Fluorescent Au@Ag Core–Shell Nanoparticles with Controlled Shell

ARTICLE pubs.acs.org/Langmuir

Fluorescent Au@Ag Core
Shell Nanoparticles with Controlled Shell Thickness and HgII Sensing Samit Guha,† Subhasish Roy,† and Arindam Banerjee* Department of Biological Chemistry, Indian Association for the Cultivation of Science, Jadavpur, Kolkata 700 032, India

bS Supporting Information ABSTRACT: Au
Ag core
shell nanoparticles have been synthesized using synthetic fluorescent dipeptide β-Ala-Trp (β-Ala is β-alanine; Trp is L-tryptophan) in water at pH 6.94 and at room temperature. The synthesis of the Au
Ag core
shell nanomaterial does not involve any external reducing and stabilizing agents, and the constituents of dipeptide β-alanine and L-tryptophan are naturally occurring. Therefore, the synthesis procedure is ecofriendly. Moreover, the shell thickness has also been controlled, and the optical property of the core
shell nanomaterial varies with the shell thickness. The core
shell nanomaterial exhibits a fascinating fluorescence property. This fluorescent Au@Ag core
shell nanoparticle can detect toxic HgII ions ultrasensitively (with a lower limit of detection of 9 nM) even in presence of ZnII, CdII, and other bivalent metal ions (CaII, MgII, NiII, MnII, BaII, SrII, PbII, and FeII). Au
Ag core
shell nanomaterials can also be reused for sensing HgII ions.

’ INTRODUCTION Colloidal dispersions of noble metals Au and Ag have been extensively studied because of their unique property of a surface plasmon absorption band in the visible region.1 In recent years, considerable attention has been directed toward the synthesis of bimetallic core
shell nanoparticles owing to their fascinating optical, electronic, magnetic, and catalytic properties, which are different from the individual metallic counterpart.2
5 These interesting physicochemical properties appear because of the combination of two kinds of metals and their fine structures, evolving new surface characteristics. Thus, bimetallic core
shell nanoparticles, composed of two different metal elements, are of intense interest compared to monometallic nanoparticles. Extensive studies have focused on the control of the composition and morphology of bimetallic core
shell nanoparticles because their properties strongly depend on the composition, shape, and size of these core
shell nanoparticles.6,7 Many methods have been developed to prepare Au@Ag core
shell nanoparticles, including citrate reduction,8 borohydride reduction,9 a microwave polyol method,10 solvent extraction
reduction,11 sonochemical methods,12 photolytic reduction,13 radiolytic reduction,14 laser ablation,15 Neem leaf broth,16 tyrosine,17 a UV-photoactivation technique,18 and metal evaporation
condensation.19 Recently, DNA-embedded Au
Ag core
shell nanoparticles have also been reported in the literature.20 Mirkin and co-workers have recently demonstrated that L-ascorbic acid acts as a reducing agent for the synthesis of Au
Ag triangular bifrustum core
shell nanocrystals.21 Yamauchi and co-workers have recently reported an autoprogramed synthesis of unique Au@Pd@Pt triple-layer core
shell-structured nanoparticles consisting of a Au core, a Pd layer, and a nanoporous Pt outer r 2011 American Chemical Society

shell.22 Shi and co-workers reported the synthesis of core
shell dual mesoporous silica spheres that possess smaller pores in the shell and larger tunable pores in the core.23 Recently, Sun et al. synthesized Ag@SiO2 cubic core
shell nanoparticles.24 To the best of our knowledge, there is no report on the preparation of core
shell Au
Ag particles using a fluorescent dipeptide. We report here the synthesis and stabilization of Au
Ag core
shell bimetallic nanoparticles using dipeptide NH2-β-Ala-L-Trp-OH (β-Ala is β-alanine; Trp is L-tryptophan) in water at room temperature and at almost neutral pH (6.94) without using any external reducing and stabilizing agents. A twostep reduction process has been applied for the synthesis of core
shell nanoparticles in which chloroaurate ions were first reduced and then AgI ions were been reduced in the presence of fluorescent dipeptide β-Ala-Trp. This synthesis procedure can be considered to be an environment friendly method using a green chemical approach because the constituent amino acid residues of the dipeptides are naturally occurring; the procedure also involves a room-temperature water medium at almost neutral pH, and it does not require any toxic stabilizing or reducing agents. Although there are several reports on the preparation of Au
Ag core
shell nanoparticles, applications of these Au
Ag core
shell nanoparticles have not been well explored. Remarkably, these Au
Ag core
shell nanoparticles are fluorescent and have more than one excitation and emission wavelength. Therefore, these types of Au
Ag core
shell nanoparticles may be regarded as a new nanomaterial that can offer an interesting Received: February 12, 2011 Revised: September 12, 2011 Published: September 14, 2011 13198

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Langmuir Scheme 1. Schematic Representation of the Synthesis of Au
Ag Core
Shell Nanoparticles

function. The outer layer thickness of the Ag shell in the Au@Ag core
shell nanomaterial has been controlled using different concentrations of the silver nanoparticle precursor. A previous example of the variation of shell thickness in the Au@Ag core
shell nanomaterial includes the fine tuning of shell thickness, well-controlled sizes, and optical properties of Au@Ag core
shell nanocubes.25 Our study also demonstrates that this Au
Ag core
shell nanomaterial can be used in the selective and ultrasensitive sensing of HgII ions and can be reused to sense HgII ions a few times. This makes this nanomaterial an interesting candidate in nanotechnology.

’ EXPERIMENTAL SECTION Synthesis of Dipeptide. The dipeptide was synthesized by conventional solution-phase methods using a racemization free fragment condensation strategy.26 The Boc group was used for N-terminal protection, and the C-terminus was protected as a methyl ester. Couplings were mediated by dicyclohexylcarbodiimide/1-hydroxybenzotriazole (DCC/HOBt). Methyl ester deprotection was performed via a saponification method, and the Boc group was deprotected by using 98% formic acid. All intermediates were characterized by 300 MHz 1 H NMR and mass spectrometry. The final compound was fully characterized by 300 MHz 1H NMR spectroscopy, 13C NMR spectroscopy, DEPT 135, mass spectrometry, and FT-IR spectroscopy. The synthesis and characterization of the dipeptide are reported in detail in the Supporting Information. Synthesis of Au
Ag Core
Shell Nanoparticles Au@Ag (1:1), Au@Ag (1:2), and Au@Ag (1:3). An aqueous solution of the

dipeptide (2 mg mL
1) was vigorously stirred with an aqueous solution of tetrachloroauric acid HAuCl4 (2 mg mL
1) at pH 6.94 at room temperature. A pink color was developed within 2 min, indicating the formation of gold nanoparticles (GNPs). These materials were centrifuged, washed several times to remove excess unadsorbed dipeptide, and dried to obtain a powder that was readily redispersible in water. Then, AgNO3 (2 mg mL
1 for 1:1 Au@Ag, 4 mg mL
1 for 1:2 Au@Ag, and 6 mg mL
1 for 1:3 Au@Ag) solution was added to it, and it was stirred for 1 h. A color change was noticed from pink to wine red (Scheme 1). Then it was centrifuged, washed several times to remove the excess unadsorbed dipeptide, and dried to obtain a powder form of the peptide/Au
Ag core
shell nanomaterial that was easily dispersed in water for use.

’ RESULTS AND DISCUSSION Noble metal gold (Au) and silver (Ag) have the same facecentered cubic (fcc) crystal structure. Their lattice constants (Au (0.408 nm) and Ag (0.409 nm)) are very similar.3 Methods for the preparation of Au
Ag core
shell nanoparticles from the corresponding metal salts can be divided into two groups: coreduction and successive reduction of two metal salts.27

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Coreduction is the simpler preparative method in which the simultaneous reduction of two metal precursors is achieved. Successive reduction is usually carried out for the preparation of core
shell bimetallic nanoparticle. In this study, we have used the successive reduction approach. Dipeptide β-Ala-Trp has a redox-active chemical moiety, tryptophan,28 that is responsible for the reduction of metal ions (AuIII/AgI) to their respective metals (Au/Ag), possibly through electron transfer. First, Au nanoparticles were synthesized using a dipeptide (β-Ala-Trp) from HAuCl4 as the gold source without any external reducing and stabilizing agents at nearly neutral pH (6.94) and room temperature. The resultant dipeptide/Au nanoparticle composite has also been characterized by UV
visible spectroscopy and transmission electron microscopy. The UV
visible spectrum (Figure 1a) of the neat β-Ala-Trp dipeptide shows a sharp absorbance peak at 280 nm, which is a characteristic of a tryptophan moiety.29 The dipeptide Au nanoparticle nanoconjugate exhibited two peaks, one at 566 nm (corresponding to Au nanoparticles) and another at 280 nm (corresponding to tryptophan) (Figure 1a(B)). However, the dipeptide-stabilized 1:1 Au
Ag core
shell nanomaterial has a peak at 550 nm, the 1:2 Au
Ag core
shell nanomaterial has a peak at 533 nm, and the 1:3 Au
Ag core
shell nanomaterial has a peak at 500 nm, and each of them also has a peak at 280 nm corresponding to the tryptophan moiety (Figure 1b). The shifted surface plasmon absorbance indicates coupling between the Au and Ag layers, and it is made of a Au core surrounded by the Ag shell.30 Because the Ag composition in the Au@Ag core
shell nanoparticles has been increased, the surface plasmon band has shifted toward the blue region. This shifting has been observed only by changing the mole fraction of the Ag precursor. A mechanistic pathway for the formation of Au nanoparticles has been proposed. TEMPO, which is a radical quencher, has been used to investigate whether the reaction passes through a radical pathway. TEMPO inhibits metal nanoparticle formation in the presence of β-Ala-Trp, whereas in the absence of TEMPO, Au nanoparticle formation occurs (Figure 1a). This phenomenon has been thoroughly investigated with UV
visible and fluorescence spectroscopy. The UV
vis study clearly indicates that no Au nanoparticles are formed in the presence of TEMPO (Figure S7, Supporting Information). Therefore, it can be stated that the reaction passes through the radical pathway; the β-Ala-tryptophile radical probably plays an important role in Au nanoparticle formation. The powder form of the dipeptide
Au nanoparticle nanoconjugate has been redispersed in Milli-Q water, and the dipeptide (β-Ala-Trp) has then been added to it. Ag nanoparticle shells have been grown on the outer surfaces of Au nanoparticles by the selective reduction of AgI ions using dipeptide β-Ala-Trp that was bound to the surfaces of Au nanoparticles. Thus, the core
shell structure is formed. The structure and composition of the dipeptide/Au
Ag core
shell composite were characterized by UV
visible spectroscopy, fluorescence spectroscopy, high-resolution transmission electron microscopy (HRTEM), selected-area electron diffraction (SAED), energy-dispersive X-ray (EDX), and X-ray powder diffraction techniques. The Au
Ag core
shell structure is further stabilized through the NH2 functional groups of the dipeptide. This core
shell nanomaterial was centrifuged at 10 000 rpm for 15 min, and it was washed several times to remove excess unadsorbed peptide and dried to get a powdered material in each case. The powder can be stored for several months, and it can be redispersed in water. 13199

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Figure 1. UV
visible absorption spectra of (a) the dipeptide (A) and dipeptide/Au nanoparticle (B) materials and (b) the dipeptide/Au
Ag core
shell nanocomposite. Plots of (c) the molar concentration of Ag vs λmax and (d) the shell thickness vs the molar concentration of the silver in the core
shell nanomaterials.

Thermogravimetric Analysis (TGA) and Differential Thermal Analysis (DTA). The β-Ala-Trp dipeptide/Au
Ag core


shell nanomaterial (1:1) has been confirmed through a TGA
DTA (TGA, thermogravimetric analysis; DTA, differential thermal analysis) experiment (Figure S8b, Supporting Information). The decomposition of the peptide begins at 280 °C, and after that the melting of the Au
Ag core
shell starts at 350 °C. The TGA and DTA profiles of the 1:2 and 1:3 nanomaterials are more or less the same as that of the 1:1 Au@Ag profile. From the control experiment using only dipeptide β-Ala-Trp, it is evident that the decomposition of the dipeptide begins at 280 °C (Figure S8a, Supporting Information). The TGA
DTA experiment confirms the presence of dipeptide molecules in the Au
Ag core
shell nanomaterial. Fourier Transform Infrared Spectroscopy (FT-IR) Study. A solid-state FT-IR experiment has been carried out to characterize the dipeptide-stabilized Au
Ag core
shell nanoparticles. The FT-IR spectrum of the β-Ala-Trp dipeptide in the KBr palate shows the amide A, amide I, and amide II bands at 3392.55, 1647.10, and 1562.23 cm
1, respectively (Figure S9, Supporting Information). The spectrum of the dipeptide/Au
Ag core
shell nanomaterial shows three prominent bands at 3442.70, 1633.59, and 1550.66 cm
1 corresponding to the bands produced by amide A, amide I, and amide II of the dipeptide, respectively (Figure S9, Supporting Information). Thus, the

presence of these bands confirms the presence of the dipeptide in the Au
Ag core
shell structure. However, broadening and slight shifting of peaks of amide A, amide I, and amide II have been observed for the dipeptide in the dipeptide/Au
Ag core
shell nanomaterial (Figure S9, Supporting Information). This indicates that the functional groups of the dipeptide play a role in the preparation and stabilization of this Au
Ag core
shell nanostructured material. X-ray Powder Diffraction (XRPD) Study. The dried powder of the Au
Ag core
shell has been examined by X-ray powder diffraction (XRPD). The XRPD patterns of the Au
Ag core
shell show sharp peaks at 2θ = 38.16° (d = 2.35404), 44.43° (d = 2.03729), 64.63° (d = 1.44085), and 77.66° (d = 1.22847), and these peaks correspond to the lattice planes 111, 200, 220, and 311 respectively, consistent with the characteristic of Au/Ag nanoparticles (Figure S10, Supporting Information).6 The sharp XRD pattern thus clearly shows that the Au
Ag core
shell nanoparticle is crystalline in nature. The XRPD pattern of the dipeptide is shown in the Supporting Information (Figure S11). UV
Vis Study. Because of the attractive plasmon absorption feature, optical properties of bimetallic nanoparticles composed of gold and silver are the subject of considerable interest in the fields of nanoscience and nanotechnology. We have studied the UV
vis spectroscopy of the peptide/Au
Ag core
shell nanomaterial. Only one plasmon band has been observed at 550, 533, 13200

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shell nanomaterials, respectively (Figure 1b), which is different from that of individual Au or Ag nanoparticles stabilized by the dipeptide. This study suggests that the optical property is dependent on the shell thickness because the plasmonic band varies from 550 to 500 nm for 1:1 to 1:3 Au@Ag core
shell nanomaterials. It is interesting that the absorption occurs at a lower wavelength (blue shift) for the core
shell nanoparticle than for the corresponding dipetide/ Au nanoparticle composite (566 nm) (Figure 1a). A peak at 280 nm is been present, and this is a characteristic peak for a tryptophan moiety present in the dipeptide (Figure 1a). The individual UV
vis spectrum of the dipeptide shows the absorption at 280 nm (Figure 1a). The consistency in shape as well as the ability to daintily tune and control the thickness of Ag shells allowed us to investigate the influence of shell thickness on the surface plasmon absorption properties of the core
shell nanoparticles systematically. Figure 1b shows the UV
vis surface plasmon absorption band in an aqueous medium for different average shell thickness of 1.34 to 4.4 to 6.07 nm. At a small shell thickness, the Au
Ag core
shell shows a characteristic absorption band at 550 nm. As the shell thickness is increased from 1.34 to 6.6 nm, the characteristic plasmon absorption band is gradually blue shifted from 550 to 500 nm. Figure 1c,d shows a plot of absorption versus the molar concentration of silver and also the shell thickness versus the molar concentration of silver. Transmission Electron Microscopy (TEM) Study. The close match in lattice constants for Au and Ag enables the formation of a Au
Ag alloy and a Au@Ag core
shell. It is very difficult to distinguish between the Au
Ag alloy and the Au@Ag core
shell formation spectroscopically because both should result in a blue shift of the surface plasmon resonance. The core
shell structure of the nanomaterial can be examined in detail using transmission electron microscopy (TEM). For Au
Ag alloy formation, one should not observe the core
shell type of structure in TEM images, where a core is located in the central portion and the shell is on the outer position occupying the periphery of the core. The core
shell structure provides an outer bright and a central dark contrast in the TEM image. It is therefore possible to detect unambiguously the finally obtained Au
Ag core
shell nanostructures by analyzing their TEM images. A TEM study has been performed using an aqueous solution of the sample (2 mg in 2.0 mL) on a carbon-coated copper grid (300 mesh) by slow evaporation and vacuum drying at 30 °C for 2 days. TEM images reveal the formation of Au
Ag core
shell nanoparticles. A clear boundary between Au and Ag elements has been distinguished by the outer bright layer and the central dark contrast in the TEM image (Figure 2).18 From the TEM image, it is evident that a majority of the core
shell nanoparticles are almost spherical in morphology (Figure 2). The size and thickness of the core and shell of the Au
Ag core
shell nanoparticles are shown in the Supporting Information (Figure S12). A TEM EDX line-scanning study has also been performed to understand the core
shell structure in detail. The TEM EDX line-scanning profile (Figure S13b,c, Supporting Information) has clearly suggested the presence of the Au core and Ag shell structure within an Au
Ag core
shell structure. High-angle annular dark field scanning transmission electronic microscopic (HAADF-STEM) imaging (Figure S13a, Supporting Information) has also shown the bisegmental feature of the Au
Ag core
shell structure. Energy-dispersive X-ray (EDX) analysis shows that elements C, N, O, Au, and Ag are present. C, N, and O come from the dipeptide (Figure S14, Supporting

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Figure 2. Transmission electron microscopy imaging of the Au
Ag core
shell nanoparticles at different compositions showing different shell thicknesses. (A, B) Au@Ag (1:1) (C, D) Au@Ag (1:2), and (E, F) Au@Ag (1:3). The left panel shows many core
shell nanoparticles, and the right panel shows only one nanoparticle at high magnification vividly exhibiting the variation of shell thickness from B to D and F.

Information). The EDX analysis has directly confirmed the formation of Au
Ag bimetallic nanoparticles (Figure S14, Supporting Information). The selected-area electron diffraction (SAED) pattern indicates sharp spots including the 111, 200, 220, 311, 420, and 422 planes, which clearly show the formation of Au/Ag nanoparticles (Figure S15, Supporting Information). Sharp spots in the electron diffraction pattern clearly indicate the crystalline nature of the nanomaterial. The HR-TEM images also indicate lattice planes corresponding to Au and Ag nanoparticles (Figure S16, Supporting Information). It is evident from the TEM image analysis (Figure 2) that the average shell thickness of silver is increased from 1.34 (for 1:1 Au@Ag) nm to 6.6 nm (for 1:3 Au@Ag). Fluorescence Study. It has been well established that tryptophan can act as a stable or transient intermediate in electron/ hydrogen transport in biological systems through tryptophan radical intermediates. The electron-transfer process in tryptophan occurs through the formation of a tryptophyl radical, which is mainly observed in some biological systems such as cytochrome c peroxidase,31 Y122F mutant Escherichia coli RNR,32 and DNA photolyase.33 Furthermore, it has been reported that the oxidation of tryptophan results in the formation of a number of byproducts such as ditryptophan, kynurenine, 3-hydroxykynurenine, N-formyl-kynurenine, and some cross-linked products.34 Each of these byproducts has a specific absorption band and they show strong emission, and this provides the basis 13201

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Langmuir for identifying a particular species (for example, tryptophan: Eex = 280 nm, Eem = 366; ditryptophan: Eex = 270
280 nm, Eem = 320; kynurenine: Eex = 365 nm, Eem = 460; cross-linked products: Eex = 410 nm, Eem = 520).34 The formation of these byproducts occurs only through the formation of the tryptophyl radical. For a control experiment, we have recorded the emission spectrum of the neat dipeptide in aqueous solution. It shows the tryptophan emission maximum at 372 nm when it is excited at 280 nm and no other emission spectrum is observed. A fluorescence spectroscopic study has also been conducted with the dipetide/Au
Ag core
shell nanomaterial. The dipeptide/ Au
Ag core
shell nanomaterial shows only the emission maximum at 372 nm when it is excited at 280 nm (Figure S17, Supporting Information). The dipeptide/Au
Ag core
shell nanomaterial does not show any emission peak corresponding to tryptophan oxidation products including ditryptophan, β-Alakynurenine, and cross-linked products. This probably happens because of the fluorescence quenching of the above-mentioned byproducts by the β-Ala-Trp/Au
Ag core
shell nanomaterial. However, we have treated the peptide/Au
Ag core
shell composite with L-cysteine for 2 h, and it has been centrifuged at 10 000 rpm for 15 min. Then it has been filtered out. The fluorescence study has been performed with this filtrate. Cysteine binds strongly on the surface of the peptide/Au
Ag core
shell nanomaterial through strong Au
S bond35 formation, and the cysteine molecule exchanges with ligands β-Ala-Trp, the ditryptophan derivative, β-Ala-kynurenine, and the cross-linked product, which are comparatively weakly bound on the surface of the Au
Ag core
shell nanomaterials. To examine whether the presence of various byproducts are present during the formation and stabilization of the core
shell nanoparticle using the fluorescent dipeptide, excitation of the core
shell nanomaterial material has been performed at different wavelengths and different emission spectra have been observed, such as (a) Eex = 280 nm, Eem = 372; (b) Eex = 320 nm, Eem = 392; (c) Eex = 365 nm, Eem = 442; (d) Eex = 410 nm, Eem = 477. This result clearly indicates that some of the β-Ala-Trp molecules have retained their native structure. This is because of the fact that there is an intense emission peak at 372 nm when it is excited at 280 nm, whereas some part of the peptide dimerizes to form the corresponding ditryptophan moiety (Eem = 392, Eex = 320 nm) and β-Ala-kynurenine (Eem = 442, Eex = 365) is also formed. Besides these fluorescent compounds, some cross-linked products can be formed, which is apparent from the emission at 477 nm upon excitation at 410 nm (Figure 3). Oxidation products of this β-Ala-Trp molecule are highly fluorescent in nature. On the basis of the fluorescence study and UV
vis absorption study, using TEMPO, a probable mechanism for the formation of Au
Ag core
shell nanomaterial (using the fluorescent dipeptide β-Ala-Trp) has been suggested. According to this mechanism, the tryptophan residue of the β-Ala-Trp dipeptide donates an electron to the metal ion and is converted to a transient β-Ala-tryptophyl radical, which eventually transforms to native β-Ala-Trp, ditryptophan, β-Ala-kynurenine, and crosslinked products of the peptide. The presence of these oxidation products of β-Ala-Trp are also confirmed from the HR-MS data (Figure S6, Supporting Information). ICP (Inductively Coupled Plasma) Study. A solution of 20 mL of the Au
Ag core
shell nanomaterials of 1:1, 1:2, and 1:3 samples with known absorbance has been carefully dried by gentle heating in a fume hood, and then the residual mass has been divided into two parts. For Au, the sample has been digested

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Figure 3. Emission spectra of the peptide/Au
Ag core
shell nanomaterial at different excitation wavelengths after treatment with cysteine. Only the different emission spectra are marked.

by 10 mL of 10% HCl and 1% HNO3, and for Ag, the sample has been digested by 10 mL of 5% HNO3 in each set. These digested samples have been transferred to two different volumetric flasks to make an aqueous solution for the ICP measurement. The ratio of Ag and Au in the Au
Ag core
shell nanomaterials of one litter has been observed from the ICP (inductively coupled plasma) experiment, and this ratio has been found to be 1: 1.1, 1:2.04, and 1:2.6 for 1:1, 1:2, and 1:3 Ag/Au core
shell nanomaterials. Metal Ion Sensing Study. The sensing of HgII ions is an important issue in current research because HgII is toxic and the threshold limit of HgII toxicity in drinking water permitted by the U.S. Environmental Protection Agency (EPA) is 10 nM.36 It is important to note that after the addition of 250 μL of HgII solution of strength 0.82 μM to the core
shell nanomaterial, a color change has been observed from wine red to faint red. The metal ion sensing experiments have been performed by taking 1:1 Au@Ag core
shell nanoparticles. It is clear from the UV
vis experiment that a 10 nm red shift from 550 to 560 nm has been observed because of the addition of HgII ions (Figure S21, Supporting Information).37 The fluorescent intensity at an emission wavelength of 372 nm (excitation at 280 nm) of the peptide/Au
Ag core
shell nanomaterial is almost quenched in the presence of HgII, and a new peak appears at an emission wavelength of 438 nm (Figure 4). This indicates that the HgII ion forms a complex with β-Ala-Trp, probably with the COO
group of the dipeptide molecule (Figure S18, Supporting Information). Hupp and co-workers have reported the sensing of heavy metal ions using 11-mercaptoundecanoic acid-functionalized GNPs.38 In their study, they have shown that the carboxylic acid groups on the gold surface form complexes with metal ions (HgII). β-AlaTrp molecules are bound on the Au
Ag core
shell surface through the N-terminus NH2 groups (evidence in the FT-IR study section) (Figure S9, Supporting Information), and the C-terminus carboxylate groups are free. When the HgII ion is added, the carboxylate group of the β-Ala-Trp molecule forms a complex with the added HgII ion (Figure S18, Supporting Information). A DLS experiment after the addition of HgII ions has also been performed in order to examine the hydrodynamic diameter of the HgII core
shell nanomaterial complex. The hydrodynamic diameter of the dipeptide/Au
Ag core
shell nanomaterial in the presence of HgII ions has been increased, and the polydispersity of the solution has also been observed to 13202

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Figure 5. (A) ZnII is not sensed by the Au
Ag core
shell nanoparticles. (B) CdII is also not sensed by the Au
Ag core
shell nanoparticles, as is evident from their respective photoluminescence spectra showing no apparent change upon the addition of ZnII and CdII ions to the core
shell nanomaterial. Figure 4. Emission spectra of Au
Ag core
shell nanoparticles in the presence of varying concentrations of HgII ( 10
6 M): (a) 0, (b) 0.01, (c) 0.12, (d) 0.24, (e) 0.40, (f) 1.23, (g) 3.04, (h) 4.22, (i) 5.38, and (j) 7.07.

increase. The fluorescence quenching and red shift may be due to complex formation between carboxylate groups of the dipeptide β-Ala-Trp-capped Au@Ag nanoparticle with the HgII ion and also may be due to the aggregation of core
shell nanoparticles in the presence of HgII (Figure S19, Supporting Information).37 Apart from the DLS results (Figure S20, Supporting Information), the TEM images are also in favor of the aggregation of the core
shell nanomaterial. The change in color in the absorption spectrum of the peptide/Au
Ag core
shell nanomaterial suspension after the addition of HgII ions indicates that this nanomaterial can be used as the HgII sensor even up to parts per billion levels in water by the naked eye. Moreover, the change in intensity and peak position in the emission spectra of the core
shell nanomaterial upon the addition of HgII ions suggest that this core
shell nanomaterial can be applied as a HgII sensor using fluorescence spectroscopy. The limit of detection (LOD) for HgII has been found to be 9 nM (1.8 ppb), which is lower than the maximum level of 10 nM (2.0 ppb) mercury in drinking water permitted by the U.S. EPA. However, this peptide/Au
Ag core
shell nanomaterial does not sense the other divalent metal ions (ZnII, CdII, CaII, MgII, NiII, MnII, BaII, SrII, PbII, and FeII ions). Other ions have not shown any significant change as HgII has (10 nm red shift) in surface plasmon resonance. This indicates that the fluorescent core
shell nanoparticle is highly selective toward the HgII ion (Figure S22, Supporting Information). The red shifting may be due to the much stronger affinity of HgII ions toward the carboxylic acid (log β4 = 17.6) part of the dipeptide-capped core
shell nanoparticle in a water medium.38 In the presence of ZnII, CdII, and other divalent metal ions, the fluorescence intensity of the peptide/Au
Ag core
shell composite at an emission wavelength of 372 nm (excited at 280 nm) remains almost unaltered even at a higher concentration of these divalent cations (Figure 5). Furthermore, the core
shell nanomaterial can also sense HgII ions in a mixture of ZnII/ HgII and CdII/ HgII. A control experiment has been performed to compare HgII sensing by using the dipeptide-capped Au@Ag core
shell nanoparticles with only dipeptide, dipeptide-capped Au nanoparticles, and dipeptide-capped Ag nanoparticles. The dipeptide has not shown any apparent change (almost no change) in fluorescence spectra with the gradual addition of HgII ions, and the dipeptide-capped Au nanoparticle has not

shown many changes in its fluorescence spectra with the stepwise addition of HgII ions (Figure S23, Supporting Information). However, the dipeptide-stabilized Ag nanoparticle has shown significant changes (with a lower limit of detection (LOD) of 79 μM) with the gradual addition of HgII ions, and the change observed for the core
shell nanoparticle is much greater (with a LOD of 9 nM) than that of the Ag nanoparticles (Figure S23, Supporting Information). Therefore, it can be stated that the dipeptide-stabilized Au@Ag core
shell nanoparticle is a much better sensor for HgII ions in water than is the dipeptidestabilized Au or Ag nanoparticle. The dipeptide-stabilized Au@Ag core
shell nanoparticle is needed for the ultrasensitive sensing of toxic HgII ions in aqueous media. The HgII ion is very toxic to living organisms, and its toxicity depends on the amount present and the form in which it is found in the environment. Hg is not biodegradable; it accumulates in the environment and can produce toxic effects even at very low concentrations. In aqueous solution, bacteria can transform the HgII ion into methylmercury, which is a potent neurotoxin that can be easily accumulated in human body through the food chain. Methylmercury triggers several serious disorders, including sensory, motor, and neurological damage. Therefore, this peptide/Au
Ag core
shell nanomaterial can be used to determine the traces of HgII ions even in a mixture of divalent metal ions (e.g., ZnII/ HgII and CdII/ HgII (Figure S24, Supporting Information). The remarkable quenching (Figure 4) of the fluorescence property of the Au
Ag core
shell nanomaterial in the presence of HgII ions has been analyzed by the Stern
Volmer equation,39 and a linear plot is obtained up to a certain range (1.0  10
8 to 0.45  10
6 M). I ¼ 1 þ K SV ½Q  I0 Figure 6 shows the resulting plot of I0/I versus the HgII ion concentration, where I0 is the fluorescence intensity of the Au
Ag core
shell nanomaterial in the absence of the HgII ion, I is the fluorescence intensity of the Au
Ag core
shell nanomaterial in the presence of the HgII ion, KSV is the Stern
Volmer fluorescence quenching constant, which measures the efficiency of the fluorescence quencher, and [Q] indicates the fluorescence quencher concentration (i.e., the concentration of HgII ions). The Stern
Volmer quenching constant has been determined from the profile; it is found to be 1.2  106 M
1. To investigate whether this type of total fluorescence quenching by HgII is selective, the fluorescence property of this Au
Ag 13203

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Figure 6. Plot of I0/I vs the concentration of HgII indicating the quenching of the fluorescence intensity of Au
Ag core
shell nanoparticles upon the gradual addition of HgII ions. (The linear plot results from the Stern
Volmer analysis.)

Figure 7. Sensitivity of the peptide/Au
Ag core
shell nanomaterial toward divalent metal ions in a water medium.

core
shell nanomaterial in the presence of other bivalent metal ions has been studied. The relative fluorescence intensities of Au
Ag core
shell nanoparticles toward a set of bivalent metal ions are shown in Figure 7. Actually, the relative fluorescence intensities of the Au
Ag core
shell nanoparticles do not significantly change in the presence of other divalent metal ions including CaII, MgII, NiII, MnII, BaII, SrII, PbII, and FeII. In this study, the concentration that has been used for all types of metal ions is the same. Although there is a slight change in the fluorescence intensity of the Au
Ag core
shell nanomaterial toward CoII and CuII ions, the extent of quenching of the fluorescence intensity of the Au
Ag core
shell toward HgII is 25 times higher than that of CuII/CoII. These observations clearly indicate that the Au
Ag core
shell nanomaterial is very selective for HgII ions. It is interesting to examine whether the Au
Ag core
shell nanomaterial can be recycled for sensing HgII ions. Therefore, we have performed a fluorescence spectrophotometric study by taking the Au
Ag core
shell nanomaterial containing HgII ions plus 0.1 mL of 1 M EDTA solution. Because carboxylic acid groups can form strong complexes with HgII ions,38 we have used EDTA to study the recovery of fluorescence intensity of the core
shell nanomaterial. It is clear from Figures 8 and 9 that the

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Figure 8. Fluorescence recovery of Au
Ag core
shell nanomaterials after the chelation of HgII with EDTA.

Figure 9. Fluorescence spectroscopic study indicating the fluorescence recovery of the Au
Ag core
shell nanomaterial after the addition of EDTA to the core
shell nanomaterial with HgII. Complete fluorescence recovery takes 6 min. EDTA forms a strong complex with HgII ions by substituting the fluorescent-peptide-stabilized core
shell nanomaterial. Thus, the fluorescent core
shell material is now available to sense HgII ions further. The origin of new peaks is due to the oxidation products of dipeptide β-Ala-Trp.

fluorescence intensity of the Au
Ag core
shell nanomaterial in the presence of HgII has been completely quenched, and this intensity has been recovered to its original position upon the addition of EDTA (0.1 mL of 1 M) to HgII ions containing the Au
Ag core
shell nanomaterial. The intensity has been changed back to its original position because of the removal of HgII ions through chelation with EDTA. The resulting solution that is left behind behaves as an original Au
Ag core
shell nanomaterial. A UV
vis study has also been performed to investigate the recovery of the core
shell nanomaterial after removing HgII ions through complexation with the EDTA chelating agent (Figure S25, Supporting Information). The red-shifted surface plasmon resonance has come back to its original position (i.e., the actual position of the core
shell nanomaterial after removing the HgII ions completely). Thus, the fluorescent Au
Ag core
shell nanomaterial in water can be reused for sensing new HgII ions because there is no effect of EDTA on the Au
Ag core
shell nanomaterial. There are many reports on the preparation of Au
Ag core
shell nanoparticles,40 and these core
shell nanoparticles have 13204

dx.doi.org/10.1021/la203077z |Langmuir 2011, 27, 13198–13205

Langmuir been used as biosensors to detect small molecules such H2O2 or metal ions. However, our reported synthesis procedure is ecofriendly because the synthesis of the Au@Ag core
shell nanoparticles does not involve any external reducing and stabilizing agents and the dipeptide with naturally occurring amino acid residues has been utilized in water to make and stabilize the core
shell nanoparticles. This fluorescent Au@Ag core
shell nanoparticle can detect the toxic HgII ion ultrasensitively (with a lower limit of detection of 9 nM) even in the presence of ZnII, CdII, and other bivalent metal ions (CaII, MgII, NiII, MnII, BaII, SrII, PbII, and FeII). Moreover, this dipeptide-capped Au
Ag core
shell nanomaterial can be reused for sensing HgII ions. An environmentally friendly method of preparing Au@Ag core
shell nanoparticles, their use in the ultrasensitive and selective sensing of toxic HgII ions in water, and also their reusability are unique. They are different from the previously reported Au@Ag core
shell nanomaterials and their sensing properties.

’ CONCLUSION This study demonstrates the synthesis and stabilization of a fluorescent Au
Ag core
shell nanoparticle at room temperature and at neutral pH using water-soluble fluorescent dipeptide β-AlaTrp. Synthetically, this method is environmentally benign because the preparation of this core
shell nanoparticle does not require any external toxic stabilizing and reducing agents. Moreover, only by changing the molar ratio of the Ag precursor has the shell layer of Ag been controlled. This newly prepared core
shell nanomaterial finds a wonderful application in sensing selectively toxic HgII ions in nanomolar quantity in the presence of ZnII, CdII, and other divalent metal ions. The fluorescence recovery of the Au
Ag core
shell nanomaterial has been achieved through the chelation of HgII ions with EDTA. This indicates that the core
shell nanomaterial can be reused for sensing new HgII ions. The variation of the shell thickness of this core
shell nanoparticle can hold future promise in investigating whether functional properties of this core
shell nanomaterial can be dependent on the shell thickness. ’ ASSOCIATED CONTENT

bS

Supporting Information. Experimental procedure for the preparation and characterization of dipeptide. UV
vis, PL, HR-TEM, EDX, SAED, TGA
DTA, and DLS for the core
shell nanomaterials. This material is available free of charge via the Internet at http://pubs.acs.org.

’ AUTHOR INFORMATION Corresponding Author

*Fax: +91-33-2473 2805. E-mail: [email protected]. Author Contributions †

These two authors contributed equally.

’ ACKNOWLEDGMENT S.R. thanks CSIR, New Delhi, India, for financial assistance. We also acknowledge Mr. Supriya Chackroborty for STEMHAADF and EDS line scanning image recording. We are very thankful to anonymous reviewers for their helpful comments. ’ REFERENCES (1) Radziuk, D. V.; Zhang, W.; Shchukin, D.; Mohwald, H. Small 2010, 6, 545–553. (2) Liu, M.; Guyot-Sionnest, P. J. Phys. Chem. B 2004, 108, 5882–5888.

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