, the electron densities for the three regions under the air/water interface correspond to gelatin concentrations of 35, 20, and 5 vol %, respectively. Integrating along the surface normal yields a total surface coverage of 1.7 mg/m2, which is quite reasonable given previous work. The values in parentheses correspond to assuming waters of hydration, and the variation between the two sets of values provides a reasonable estimate of the uncertainty in our ability to extract polymer concentrations from measurements of electron densities. Discussion and Conclusions The most significant finding reported here concerns the densely packed gelatin chains, which form a monolayer at the water surface. Additionally, independent of the nature of the surface, we find an extended dilute gelatin layer. In the case of a hydrophilic, like-charged surface, the binding disturbs the solution chain geometry only locally. In contrast, to bind at the hydrophobic water surface, the coils deform substantially. For reference we note that a free coil in solution has Rg ) 200 Å, which results in a gelatin volume fraction of ∼0.4%. A relevant question is, does deformation upon adsorption occur only for gelatin or is it a more general property of polyampholytes? In the protein literature there is precedent for such observations. Measurements of the β-casein structure adsorbed from water onto hydrophobic surfaces show a dense hydrophobic adsorbed layer with charged trains extending into the aqueous phase as a diffuse layer.5 Polyelectrolyte brushes anchored to a hydrophobic surface form an interfacial layer of flat, closepacked chains, as has been reported for poly(styrene-

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Figure 3. Electron density profile of (top) gelatin at the air/ water interface ((a) as adsorbed and (b) compressed to 1/7 of the surface area) and (bottom) of (c) pure arachidic acid and of (d) arachidic acid with adsorbed gelatin.

Figure 2. Normalized X-ray reflectivity curves of gelatin (top, a) adsorbed onto clean water and (b) compressed to 1/7 of the surface area and of (bottom) arachidic acid compressed to 18 mN/m (c) without and (d) with adsorbed gelatin. Bulk phase conditions: 0.1 M CsCl, T ) 30 °C. Solid lines are fits to the model.

sulfonic acid).9 Indeed most of the observations in the literature on polyion adsorption to amphiphilic monolayers report an additional lateral repulsion that could not be explained purely by electrostatics.18 Thus, because adsorption occurs in the expanded state of the monolayer, the extra repulsion is probably a competition between the

amphiphiles and the polyions for binding places at the water surface. Differences in the absolute gelatin concentrations are observed when comparing our data to the adsorbed layer structure recently measured at the gelatin/polystyrene interface by Turner et al.8 They report a single diffuse layer of approximately 50 Å thickness containing 70 vol % gelatin. This is a higher adsorbed amount and shorter layer thickness than we observe for our hydrophobic air/ water interface but a comparable layer thickness and significantly higher concentration than our results for the charged, hydrophilic arachic acid interface. The values for the adsorbed layer thickness reported here are comparable to those determined for the adsorbed gelatin corona by small angle neutron scattering on acrylic3 and polystyrene lattices.4 The overall layer densities and adsorbed amounts are also comparable. However, our layer sizes are significantly smaller than those determined previously by measuring hydrodynamic radii by DLS on a colloidal latex or by surface forces measurements (see Table 8 in ref 3). These methods yield adsorbed layers of several hundreds of angstroms with average densities of ∼10-100 mg/cm3 adsorbed onto like-charged surfaces. X-ray reflectometry is able to measure the adsorbed gelatin structure with much higher resolution near the interface but cannot access these long length scales. Thus, comparing these results to those of neutron scattering and static and dynamic light scattering, one finds that DLS is most sensitive to the long tails, while SANS determines a mass(18) de Meijere, K.; Brezesinski, G.; Mo¨hwald, H. Macromolecules 1997, 30, 2337.

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Langmuir, Vol. 15, No. 14, 1999 4689 Table 2. Parameters for the Gelatin Adsorbed to the Water Surface and to Arachidic Acid Water Surfacea

L1 (Å) 14 ( 2

F1

(e-/Å3)

0.41 ( 0.01

F2

L2 (Å) 14 ( 7

(e-/Å3)

0.38 ( 0.01

L3 (Å)

F3 (e-/Å3)

σ23 (Å)

σ3sub (Å)

94 ( 32

0.348 ( 0.005

21 ( 3

42 ( 13

Arachidic Acidb L1 (Å)

F1 (e-/Å3)

σ (Å)

47 ( 10

0.346 ( 0.002

56 ( 26

a Values averaged across six experiments. Not given is the roughness at the air/polymer interface σ , because it depends on the state air of compression. b The fatty acid layer was unchanged.

Table 3. Parameters Derived from X-ray Reflectometrya arachidic acid interface extended region adsorbed gelatin (mg/m2) region thickness (Å) gelatin volume % a

0.26 (0.37) 47 4 (6)

air interface dense region1 14 35 (49)

dense region2 1.7 (2.3) 14 20 (29)

extended region 94 5 (8)

Numbers in parentheses assume full hydration.

averaged layer profile, but neither is especially sensitive to adsorbed layers of molecular scale. X-ray reflectometry with lower contrast and with roughness at the polymer/ water interface is not able to resolve the extended tails penetrating into solution. However, combined with “labeling” of the polyampholyte with heavy counterions, this technique provides an alternative method for examining the molecular-level structure in the adsorbed layer near the interface.

Acknowledgment. We appreciate the financial support of the GARN program. Also, the Deutsche Forschungsgemeinschaft (He 1616/5-2) was helpful. N.J.W. and K.A.V. acknowledge the financial and material support of Eastman Kodak Co. We also thank A. Howe and A. Rennie for providing preprints of their recent measurements. LA990011+