© Copyright 2001 American Chemical Society
FEBRUARY 20, 2001 VOLUME 17, NUMBER 4
Letters Plasmon Resonance Measurements of the Adsorption and Adsorption Kinetics of a Biopolymer onto Gold Nanocolloids Dirk Eck and Christiane A. Helm* Institut fu¨ r Physikalische Chemie, Johannes Gutenberg-Universita¨ t, D-55099 Mainz, Germany
Norman J. Wagner and K. Abraham Vaynberg† Center for Molecular and Engineering Thermodynamics, Department of Chemical Engineering, University of Delaware, Newark, Delaware 19716 Received August 7, 2000. In Final Form: November 29, 2000 A shift in plasmon surface resonance is measured for colloidal solutions of gold nanoparticles upon adsorption of polyampholyte (gelatin). It is demonstrated that the shift in wavelength of the absorption maximum can quantitatively yield measurements of the adsorbed amount as well as information about the structure of the adsorbed polymer layer. Semiquantitative agreement is found with calculations based on the Mie theory in the quasi-static limit. The method is demonstrated to provide quantitative measurements of the kinetics of biopolymer adsorption onto colloidal nanoparticles in solution.
In his 1857 Bakerian lecture, Michael Faraday described a comprehensive experimental investigation into the optical properties of colloidal gold sols and films obtained from such sols.1 Of particular interest was the color change observed upon drying “ruby jelly”, from ruby to amethyst. This change was noted to be reversible such that wetting the dried material returned it to a ruby color. Dissolution of the jelly by heating also showed that the gold colloids did not aggregate in the presence of the jelly, as evidenced by the solution color and sedimentation rate. On the basis of these observations (and many other observations of gold sols in the presence of other salts and “animal * Corresponding author. E-mail:
[email protected]. de. Current address: Ernst Moritz Arndt Universita¨t Greifswald, Institute of Physics: Angewandte Physik, Friedrich-Ludwig-JahnStr. 16, D-17487 Greifswald, Germany † Current address: Hercules Inc., Wilmington, DE. (1) Faraday, M. The bakerian lecture: Experimental relations of gold (and other metals) to light. Philos. Trans. R. Soc. London 1857, 147, 145-181.
substance”), Faraday deduced that the color change is “a striking case of the joint effect of the media and the gold in their action on the rays of light, and the most striking case amongst those where the medium may be changed to and fro.” This effect is quantitatively understood within the context of Mie scattering theory.2-4 Here, we exploit Faraday’s observations on the properties of ruby jelly, namely, the shift in optical absorbance of gold colloids with changes in the chemical composition of the surrounding medium, as a means to quantify the amount and kinetics of low levels of biopolymer adsorption. The adsorption of polymers onto a wide variety of colloids (2) Kreibig, U.; Vollmer, M. Optical Properties of Metal Clusters; Springer-Verlag: Heidelberg, Germany, 1995. (3) Lyon, L. A.; Pena, D. J.; Natan, M. J. Surface plasmon resonance of au colloid-modified au films: Particle size dependence. J. Phys. Chem. B 1999, 103, 5826-5831. (4) Schmitt, J.; Ma¨chtle, P.; Eck, D.; Mo¨hwald, H.; Helm, C. A. Preparation and optical properties of colloidal gold monolayers. Langmuir 1999, 15, 3256-3266.
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is of broad industrial relevance in such areas as pharmaceuticals, photography, paints and coatings, foods, wastewater treatment, rubbers, and composite materials and is ubiquitous in natural aqueous environments as well as in many biological organisms. Polymer adsorption and its consequences on colloidal stability and rheology is a well-investigated topic.5-7 Small angle neutron scattering with deuterium labeling and contrast matching is the most promising measurement method to date,8,9 with X-ray scattering10 and light scattering9 useful in some applications. All of these methods are severely limited, however, in their ability to study adsorption kinetics and become progessively more difficult for nanoparticles. As noted by Faraday, changes in the surrounding medium create measurable shifts in the plasmon resonance of gold nanoparticles, as measured by UV-visible spectrophotometry. This can be exploited to yield real-time measurements of polymer adsorption as well as information about the structure of the adsorbed biopolymer layer, as will be illustrated here. We note that extensive studies of UV-visible absorption have been conducted recently on gold particles and aggregates4 and have been employed to study alignment of rodlike gold nanoparticles. The method of surface plasmon resonance has been used extensively to investigate polymer and biopolymer adsorption onto gold films and gold nanoparticles anchored to gold films with applications to biosensors (see Lyon et al. and references therein for a review3). Our focus here, however, is the novel elucidation of the kinetics of adsorption of biopolymers to nanocolloidal gold free in solution. As a demonstration, we focus in this letter on aqueous suspensions of colloidal gold particles of 15.2 ( 1.9 nm diameter (as measured by transmission electron microscopy). Previous work has demonstrated the effects of surface modification on the optical properties of gold nanocolloids.2,3 As a candidate biomacromolecule, we investigate photographic grade gelatin (Kodak, Mn ) 100 000, Mw ) 160 000, IEP 5.1) derived from lime processed deionized bone gelatin. This polyampholyte is a purified polypeptide and has a known amino acid content. Here, all of the measurements are performed in a buffer of 0.01 M sodium acetate, which maintains the pH at 5.7. At this pH, there are both positively and negatively dissociated amino acids, such that the net charge on the polyampholyte is negative. In solution, the measured hydrodynamic radius is 20 nm by dynamic light scattering, with a radius of gyration of 16 nm as determined by small angle neutron scattering.9 Solutions were prepared by mixing solutions of stable colloidal gold, prepared according to standard precipitation methods,4,11 with dilute solutions of the gelatin, also prepared according to standard methods.9 All adsorption measurements were performed at constant solution (5) Russel, W. B.; Saville, D. A.; Schowalter, W. R. Colloidal Dispersions; Cambridge University Press: Cambridge, 1989. (6) Fleer, G. J.; Cohen, S. M. A.; Scheutjens, J. M. H. M.; Cosgrove, T.; Vincent, B. Polymers at Interfaces; Chapman and Hall: Cambridge, 1993. (7) Hunter, R. J. Foundations of Colloid Science I; Clarendon Press: Oxford, 1986. (8) Cosgrove, T.; Hone, J. H. E.; Howe, A. M.; Heenan, R. K. A small angle neutron scattering study of the structure of gelatin at the surface of polystyrene latex particles. Langmuir 1998, 14, 5376-5383. (9) Vaynberg, K. A.; Wagner, N. J.; Sharma, R.; Martic, P. Structure and extent of adsorbed gelatin on acrylic latex and polystyrene colloidal particles. JCIS 1998, 205, 131-140. (10) Seelenmeyer, S.; Ballauff, M. Investigation of the adsorption of surfactants on the poly(styrene) latex particles by small-angle X-ray scattering. Macromol. Symp. 1999, 145, 9-20. (11) Chow, M. K.; Zukoski, C. F. Gold sol formation mechanisms-role of colloidal stability. JCIS 1994, 165, 97-109.
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conditions (10 mM sodium acetate buffer, pH ) 5.7), as polymer adsorption can be kinetically limited.6 Precise measurements of adsorbed amounts were afforded by fluorescent labeling of the gelatin with fluorescein-5isothiocyanate (FITC, Molecular Probes), as described previously.9 Labeling was shown to have no effect on the adsorption.9 Absorption measurements in the UV-visible were made on a Perkin-Elmer Lambda 17 spectrophotometer at 22 °C. The interaction of the electromagnetic waves with metal nanocolloids is described with the Mie theory in the quasistatic limit, which is justified by the small size of the nanoparticles (radius R) respective to the optical wavelength λ (2R , λ).4,12 The polarizability R of isolated nanocolloids in a medium (dielectric constant m) is given by
(λ) - m R(λ) ) 4π0mR3 (λ) + 2m
(1)
with 0 representing the dielectric constant of vacuum and (λ) representing the complex dielectric constant of the gold, which depends strongly on the wavelength. If the surrounding medium is not homogeneous, it is described by an effective dielectric constant eff and a normalized polarizibility Λ(λ).
8 1 + NπR3 Λ(λ) 3 eff(λ) ) m 4 1 - NπR3 Λ(λ) 3
(2)
For the case of isolated particles, this reduces to Λ ) R/4π0mR3. However, if the colloids are encapsulated by a shell of thickness d and dielectric constant s, Λ becomes (for d , λ)13
Λ(λ) )
[R R+ d] ((λ) - )( + 2 ) R ( + 2 )((λ) + 2 ) + 2[ ((λ) - )( - ) R + d] (s - m)((λ) + 2s) +
3
s
m
s
3
s
m
s
s
s
m
(3) The experimentally accessible quantity is the absorption coefficient γ, as measured in UV-vis experiments; it is calculated from eff(λ) ) n˜ 2, with n˜ ) n + iκ as the complex index of refraction, using the relationship γ ) (4π/λ)κ. The adsorption isotherm at pH ) 5.7 and 22 °C is shown in Figure 1. Note first that the net negatively charged polyampholyte adsorbs onto negatively charged gold with a saturation coverage of ∼6 mg/m2. Typical values for gelatin adsorption range from ∼1-2 mg/m2 onto negatively charged 60 nm colloidal latex particles9,8 to ∼6 mg/m2 onto negatively charged mica cylinders used in surface force measurements.14 The adsorption isotherm can be fit by a simple Langmuirian form to yield (see inset) an estimate for the energy of adsorption of 5.5 kJ/mol, or about 1.9 kT/molecule (see ref 6 for the complexities of (12) Link, S.; El-Sayed, M. A. Spectral properties and relaxation dynamics of surface plasmon electronic oscillations in gold and silver nanodots and nanorods. J. Phys. Chem. B 1999, 103, 8410-8426. (13) Guettler, A. Die Miesche Theorie der Beugung durch elektrische Kugeln mit adsorbierendem Kern und ihre Bedeutung fu¨r Probleme der interstellaren Materie und des atmosphaerishen Aerosols. Ann. Phys. 1952, 11, 5. (14) Kamiyama, Y.; Israelachvili, J. Effect of pH and salt on the adsorption and interaction of an amphoteric polyelectrolyte. Macromolecules 1992, 25, 5081-5088.
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Figure 1. Polyampholyte adsorption isotherm for pH ) 5.7 at 22 °C as measured by solution depletion with fluorometry. The inset plot shows the fit to a Langmuir isotherm used to determine the adsorption energy.
this interpretation). This value is consistent with the mechanism of electrostatic polarization,9,15,16 whereby the net negatively charged polyampholyte adsorbs to the surface of the negatively charged gold colloid. This adsorption energy is weak enough that little macromolecular rearrangement is expected upon adsorption. The saturation coverage corresponds to approximately 12 adsorbed biopolymers per gold nanoparticle. Considering that the polyampholyte in solution has a hydrodynamic diameter nearly 3 times the diameter of the gold nanoparticle, 12 adsorbed gelatins could be arranged in an approximately face-centered lattice to surround the gold colloid. The UV-visible adsorption spectra for the gold sols in solution are shown in Figure 2. When the surface of the gold is saturated with gelatin (corresponding to a free gelatin concentration of 25 µg/mL in solution), a shift is observed, with the central absorption peak moving to higher wavelengths without broadening. The calculated spectra are shown for comparison in Figure 2b, where a uniform layer of gelatin is assumed. In the calculation, the refractive index of the solvent is taken to be that of water (1.33), whereas the value for the gelatin corona (1.3548) was determined by differential diffractometry on gelatin solutions corresponding to concentrations estimated for the corona. As the thickness of the corona was estimated to be 19 nm9,17 and the polymer concentration in the corona is assumed to be uniform, the comparison between calculation and experiment can be considered only qualitative. The shift in the peak wavelength with biopolymer adsorption can be used to quantitatively detect adsorption, as shown in Figure 3. The measured wavelength at the absorption maximum increases by approximately 1% immediately with adsorption of polymer corresponding to roughly one adsorbed macromolecule per nanocolloid and then saturates thereafter (the instrument resolution is (15) Dobrynin, A. V.; Rubinstein, M.; Joanny, J.-F. Adsorption of a polyampholyte chain on a charged surface. Macromolecules 1997, 30, 4332-4341. (16) Vaynberg, K. A.; Wagner, N. J.; Sharma, R. Polyampholyte (gelatin) adsorption to colloidal latex: pH and electrolyte effects on acrylic and polystyrene latex. Biomacromolecules, submitted. (17) Likos, C. N.; Vaynberg, K. A.; Lo¨wen, H.; Wagner, N. J. Colloidal stabilization by adsorbed gelatin. Langmuir 2000, 16, 4100-4108.
Figure 2. UV-visible absorption spectra (a) for gold colloids with (dashed line) and without (solid line) saturated adsorption of polymer and (b) corresponding to theoretical calculation. For comparison, the curves have been normalized at 400 nm.
Figure 3. Normalized percentage shift in the wavelength of maximum absorption as measured by UV-visible spectrophotometry as a function of adsorbed amount. The inset shows the model calculations corresponding to a fixed corona thickness (9) and a fixed corona refractive index (O).
0.5 nm and the standard error of the peak maximum in fits is 0.15 nm according to the Marquardt-Levenberg algorithm). This adsorption “fingerprint” can elucidate the structure of the adsorbed polymer layer. The inset of
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Figure 4. Measured adsorbed amount of polymer as a function of time from exposure as measured by UV-visible spectrophotometry. The inset shows the initial rate scales with xt, indicating perikinetic aggregation.
Figure 3 shows theoretical calculations for two simple scenarios: model a corresponds to a spherical corona of constant refractive index growing uniformly in thickness with additional adsorption (i.e., constant adsorbed polymer concentration) and model b corresponds to a constant adsorbed layer thickness with increasing refractive index with adsorption (i.e., a simplified calculation assuming the polymer adsorbs unevenly). More extensive investigations suggest that the adsorbed layer structure is more complex,8,9,18,19 but for the first-order analysis considered here we will assume a uniform adsorbed corona. Comparison with the data demonstrates that the adsorbed polymer layer behaves as if it adsorbed with constant density. Further, the measured relative shift in wavelength upon adsorption is twice that calculated using the best-guess parameters, indicating that the measurements are easily sensitive enough to detect coverages of only one polymer per nanocolloid. This demonstrated sensitivity to polymer adsorption and the rapidity of data collection enables real-time measurements of the adsorption kinetics, which is not possible by the scattering methods employed to date or by solution depletion and fluorometry. Figure 4 displays the shift in the absorption maximum with time following introduction of the polyampholyte into the gold suspension. The signal saturates after approximately 20 min. Converting the wavelength shift to adsorbed amount yields the inset plot, which is plotted according to the following relation for diffusion-dominated adsorption:
xDtπ
Γ(t) ) 2c0
(4)
where Γ is the adsorbed amount, c0 is the initial concentration of polymer, and D is the polymer diffusivity. As seen, the initial uptake scales as xt and analysis of the slope yields a value of D ) 1.25 × 10-12 m2/s. This is to be compared to the calculated diffusivity of the gold colloid (2.79 × 10-11 m2/s) and that of gelatin (1.05 × 10-11 m2/s). Analysis of perikinetic heteroaggregation20 would stipulate
that a number concentration weighted inverse of the diffusivities should be observed in the absence of any potential energy barriers and hydrodynamic interactions (the above derivation is for immobile substrates). That the observed diffusivity is an order of magnitude slower than either component suggests that there is an energy barrier to adsorption on the order of 2 kT, which is not unreasonable given that a negative polyampholyte is adsorbing to a negatively charged colloid. That this energy is on the order of the adsorption energy probably reflects the energy barrier required to rearrange, and thus polarize, the polyampholyte in response to a weak external electric field. The data suggest the following as a possible scenario for gelatin adsorption to colloidal gold for the conditions studied here (i.e., both polyampholyte and colloid are negatively charged). The first macromolecules to adsorb encapsulate the nanoparticle without large-scale deformation of the polyampholyte. We, like Faraday, observe that this gold/gelatin complex is stable against both perikinetic and orthokinetic aggregation, further supporting this physical arrangement. Additional macromolecules can adsorb to the gold colloid, resulting in a saturated coverage corresponding to a gold nanocolloid surrounded by a tightly packed cluster of macromolecules. The shifts in optical absorption to longer wavelengths upon biopolymer adsorption, however, are small for gelatin on gold. Calculations suggest that the observed shift in color from ruby to amethyst and back upon drying and wetting the ruby jelly are more a consequence of gel collapse bringing the gold colloids into closer proximity than the change in media refractive index. Shifts due to gold particle aggregation21,12 as well as merely bringing particles into close proximity4 are known to be much larger. Our demonstration of the sensitivity of plasmon resonance in the UV-visible wavelengths to biopolymer adsorption and the ability to collect real-time adsorption kinetics suggests that this technique is suited for applications requiring detection of biopolymer adsorption from solution onto suspended colloidal particles that is highly selective and in real time. The ability to control the size and polydispersity of gold nanocolloids,22 the ability to functionalize the surface of the gold colloid with biologically or chemically specific receptors, and their demonstrated uses in biosensors based on surface plasmon resonance12,3 suggest a wealth of technological and scientific applications. Acknowledgment. Discussions with U. Kreibig are gratefully acknowledged. The financial support of the German American Research Networking program (of the Alexander von Humboldt-Stiftung and the National Academy of Sciences) and the German Science Foundation (HE 1616/10-1) is appreciated. We acknowledge Eastman Kodak Corporation for supplying the gelatin used in this study and Dr. Ravi Sharma of Eastman Kodak Corporation for scientific assistance. LA001142+
(18) Howe, A. M.; Clark, A.; Whitesides, T. H. Viscosity of emulsions of polydisperse drops with a thick adsorbed layer. Langmuir 1997, 13, 2617-2626. (19) Vaynberg, K. A.; Wagner, N. J. Rheology of polyampholyte (gelatin) stabilized colloidal dispersions: The tertiary electroviscous effect. J. Rheol., submitted. (20) Hanus, L.; Wagner, N. J. Heteroflocculation of polydisperse colloidal dispersions. JCIS, in preparation.
(21) Westcott, S. L.; Oldenburg, S. J.; Lee, T. R.; Halas, N. J. Construction of simple gold nanoparticle aggregates with controlled plasmon-plasmon interactions. Chem. Phys. Lett. 1999, 300, 651-655. (22) Brown, K. R.; Walter, D. G.; Natan, M. J. Seeding of colloidal au nanoparticle solutions. 2: Improved control of particle size and shape. Chem. Mater. 2000, 12, 306-313.
