Silanization of Polyelectrolyte-Coated Particles: An Effective Route to

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Silanization of Polyelectrolyte-Coated Particles: An Effective Route to Stabilize Raman Tagging Molecules Adsorbed on Micrometer-Sized Silver Particles Kwan Kim,*,† Hyang Bong Lee,† and Kuan Soo Shin*,‡ Department of Chemistry, Seoul National UniVersity, Seoul 151-742, Korea, and Department of Chemistry, Soongsil UniVersity, Seoul 156-743, Korea ReceiVed January 24, 2008. ReVised Manuscript ReceiVed March 5, 2008 Micrometer-sized Ag (µAg) powders are very efficient surface-enhanced Raman scattering (SERS) substrates. To use µAg powders as a core material for molecular sensors operating via SERS, it is necessary to stabilize the tagging (i.e., SERS-marker) molecules adsorbed onto them. We demonstrate in this work that once the tagging molecules are coated with aliphatic polyelectrolytes such as poly(allylamine hydrochloride), the base-catalyzed silanization can be readily carried out to form stable silica shells around the polyelectrolyte layers by a biomimetic process; any particle can therefore be coated with silica since polyelectrolytes can be deposited beforehand via a layer-by-layer deposition method. Even after silanization, the SERS peaks of marker molecules on µAg particles are the only observable peaks since aliphatic polyelectrolytes, as well as silica shells, are intrinsically weak Raman scatterers, and more importantly, the SERS signals must be derived mostly from the first layer of the adsorbates (i.e., the marker molecules) in direct contact with the µAg particles. Silica shells, once fabricated, can further be derivatized to possess biofunctional groups; therefore, the modified µAg particles can be used as platforms of highly stable SERS-based biological sensors, as well as barcoding materials.

Introduction Surface-enhanced Raman scattering (SERS) is an abnormal surface optical phenomenon resulting in strongly increased Raman signals from molecules attached to nanostructured metal substrates.1,2 Raman signals arising from molecules adsorbed on Ag nanoparticles can be enhanced up to 1014 or 1015 times, allowing for the detection of single molecules.3–7 Since its first discovery in the 1970s,8 SERS has thus been the subject of great interest in many areas of science and technology including chemical analysis, corrosion, lubrication, heterogeneous catalysis, biological sensors, and molecular electronics.9–13 We recently demonstrated that commercially available 2 µmsized silver (µAg) powders are substrates that are effective for both infrared and Raman spectroscopic characterizations of * Corresponding authors. Tel.: +82-2-8806651 (K.K.) and +82-28200436 (K.S.S.). Fax: +82-2-8891568 (K.K.) and +82-2-8244383 (K.S.S.). E-mail: [email protected] (K.K.) and [email protected] (K.S.S.). † Seoul National University. ‡ Soongsil University. (1) Chang, R. K.; Furtak, T. E. Surface Enhanced Raman Scattering; Plenum Press: New York, 1982. (2) Moskovits, M. ReV. Mod. Phys. 1985, 57, 783. (3) Nie, S.; Emory, S. R. Science (Washington, DC, U.S.) 1997, 275, 1102. (4) Kneipp, K.; Wang, Y.; Kneipp, H.; Perelman, L. T.; Itzkan, I.; Dasari, R. R.; Feld, M. S. Phys. ReV. Lett. 1997, 78, 1667. (5) Kneipp, K.; Kneipp, H.; Itzkan, I.; Dasari, R. R.; Feld, M. S. Chem. ReV. 1999, 99, 2957. (6) Xu, H.; Bjerneld, E. J.; Käll, M.; Börjesson, L. Phys. ReV. Lett. 1999, 83, 4357. (7) Futamata, M.; Maruyama, Y.; Ishikawa, M. Vib. Spectrosc. 2002, 30, 17. (8) Fleischmann, M.; Hendra, P. J.; McQuillan, A. J. Chem. Phys. Lett. 1974, 26, 163. (9) Ni, J.; Lipert, R. J.; Dawson, G. B.; Porter, M. D. Anal. Chem. 1999, 71, 4903. (10) Kim, N. H.; Lee, S. J.; Kim, K. Chem. Commun. (Cambridge, U.K.) 2003, 724. (11) Cao, P.; Gu, R.; Tian, Z. Q. Langmuir 2002, 18, 7609. (12) Chu, W.; LeBlanc, R. J.; Williams, C. T.; Kubota, J.; Zaera, F. J. Phys. Chem. B 2003, 107, 14365. (13) Tian, Z. Q.; Ren, B.; Wu, D. T. J. Phys. Chem. B 2002, 106, 9463.

molecular adsorbates on silver surfaces.14–17 The Raman spectrum of organic monolayers on µAg powders is a SERS spectrum. Accordingly, µAg powders can be used as core materials for constructing molecular sensing/recognition units operating via SERS.15,18 One difficulty in the latter application is the necessity of the stabilization of SERS marker molecules adsorbed on µAg particles. It is well-known that silica shells are biocompatible and stable.19–21 The fabrication of silica shells would then be an effective strategy to protect SERS marker molecules. Silica shells, once fabricated, can further be derivatized to possess biofunctional groups.22,23 Silica-coating procedures reported in the literature generally involve surfaces with a significant chemical or electrostatic affinity for silica, such as clay minerals, hematite, zirconia, and titania.24–26 We demonstrated recently, by choosing 4-mercaptophenol (4-MPH) as a model SERS-marker molecule, that silica could be deposited onto 4-MPH on Ag by the basecatalyzed hydrolysis of tetraethyl orthosilicate (TEOS).27 The thickness of the silica shells could be tuned by adjusting (14) Kim, K.; Lee, H. S.; Yu, H. D.; Park, H. K.; Kim, N. H. Colloids Surf., A 2008, 316, 1. (15) Kim, K.; Park, H. K.; Kim, N. H. Langmuir 2006, 22, 3421. (16) Kim, K.; Lee, H. S.; Kim, N. H. Anal. Bioanal. Chem. 2007, 388, 81. (17) Han, H. S.; Han, S. W.; Joo, S. W.; Kim, K. Langmuir 1999, 15, 6868. (18) Kim, K.; Kim, N. H.; Park, H. K. Biosens. Bioelectron. 2007, 22, 1000. (19) Barbé, C.; Bartlett, J.; Kong, L.; Finnie, K.; Lin, H. Q.; Larkin, M.; Calleja, S.; Bush, A.; Calleja, G. AdV. Mater. 2004, 16, 1959. (20) Harrell, T. M.; Hosticka, B.; Power, M. E.; Cemke, L.; Hull, R.; Norris, P. M. J. Sol-Gel Sci. Technol. 2004, 31, 349. (21) Kros, A.; Gerritsen, M.; Sprakel, I. V. S.; Sommerdijk, N. A. J. M.; Jansen, J. A.; Nolte, R. J. M. Sens. Actuators, B 2001, 81, 68. (22) Green, M.; Harries, J.; Wakefield, G.; Taylor, R. J. Am. Chem. Soc. 2005, 127, 12812. (23) Philipse, A. P.; van Bruggen, M. P. B.; Pathmamanoharan, C. Langmuir 1994, 10, 92. (24) Kobayashi, Y.; Misawa, K.; Kobayashi, M.; Takeda, M.; Konno, M.; Satake, M.; Kawazoe, Y.; Ohuchi, N.; Kasuya, A. Colloids Surf. A 2004, 242, 47. (25) Ryan, J. N.; Elimelech, M.; Baeseman, J. L.; Magelky, R. D. EnViron. Sci. Technol. 2000, 34, 2000. (26) Ohmori, M.; Matijeviæ, E. J. Colloid Interface Sci. 1992, 150, 594. (27) Xia, L.; Kim, N. H.; Kim, K. J. Colloid Interface Sci. 2007, 306, 50.

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the silanization reaction time. However, due to the reaction of the hydroxyl groups of 4-MPH on Ag with TEOS to form O-Si bonds,28 the intrinsic SERS spectral feature of 4-MPH was severely distorted, obviously diminishing the effectiveness of the SERS-marker molecule. However, when thiol molecules without pendent hydroxyl groups, such as benzenethiol, were used as SERS-markers, TEOS did not form silica shells. Over two decades, a tremendous amount of research has been carried out on biomineralization not only for purely scientific interest but also with regards to prospective applications in the area of materials science for the facile production of biocompatible nano- and biomaterials in mild conditions.29 Biomineralization is the synthesis of minerals from simple compounds carried out by organisms.30 Biosilicification, as seen in diatoms, sponges, and grasses, 31 is one example of biomineralization, demonstrating a wide variety of natural purposes for this type of process. Various proteins have been isolated from such biological systems, and their ability to form silica in vitro from various silica precursors has been demonstrated.32 By sequencing the extracted proteins, it was postulated that amino acids such as lysine, histidine, arginine, proline, cysteine, and serine are important in biosilicification. Homopolymers of lysine (poly-L-lysine), arginine (poly-L-arginine), and proline (poly-L-proline) have been shown to produce silica from various silica precursors in vitro at, or close to, neutral pH values.33–39 The information gained from such bioinspired silica investigations has been used to develop silica films, patterned surfaces, and surface coatings via soft matter routes. In particular, Pogula et al.40 recently succeeded in forming continuous silica coatings on glass fibers using aminecontaining polymeric templates under mild conditions at room temperature. On the other hand, Sun et al.41 demonstrated that silica coating can be accomplished even onto negatively charged surfaces of polystyrene particles and latex nanocapsules by depositing layered polyelectrolytes on them beforehand. We found recently that aliphatic polyelectrolytes such as anionic poly(acrylic acid) (PAA) and cationic poly(allylamine hydrochloride) (PAH) can be deposited consecutively not only on polar surfaces but also on nonpolar surfaces.15,16 In a separate experiment, the SERS spectra of aromatic thiols adsorbed on µAg particles were found not to be affected by the deposition on them of those polyelectrolytes. Referring to biosilicification in living systems, we thus conducted silanization and found that silica shells can be readily fabricated even onto the PAA/PAH layers. The Raman spectral features of the thiol tags in direct contact with µAg particles were not distorted by the silanization nor by the polyelectrolyte layers. Since the tagging molecules do not take part in the silanization reaction, any aromatic thiols and their mixtures that possess relatively large Raman crosssections can be used as SERS markers. Multiple bioassays via (28) Torimoto, T.; Reyes, J. P.; Murakami, S.-Y.; Pal, B. J. Photochem. Photobiol. 2003, 160, 69. (29) Veronese, F. M.; Morpurgo, M. Farmaco 1999, 54, 497. (30) Lia, A.; Stephen, W. Angew. Chem., Int. Ed. Engl. 1992, 31, 153. (31) Simpson, T. L.; Volcani, B. E. Silicon and Siliceous Structures in Biological Systems; Springer-Verlag: New York, 1981. (32) Cheryl, W. P. F.; Jia, H.; David, L. K. Trends Biotechnol. 2004, 22, 577. (33) Patwardhan, S. V.; Mukherjee, N.; Clarson, S. J. J. Inorg. Organomet. Polym. 2001, 11, 193. (34) Siddharth, V. P.; Stephen, J. C. J. Inorg. Organomet. Polym. 2003, 13, 49. (35) Patwardhan, S. V.; Clarson, S. J. Silicon Chem. 2002, 1, 207. (36) Coradin, T.; Livage, J. Colloids Surf., B 2001, 21, 329. (37) Coradin, T.; Durupthy, O.; Livage, J. Langmuir 2002, 18, 2331. (38) Coradin, T.; Roux, C.; Livage, J. J. Mater. Chem. 2002, 12, 1242. (39) Sudheendra, L.; Raju, A. R. Mater. Res. Bull. 2002, 37, 151. (40) Pogula, S. D.; Patwardhan, S. V.; Perry, C. C.; Gillespie, J. W., Jr.; Yarlagadda, S.; Kiick, K. L. Langmuir 2007, 23, 6677. (41) Sun, B.; Mutch, S. A.; Lorenz, R. M.; Chiu, D. T. Langmuir 2005, 21, 10763.

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SERS can then be accomplished after attaching proper ligands (or proteins) onto the silica shells. They also may serve as useful barcode materials.

Experimental Procedures 4-MPH (97%), benzenethiol (BT, 99+%), 4-mercaptotoluene (4MT, 98%), 4-aminobenzenethiol (4-ABT, 90%), rhodamine B isothiocyanate (RhBITC, 97%), PAA (MW ∼450 kDa), PAH (MW ∼70 kDa), TEOS (98%), and silver powder (µAg, 99.9+%) with a particle size of 2.0–3.5 µm were purchased from Aldrich and used as received. An ammonia solution (28.0–30.0 wt %) was obtained from Sanchun Pure Chemical Company. 2-Propanol (99+%) was acquired from Daejung Chemicals and Metals Company. Other chemicals, unless specified, were of reagent grade and also used as received. Highly pure water, of a resistivity greater than 18.0 MΩ cm (Millipore Milli Q plus system), was used throughout the study. To adsorb 4-MPH (or BT, 4-MT, or 4-ABT) onto µAg powders, 0.050 g of silver powder was placed in a small vial into which 2 mL of 1 mM ethanolic solution of 4-MPH (or BT, 4-MT, or 4-ABT) was added. After 30 min, the solution phase was decanted, and the remaining solid particles were left to dry in ambient conditions. The layer-by-layer (LbL) deposition of polyelectrolytes was subsequently conducted by alternate immersion of the modified µAg powders into the PAA and PAH solutions (0.1 mg/mL) for 10 min at a time at room temperature; before changing the polyelectrolyte solution, the µAg powders were intensively rinsed with water. After the deposition of five bilayers of PAA/PAH, the modified µAg powders were vacuum-dried for 4 h and then subjected to silanization. Silanization was performed using a method similar to that described by Xia and co-workers.27 Initially, 0.05 g of modified µAg powder was weighed and then poured, under continuous magnetic stirring, into a mixture composed of 4 mL of water and 25 mL of 2-propanol. To this mixture were added ammonia (28.0–30.0 wt % solution) and TEOS, to a final concentration of 0.15 and 0.1 M, respectively, and then the whole mixture was left to react for 2 h at room temperature under continuous magnetic stirring. The reacted µAg particles were finally allowed to fall down to the bottom of the reaction vessel, and the solution phase was decanted. After a thorough washing with water and ethanol, the powdered products were vacuum-dried for 4 h. Infrared spectra were obtained using a Bruker IFS 113v FT-IR spectrometer equipped with a Globar light source and a liquid N2cooled wide-band mercury cadmium telluride detector. Raman spectra were obtained using a Renishaw Model 2000 Raman spectrometer equipped with an integral microscope (Olympus BH2-UMA). Radiation of 632.8 nm from a 17 mW helium-neon laser (SpectraPhysics Model 127) was used as the excitation source, and Raman scattering was detected over 180° using a Peltier cooled (-70 °C) charge-coupled device (CCD) camera (400 pixels × 600 pixels). Field emission scanning electron microscopy (FE-SEM) images were obtained with a JSM-6700F FE-SEM instrument operated at 5.0 kV. X-ray photoelectron spectra were obtained on an AXIS-Krato instrument using Mg KR radiation.

Results and Discussion Figure 1 shows a schematic diagram of the stabilization of SERS-marker molecules on µAg particles via silanization following the deposition of aliphatic polyelectrolytes. Initially, aromatic thiol molecules are adsorbed on Ag by forming an Ag-S bond after deprotonation of the thiol group. Polyelectrolytes of PAA and PAH are then deposited onto the modified µAg particles. Subsequently, silica sols generated by the hydrolysis of TEOS are condensed onto the PAA/PAH layers. The latter layers serve as a template for silanization, and ammonia acts as a catalyst to speed up the hydrolysis of TEOS to form crosslinked Si-O-Si bonds. These scenarios were verified using infrared and Raman spectroscopy, XPS, and FE-SEM.

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Figure 1. Schematic diagram of stabilization of SERS-marker molecules on µAg particles via silanization following deposition of polyelectrolytes.

Figure 2. Infrared spectra of (a) pure 4-MPH in the powdered state and (b) 4-MPH adsorbed onto µAg powders. Infrared spectrum (c) taken after five bilayers of PAA/PAH were deposited onto the 4-MPH adsorbed µAg powders and (d) that taken after silanization onto those PAA/PAH bilayers.

First, we examined the infrared spectral features. Figure 2a,b shows, respectively, the infrared spectrum of pure 4-MPH in the powdered state and that of 4-MPH adsorbed onto µAg powders. It is clear immediately that the S-H stretching peak appearing at 2558 cm-1 in Figure 2a is completely absent in Figure 2b, as expected due to the formation of Ag-S bonds. A very broad band appears in the region of 3100 to ∼3600 cm-1 for pure 4-MPH in Figure 2a, probably due to the formation of intermolecular H-bonding. Otherwise, the infrared spectral features in Figure 2a,b correlate closely with each other. The strong peak at 1246 cm-1 in Figure 2b can be attributed to the C-O stretching vibration of 4-MPH, while the bands at 1367 and 1095 cm-1 are due, respectively, to in-plane O-H bending and a ring vibration possessing C-S stretching characteris-

tics.42–44 Other bands can be assigned to proper ring modes. Figure 2c shows the infrared spectrum taken after five bilayers of PAA/PAH were deposited onto the 4-MPH adsorbed µAg powders. Since aliphatic polyelectrolytes are stronger infrared absorbers, there are some quite intense new peaks, for instance, at 1713, 1567, and 1397 cm-1, which can be attributed to the CdO stretching band of the carboxylic group (-COOH), and the antisymmetric and symmetric stretching bands of the carboxylate group (-COO-) of PAA, respectively.45,46 Figure 2d shows the infrared spectrum taken after silanization onto the PAA/PAH layer. Strong new bands appear in the region of 1200-900 cm-1. The three peaks at around 1230, 1120, and 790 cm-1 can be attributed to the vibrational modes involving the bridging oxygen atoms in Si-O-Si moieties, while the peak at 940 cm-1 is due to the Si-O stretching vibration of the Si-OH bonds.47,48 These infrared spectral features clearly indicate that silica shells are formed successfully over the PAA/PAH layers on µAg powders. As mentioned briefly in the Introduction, Sun et al.41 recently demonstrated the ability to grow silica directly on a deposited surface of polyelectrolyte. Pogula et al.40 also demonstrated recently that silica coatings could be made not only on PAHcoated fibers but also on glass fibers onto which poly-L-lysine (PLL) and poly(L-lysine-tyrosine) (PLT) had previously been deposited. The claims in earlier publications are that electrostatic interactions between organic polycations and anionic silica species are one of the main factors controlling silicification. It was suggested that the formation of silica shells onto the PAA/PAHcapped µAg powders proceeds similarly. The formation of silica shells must proceed by adsorption of nuclei and small particles (42) Socrates, G. Infrared Characteristic Group Frequencies: Tables and Charts; John Wiley and Sons: Chichester, U.K., 1994. (43) Smith, B. C. Infrared Spectral Interpretation: A Systematic Approach; CRC Press: Boca Raton, FL, 1998. (44) Lee, H. M.; Kim, M. S.; Kim, K. Vib. Spectrosc. 1994, 6, 205. (45) Choi, J.; Rubner, M. F. Macromolecules 2005, 38, 116. (46) Wang, Y.; Yu, A.; Caruso, F. Angew. Chem., Int. Ed. 2005, 44, 2888. (47) Tomozawa, M.; Hong, J.-W.; Ryu, S.-R. J. Non-Cryst. Solids 2005, 351, 1054. (48) Balamurugan, A.; Sockalingum, G.; Michel, J.; Fauré, J.; Banchet, V.; Wortham, L.; Bouthors, S.; Laurent-Maquin, D.; Balossier, G. Mater. Lett. 2006, 60, 3752.

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Figure 3. (a) Infrared spectrum of BT adsorbed on µAg powder. Infrared spectra (b) taken after five bilayers of PAA/PAH were deposited onto the BT adsorbed µAg powders and (c) taken after silanization onto those PAA/PAH bilayers.

from solution onto the µAg particles due to the electrostatic attraction between the positively charged amine groups of PAH and the anionic silica species, followed by particle growth. As another example, Figure 3a shows the infrared spectrum of BT adsorbed on µAg powders. The peaks at 1473 cm-1 (ν(CC), a1), 1437 cm-1 (ν(CC), b2), 1181 cm-1 (β(CH), a1), 1081 cm-1 (β(CCC) + ν(CS), a1), 1022 cm-1 (β(CH), a1), 1000 cm -1 (β(CCC), a1), 735 cm-1 (γ(CH), b1), and 689 cm-1 (γ(CCC), b1) are clearly due to BT on µAg particles.49 Figure 3b shows the infrared spectrum taken after five bilayers of PAA/PAH were deposited onto the BT molecules on µAg powders. The strong peaks at 1713, 1567, and 1397 cm-1 can be attributed to the CdO stretching band of the carboxylic group (-COOH) and the antisymmetric and symmetric stretching bands of the carboxylate group (-COO-) of PAA, respectively,45,46 implying that the PAA/ PAH bilayers were successfully deposited onto the BT molecules. Figure 3c shows the infrared spectrum taken after silanization onto the PAA/PAH layer. The infrared spectral features in Figure 3b,c are comparable to those in Figure 2c,d, respectively. Much the same infrared spectral patterns were obtained from silanized Ag powders that had initially been derivatized with 4-mercaptotoluene and 4-aminobenzenethiol. On the basis of the infrared spectral data, we subsequently examined the Raman spectral features. Figure 4a,b shows the Raman spectra of pure 4-MPH in the powdered state and 4-MPH adsorbed onto µAg powders, respectively, taken using 632.8 nm radiation as the excitation source. In agreement with the infrared spectral data, the S-H stretching band is seen at 2558 cm-1 in the normal Raman spectrum in Figure 4a but is completely missing in the SERS spectrum in Figure 4b; we nonetheless omitted the assignment of the Ag-S stretching band around 230 cm-1 due to its extreme weakness.44,50 Figure 4c shows the SERS spectrum taken after five bilayers of PAA/PAH were deposited onto the 4-MPH adsorbed µAg powders. The spectral pattern in Figure 4c is hardly different from that in Figure 4b. In addition, the SERS signal of 4-MPH is maintained even after the deposition of five bilayers of PAA/ PAH. This is not surprising because aliphatic polyelectrolytes are weak Raman scatterers,51 and the SERS signal must be derived mostly from the adsorbate that is in direct contact with the SERS substrates in accordance with electromagnetic and chemical (49) Dai, Z.; Voigt, A.; Leporatti, S.; Donath, E.; Dähne, L.; Möhwald, H. AdV. Mater. 2001, 13, 1339. (50) Joo, T. H.; Kim, M. S.; Kim, K. J. Raman Spectrosc. 1987, 18, 57. (51) Mulvaney, S. P.; Musick, M. D.; Keating, C. D.; Natan, M. J. Langmuir 2003, 19, 4784.

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Figure 4. Raman spectra of (a) pure 4-MPH in the powdered state and (b) 4-MPH adsorbed onto µAg powders. Raman spectra (c) taken after five bilayers of PAA/PAH were deposited onto the 4-MPH adsorbed µAg powders and (d) taken after silanization onto those PAA/PAH bilayers. (e) Raman spectrum taken after silanization directly onto 4-MPH on µAg powders.

enhancement mechanisms.1,2 The present observation also suggests that any electrostatic as well as hydrophobic and van der Waals interactions acting between 4-MPH and PAA/PAH are not strong enough to deteriorate the SERS features of 4-MPH. Figure 4d shows the SERS spectrum taken after silanization; the sample used to take the SERS spectrum was the same one used to take the infrared spectrum in Figure 2d. The peak intensities as well as the peak positions in Figure 4d are barely different from that in Figure 4b or c. The present observation is in obvious contrast to the earlier report27 that the SERS spectral feature of 4-MPH is affected substantially when silica shells are formed directly onto 4-MPH. For reference, the SERS spectrum reproduced in Figure 4e was acquired after a direct silanization onto the self-assembled monolayers (SAMs) of 4-MPH on µAg powders. Although all the peaks in Figure 4e can be attributed to 4-MPH on µAg particles, the ring breathing band of 4-MPH at 1080 cm-1 in Figure 4e is far weaker than those in Figures 4b-d, while the ring 8a band around ∼1600 cm-1 is much broader than those in Figures 4b-d; this must be due to the reaction of the hydroxyl group of 4-MPH with TEOS. When 4-MPH is coated with polyelectrolytes, the SERS peaks of 4-MPH on µAg powders are hardly affected by the silanization. As mentioned previously, this reflects the fact that the interaction of 4-MPH with PAA/PAH is not strong enough to change the SERS of 4-MPH on µAg particles. The absence of peaks related to Si-O groups in the SERS spectrum in Figure 4d is not unreasonable since silica itself is a weak Raman scatterer,51 and, as mentioned previously, the SERS signals must be derived from the first layer of the adsorbates (i.e., 4-MPH).1,2 As another example, Figure 5a shows the SERS spectrum of BT adsorbed on µAg powders taken using 632.8 nm radiation as the excitation source. The five strong peaks at 1573, 1072, 1022, 999, and 691 cm-1 are due to the ring 8a, 1, 18a, 12, and 6a modes of BT, respectively.50,52 Figure 5b shows the SERS spectrum measured after the deposition of five PAA/PAH bilayers onto the BT adsorbed µAg powders. We notice that the spectral pattern in Figure 5b is the same as that in Figure 5a. Not only the peak positions but also the peak intensities of BT hardly differ from each other. Figure 5c shows the SERS spectrum taken after silanization onto the PAA/PAH layer; the sample used to take the SERS spectrum is the same one used to take the infrared spectrum in Figure 3c. It is seen that the spectral feature (52) Han, S. W.; Lee, S. J.; Kim, K. Langmuir 2001, 17, 6981.

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Figure 5. (a) Raman spectrum of BT adsorbed on µAg powder. Raman spectra (b) taken after five bilayers of PAA/PAH were deposited onto the BT adsorbed µAg powders and (c) taken after silanization onto those PAA/PAH bilayers.

in Figure 5c is the same as that in Figure 5a,b. Much the same observation is made using 4-MT and 4-ABT derivatized Ag powders. In addition, the SERS spectral features were hardly affected even after the silanized µAg powders were left for an extended time period in air or soaked in water, ethanol, and PBS solution. This proves that the PAA/PAH five layers are, in fact, very effective for the silanization of SERS marker molecules assembled on µAg powders. The formation of silanized µAg particles also can be confirmed by taking XPS spectra after silanization. The characteristic XPS peak of silica particles is known to appear at ∼103 eV.53 The same peak is observed for all samples in this work (data not shown). This is coincident with the infrared spectral data, indicating that silica shells are readily formed over the PAA/PAH layer assembled on µAg powders. Nonetheless, it is not yet clear as to whether µAg powders are enclosed by silica shells. Discerning the formation of silica shells by FE-SEM is difficult in the intact state, owing to the intrinsically irregular shape of µAg powders. To identify the silica shells by FE-SEM, we thus removed silver from the silanized µAg powders. Specifically, 50 mg of silanized µAg powder (the same one used in taking IR and Raman spectra) was poured into 10 mL of 1 M aqueous HNO3 solution and then ultrasonicated for 1 h. When the resulting mixture was left for an additional 12 h without sonication, silver particles were no longer identifiable. Upon centrifugation of the solution at 3000 rpm, white silica powder could be collected at the bottom. Figure 6 shows the FE-SEM image of the recovered silica. It is evident that silica shells are, in fact, formed onto µAg powders. The morphology of the silanized Ag powders is seen to be rougher than that before silanization, probably due to the deposition of polyelectrolytes, but the thickness of silica shells appears to be quite homogeneous; the thickness is estimated to be at least ∼80 nm when silanization is conducted at 0.1 M TEOS in the presence of 0.15 M ammonia for 2 h. Consulting the infrared, XPS, and FE-SEM data, enough silica shells must have been formed over the µAg powders.54,55 Nonetheless, it is not evident as to whether the µAg powders were covered fully with silica shells without any defect. We thus conducted a place exchange reaction to occur on µAg powders with rhodamine B isothiocyanate (RhBITC). RhBITC is a very (53) Chastain, J., Jr. Handbook of X-Ray Photoelectron Spectroscopy; Physical Electronics, Inc.: Eden Prairie, MN, 1995. (54) Veiseh, M.; Zhang, M. J. Am. Chem. Soc. 2006, 128, 1197. (55) Guczi, L.; Frey, K.; Beck, A.; Petõ, G.; Daróczi, C. S.; Kruse, N.; Chenakin, S. Appl. Catal., A 2005, 291, 116.

Figure 6. FE-SEM image of silica shells taken after dissolving out core silver from silanized µAg powders.

Figure 7. Raman spectra taken after samples silanized initially onto five bilayers of PAA/PAH deposited on µAg powders containing the SAMs of (a) 4-MPH, (b) BT, (c) 4-MT, and (d) 4-ABT were soaked in 0.1 mM ethanolic RhBITC solution for 24 h. (e) Raman spectrum of RhBITC adsorbed on µAg powder.

strong Raman scatterer and is also known to adsorb to such an extent on Ag that 4-MPH on µAg powders, for instance, can be exchanged with RhBITC.27 The RhBITC peaks were identified when PAA/PAH stabilized 4-MPH/µAg powders were soaked in the dye solution for 24 h. However, we did not observe any RhBITC peak when silanized µAg powder was subjected to the place exchange reaction with RhBITC. Figure 7 shows the Raman spectra taken after the silanized µAg powders were soaked in RhBITC solution for 24 h. The observed spectra all were due to SERS-marker molecules used initially to modify µAg powders. These Raman spectral data clearly illustrate that all µAg powders were fully coated with silica shells without any defect.

Conclusion We demonstrated using infrared spectroscopy, Raman spectroscopy, XPS, and FE-SEM that SERS-marker molecules adsorbed on µAg powders can be readily coated with silica shells following the LbL deposition of PAA and PAH polyelectrolytes. We were able to carry out the LbL deposition of PAA and PAH and the subsequent silanization under mild conditions at room temperature as in living systems, irrespective of the kind of SERS-marker molecules adsorbed initially on the µAg powders. We suggest that the formation of silica

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shells proceeds by the adsorption of nuclei and small silica particles from solution onto the µAg particles due to the electrostatic attraction between the positively charged amine groups of PAH and the anionic silica species, followed by particle growth. The cross-linked silica shells prevented the SERS-marker molecules from being liberated from the surface of the µAg powder. Since silica shells can be further derivatized with diverse biological molecules, µAg particles capped with

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different SERS-marker molecules can be used in multiple molecular recognition, as well as in barcoding. Acknowledgment. This work was supported by the Korea Science and Engineering Foundation (Grants R01-2006-00010017-0 and R11-2007-012-02002-0). K.S.S. also was supported by the Soongsil University Research Fund. LA800251T