Langmuir 1992,8, 390-396
390
2HNMR Technique Sensitive to Surface Electrostatic Charge in Latex Dispersions Yihua Yue, John R. Rydall, and Peter M. Macdonald* Department of Chemistry and Erindale College, University of Toronto, 80 St. George Street, Toronto, Ontario, Canada M5A 1A2 Received May 20,1991. In Final Form: September 5, 1991
We have developed a new technique for measuring surface electrostatics in latex dispersions. It is based on the response of the quadrupole splitting from the 2H NMR spectrum of the zwitterionic surfactant hexadecylphosphocholine(HDPC),deuterated in the methyls of the choline quaternary nitrogen (HDPCy d e ) and bound as a monolayer at the surface of latex particles. When ionic surfactants are blended into the surface monolayer, the presence of cationic surfactants causes the 2H NMR quadrupole splitting to increase, while anionic surfactants cause it to decrease. Identical effects are observed when cationic or anionic ions and polyelectrolyte flocculants are adsorbed from aqueous solution onto the surface of the surfactant monolayer. In cases where two sources of surface charge are present simultaneously, the quadrupole splitting reports on the net surface charge. The results suggest that the choline group of HDPC undergoes a conformation change in response to surface charge similar to that of phosphatidylcholine in lipid bilayer membranes. We conclude that 2H NMR can be employed to quantitate latex surface electrostatics even in difficult circumstances such as flocculated colloidal dispersions. Introduction Surface electrostatics play a decisive role in determining the stability of colloidal dispersions.1>2Manipulation of colloid stability is achieved by perturbing the surface electrostatic potential. For instance, coagulated or flocculated colloids are produced by the addition of agents which act to reduce the surface potential, thereby permitting individual particles to approach one another more readily. Unfortunately, a detailed understanding of the effects of flocculants and coagulants on surface electrostatics is currently lacking. This is due, primarily, to the difficulties of obtaining meaningful measurements of surface charge in condensed as compared to dispersed particulate states. It is well established that deuterium nuclear magnetic resonance (2H NMR) spectroscopy provides a means to measure surface charge in lipid bilayer membrane vesicles, which likewise constitute a condensed particulate state. Seeligand co-workers3have shown that the phosphocholine head group of the naturally occurring membrane lipid phosphatidylcholine (lecithin) behaves like a "molecular voltmeter" in that it senses and responds to membrane surface charge by altering its conformation. This response can be monitored through the changes it produces in the quadrupole splitting measured from the 2HNh4R spectrum obtained when deuterium labels are placed on the phosphocholine group. It seems apparent that a transfer of the "molecular voltmeter" technology developed in lipid bilayer membranes to the study of surface electrostatics in colloid dispersions would be desirable. In order to accomplish such a technologytransfer, the phosphocholine group must first be localized to the surface of the colloidal particle, and once localized it must provide a 2H NMR spectrum capable of reflecting changes in the choline conformation. Recently, we have synthesized and characterized a homologous series of zwitterionic surfactants consisting
* To whom correspondence should be addressed.
(1) Derjaguin, B. V.; Landau, L. Acta Physicochim. URSS 1941, 14, 633. (2) Verwey, E. J. W.; Overbeek, J. Th. G. Theory of Stability of Lyophobic Colloids; Elsevier: Amsterdam, 1948. (3) Seelig,J.;Macdonald, P. M.; Scherer,P. G. Biochemistry 1987,26, 7535.
0743-7463/92/2408-0390$03.00/0
of an n-alkyl chain esterified to phosph~choline.~ When packaged in surfactant form phosphocholine preferentially binds to the hydrophobic surface of latex particles. Hexadecylphosphocholine (HDPC) was shown to adsorb to polystyrene particles with especial avidity and to form a monolayer of approximately erect surfactant molecules at the solid/water interface. The bound HDPC, when deuterolabeled at the methyls of the choline quaternary nitrogen (HDPC-r-de), provides a 2H NMR spectrum consisting of a Pake doublet with a characteristic quadrupole ~plitting.~ Thus, the necessary prerequisites to a transfer of the "molecular voltmeter" technology to colloid dispersions have been satisfied. However, it has not been proven yet that the 2H NMR spectra, and by implication the phosphocholine conformation, undergo any specific response to surface charges when HDPC is bound a t the particle surface. In this report we demonstrate the sensitivity to surface charge of the quadrupole splitting in the 2HNMR spectrum from surface-bound HDPC-yde. Two distinct tactics were employed to manipulate the surface charge. In the first instance HDPC was mixed with various proportions of either a cationic or an anionic surfactant and the resulting mixture was adsorbed to the surface of polystyrene particles. Alternately, charged ligands were allowed to adsorb from aqueous solution onto the surface of an HDPC monolayer preformed at the polystyrene particle surface. In both cases, the changes produced in the surface charge density were accompanied by corresponding changes in the 2HNMR quadrupole splitting. These were such as to indicate that the "molecular voltmeter" is fully operational in latex dispersions. Materials and Methods The synthesis of HDPC has been described previously? Deuteron labels were introduced into the methyl groups of the choline quaternary nitrogen by replacing methyl iodide with methyl-& iodide (Aldrich Chemicals, Milwaukee, WI) in the final reaction step, thereby yielding HDPC-yds. Homodisperse, emulsifier-free polystyrene latex was made according to the method of Kotera et a1.6in a single stage process, (4) Macdonald, P. M.; Rydall, J. R.; Kuebler, S. C.; Winnik, F. M. Langmuir 1991, 7, 2602. (5) Macdonald, P. M.; Yue, Y.; Rydall, J. R. Langmuir 1992,8, 164.
0 1992 American Chemical Society
Langmuir, Vol. 8, No. 2, 1992 391
Surface Electrostatic Charge in Latex Dispersions
using K2S208as the initiator. The styrenemonomer was vacuum distilledprior to use. After reactionthe formed latex was cleaned using ten cycles of centrifugation and redispersion (including sonication). The number-average particle diameter was determined to be 520 10 nm from scanning electron micrographs. The particle surface charge density was determined to be -2.2 0.1 pC/cm2from conductometric titrations. Surfactantbindingto the polystyreneparticleswas performed as follows. Stock solutions of the pure surfactants in organic solvent were mixed in the proportions appropriate to yield the desired surfactant composition,the solvent was removed under a stream of nitrogen gas followed by high vacuum, and the surfactantawere f i i y diasolved in water. Aliquota of these aqueous solutions of surfactant were added to a 16 wt 7’% polystyrene latex at 23 O C (totalweight of latex was typically 0.7 g) such that the desired final surfactant concentrationwas obtained (8 X M for saturation binding). During the binding period the surfactant/polymer mixtures were periodically gently vortexed to maintain homogeneity. After equilibration for 4 days: the polystyrene particles were separated from the aqueous supernatant by centrifugation (7OOOg, 10 min), the supernatant was removed, and the condensed latex was transferred to an NMR tube for measurementas described below. The concentrationof polystyrene particles in the pellet fractions was 42 wt % For the adsorption of flocculanta and aqueous ions, an aliquot of an aqueous stock solution of the desired agent was added to polystyrene latex preequilibrated for 4 days with HDPC, and following a further 2 h of equilibration,the polystyrene particles were separated from the aqueous supernatant by centrifugation as described above and transferred to an NMR tube. 2H NMR spectra were obtained at 45.98 MHz on a Chemagnetics CMX300 NMR spectrometer. The quadrupole echo technique’ was employed for the polystyrenepellet using quadrature detection and complete cycling of the pulse pairs! Particulars regarding the 90° pulse length (2.0 ps), the interpulee delay (40 ps), the recycle delay (100ms), the spectral width (10 kHz),the data size (2K),and the number of acquisitions (16 OOO) are those noted in parentheses. Cetyltrimethylammoniumbromide (CTAB), sodium hexadecyl sulfate (SHDS),poly(sodium 4-styrenesulfonate)(PSSS) (molecularweight -70 OOO), and polyethylenimine(PEI) (50w t % in water, molecular weight 50000-60000) were purchased from Aldrich Chemicals (MilwaukeeWI). All other chemicals were of reagent grade or better.
*
.
Results The 2HNMR spectrum of HDPC-y-ds bound as a monolayer at the surface of polystyrene particles is shown in Figure 1. The spectrum consists of a pair of narrow resonance lines superimposed upon a broad, axially symmetric Pake pattern. The narrow resonance line at 0 Hz arises from the natural abundance deuterium in water, since the spectrometer frequency was referenced to HDO. The narrow resonance at -79 Hz originates from “free” HDpc-7-d~present in the aqueous interstices between particles. At the HDPC concentration used here, the surface of the polystyrene particle is saturated with bound surfactant, leaving a small excess in solution. The broad Pake pattern arises from “bound” HDpc-7-d~. This spectral lineshape is characteristic of deuterons in a motionally restricted environmentgsuch as that of the closely packed HDPC surface monolayer. At saturation-coverage of the polystyrene particles, each HDPC molecule occupies approximately 35 A2of surface, which is not much greater than the transverse cross-sectional area of HDPC (25 A2) estimated from CPK model^.^ The quadrupole splitting ~~
(6) Kotera, A.; Furusawa, K.; Takeda, Y. Kolloid 2.2.Polym. 1970, 239, 677.
(7) Davis, J.; Jeffrey, K. R.; Bloom, M.;Valic, M. I.; Higgs,T. P. Chem. Phys. Lett. 1976, 42, 390. (8) Griffin, R. G. Methods Enrymol. 1981, 72, 108. (9) Seelig, J. Q.Reu. Biophys. 1977, 10, 353.
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Figure 1. Deuterium NMR spectrum of HDPC-y-de bound to polystyrene particles as a surface monolayer. The two narrow resonance lines at 0 Hz and -79 Hz correspond to residual deuteriumin water (HDO)and to ’free” HDPC-y-de,respectively, present in the aqueous interstices between particles. The broad Pake pattern is attributed to ‘bound” HDPC-y-de.
corresponds to the separation, in Hz, between the two maxima in the Pake pattern and, in this instance, equals 574 Hz. Note that the frequencies of these two maxima (+208 and -366 Hz) are such that the Pake pattern is positioned symmetrically with respect to the narrow resonance at -79 Hz. Under the conditions employed to acquire such 2HNMR spectra,the intensitiesof the signals from “bound” and “free” HDPC-y-ds are not proportional to their relative amounts present in the sample. This is a consequence of the fact that the recycle delay usedduring spectral acquisition (100 ms) was short relative to the longitudinal (2’1) relaxation time of the HDO or the “free” HDPC-y-d6.10 Under “fully relaxed” conditions (recycle delay > 5 times 2’1) the intensity of the “free” HDPC-r-ds resonance is indeed proportional to amount of free surfactant and in fact may be used to generate equilibrium binding isotherm^.^ Figure 2 shows a series of 2HNMR spectra from HDPCy d 6 bound to polystyrene particles as a mixture containing various proportions of either CTAB, a cationic surfactant of equivalent alkyl chain length, or SHDS, an anionic surfactant of equivalent alkyl chain length. In each instance the 2H NMR spectrum consists of a superposition of the two narrow resonances assigned to HDO and to “free” HDPC-y-ds upon the broad Pake pattern attributed to “surface-bound” HDPC-y-de. The effect of blending in an increasing mole fraction of the cationic CTAB was to increase the quadrupole splitting from HDPC-yds, as shown in the upper two spectra in the figure. In contrast, the effect of an increasing mole fraction of the anionic SHDS was t o decrease the quadrupole splitting from (10) Kuebler, S.C.; Macdonald, P. M. Langmuir 1992,8, 397.
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Yue et al.
392 Langmuir, Vol. 8, No. 2, 1992
Table I. Effects of Electrolytes and Polyelectrolytes on the Deuterium NMR Quadrupole Splitting from HDPC-r-ds Bound as a Surface Monolayer on Polystyrene Particles additive quadrupole monolayer composition splitting, Hz 100% HDPC 574 NaCl 26mM 560 KCl 26mM 556 NaC104 25mM 535 0.5 mM CaClz 60 1
49% HDPC + 51% SHDS 48% HDPC + 52% CTAB
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function of the mole fraction of admixed anionic or cationic surfactant: CTAB (circles),SHDS (squares),pure HDPC (triangle); closed symbols,HDPC plus CTABor SHDSmaintained constant; open symbols, HDPC maintained constant, CTAB or SHDS added in excess. HDPC-746, as shown in the lower two spectra. These observations suggest a specific response of the quadrupole splittings to alterations of the surface charge as imposed by the presence in the HDPC monolayer of added CTAB or SHDS. Note that only a single quadrupole splitting is observed in each situation. Figure 3 illustrates the manner in which the quadrupole splitting from "surface-bound*HDPC-y-ds varies with
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surface charge for an entire series of different mole fractions of CTAB or SHDS. The changes in the quadrupole splitting are progressive with increasing mole fraction of charged surfactants, and the direction of the change is opposite for cationic versus anionic surfactants. The largest mole fraction of CTAB which was tested (over 0.8) caused the quadrupole splitting to more than double relative to its value in the absence of CTAB. A similar mole fraction of SHDS caused the quadrupole splitting to collapse virtually to zero. In one series of experiments the l l l ~ l l l l total surfactant concentration was maintained at a constant saturating level while the proportion of HDPC to charged surfactant was varied. In a separate series of experiments the concentration of HDPC was maintained at a constant saturating level and the charged surfactants were coadsorbed in excess. As illustrated in Figure 3, the two experiments yielded virtually identical results when the quadrupole splittings are plotted as a function of the overall mole fraction of charged surfactant. Thus, there appears to be no preferential binding of HDPC versus CTAB or SHDS. Under the conditions used here, all surfactants were present at concentrations above their respective critical micelle concentrations, and the total surfactant concentration was always sufficient to achieve saturation of surface binding. We assume that the composition of any one surfactant micelle mirrors the overall surfactant composition and that formation of a surfactant layer at the surface of a polystyrene particle occurs followingadsorption of a surfactant micelle. Hence, there is no reason to expect a priori any difference between the mole fraction of charged surfactant added and the mole fraction present at the particle surface. The main factor determining the quadrupole splitting, provided surfactant adsorption is saturating, is then the mole fraction of charged surfactant present at the surface. The quadrupole splitting is not linearly related to the mole fraction of charged surfactant but instead appears to approach a limiting value at high mole fractions. In order to better define the range of surface charge effects to which HDPC responds, we tested a number of electrolytes and polyelectrolytes for their influence on the 2HNMR quadrupole splitting from surface-bound HDPC7 d 6 . These agents may influence the surface electrostatics by binding to the particle surface, where their presence imparts a surface charge proportional to the extent of binding and the valence charge of the particular ion. The results are tabulated in Table I. The 1:l electrolytes sodium chloride, potassium chloride, and sodium perchlorate, at concentrations of 25 mM, each cause the quadrupole splitting to decrease somewhat relative to its value in their absence, with sodium perchlorate displaying the
Langmuir, Vol. 8, No. 2, 1992 393
Surface Electrostatic Charge in Latex Dispersions
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largest effect. The 1:2 electrolyte calcium chloride causes a far greater response, resulting in an increase in the quadrupole splitting. The effect depends on the concentration of calcium chloride and leads to an increase a t 500 mM of 115Hz over the value measured in the absence of added salt. The anionic polyelectrolyte poly(sodium styrenesulfonate) (PSSS) and the cationic polyelectrolyte polyethylenimine (PEI) have relatively modest effects on the quadrupole splitting, and both lead to small decreases. The results described above demonstrate that the quadrupole splitting from “surface-bound” HDPC-y-ds responds to surface charges whether their origin is intrinsic to the monolayer, as is the case for ionic surfactants, or extrinsic to the monolayer, as is the case for adsorbed electrolytes. I t is desirable to demonstrate that the response of the quadrupole splittings to these two types of surface agents proceeds through a common physical mechanism, i.e., through changes in the surface charge density. Figure 4 shows the sensitivity of the 2H NMR spectrum of surface-bound HDPC-r-ds to the interplay between two different sources of surface charge. The top spectrum (A) in the figure was obtained with a 100% HDPC-r-ds monolayer exposed to the anionic polyelectrolyte PSSS. The resulting quadrupole splitting is only marginally reduced relative to ita value in the absence of PSSS, indicating little adsorption of PSSS to the surface of the HDPC monolayer. In the middle spectrum (B), the HDPC-yd6 had been mixed with 50 mol % CTAB and the mixture allowed to adsorb to the polystyrene particle surface. As expected in the presence of positive surface charge, the resulting quadrupole splitting increased substantially relative to its value in the absence of CTAB. In the lower spectrum (C),PSSS was added to the 1:l HDPC/ CTAB saturated particles. In this instance PSSS had a
large effect on the quadrupole splitting, reducing its size and counteracting the influence of the positive surface charge originating from the CTAB. Evidently PSSS displays a preference for binding to the positively charged CTAB-containing surfactant monolayer, as may be discerned by comparing its effects on the quadrupole splitting from 100% H D p c - 7 - d ~versus 1:l HDPC-y-dslCTAB monolayers. It is also evident that in all cases only a single quadrupole splitting is observed in the 2HNMR spectrum, so that 2HNMR appears to report on the net surface charge averaged over all charged species present at the monolayer surface. The actual values of the quadrupole splittings measured for these experiments are listed in Table I. We also performed the mirror-image experiment (from an electrical charge perspective) in which the cationic polyelectrolyte PEI was adsorbed to a particle surface monolayer consisting of 1:lHDPC-yd6ISHDS. The results of this experiment are listed in Table I. The preference of cationic PEI for the monolayer surface containing anionic SHDS is evident from a comparison of its effects on the quadrupole splitting from 100% HDPC-yd6 versus 1:l HDPC-ydelSHDS monolayers. In the latter case, the effect of PEI on the quadrupole splitting was to counteract the effect of SHDS. Thus is would appear that the response of the quadrupole splitting to these various agents is the result of their combined effects on one particular feature of the particle surface, its electrical charge.
Discussion The “molecular voltmeter” concept arose from 2HNMR studies of the effects of surface charge on the conformation of the cholinehead group of the bilayer-membrane-forming lipid pho~phatidylcholine.~ The evidence suggests that surface charges exert a torque on the choline group such that its angle of tilt with respect to the plane of the surface is altered.11-13 A schematic diagram of the “choline-tilt” model is shown in Figure 5. The torque arises because the quaternary nitrogen of the choline group is either attracted to or repelled by opposite or like surface charges, respectively. When deuteron labels are placed on the choline group, the 2H NMR spectrum reflects the altered choline tilt through changes in the quadrupole splitting. In this respect the choline group behaves like a “molecular voltmeter”, responding to and reporting on surface electrostatics. The quadrupole splitting is a sensitive monitor of local molecular geometry through its angular dependence which is embodied in the expression AUi
-= $0 cos2p - 1)SJ
AUQ where AUQis the static quadrupole splitting (125 kHz), B (11) Scherer, P.G.;Seelig, J. Biochemistry 1989, 28, 7720. (12) Rous, M.; Neumann, J. M.; Hodges, R. S.;Devaux, P.;Bloom, M. Biochemistry 1989,28, 2313. (13) Macdonald, P. M.; Leisen, J.; Marassi, F. M. Biochemistry 1991, 30, 3558.
Yue et al.
394 Langmuir, Vol. 8, No. 2, 1992
is the angle between the C-D bond vector and the axis of motional averaging (taken to be the long molecular axis of the phospholipid), and Sf is an order parameter representing the degree of off-axis wobbling of the choline group.9 There are three possible deuterolabeling positions in the choline group as shown: the methylene next to the phosphate (referred to as the a position), the methylene next to the nitrogen (referred to as the j3 position), and the methyls attached to the nitrogen (referred to as the y position)
Each of these positions exhibits a particular dependence of the quadrupole splitting on the surface charge.3 Specifically, the quadrupole splitting from the a position increases with added negative surface charge and decreases with added positive surface charge. The effects are opposite for both the p and the y positions, so that negative surface charges cause their quadrupole splittings to decrease while positive surface charges cause their quadrupole splittings to increase. This counterdirectional effect of surface charge on the quadrupole splittings from the a versus the j3 or y positions is evidence that the choline group undergoes a concerted conformational change in response to the influence of surface charge. The strength of the "molecular voltmeter" concept is that it provides a unifying framework within which to understand the response of the choline group to an array of chemicallydiverse agents, all of which have one property in common-they alter the charge present a t the surface of bilayer membranes. These include charged amphiphiles such as phospholipids, anaesthetics, and peptides, which physically intercalate among the lipid molecules and become an intrinsic part of the bilayer proper. Included as well are charged species such as aqueous ions and peptides and proteins which bind only to the external membrane surface but which, in the process, produce a surface charge. The physical analogiesbetween a lipid bilayer membrane and a monolayer of surfactant adsorbed at the surface of a polymer particle suggest that the "molecular voltmeter" could be operative in both situations. The results reported here represent the first indications that choline is able to respond to the influence of surface charge in a system other than a bilayer membrane and that there exists a counterpart in the surfactant monolayers to many of the effects which have been observed in lipid bilayers. The fundamental finding is that the quadrupole splitting from surface-bound H D p c - y - d ~responds in a direct fashion to the presence of charged surfactants in the surface monolayer. The presence of such charged surfactants directly alters the monolayer surface charge in proportion to the mole fraction added. Since the effects of anionic and cationic surfactant on the quadrupole splitting are counterdirectional, this suggests a specific response of the choline conformation to the surface charge, rather than some nonspecific alternate effect such as an increase or decrease in overall mobility of the choline group. The direction of the response to negative surface charge (quadrupole splitting decreases) and to positive surface charge (quadrupole splitting increases) is precisely that which is observed for y-deuterated choline in phosphatidylcholine bilayer membranes,13again suggesting that a mechanism similar to that deduced in bilayer membranes is operating in the HDPC surface monolayer. The surface charge
density, u,may be calculated using the following expression I
u=e-
CXiSi i where Xi is the mole fraction of a particular surfactant bearing a charge Zi and occupying a surface area Si, while e is the elementary charge (1.6022 X 10-19 C). The summation is over all species present at the surface, whether intrinsic or extrinsic. For binary mixtures of HDPC with either CTAB or SHDS, and assuming identical cross-sectionalareas of 35A2for HDPC, CTAB, and SHDS, the surface charge density will depend only on the mole fraction, Xi, of added CTAB or SHDS as follows u = Xi(0.4578C m-')
(3) The size of the change in the quadrupole splittings for the maximum surface charge density achievable with CTAB or SHDS (h600Hz for u = f0.4 C m+) is comparable to the size of the changes obtained for comparable surface charge densities in phospholipid bi1a~ers.l~Hence the sensitivity of the choline group to surface charge density appears to be similar in both bilayer membranes and HDPC monolayers. In the above expression we have ignored the intrinsic negative surface charge of the latex particles themselves. It is not yet clear how such charges will influence the 2H NMR spectrum of surface-bound HDpc-y-d~,but it is likely that a perturbation not only of the "choline-tilt" but also of the overall monolayer structural ordering needs to be considered. At this juncture eqs 2 and 3 must be taken, therefore, to represent relative rather than absolute surface charge densities. We note, however, that the charge density of the latex particles alone is roughly equivalent to that expected in the presence of a mole fraction of SHDS of only 0.05. The nonlinear relationship observed here between the quadrupole splitting and the mole fraction of added charged surfactant reproduces similar nonlinearities observed in bilayer membranes. They appear to stem from the harmonic dependence of the torque-countertorque pair which, according to the "choline-tilt" model, dictate the equilibrium choline tilt angle for a given surface charge density.11-13 At extreme surface charge densities, the electrical field torque falls off harmonically while the internal-resistance countertorque increases harmonically, leading to a decreased change in the quadrupole splitting per unit increase in surface charge density. In that portion of the curve where the quadrupole splittings are approximately linearly related to the mole fraction of charged surfactant, we can express their interdependence in an equation of the form Avi = mixi + Avo
(4) where Aui is the quadrupole splitting measured a t a particular mole fraction Xi of species i, mi is the slope of the line for a particular species, and AVOis the quadrupole splitting in the absence of added charged surfactant. For CTAB the slope in eq 4 was found to equal +1264 Hz, while for SHDS the slope equaled -930 Hz. For purposes of comparison we note that the negatively charged phospholipid dimyristoylphosphatidylglycerol(DMPG) upon addition to dimyristoylphosphatidylcholine (DMPC-yde) bilayers yielded a slope from eq 4 equal to -800 H z , ~ ~ indicating again the similarities between the behavior of the quadrupole splittings in the two systems. Yet another similarity is the somewhat greater sensitivity of the qua-
Surface Electrostatic Charge in Later Dispersions
drupolesplittings to positive as opposed to negative surface charges as expressed in the slopes mi. In bilayer membranes the slopes of such lines as represented by eq 4 are generally larger for positive charges than for negative charges, differing by a factor of 2 for deuterons at the a! and B position^.'^ The approximately 30% difference in slope measured here for positive versus negative surface charges is in agreement with the trends established from bilayer membranes and constitutes one more piece of evidence that the two systems are behaving very similarly. The “molecular voltmeter” must be responsive to both intrinsic and extrinsic modifiers of surface charge. The resulta listed in Table I indicate that extrinsic agents known to alter the surface charge of bilayer membranes also influence the response of the quadrupole splitting from surface-bound HDpc-y-d~.The 1:l electrolytes sodium chloride and potassiumchloride have little effect on surface charge because they bind with only low affinity to bilayer membra ne^'^.'^ and the same appears to be the case for HDPC monolayers. Sodium perchlorate is an interesting case because the perchlorate ion is representative of a class of aqueous anions known to bind with relatively high affinity to phosphatidylcholine bilayers and which thereby impose a negative surface ~ h a r g e . ’ ~The J ~ change in the quadrupole splitting observed for the HDPC monolayer in the presence of perchlorate indicates the accumulation of negative surface charges which are likely the result of perchlorate binding. Some of the largest effects on the quadrupole splittings were obtained in the presence of the 1:2 electrolyte calcium chloride. Calcium binds with good affinity to bilayer m e m b r a n e ~ ’ ~ and J ~ *in ~ ~so doing creates a positive surface charge density. The direction of the change in quadrupole splitting observed here in the presence of calcium is consistent with an accumulation of positive charges a t the surface of the HDPC monolayer. We note that the effects of these ions are the result of equilibrium binding and that the quadrupole splittings are sensitive to the amounts of bound ions. The level of binding is a function of both chemical affinity considerations and electrostatic considerations. For an initially neutral surface, as binding levels increase, a surface charge accumulates which tends to inhibit further binding because of its influence on the concentration of ions near the surface.21 This appears to be the case for calcium chloride where increasing the concentration beyond about 5.0 mM has little further effect upon the quadrupole splitting. Polyelectrolytes are employed as flocculants of colloid dispersions.22 The analogous biological structures pertinent to bilayer membranes are the extrinsic charged polypeptides which preferentially bind to membrane surfaces and modify the surface charge as monitored via the 2H NMR quadrupole ~p1ittings.l~ The cationic polyelectrolyte PEI and the anionic polyelectrolyte PSSS had little or no effect on the quadrupole splitting from HDPC-y-ds when the particle surface monolayer consisted purely of HDPC. It appears that the chemical affinity of such polyelectrolytes for a zwitterionic surfactant monolayer is not appreciable. In contrast, the presence of charged sur(14)Beschiaschvili, G.; Seelig, J. Biochim. Biophys. Acta 1991,1061, 78. (15) Macdonald, P.M.; Seelig, J. Biochemistry 1987, 26, 1231. (16) Lau, A.; McLaughlin, A.; McLaughlin, S.Biochim. Biophys. Acta 1981,645, 279. (17) Macdonald, P.M.; Seelig, J. Biochemistry 1988, 27, 6769. (18) Rydall, J. R.; Macdonald, P. M. Biochemistry, in press. (19) Altenbach, C.; Seelig, J. Biochemistry 1984,23, 3913. (20) Macdonald, P.M.; Seelig, J. Biochemistry 1987,26, 6292. (21) Aveyard, R.;Haydon, D. A. An Introduction to the Principles of Surface Chemistry; Cambridge University Press: London, 1973. (22) Gregory, J. In SolidlLiquid Dispersions; Tadros, T. F., Ed.; Academic Press: London, 1987; Chapter 8.
Langmuir, Vol. 8, No. 2, 1992 395
factants in the surface monolayer lead to large changes in the quadrupole splitting when the latex was exposed to a polyelectrolyte of opposite charge. This is a clear illustration of the principle that not only does ligand binding influence surface charge, but that surface charge must also influence ligand binding. Apparently in the caae of polyelectrolytes surface charge can represent the decisive factor. These results indicate that it should be possible using the “molecular voltmeter” technology demonstrated here to study in detail the interdependence of polyelectrolyte concentration, particle surface electrostatics, and flocculation. The results described in this report leave little doubt that 2HNMR may be used to monitor surface electrostatics in latex dispersions. They do not prove, however, that the “choline-tilt” model applies or even that the choline group is undergoing a concerted conformational change in response to surface charge effects. It is not yet certain even that the surfactant molecules undergo anisotropic long-axis rotations, a condition necessary for the interpretation of quadrupole splitting data in terms of a tilt of the choline group. In order to address these questions, the dependence of the quadrupole splittings on surface charge from HDPC labeled at other positions, a! and j3,in the choline group must also be investigated. Furthermore, experiments to probe the orientation dependence of the quadrupole splittings in surface monolayers bound to oriented polymer films are required before conclusions can be drawn concerning overall motional averaging in the surfactant monolayer. One limitation of the “molecular voltmeter” method in the investigation of latex stability has become evident here. Coagulation of latex is often affected by simply adding salt to the dispersion. The role of the salt is to decrease the Debye length of the electrostatic potential extending out from the particle surface, thereby permitting closer approach of the particles.23 Thus the salt need not affect the surface charge but can still affect latex stability. The “molecularvoltmeter” does not respond in such a situation. One advantage of the “molecular voltmeter” method is that in addition to its sensitivity to surface charge the 2H NMR spectrum also provides insights into the physical properties of the groups bound at the surface. For instance, the size of the quadrupole splitting measured here is only a fraction of the static splitting (125 kHz) expected for aliphatic deuterons? This indicates that the H D p c - 7 - d ~ deuterons experiencerapid but anisotropic diffusional reorientations. In the restricted motional environment of the HDPC monolayer, any residual motion must be anisotropic. It is likely that the motional averaging which has narrowed the 2H NMR spectrum is a combination of overalllong-axis rotations of the entire surfactant molecule and individualbond rotations internal to the choline group. Since distinct signals are observed for “free”and “surfacebound” HDPC-y-ds, the exchange of surfactant between these two populations must be slow on a time scale of the inverse of the quadrupole splitting (2 X 10-3 s). Furthermore, only a single quadrupole splitting is observed regardless of the mole fraction of ionic surfactant and regardless of the presence of multiple charged species at the surface. This indicates that the 2H NMR spectrum is reporting a statistically averaged surface charge. For ionic surfactants coadsorbed onto the particle surface and mixing ideally with the surface-bound HDPC monolayer, individual HDPC molecules will experience somewhat (23) Bijsterbosch, B. H. In SolidlLipuid Dispersions; Tadros, T. F., Ed.; Academic Press: London, 1987; Chapter 4.
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different charged environments due to the dispersion at a local level of the mole fraction of ionic surfactant relative to the global average. Since we observe only a single welldefined quadrupole splitting for HDPC-yd,, there must exist some mechanism acting to average the local charge environments. One such mechanism would be rapid translational diffusion of the surfactant molecules within the plane of the surface monolayer, analogous to the lateral diffusion of lipids in bilayer membranes.24 For aqueous ions and polyelectrolytes binding to the surface, the fact that we observe only a single quadrupole splitting in all cases indicates two particulars regarding the surface structure. First, the exchange of surfactant between the “ion-bound” and “ion-free” states must be fast on the 2H NMR time scale. In other words, the rate of associationdissociation of the ions with the surface is rapid. Second, there is no portion of the surfactant population sequestered from exposure to the aqueous ions or polyelectrolytes.Only amonolayer arrangement of the surfactants at the particle surface with their hydrophilic head groups oriented outward toward the aqueous bathing media is consistent with the observation of a single quadrupole splitting. Rupprecht and G u have ~ ~ recently reviewed the evidence supporting a bilayer arrangement for surfactants bound to hydrophilic (mainly silica) surfaces, where the surface/ adsorbant interactions are predominantly electrostatic in nature. Hydrophobic surfaces, such as that of polystyrene, bind surfactants predominantly via hydrophobic interactions, and one predicts a monolayer arrangement of surfactant. In a continuous bilayer architecture roughly half of the surfactant should remain oblivious to presence of charged ions or polyelectrolytes added into the external aqueous medium, leading to the appearance of two qua(24) Silver, B. L. The Physical Chemistry of Membranes; Solomon Press: New York, 1985; Chapter 10. (25) Rupprecht, H.; Gu, T. Colloid Polym. Sci. 1991, 269, 506-522.
drupole splittings for the two environments. Even a discontinuous bilayer, having pores which permit complete equilibration of small ions in the water layer separating the particle surface and the inner surface of the bilayer with the external solution, as envisaged by Rupprecht and G u , is~ inconsistent ~ with our observations. Specifically, it is unlikely that large polyelectrolytes such as PSSS (70 OOO molecular weight) or PEI (60 OOO) could equilibrate rapidly between the inner and outer environments unless the “pores” were so large that the surfactant aggregates consisted of Ypatches”. The latter arrangement of surfactants would permit near isotropic motional averaging of the surfactant molecules and no quadrupole splitting would be observed whatsoever.
Summary and Conclusions We have demonstrated that the 2H NMR quadrupole splitting from the phosphocholine-containing surfactant HDPC, deuterolabeled in the methyls of the choline quaternary nitrogen and bound to the surface of polystyrene particles, is sensitive to the particle surface charge. The source of the surface charge may be either intrinsic or extrinsic to the surface. The behavior of the quadrupole splittings suggests a specific response of the choline conformation under the influence of latex particle surface charge which is similar to that which has been observed in bilayer lipid membranes. This “molecular voltmeter” effect should enable us to quantitate latex surface electrostatics even in difficult circumstances such as flocculated colloidal dispersions.
Acknowledgment. This work was supported by granta from the National Science and Engineering Research Council (NSERC) of Canada and the Ontario Centre for Materials Research (OCMR).
