Adsorption of Polyelectrolyte versus Surface Charge: in Situ Single

We have studied the effect of the pH and surface charge of mica on the adsorption of the positively charged weak polyelectrolyte (PE) poly(2-vinylpyri...
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J. Phys. Chem. B 2007, 111, 8597-8604

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Adsorption of Polyelectrolyte versus Surface Charge: in Situ Single-Molecule Atomic Force Microscopy Experiments on Similarly, Oppositely, and Heterogeneously Charged Surfaces† Yuri Roiter and Sergiy Minko* Clarkson UniVersity, Department of Chemistry and Biomolecular Science, 8 Clarkson AVenue, Potsdam, New York 13699-5810 ReceiVed: January 21, 2007; In Final Form: April 16, 2007

We have studied the effect of the pH and surface charge of mica on the adsorption of the positively charged weak polyelectrolyte (PE) poly(2-vinylpyridine) (P2VP) using atomic force microscopy (AFM) single-molecule experiments. These AFM experiments were performed in situ directly under aqueous media. If the mica’s surface and the PE are oppositely charged (pH > 3), the PE forms a flat adsorbed layer of two-dimensionally (2D) equilibrated self-avoiding random walk coils. The adsorbed layer’s structure remains almost unchanged if the pH is decreased to pH 3 (the mica’s surface is weakly charged). At pH 2 (the mica surface is decorated by spots of different electrical charges), the polyelectrolyte chains take the form of a 2D compressed coil. In this pH range, at an increased P2VP concentration in solution, the PE segments preferentially adsorb onto the top of previously adsorbed segments, rather than onto an unoccupied surface. We explain this behavior as being caused by the heterogeneous character of the charged surface and the competitive adsorption of hydronium ions. The further increase of polymer concentration results in a complete coverage of the mica substrate and the charge overcompensation by P2VP chains adsorbed on the similarly charged substrate, due to van der Waals forces.

Introduction It is well-known that polycationic polyelectrolytes (PE) such as poly(ethylene imine), polyamines, poly(2-vinylpyridine) (P2VP), etc., can adsorb strongly onto neutral and even onto similarly charged surfaces when van der Waals attraction of the PE chains to the substrate dominates the electrostatic repulsion between the PE chains and the similarly charged substrate. This behavior of PEs is broadly used for colloidal stabilization, adhesives, coatings, and the fabrication of thin films using the LBL method. The phenomenon occurs in different situations where proteins and cells interact with materials and where the overall charge of the protein globule (polyampholytic and polyamphiphilic in nature) may be similar to the charge of the substrate. However, PE’s adsorption onto weakly charged and similarly charged surfaces has not been fully investigated. The amount of PE adsorbed onto similarly charged surfaces is typically small, so many experimental methods fail to obtain results in this case. Although adsorption of PE is an old and well-established field continuously attracting the interest of researchers and industry, many gaps remain in our understanding of the phenomenon. Most experimental results obtained from measurements of adsorption excess, force-distance relationships, spectroscopy of adsorbed chains, scattering by adsorbed PE layers, and other techniques have been deeply reviewed in the literature.1-14 These methods deliver indirect information concerning the conformation of adsorbed PE chains so that the conformation can be reconstructed using different models. Direct analysis of the adsorbed PE chains’ conformation visualized in the air atmosphere brings an additional valuable option for the study of PE adsorption.15-29 Atomic force microscopy (AFM) experiments performed in situ under aqueous solutions in the attempt to visualize adsorbed small PE molecules30,31 and giant PE † Part of the special issue “International Symposium on Polyelectrolytes (2006)”. * Corresponding author. Phone: (315) 268-3807. Fax: (315) 268-6610. E-mail: [email protected].

chains,32-45 and to study PE using a single-molecule force spectroscopy,46-54 are of special interest because they allow for a statistical analysis of the adsorbed PE coils and a comparison of the experimental results with well-developed theories55-57 of PE adsorption. AFM experiments work well for dilute and semidilute surface concentration regimes. Sheiko and co-workers have recently suggested an interesting application for single polymer molecule experiments.58,59 A single polymer chain can be used as a miniaturized probe (sensor) to study its environment. The PE molecule is a promising candidate for such an application to study local interactions at the solid-liquid interface. Recently, we reported several in situ studies of the conformations of adsorbed chains of cationic PE on mica substrates. We followed the coil-to-globule transition of the adsorbed hydrophobic PE upon changes in the PE chains’ charge density30 and the transition of adsorbed PE into necklace structures in the presence of polyvalent counterions.31 In those experiments, we studied PE adsorption onto oppositely charged surfaces as a function of the charge density on P2VP chains. Here, we study the conformation and structure of PE adsorbed onto charged surfaces as a function of a charge on the adsorbing surface. The major findings of this work are as follows: (1) in a dilute surface concentration regime, adsorbed P2VP adapts conformation of equilibrated two-dimensional (2D) coils, although the chains are formally neither in a local equilibrium nor in a global equilibrium condition, (2) the statistics of the PE coil on oppositely and weakly charged surfaces perfectly fits the 2D self-avoiding random walk (SAW) model, and (3) an increase in the PE concentration at low pH results in adsorbed structures in which the PE segments preferentially adsorb onto the top of previously adsorbed segments (abbreviated here SOAS), rather than onto an unoccupied surface. Experimental Methods Materials. Poly(2-vinylpyridine) (P2VP) of Mw 159 000 and Mw/Mn 1.05 (secondary chromatography standard, Aldrich, MO)

10.1021/jp070518q CCC: $37.00 © 2007 American Chemical Society Published on Web 06/08/2007

8598 J. Phys. Chem. B, Vol. 111, No. 29, 2007 was used in the experiments. Hydrochloric acid (36.5-38%, EM Science, NJ) was used to prepare the P2VP solutions in Millipore water (18.3 MΩ‚cm) and to adjust the medium pH. V-1 grade muscovite mica, KAl2(Al,Si3)O10(OH)2,60 (Structure Probe, PA) was used as a substrate (discs of 15 mm diameter). Solutions. A stock P2VP solution (0.1 g/L) was prepared in Millipore water at pH 2.5 (HCl). Then the P2VP solutions for the experiments (working solutions) were prepared by diluting the stock solution with HCl solutions to adjust the pH to the desired value. The P2VP concentration was adjusted to 10-3 g/L or 10-4 g/L (monomer units concentration [2VP] ) 9.51 × 10-6 g‚equiv/L or 9.51 × 10-7 g‚equiv/L, respectively) by mixing P2VP solution with corresponding HCl solution. The working solutions’ pH values (pH meter 345, Corning Incorporated, Corning, NY) were measured after the completion of the AFM experiments (to avoid the working solution contacting the glass electrode). We excluded glass instruments and containers (mainly polyethylene materials were used) from all preparatory steps to avoid the adsorption of polymer molecules, which could change the concentration of solutions. All solutions were filtered using a Millex-LCR 0.45 µm (Millipore, MA). Unfortunately, it was not possible to determine precisely the degree of P2VP protonation because of its very low concentration (similar difficulties were reported earlier for P2VP concentrations over 5-50 times higher13,61). Puterman and coworkers62,63 have shown that P2VP’s coil-to-globule transition occurs at a degree of protonation equal to 0.5. The AFM experiments have shown that this narrow transition region was approached at pH 4.30 Further lowering of the medium’s pH increases P2VP’s protonation and stretching due to intramolecular repulsion. At pH 3.89 and lower, P2VP chains exhibit extended coil conformation.30 We performed an estimation of the P2VP charge by titration of the solution at 100 times greater concentration (0.1 g/L) of the polymer. AFM Experiments. AFM images were recorded using a MultiMode scanning probe microscope (Veeco Instruments, NY) operated in tapping mode. The samples were scanned using NP silicon nitride probes (Veeco Instruments, NY) with a radius of 20 nm, a spring constant of 0.32 N/m, and a resonance frequency in aqueous media of ∼9 kHz at an amplitude set point in the range from 0.6 to 2.2 V and a tapping force of about 98% of the set point at 1-2 Hz at ∼28 °C (temperature of the fluid cell in the MultiMode microscope, measured by a calibrated thermistor placed into the fluid cell). The muscovite mica discs were glued to metal supporting disks 15 mm in diameter (Structure Probe, PA) with a 2-Ton epoxy composition (Devcon Consumer Products, FL). Substrates were cleaved and carefully checked for surface quality and uniformity (no aging effect was observed for the mica samples several hours after cleaving). The cleaved mica disks were mounted in the liquid cell of the MultiMode microscope. Mechanical drift induced by the O-ring was decreased by precisely placing the O-ring to minimize lateral stresses. The polymer solutions were injected directly into the fluid cell, and technically (because of the sequence of the steps, i.e., adjustment of the laser beam’s reflection, the first manual search of resonance frequency, the tip’s first “rough” approach to the surface, the tip’s withdrawal, the second adjustment of the resonance frequency near the surface, and the second “fine” approach to the surface), it was possible to start the measurements about 7 min after the injection of the polymer solution. The thermal drift of the microscope scanner was minimized by incubating the assembled cell with the mounted mica disc for 0.5-2.5 h to equilibrate the heat flow in the cell induced by the instrument. We have demon-

Roiter and Minko strated that in the case of single-molecule studies under these conditions, the probe does not affect the sample, and the scanned molecules’ conformation remains unchanged for several hours.30 The actual time of the image recording varied from 7 min to 3 h after the polymer solution injection. The study of surface charges of mica versus pH was performed using functionalized AFM tips with primary amino groups (CT.AU.NH3.SNNovascan Technologies, Inc., IA) of 42 nm curvature radius and 0.12 N/m spring constant. For each set of conditions, we have recorded about 250 force-distance curves. Data Processing. Self-coded software was used to process the images.64 The chains’ coordinates were recorded by dragging a cursor along the zoomed contours of clearly isolated chains. The recorded coordinates were used to estimate the experimental values of the root-mean-square (rms) end-to-end distances, 〈r2〉1/2, and rms radii of gyration, 〈s2〉1/2. End-to-end distances were recorded only if both ends of the molecule were clearly distinguishable. About 100 molecules were processed for each pH value during 〈r2〉1/2 measurements. P2VP’s average contour length, LC, was estimated from the measurements of 530 extended chains, providing the same LC value as in previous studies,30 204 ( 91 nm. The fraction of monomer units in the second and third layers of P2VP on the surface was estimated at 10-3 g/L P2VP concentration by gradually flooding the AFM images using WSxM software.65 Flooding to a 0.2 nm image height determined the area occupied by the monolayer (A1), and then flooding to 0.6 and 1.0 nm heights provided the areas of the bilayers (A2) and trilayers (A3), respectively. The data for each set of conditions were calculated from 4 to 5 µm2 of corresponding images. The ratio of bilayer and trilayer areas to their sum with the monolayer area provided the fraction of monomer units in second, A2/(A1 + A2 + A3), and third, A3/(A1 + A2 + A3), layers, respectively. Results and Discussion Surface Charge: No Polymer Adsorbed. Mica consists of aluminosilicate layers ionically bonded by potassium ions. Muscovite cleaves along the potassium-oxygen layer. Mica’s surface is negatively charged in contact with water due to the loss of K+ ions from its surface into water. If pH decreases in the presence of a background electrolyte (∼1 mM), mica passes its isoelectric point (IEP) near pH 3 with the charge compensated mainly by the adsorption of hydronium ions (H3O+).3,28,66 Electrokinetic measurements in the presence of a buffer electrolyte show that a further decrease in pH charges the mica’s surface positively. However, this result disagrees with surface force measurements,13 which exhibit a relationship for negative charge versus pH with a minimum around pH 3. Since results reported in the literature differ by concentrations of buffer electrolyte solutions, we performed an evaluation of the mica surface for the same solutions which were used for the singlemolecule experiments with no added electrolytes and polymer. The mica surface charges were probed with an NH2-modified AFM tip. The results obtained (Figure 1a) suggest that mica is negatively charged in the studied range of pH values (2 < pH < 4): the positively charged tip jumps into contact with the mica surface in all experiments. The lowest density of negative charges on the mica surface was obtained at pH 3. This result is in good accord with ref 13. A further increase of the negative charge on the mica surface at pH levels below pH 3 was explained in the literature by the extraction of aluminum and potassium ions from the mineral surface in acidic aqueous solutions.

Single-Molecule AFM Experiments on Charged Surfaces

Figure 1. Force-distance curves recorded at different pH values with the NH2-modified tip (k ) 0.12 N/m) on the bare mica surface (a) and on the mica surface with P2VP molecules adsorbed at pH 2 from 10-4 g/L (b) and 10-3 g/L (c) solutions. The plots are shifted on the y-axis for convenience.

Using AFM, we probed the mica surface to determine whether the charges decorated the surface homogeneously. We have recorded the phase images of mica using an NH2-modified tip in tapping mode. At pH 3.00 and pH 3.57, we were not able to achieve a phase contrast, which suggests a homogeneous distribution of negative mica surface charges. However, it can be clearly seen from Figure 2a that mica does demonstrate a heterogeneous surface at pH 2.00. We may speculate that the phase contrast image is obtained when the positively charged tip is repelled from the sites with positive charges on the mica (red-colored spots) and attracted to sites with negative charges (blue-colored spots). The decision concerning the sign of the charges was made based on the phase contrast images obtained for adsorbed P2VP molecules (discussed below). Surface Charge: Polymer Adsorbed. We used the abovementioned approach to probe the mica surface charges after adsorption of single P2VP molecules from 10-3 g/L and 10-4 g/L solutions at pH 2. The same samples with the P2VP molecules adsorbed at pH 2 were used to measure forcedistance curves at different pH (Figure 1, parts b and c). It is

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Figure 2. Phase images recorded with the NH2-modified tip on the bare mica surface under a pH 2.00 aqueous medium (a) and on mica surface with P2VP molecules (b-d) adsorbed from 10-4 g/L solution at pH 2 (b), 10-3 g/L solution at pH 2 (c), and 10-4 g/L solution at pH 3.57 (d). The insets represent corresponding topographical images with a 2 nm Z-scale.

noteworthy that AFM scans of the surface visualizing the adsorbed chains proved that changes in pH caused no changes in the adsorbed amount of P2VP (evaluated to be 6.2 × 1013 chains/m2 for adsorption from 10-4 g/L solution and 3.2 × 1014 chains/m2 for adsorption from 10-3 g/L solution). The obtained results clearly demonstrate that the adsorption of P2VP resulted in a partial compensation of the mica surface charges for adsorption from the 10-4 g/L solution (Figure 1b). For the 10-3 g/L solution we observed a partial compensation of the mica surface charges at pH 3.57, a complete compensation of the mica charge at pH 3, and overcompensation (switch from negative to positive surface charge) of the mica charge at pH 2 (Figure 1c). The phase image for the latter sample (Figure 2c) demonstrates a substantially decreased phase contrast, supporting the conclusion of the force-distance measurements and suggesting that the positive charges decorate the surface almost homogeneously. At this point, we focus on the sample prepared from 10-4 g/L P2VP solution where the effect of P2VP adsorption on the surface charge is not very strong. The phase image of the sample at pH 3.57 (Figure 2d) shows a heterogeneous charge distribution due to the local compensation of the negative mica surface

8600 J. Phys. Chem. B, Vol. 111, No. 29, 2007 charges by positively charged P2VP. Comparison of the phase image with the topographical image (inset in Figure 2d) provides evidence for the charge compensation in areas with adsorbed P2VP chains. Thus, we concluded that a positive phase shift was caused by positively charged spots on the mica surface. The same sample shows a much higher surface density of positively charged spots at pH 2 (Figure 2b). Consequently, the mica surface is heterogeneously charged and appears as a surface patterned by charges with positive and negative signs. Only minor changes in the surface charges (as compared to the bare mica surface) were introduced due to the small surface concentration (6.2 × 1013 chains/m2) of adsorbed P2VP molecules (Figure 2b). Thus, in this article we distinguish four major regimes of adsorbed P2VP layers (in a dilute surface concentration regime) in terms of mica’s surface charge under aqueous solutions at different pH levels: (1) pH > 3snegatively charged surface. With no P2VP adsorbed, this regime was well documented with electrokinetic experiments and measurements of interaction forces between two surfaces in close proximity to each other as a function of separation13 using a surface force apparatus67 and colloidal probe microscopy (CPM),3 which revealed strong repulsion from the mica surface. Adsorption of a small amount (6.2 × 1013 chains/m2) of P2VP does not affect the mica surface charge. (2) Around pH 3, mica’s surface is weakly charged (a negative charge compensated by adsorbed cations). With no PE adsorbed, electrokinetic experiments showed a streaming potential which was significantly decreased, while CPM detected a transition from repulsion to attraction. The latter indicated a decrease in electrostatic repulsion and a switch from repulsion to attraction due to van der Waals forces. Only a small change in the surface charge was observed at 6.2 × 1013 chains/m2 surface concentration of P2VP, while 3.2 × 1014 chains/m2 may compensate the negative charge on the mica surface. (3) pH < 3 heterogeneously charged bare mica surface. With no P2VP adsorbed, the overall surface charge is negative; however, the surface of mica is decorated by negatively and positively charged spots. In this case, the adsorption of P2VP 6.2 × 1013 chains/ m2 resulted in a small decrease in the overall negative surface charge. (4) pH < 3 and a high adsorbed amount of P2VP. The mica’s surface is positively charged because of P2VP adsorption (3.2 × 1014 chains/m2) and the adsorption of hydronium ions, which competes with the P2VP adsorption. Regime 3 is clearly distinguished from the other regimes in the adsorption experiments as discussed below. Charge on Polyelectrolyte Chain. We have recently shown that adsorbed P2VP’s coil-to-globule transition occurs in the narrow pH range from pH 3.9-4.2.30 At pH below 3.9, P2VP chains pose extended conformation, and thus, they are strongly charged. Since we used very dilute solutions for the singlemolecule experiments, it was impossible to perform direct evaluation of the degree of protonation based on titration curves in this concentration regime. Nevertheless, we performed titration for a P2VP concentration 100 times greater (0.1 g/L) and plotted the degree of protonation versus pH (Figure 3). The plot shows that the protonation degree of P2VP at 0.1 g/L changes from about 60% to 80% as pH decreases from pH 3.6 to pH 2. We may expect some shift toward greater pH values of the relationship obtained when the P2VP concentrations are lower. Hence, in all four regimes listed above, P2VP will be highly charged with a small increase of the degree of protonation at pH < 3.5. Consequently, the major effect on P2VP adsorption at pH < 3.5 is due to the mica’s surface charge. Equilibrium versus Kinetically Trapped Conformations. Whether the adsorbed chain conformation is in equilibrium or reflects a kinetically trapped structure is the major concern in polymer adsorption experiments. Numerous experiments have

Roiter and Minko

Figure 3. P2VP protonation degree vs pH for titration of 0.1 g/L P2VP solution.

assumed that the adsorbed layer is in an equilibrium state because the system was allowed to approach equilibrium for a long time. However, many experiments have demonstrated that the adsorption kinetics can be very slow in some situations.68,69 The decision as to whether the adsorbed polymer is in the equilibrium state can be made based on the following: (1) the dynamic exchange between adsorbed molecules and molecules in solution. In many cases, adsorption of polymers is quasiirreversible with respect to the exchange between adsorbed polymer chains and molecules of solvent (this process is very slow because of the very low probability of detaching all adsorbed segments from the substrate). Adsorbed polymers can be exchanged by polymer chains arriving from the bulk solution and competing for the same sites on the substrate surface. Using isotope-labeled polymers, Pefferkorn and co-workers. have investigated the exchange between adsorbed and bulk molecules for different types of PEs.7,14,70 These experiments have demonstrated a high exchange velocity, giving evidence for the dynamic character of the adsorption-desorption processes. However, exchange was not always observed for 100% of the adsorbed chains. An adsorption overshoot and oscillations in the adsorbed amount were observed in very dilute concentrations. These effects were not clearly explained and were considered as kinetic effects caused by a range of consecutive adsorption/desorption steps approaching the equilibrium state.4,71 The criterion for the dynamic character of adsorption cannot be applied to very dilute polymer solutions, because their exchange kinetics are very slow. (2) Whether the adsorbed polymer is in an equilibrium state can also be determined based on an analysis of the conformation of adsorbed chains for the regime of the dilute 2D adsorbed layer: we use this criterion in this paper. We compare the experimental sizes of the adsorbed polymer coils to the theoretical predictions of the 2D SAW model. Figure 4 shows representative images of P2VP chains adsorbed from very dilute solutions at different pH values. In all cases, the adsorbed layer can be represented as a dilute 2D adsorbed layer. A statistical analysis of about 100 molecules of each case allowed us to determine the rms end-to-end distances and rms radii of gyration (Figure 5). Additional experiments were performed with added KCl salt (Figure 6a) to study the effect of ionic strength. Here, and in all other experiments with KCl, we used 20 mM solutions of the salt, which had an ionic strength twice that of HCl solution at pH 2. The addition of KCl results in some decrease in the chains’ dimensions due to the electrostatic screening effect. However, 〈r2〉 at pH 2 is smaller than 〈r2〉 obtained at pH 3 with KCl.

Single-Molecule AFM Experiments on Charged Surfaces

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Figure 6. Adsorption of P2VP extended coils onto mica from a 20 mM KCl solution at pH 3 and P2VP concentrations of 10-4 g/L (a) and 10-3 g/L (b), where 30.2% of the mica’s surface is covered by P2VP.

Figure 4. Adsorption of P2VP extended coils onto mica from 10-4 g/L solutions at different pH values: (a) pH 2, the mica surface is heterogeneously charged; (b) pH 3, the mica surface is neutral; (c) pH 3.57, the mica surface is negatively charged.

Figure 7. Diagram for scaling factor ν determined from the dependence 〈r2〉1/2 ∼ LCν and 〈s2〉/〈r2〉 ratio vs pH. Lines 1a-1c and 2a-2c represent denoted ν values (refs 72-75) and 〈s2〉/〈r2〉 ratios (SAW I (ref 74) and SAW II (ref 76)), respectively, for different models. Filled symbols correspond to the experiments without salt, and symbol ] represents results for solutions with 20 mM KCl.

Figure 5. Root-mean-square end-to-end distance (1) and rms radius of gyration (2) of adsorbed P2VP molecules vs pH. Symbol ] represents results for P2VP molecules adsorbed from 10-4 g/L solutions at pH 3 with 20 mM KCl. Lines are given for convenience to guide the reader. Deviation bars are estimations of characteristic distributions.

Thus, not only ionic strength, but also surface charge density, affects the conformation of the adsorbed P2VP chains at pH 2. Further analysis of statistical data has shown (Figure 7) that at pH 3 with no added salt, the molecules were 2D equilibrated in good agreement with the 2D SAW model for swollen polymer chains (scaling factor ν ) 0.76,72-75 ratio 〈s2〉/〈r2〉 ) 0.140),74 and that they were 2D equilibrated at pH 3 in the presence of 20 mM KCl (ν ) 0.75, 〈s2〉/〈r2〉 ) 0.138). The scaling factor and the ratio 〈s2〉/〈r2〉 for different models are depicted in the diagram in Figure 7. Now we have a good starting point for the discussion because the data obtained at pH 3 and pH 3.57 provide clear evidence that the adsorbed P2VP chains possess an equilibrium confor-

mation of swollen 2D coils. This equilibrium conformation is reached immediately upon adsorption. Also, it is unlikely that the AFM probe affected the conformation of polymer chains. The deviation obtained for the adsorbed P2VP chains at pH 2 is discussed below. The second question we address here is whether the adsorbed coils are in the “true” equilibrium state or were simply trapped by the adsorbent surface in a 2D “equilibrium” conformation. The answer was obtained from a simple experiment. P2VP single chains were adsorbed and visualized at pH 2. After this, we substituted the pH 2 aqueous solution with a pH 3.5 aqueous solution (by injection into the cell with the pH 2 solution of an excess of the pH 3.5 solution) and visualized the adsorbed molecules. Even after several hours of incubation in the pH 3.5 solution, no change in the conformation was observed, suggesting that the adsorbed chains are kinetically trapped. Consequently, we observe a very interesting behavior of polymer adsorption here. The adsorbed polymer chains are neither in global equilibrium (since the solution is very dilute and it may take very long period of time to approach the

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Figure 9. Fraction of monomer units in the second (1) and third (2) layers of P2VP adsorbed from 10-3 g/L solutions vs pH. Symbol ] represents the results for P2VP molecules adsorbed from 10-3 g/L solutions at pH 3 with 20 mM KCl added. Lines are given to guide the reader.

Figure 8. Adsorption of P2VP extended coils onto mica from 10-3 g/L solutions at different pH values: (a) pH 2, the mica surface is positively charged and partly covered with P2VP; (b) pH 3, the mica surface is neutral; (c) pH 3.57, the mica surface is negatively charged.

equilibration with respect to all molecules in the solution) nor in a local equilibrium state (since the chains are kinetically trapped). Nevertheless, the adsorbed chains’ statistics are consistent with the statistics of an equilibrated 2D SAW chain, implying a definition of the adsorbed molecule state in this case as a quasi-equilibrium state. This behavior of polymer adsorption is due to a very high activation energy for desorption. It is important to distinguish this quasi-equilibrium state from a kinetically trapped chain in the conformation of 3D to 2D projected coils, which can be obtained by a very rapid deposition of the polymer and evaporation of the solvent.31 Thus, the adsorbed P2VP chains are kinetically trapped; however, their conformations were rearranged near the adsorbent surface and correspond to the equilibrium 2D chain statistics. The latter conclusion is very important since we may use the chain dimensions as those which reflect the equilibrium conformation, and consequently, we may study the effect of the surrounding environment on the equilibrium conformation if all the changes in the surrounding environment occurred before adsorption of the polymer. Adsorbed Layer Structure versus Surface Charge. Very interesting results were obtained for the increased concentration of P2VP (10-3 g/L) when we approached a semidiluted surface concentration regime. The structure of the adsorbed layers obtained at different pH values is shown in Figure 8. Adsorbed P2VP chains overlap at this concentration. The adsorbed layer formed at pH 2 (regime 3) differs dramatically from the layers obtained at pH g 3 (regimes 1 and 2). At pH 2, the adsorbed P2VP layer has an SOAS structure, although many spots on the mica surface remain unoccupied because new chains approaching the surface prefer to adsorb onto previously adsorbed P2VP segments. In contrast to this, P2VP molecules at pH 3.57 tend to adsorb homogeneously onto the mica surface, bearing negative charges at pH 3.57. At pH 3, we observe an intermediate situation where some segments adsorb on top of

Figure 10. Adsorption of P2VP onto mica from 2.5 × 10-3 g/L solution (a) and 5 × 10-3 g/L solution (b) at pH2.

previously adsorbed segments, but the mica surface is homogeneously covered by PE. In order to test whether such behavior was caused by the screening effect in the presence of the HCl electrolyte, we performed a corresponding experiment in a pH 3 aqueous solution with 20 mM KCl added (Figure 6b). The results of this experiment are shown in Figure 9. Although the fraction of P2VP segments in the second and third layers increased compared to that of the adsorption experiment in solution at pH 3 with no salt added, the values are far below those obtained at pH 2. This result provides clear evidence that the behavior of adsorbed P2VP chains at pH 2 was not affected only by the increase in the solution’s ionic strength, but also by its increased acidity and, consequently, by changes in the mica surface charge. It is noteworthy that the thickness of the P2VP adsorbed layers was measured to be 0.4 nm. In an earlier experiment, Marra and Hair13 showed, using a surface forces apparatus, that the separation between two mica surfaces with adsorbed P2VP at pH 2 is 2.5 nm. This value agrees well with the SOAS structure (three layers of segments) shown in Figure 8a and

Single-Molecule AFM Experiments on Charged Surfaces

J. Phys. Chem. B, Vol. 111, No. 29, 2007 8603 interaction between charged P2VP chains and weakly charged mica when the electrostatic interaction with the substrate is less important. A further decrease in pH dramatically changes the structure of the adsorption layer (Figure 11a). The P2VP chains remain 2D coils, but are compressed. Newly arriving P2VP segments adsorb preferentially on top of adsorbed P2VP segments, rather than onto the mica’s PE-free surface, and form the SOAS structures. A decrease in pH may (1) increase charge density on P2VP chains, (2) increase screening electrostatic interactions, and (3) create a heterogeneously charged mica surface. Effect 1 cannot bring about the coils’ compression or create SOAS structures. Effect 2 is not strong enough to compress the chains, as can be concluded from reference experiments with KCl. In addition, effect 2 may not prevent adsorption of P2VP onto unoccupied spots on the mica. Effect 3 is just a hypothesis that arises from the AFM mapping of the mica surface. Literature reports on streaming potential measurements on mica are contradictory. Different authors report either remarkably negative values -60 mV77 or an IEP at pH 3.78 Our results are consistent with the recent report suggesting an inhomogeneous charge distribution on the mica substrate.79 Our speculative mechanism considers competition between the adsorption of P2VP and of hydronium ions. In this case, the coil size is decreased due to screening of electrostatic repulsion between charged P2VP segments because of a high local concentration of positively charged hydronium ions. At the same time, the adsorbed hydronium ions repel P2VP molecules from the mica surface, inducing the arriving molecules to adsorb initially on the negatively charged spots and then on the previously adsorbed segments. Finally, at the increased surface concentration of P2VP, the mica surface charge is overcompensated and arriving molecules are adsorbed on the similarly charged mica surface due to the van der Waals forces (Figure 10). Conclusions

Figure 11. Schematics of adsorption in terms of the surface charge effect on the conformation of adsorbed P2VP molecules: (a) at pH 2 chains are compressed due to the screening effect (high concentration of hydronium ions at the interface), and multilayered adsorption is well expressed; (b) at pH 3 with 20 mM KCl added the PE molecules are 2D equilibrated and compressed due to the screening effect (high salt concentration); (c) at pH 3 PE molecules are 2D equilibrated; (d) at pH 3.57 PE molecules are 2D equilibrated, but in a somewhat more extended conformation than in (c). Insets display representative images of AFM-visualized P2VP individual molecules.

Figure 9. Further increase in P2VP concentration results in a homogeneous coverage (Figure 10). Adsorption Mechanism. Here we summarize the experimental data. Our speculations about the adsorption mechanism are illustrated in Figure 11. Adsorption of highly charged PE onto an oppositely charged surface in dilute and semidilute surface concentration regimes results in 2D adsorbed layers (Figure 11d). In a dilute surface concentration regime the layer consists of 2D SAW coils. A decrease in surface charge (generated by decreasing pH) compresses the coils somewhat, but they remain equilibrated 2D SAW coils (Figure 11c). An increase in ionic strength (screening of electrostatic interactions) causes some decrease in the coils’ size and a small increase in the fraction of segments in the second and third layers (Figure 11b). This can be explained by the dominant van der Waals

Under opposite surface and PE charges, P2VP tends to adsorb onto a bare (unoccupied) surface. The adsorbed chains form equilibrated 2D SAW coils. This mechanism remains almost unchanged if pH decreases and approaches the substrate’s IEP. Around the IEP, van der Waals interactions with the substrate dominate electrostatic interactions. A further decrease in pH compresses the chains (2D laterally) and causes their segmenton-top-of-adsorbed-segment adsorption. We explain this effect speculatively by suggesting that (1) adsorption takes place on the heterogeneously charged mica surface and the competitive adsorption of hydronium ions, which in turn (2) increase electrostatic screening locally and (3) repel P2VP chains from unoccupied spots on the mica surface. The polymer segments in the second and third layers do not adhere strongly to the mica surface. This may cause the experimentally observed effect of adsorption overshoot or oscillations in the adsorption amount. Acknowledgment. This work was supported by the North Atlantic Treaty Organization under NSF Grant DGE-0411649 awarded in 2004. References and Notes (1) Messina, R.; Holm, C.; Kremer, K. J. Polym. Sci., Part B: Polym. Phys. 2004, 42, 3557. (2) Lichtenfeld, H.; Stechemesser, H.; Moehwald, H. J. Colloid Interface Sci. 2004, 276, 97. (3) Notley, S. M.; Biggs, S.; Craig, V. S. J.; Wagberg, L. Phys. Chem. Chem. Phys. 2004, 6, 2379.

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