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sensing of l -methionine
Materials Science and Engineering C 70 (2017) 656–664
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Synthesis, characterization of Ag-Au core-shell bimetal nanoparticles and its application for electrocatalytic oxidation/sensing of L-methionine M. Murugavelu, B. Karthikeyan ⁎ Department of Chemistry, Annamalai University, Annamalainagar 608 002, Tamil Nadu, India
a r t i c l e
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Article history: Received 8 May 2016 Received in revised form 30 August 2016 Accepted 21 September 2016 Available online 23 September 2016 Keywords: Ag-Au core-shell Cyclic voltammetry Electrocatalytic L-methionine
a b s t r a c t The Ag-Au core-shell bimetal nanoparticles (BNPs) was prepared using chemical reduction method. The prepared Ag-Au core-shell BNPs were characterized by UV–Visible (UV–Vis) spectroscopy, field emission scanning electron microscopy (FE-SEM), transmission electron microscopy (TEM), high resolution transmission electron microscopy (HR-TEM), X-ray diffraction (XRD), atomic force microscopy (AFM) and X-ray photoelectron spectroscopy (XPS) pattern. These results showed the Ag-Au BNPs exhibited core-shell shape. The Ag-Au core-shell BNPs was examined towards electrocatalytic oxidation of L-methionine (L-Met) by cyclic voltammetry (CV), linear sweep voltammetry (LSV) and chronoamperometry. According to the results, L-Met is determined with detection limit of 30 μM. Interference studies in biological buffer was also studied. © 2016 Elsevier B.V. All rights reserved.
1. Introduction The preparation of bimetallic nanoparticles (BNPs) have great significance because of the modification of physical and chemical properties, compared with monometallic nanoparticles (NPs) due to size effect. Enhanced electrocatalytic activity, catalysis, result from the combination of different metals are also taken place [1–5]. These BNPs can retain the functional properties of each component and possibly offer synergistic effects via cooperative interactions, resulting in important features such as increased surface area, enhanced electrocatalytic activity, improved biocompatibility, promoted electron transfer, and better invulnerability against intermediate species. During the past years, such bimetallic nanostructured materials were extensively developed and reported by many scientists [6,7]. Metal NPs offer excellent applications for chemical and biological sensing because of their unique optical and electrical properties [8]. Especially Au nanoparticle-based electrochemical stripping detection of DNA hybridization reported by Authier et al. [9,10] and signal amplification with Ag enhancement [11] for gene analysis may be noted. In the case of BNPs, when a new nanoparticle composition is designed, new modification methods must be developed for immobilizing biomolecules on the surface of the particles of interest. The methods for modifying Au NPs have now been optimized and generalized for a wide range of particle sizes and surface compositions, including spheres and rods [10,11]. From the wide literature review, we have picked to prepare Ag-Au BNPs and its synergetic effect in a modified electrodes for determination of L-methionine. ⁎ Corresponding author at: Department of Chemistry, Annamalai University, Annamalainagar 608 002, Tamil Nadu, India. E-mail address: [email protected] (B. Karthikeyan).
http://dx.doi.org/10.1016/j.msec.2016.09.046 0928-4931/© 2016 Elsevier B.V. All rights reserved.
Methionine is one of the essential amino acid among the 20 amino acids, it occurs naturally as L-methionine (L-Met), and is a major source of sulfur, which is required in the human diet for normal metabolism and growth, since it is not synthesized in humans. Methionine is often used as an additive in animal feedstuffs, and it is also used in production of medicines and active pharmaceutical ingredients, and as a precursor to other amino acids [12]. It governs the main supply of sulfur in the diet, and also prevents disorders in hair and skin. It acts as an essential amino acid with the key role in biological methylation. L-Met helps reducing cholesterol level by increasing lecithin production in liver and maintains normal growth of cells [13]. Methionine is easily oxidized because of the presence of sulfur in its side chain and its redox chemistry has attracted a lot of interest because of its postulated link with the pathogenesis of diseases like Alzheimer's [14–16] and Parkinson's [17]. Alzheimer's disease (AD) is a neurodegenerative disorder. Consequently the exclusive determination of L-Met is promising from the clinical point of view. In recent years, the electrochemical detection has gained considerable interest for electroactive compounds because of its simpleness, low-cost, sensitivity, selectivity, and reproducibility. So determination of L-Met using modified electrodes is an attractive idea. Eventhough there are some reports are available in the literature for the determination of L-Met using modified electrodes, like doped with Ru(II) metallodendrimer [18], carbon electrode modified with ruthenium metallodendrimer multi layers [19], colloidal gold-cysteamine modified carbon paste electrode [20], platinum/poly(methyl violet) modified electrode [21], nickel powder doped carbon ceramic electrode [13], fullerene-C60 modified gold electrode [22], screen printed graphite macro electrode [23], boron doped diamond and glassy carbon electrodes [24], 1,8,15,22-tetraaminophthalocyanato-copper(II) modified glassy carbon electrode [25], Pt doped TiO2 nanoparticles and carbon
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nanotubes composite on glassy carbon electrode (Pt-TiO2/CNT/GCE) [26], multiwalled carbon nanotubes-based pencil graphite electrode modified with an electrosynthesized molecularly imprinted nanofilm [27], A L-tryptophan-Cu(II)based fluorescence turn-on probe [28], 4Amino nicotinic acid mediated synthesis of gold nanoparticles [29], Electrochemically controlling oxygen functional groups in grapheme oxide for the optimization in the electro-catalytic oxidation of L-methionine [30], modified carbon paste electrode [31]. However, these electrodes have several drawbacks including tedious preparation. In the present work, we focused on synthesis and catalytic activity of Ag-Au bimetal nanoparticles (BNPs) system. The Ag-Au BNPs modified glassy carbon electrode was investigated for the electrochemical oxidation of L-Met using cyclic voltammetry, linear sweep voltammetry and chronoamperometry. The present modified electrode shows good results for the determination of L-Met. 2. Experimental
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(CV), linear sweep voltammetry (LSV) and chronoamperometry were performed with a CHI604C electrochemical workstation (CH Instruments, USA). 2.3. Synthesis of Ag-Au BNPs Bimetal colloidal solution of Ag-Au was synthesized using reducing and stabilizing agent by chemical reduction method. 50 mL of sterile double distilled water containing 0.0158 g of AgNO3 (0.00005 mol) was added and refluxed using oil path for 1 h at 90–100 °C. Then 5 mL of 1% Trisodium citrate (TSC) was slowly added to the reaction mixture. Initially the visible change of the solution from colourless to greenish yellow colour shows the Ag nanoparticle formed. After 30 min of boiling 0.0172 g of HAuCl4 (0.00004 mol) was added dropwise directly to the reaction mixture which is in the round bottom flask and heating was continued with 1.30 h. The reaction mixtures colour changed to dark blue and after completion to dark pink colour solution which indicates the formation of Ag-Au BNPs.
2.1. Reagents and materials 2.4. Preparation of modified electrode Silver nitrate (AgNO3), gold(III) chloride (HAuCl4), trisodium citrate, L-methionine and 5% Nafion-117 were purchased from Sigma-Aldrich and were used without further purification. All aqueous solutions were prepared with deionized (DI) water. Double distilled water was used throughout the studies. All other reagents were of analytical grade. The 0.1 mol L−1 phosphate buffer solutions (PBS), which was made from Na2HPO4, NaH2PO4, and H3PO4 was employed as a supporting electrolyte in the electrochemical measurements.
2.2. Characterization Optical absorbance studies were recorded on a SHIMADZU dualbeam spectrometer (Model UV-1650 PC) operated at a resolution of 1 nm. The morphologies were characterized using a field emission scanning electron microscopy (Hitachi FE-SEMSU6600) and transmission electron microscopy (TEM, JEOL2000). XRD were acquired on an X'Pert X-ray diffraction spectrometer (Philips, USA). The particle size was calculated by using X-ray line broadening technique by employing DebyeScherrer equation. t¼
0:9 λ βcos θ
where, t = particle diameter (in Å), λ = wavelength of the radiation (in Å), β = full-width at half maximum (FWHM, in radians) and θ is the Bragg diffraction angle (in degrees). XPS measurements were performed with Omicron nanotechnology, GMBH, Germany XM1000monochromator with Al Kα radiation of 1483 eV operated at 300 W (20 mA emission current, 15 kV) and a base pressure of 5 × 10−5 mbar. The survey scan was performed with a step size of 0.5 eV along with 50 eV as the pass energy. The high-resolution scan was done with 0.03 eV as the step size and 20 eV as the pass energy with three sweep segments. Zeta-potential measurements were carried out using a Malvern Zetasizer Ver. 6.32 in disposable capillary cells with a volume of 750 mL. The samples were equilibrated for 2 h at 25 °C and measurements of each sample were repeated three times. The zeta-potential distributions were calculated applying standard Smoluchowski fitting. Interpretation was performed by determining the mean values of the resulting peak. If samples having multiple peaks, average mean value, weighted by the peak integrals, were calculated. Dynamic light scattering (DLS) was used to measure the hydrodynamic diameter and the particle size distribution of the as-synthesized colloidal sols. Measurements were carried out by a Malvern Zetasizer Ver. 6.32 working at a fixed angle of 173°. Synthesized sols were properly diluted with water and poured in a polystyrene cuvette before measurement. Data obtained were analyzed using Zetasizer software. Cyclic voltammetry
Electrochemical activity of Ag-Au BNPs was measured in a conventional three electrode cell. A platinum wire served as a counter electrode, Ag/AgCl as a reference electrode and glassy carbon (GC) coated with Ag-Au BNPs as a working electrode. Prior to modification, the glassy carbon electrode (GCE) was polished with 1, 0.3 and 0.05 μm alumina slurry, rinsed thoroughly with ultra pure water between each polishing step, then washed successively with 1:1 nitric acid, acetone and ultra pure water in an ultrasonic bath and dried in air. To fabricate working electrode, 0.5 mL BNPs were mixed with 1 mL of ethanol and sonicated for 30 min. 20 μL of this solution was pipetted onto the surface of GC electrode and dried for 15 min. While ethanol was evaporated, the electrode was coated with a layer of the Ag-Au BNPs. 10 μL of 5% Nafion117 solution was then placed onto the GC electrode and dried for 15 min to form a membrane on the top. The Nafion membrane facilitates Ag-Au BNPs to stick on the electrode surface and works as a proton exchange membrane. Finally, the obtained modified electrode was activated by several successive scans with a scan rate of 50 mV s−1 in phosphate buffer solution (pH 7.4) until a steady voltammogram was obtained. The modified electrode was stored at 4 °C when not in use. 3. Results and discussion 3.1. Characterization of Ag-Au core-shell BNPs 3.1.1. UV–Vis analysis Fig. 1 shows the UV–Vis absorption spectra for the Ag-Au core-shell BNPs, Ag and Au nanoparticles (NPs), respectively. The Ag-Au core-shell BNPs show a characteristic Plasmon peak at 540 nm which is broad and red shifted compared with the characteristic peaks of pure Au and pure Ag NPs. The pure Au and Ag NPs show a characteristic peak at around 530 nm and 410 nm (Fig. 1a and b), respectively. The results accord with previous results [32]. The core-shell formation is concluded from the fact that the optical absorption spectrum shows only one plasmon band [33]. When the solution contains only Ag or Au NPs, absorption peak around 400 or 520 nm should be observed. But Fig. 1c shows there is no such absorption peaks. So the results suggested that there is the Ag-Au core-shell formation. 3.1.2. Zeta potential and dynamic light scattering Zeta potential (ZP) values reveal details about the surface charge and stability of the synthesized Ag-Au core-shell BNPs. The ZP value was found to be _26.6 mV for particles with an average size 40 nm (Fig. 2a). This can be attributed to extremely high surface energies of nanoparticle suspensions. The stability of the Ag-Au core-shell BNPs was tested after repeated intervals (1, 3 and 6 months) using UV–Vis
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the Ag-Au core-shell BNPs was measured by DLS. DLS measures the scattering intensity based on Rayleigh scattering [34]. DLS gives the average particle size of the synthesized Ag-Au core-shell BNPs and found to be 40 nm as shown in Fig. 2b. Polydispersity index of the Ag-Au core-shell BNPs was found to be 0.539, which indicates that the BNPs are generally polydispersed in nature. 3.1.3. XRD analysis Fig. 3 shows the XRD patterns of the prepared Ag-Au (1:1) core-shell BNPs. The peaks were characterized by (111), (200), (220), (311) and (222) peaks corresponding to 2θ values of 38.2, 44.2, 64.6, 77.4, and 81.6 can be assigned to the face-centered cubic (fcc) lattice planes of respectively. By using the Scherrer equation [35], the average particles size is calculate and found to be of 25 nm. Here, the (200) reflection of Ag-Au core-shell BNPs was used to calculate the crystallite size. Note that the diffraction peaks of the Ag-Au core-shell BNPs are much broader compared with those of the monometal Ag and Au NPs owing to the fact that the BNPs are smaller than the monometal NPs. Fig. 1. UV–Vis absorption spectra of (a) Ag NPs, (b) Au NPs and (c) Ag-Au core-shell BNPs.
and ZP measurements and no significant shift either in absorption peak or ZP value was observed. This implies that Ag-Au core-shell BNPs prepared using this protocol can be stored for relatively large periods without compromising their stability. Dynamic light scattering (DLS) is a technique, which determines polydispersity, hydrodynamic sizes and aggregation of particles in suspension. The hydrodynamic diameter of
3.1.4. FE-SEM/EDX analysis FE-SEM image (Fig. 4a) provides more detailed information involving in the formation of Ag-Au core-shell BNPs as spherical morphology. It can be clearly seen that the BNPs are distributed uniformly and having average size of 30 nm in diameter. To further demonstrate the Ag-Au core-shell BNPs, their EDX was also analyzed as shown in Fig. 4b, it is obvious that the elements Ag and Au were present. 3.1.5. TEM and HR-TEM studies Fig. 5a and b illustrate TEM images and size distributions of the AgAu core-shell BNPs prepared by chemical reduction method. One can see that the Ag-Au core-shell BNPs are clearly separated, which shows that the Ag-Au core-shell BNPs exhibit the in homogeneous morphology and showed that the central portions of the particles appeared to be lighter than their edges, denoting the formation of core-shell structures. The HR-TEM of an individual Ag and Au NPs in Ag-Au BNPs and its fast Fourier transform (FFT) image (inset) are presented in Fig. 5c and Fig. 5d, different growth directions are present within an Ag-Au coreshell like structure. The multiple lattice fringes with an interplanar spacing of 0.235 and 0.234 nm were observed, all of which is consistent with the interplanar distance of Ag and Au (111) planes fcc Ag and Au. The electron diffraction pattern indicates that the crystal structure is fcc (Fig. 5e). The size distribution histogram and the BNPs occupied
Fig. 2. Representative (a) zeta potential and (b) dynamic light scattering for Ag-Au coreshell BNPs.
Fig. 3. XRD pattern of Ag-Au core-shell BNPs.
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respectively. The XPS result indicates the presence of Ag and Au atoms in the core-shell BNPs. 3.2. Applications of Ag-Au core-shell BNPs 3.2.1. Electrochemisty of Ag-Au/GC modified electrodes We have examined the electrocatalytic activity of Ag-Au on GCE towards L-Met electrooxidation in buffer solution (pH 7.00) using cyclic voltammetry (Fig. 8). We obtained higher oxidation current with less positive potential for L-Met at pH 7.00. Bare GCE shows an oxidation wave at 0.756 V for L-Met (curve a). On the other hand, the Ag/GCE modified GCE exhibits an oxidation wave for L-Met at 0.764 (curve b). A comparison of the cyclic voltammograms of L-Met at bare GCE (curve b), Au/GCE (curve 8c) demonstrates the oxidation peak potential of L-Met at Ag-Au/GCE occurs at a potential about 119 mV less positive than at a bare GCE (potential shift from 0.932 to 0.637 V). The clear voltammetric signal with higher oxidation current and less positive potential for L-Met at Ag-Au/GCE was attributed to the oxidation of L-Met catalyzed by Ag-Au/GCE. We obtained higher oxidation current with less positive potential for L-Met at pH 7.0. L-Met containing \\SH group selectively bind to the nanorods and the other end containing \\NH2 and\\COOH are free for further interaction form zwitterions at its electrostatic point and form end to end self assembly. Thiol group selectively binds to the nanorods and\\COOH group forming zwitterions at a particular pH [36,37]. The same mechanism follows the oxidation of L-Met on Ag-Au core-shell BNPs. The oxidation peak current of L-Met on Ag-Au core-shell BNPs is higher than that of Ag/GCE and Au/GCE, which might be attributed to the synergistic effect of Ag and Au NPs present in the BNPs.
Fig. 4. (a) FE-SEM image and (b) EDX of Ag-Au core-shell BNPs.
surface volume are given in Fig. 5f and Fig. 5g. The colloid is composed of Ag-Au core-shell BNPs and the average particle size is 30 nm. The energy dispersive X-ray analysis (EDX) (Fig. 5h) confirms that the nanoparticles observed in the TEM images consist of Ag and Au atoms.
3.1.6. AFM morphology studies The shape of Ag-Au core-shell BNPs was then characterized by the AFM. The Ag-Au bimetal colloid for AFM experiments were prepared by depositing a drop of the colloid containing Ag-Au core-shell BNPs onto mica flakes and letting them dry completely at room temperature. Fig. 6 shows the noncontact AFM image of the Ag-Au core-shell BNPs. Two dimensional (Fig. 6a) and three dimensional (Fig. 6b) AFM image of Ag-Au core-shell BNPs indicated that nearly spherical structure. Each particle appears to be an agglomeration of several smaller particles. The diameter of each particle was estimated by examining the height profile or bearing ratio mapped by the AFM tip. The particles size data are shown in Fig. 6c, it is in good agreement with the measurements that done on the similar surfaces by TEM. The particle aggregation is still small and the majority of the particles are well separated.
3.1.7. XPS studies XPS can provide with further information regarding the chemical state of the Ag-Au core-shell BNPs. Fig. 7a presents XPS survey scan of the Ag-Au sample and high resolution XPS shows the Ag3d (Fig. 7b) and Au4f (Fig. 7c) spectral regions. The high resolution XPS from 368 to 372 eV (Fig. 7b) shows the binding energies of Ag3d5/2 at 368 eV and Ag3d3/2 at 374.29 eV and the splitting of the 3d doublet (6 eV). These binding energies reveal that Ag is present in the metallic state. Au4f regions for Ag-Au can be fitted into two sets of spin-orbit doublets. In Fig. 7c, Au4f7/2 and Au4f5/2 lines appears at 83.93 and 87.79 eV,
3.2.2. Effect of scan rate The cyclic voltammograms of Ag-Au core-shell BNPs modified GC electrode at various scan rates were also investigated (Fig. 9a). The effect of different scan rates (ν ranging from 25 to 150 mV s−1) on the current response of L-Met (1.0 × 10−4 mol L−1) on Ag-Au/GCE in phosphate buffer (pH 7.0) was studied and their corresponding expansion spectrum shown in Fig. 9b, a plot of current (ipa) versus square root of scan rate (ν1/2) gave a straight line relationship (Fig. 9c) and a plot of potential versus scan rate give a straight line (Fig. 9d). This revealed that the linearity of the relationship was realized up to a scan rate of 150 mV s−1. This also indicated that the charge transfer was under diffusion control. A good linear relationship was found for the oxidation peak currents and potentials at different scan rates. The oxidation peak currents increased linearly with the linear regression equations as ipa (μA) = 0.34527 (mV) + 0.00915, r2 = 0.995 suggesting that the reaction is a diffusion-controlled electrode reaction. The surface concentration of the Ag-Au/GCE can be estimated by using Brown-Anson model [38]. The peak current is related to the surface concentration of electroactive species, by following equation: Ip ¼
n2 F 2 AI ν 4RT
where, n is the number of electron involved in the reaction, F is the Faraday constant (96.485C mol−1), A is the surface geometrical area of the electrode (0.07 cm2), I* is the surface concentration of the electrode (mol cm−2), ν is the scan rate (50 mV s− 1), R is the gas constant (8.314 J mol−1 K−1) and T is the absolute temperature (298 K). From the slope of peak current versus scan rate (Fig. 9c) the surface concentration of the Ag-Au BNPs modified electrode has been calculated 3.19 × 10−9 mol cm−2. 3.2.3. Effect of pH on L-met oxidation It well known, the electrochemical behaviour of L-Met is dependent on the pH value of the aqueous solution [20,39]. The effect of pH value on the electro oxidation of L-Met at the surface of Ag-Au/GCE was
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Fig. 5. Representative (a, b) TEM images, (c, d) HR-TEM images (inset figure FFT), (e) corresponding SAED pattern, (f) distribution histogram, (g) surface plot and (h) corresponding EDX of Ag-Au core-shell BNPs.
investigated by using of different phosphate buffer solutions (pH 3.00– 11.00). The current response of L-Met at Ag-Au/GCE increases from pH 3.00 to 7.00 and then a decrease is noted at pH values higher than 7.00 (not shown). The electrooxidation of L-Met at a GCE occurs in two steps, corresponding to the formation of sulfoxide and sulfone species [40], involving the adsorption and the protonation/deprotonation of the thiol group, followed by electrochemical oxidation. Pingarron's group have reported that at Au modified carbon paste electrodes preadsorption on the Au surface occurs followed by the oxide-catalyzed oxidation to the sulfone occurs where the mechanism has a pH dependent
response involving 4 electrons and 4 protons [20]. In our case in Ag-Au/ GCE due to the negligible adsorption, L-Met undergoes a one step oxidation reaction with formation of L-Met sulfone (Scheme 1). 3.2.4. Electrochemical impedance spectroscopy (EIS) EIS is another powerful tool for studying the interface properties of surface-modified electrodes. The Nyquist plot of the EIS includes a semicircular portion and a linear portion. The semicircle portion at high frequencies corresponds to electron transfer limited process, and linear portion at lower frequencies corresponds to diffusion process [41].
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Fig. 7. XPS of Ag-Au coresheel BNPs: (a) survey scan, (b) high resolution scans for Ag3d and (c) Au4f. Fig. 6. Topography AFM images of Ag-Au core-shell BNPs: (a) 2D, (b) 3D and (c) height profile.
Fig. 10 shows the EIS curves of the bare and modified GCEs. The values of charge-transfer resistance (RCT) for bare GCE, Ag/GCE, Au/GCE, and AgAu/GCE modified electrodes have been estimated as 256, 238, 213 and 179 Ω cm−2, respectively. The RCT value of Ag-Au/GCE modified GCE is much lower than that of the other modified GCEs. Fig. 10 presents the envoy impedance spectrum of the bare GC electrode (curve a), as GCE was modified with Ag NPs (curve b), the RCT significantly decreased
compared to that of bare GCE. This indicates that the NPs formed an interpenetrating network in favor of diffusion probes and interfacial electron transfers. RCT continued to decrease with modifications with Au and Ag-Au BNPs, respectively (curves c and d). This result shows AgAu core-shell BNPs provides better electron transfer interface between electrode surface and electrolyte solution and also between electro active sites of immobilized L-Met and electrode. Compared to monometal NPs, bimetal Ag-Au core-shell BNPs is an ideal platform for sensing LMet and acts as better biosensors.
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Scheme 1. Representative electrooxidation of L-methionine.
positive potential shift in the oxidation peak potential upon each increment of 50 μM. The oxidation currents had a linear relationship with the concentration of L-Met and the linear regression equation is I (μA) = 1.241 + 0.00108 (μM), with a correlation coefficient of r2 = 0.995 (Fig. 11b). Fig. 8. Cyclic voltammograms of phosphate buffer solution (pH 7.0) at scan rate of 50 mV s−1 with (a) bare GCE, (b) Ag/GCE, (c) Au/GCE and (d) Ag-Au/GCE.
3.2.5. Linear sweep voltammetry obtained for L-Met Fig. 11a shows the linear sweep voltammograms (LSVs) obtained for in the concentration range of 50–500 μM at the Ag-Au/GCE modified electrode. The oxidation current of L-Met increased with a slight
L-Met
3.2.6. Amperometric detection of L-Met We have also carried out the amperometric method to examine the sensitivity of Ag-Au/GCE towards the detection of L-Met individually. Fig. 12 shows the amperometric i–t curve for L-Met with different concentrations using the modified electrode in a stirred 0.1 M PBS at the fixed potential of 0.63 V. It is seen that the current response increases slightly for initial addition of 50 μM and increases significantly for the further addition of 50 μM L-Met with a time interval of 100 s. The
Fig. 9. (a) Cyclic voltammograms of 0.5 mM L-Met at Ag-Au/GCE at different scan rate in 0.1 M phosphate buffer solution (pH 7.0): (i–vi) 25, 50, 75, 100, 125 and 150 mV s−1, respectively, (b) corresponding expand of potential window spectrum for different scan rate, (c) variation of the electrocatalytic currents versus the square root of scan rate and (d) variation of the potential versus the square root of scan rate.
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Fig. 10. Electrochemical impedance spectra recorded at 0.1 M phosphate buffer solution (pH 7.0) with (a) bare GCE, (b) Ag/GCE, (c) Au/GCE and (d) Ag-Au/GCE.
calibration curve of Fig. 12 depicts the linear relationship between the amperometric current and the concentration of L-Met from 50 to 500 μM at Ag-Au/GCE.
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Fig. 12. Amperometric responses of the Ag-Au/GCE after the subsequent addition of L-Met solution in a 0.1 M PBS (pH 7.0) at 0.63 V.
3.2.7. Interference studies The practical application of Ag-Au/GCE was tested by measuring the concentration of L-Met in human blood serum samples. It is very important to determine the concentration of L-Met in the presence high concentration of Glutamine (Glu) because its concentration is generally higher than L-Met in human fluids such as blood serum, plasma and urine. Fig. 13 shows the LSVs obtained for 5 mM and 0.5 mM Glu of each L-Cystine (L-Cys) and L-Met at Ag-Au BNPs modified GCE in 1 M PBS (pH 7.0). It can be seen from Fig. 13 that a very clear voltammetric signal was found for L-Met at 0.75 (±0.03) V even in the presence of 10fold higher concentration of Glu and an equal concentrations of L-Cys. This result reveals that the present modified electrode could be applied to determine of L-Met even in the presence of high concentration of Glu. 3.2.8. Reproducibility and stability of the Ag-Au/GCE In order to investigate the stability of the Ag-Au/GCE modified electrode by CV in PBS (pH 7.0) containing 0.5 mM of L-Met; 25 continuous
Fig. 11. (a) LSVs obtained for L-Met in the concentrations ranging from 50 to 500 mM. LMet was added in steps of 50 mM each at the Ag-Au/GCE modified electrode in PBS (pH 7.0) and (b) represent linearity curve at different concentrations.
Fig. 13. LSVs obtained for 5 mM of Glu, and 0.5 mM each L-Cys and L-Met at Ag-Au/GCE in 0.1 M PBS (pH 7.0).
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project research F. No. 37-33/2010. One of the authors M. Murugavelu acknowledges UGC Networking Resource Centre, University of Hyderabad for providing the characterization facility and is thankful to Dr. Tushar Jana, School of Chemistry, University of Hyderabad for the laboratory facility.
References
Fig. 14. Cyclic voltammetric stability of Ag-Au/GCE modified GCE in PBS containing 0.5 mM of L-Met at a scan rate of 50 mV s−1.
cycles were carried out at a scan rate of 50 mV s−1. The results show only 1.71% loss from the initial determination value (Fig. 14). This result indicates that Ag-Au/GCE modified electrode has a good stability, reproducibility and does not undergo surface fouling. The storage stability of the sensors was investigated over a period of 2 weeks by storing the AgAu/GCE in 0.1 M PBS (pH 7.0) at room temperature. No obvious changes were observed during the first week and the response maintained up to 96.0%. This result showed that the stability of the Ag-Au/GCE was satisfactory. Thus it has good stability, excellent reproducibility and stability and may be mainly ascribed to the noble metals Ag and Au in the modified electrode. 4. Conclusion In this report, the Ag-Au core-shell BNPs have been synthesized and characterized using various methods. Ag-Au core-shell BNPs modified GC electrode (Ag-Au/GCE) was fabricated by drop casting method. The as-prepared electrode exhibited a high catalytic efficiency to the oxidation of L-Met due to the high surface area of the Ag-Au core-shell structure. Ag-Au/GCE shows good characteristics, such as a large determination range (50 μM – 1 mM), low detection limit (30 μM), under the optimum conditions. The constructed Ag-Au/GCE was established for the sensing of L-Met with robust stability and good reproducibility. Acknowledgements The author (BK) thanks University Grants Commission (UGC), New Delhi, for the granting financial support through a major research
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Materials Science and Engineering C journal homepage: www.elsevier.com/locate/msec
Synthesis, characterization of Ag-Au core-shell bimetal nanoparticles and its application for electrocatalytic oxidation/sensing of L-methionine M. Murugavelu, B. Karthikeyan ⁎ Department of Chemistry, Annamalai University, Annamalainagar 608 002, Tamil Nadu, India
a r t i c l e
i n f o
Article history: Received 8 May 2016 Received in revised form 30 August 2016 Accepted 21 September 2016 Available online 23 September 2016 Keywords: Ag-Au core-shell Cyclic voltammetry Electrocatalytic L-methionine
a b s t r a c t The Ag-Au core-shell bimetal nanoparticles (BNPs) was prepared using chemical reduction method. The prepared Ag-Au core-shell BNPs were characterized by UV–Visible (UV–Vis) spectroscopy, field emission scanning electron microscopy (FE-SEM), transmission electron microscopy (TEM), high resolution transmission electron microscopy (HR-TEM), X-ray diffraction (XRD), atomic force microscopy (AFM) and X-ray photoelectron spectroscopy (XPS) pattern. These results showed the Ag-Au BNPs exhibited core-shell shape. The Ag-Au core-shell BNPs was examined towards electrocatalytic oxidation of L-methionine (L-Met) by cyclic voltammetry (CV), linear sweep voltammetry (LSV) and chronoamperometry. According to the results, L-Met is determined with detection limit of 30 μM. Interference studies in biological buffer was also studied. © 2016 Elsevier B.V. All rights reserved.
1. Introduction The preparation of bimetallic nanoparticles (BNPs) have great significance because of the modification of physical and chemical properties, compared with monometallic nanoparticles (NPs) due to size effect. Enhanced electrocatalytic activity, catalysis, result from the combination of different metals are also taken place [1–5]. These BNPs can retain the functional properties of each component and possibly offer synergistic effects via cooperative interactions, resulting in important features such as increased surface area, enhanced electrocatalytic activity, improved biocompatibility, promoted electron transfer, and better invulnerability against intermediate species. During the past years, such bimetallic nanostructured materials were extensively developed and reported by many scientists [6,7]. Metal NPs offer excellent applications for chemical and biological sensing because of their unique optical and electrical properties [8]. Especially Au nanoparticle-based electrochemical stripping detection of DNA hybridization reported by Authier et al. [9,10] and signal amplification with Ag enhancement [11] for gene analysis may be noted. In the case of BNPs, when a new nanoparticle composition is designed, new modification methods must be developed for immobilizing biomolecules on the surface of the particles of interest. The methods for modifying Au NPs have now been optimized and generalized for a wide range of particle sizes and surface compositions, including spheres and rods [10,11]. From the wide literature review, we have picked to prepare Ag-Au BNPs and its synergetic effect in a modified electrodes for determination of L-methionine. ⁎ Corresponding author at: Department of Chemistry, Annamalai University, Annamalainagar 608 002, Tamil Nadu, India. E-mail address: [email protected] (B. Karthikeyan).
http://dx.doi.org/10.1016/j.msec.2016.09.046 0928-4931/© 2016 Elsevier B.V. All rights reserved.
Methionine is one of the essential amino acid among the 20 amino acids, it occurs naturally as L-methionine (L-Met), and is a major source of sulfur, which is required in the human diet for normal metabolism and growth, since it is not synthesized in humans. Methionine is often used as an additive in animal feedstuffs, and it is also used in production of medicines and active pharmaceutical ingredients, and as a precursor to other amino acids [12]. It governs the main supply of sulfur in the diet, and also prevents disorders in hair and skin. It acts as an essential amino acid with the key role in biological methylation. L-Met helps reducing cholesterol level by increasing lecithin production in liver and maintains normal growth of cells [13]. Methionine is easily oxidized because of the presence of sulfur in its side chain and its redox chemistry has attracted a lot of interest because of its postulated link with the pathogenesis of diseases like Alzheimer's [14–16] and Parkinson's [17]. Alzheimer's disease (AD) is a neurodegenerative disorder. Consequently the exclusive determination of L-Met is promising from the clinical point of view. In recent years, the electrochemical detection has gained considerable interest for electroactive compounds because of its simpleness, low-cost, sensitivity, selectivity, and reproducibility. So determination of L-Met using modified electrodes is an attractive idea. Eventhough there are some reports are available in the literature for the determination of L-Met using modified electrodes, like doped with Ru(II) metallodendrimer [18], carbon electrode modified with ruthenium metallodendrimer multi layers [19], colloidal gold-cysteamine modified carbon paste electrode [20], platinum/poly(methyl violet) modified electrode [21], nickel powder doped carbon ceramic electrode [13], fullerene-C60 modified gold electrode [22], screen printed graphite macro electrode [23], boron doped diamond and glassy carbon electrodes [24], 1,8,15,22-tetraaminophthalocyanato-copper(II) modified glassy carbon electrode [25], Pt doped TiO2 nanoparticles and carbon
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nanotubes composite on glassy carbon electrode (Pt-TiO2/CNT/GCE) [26], multiwalled carbon nanotubes-based pencil graphite electrode modified with an electrosynthesized molecularly imprinted nanofilm [27], A L-tryptophan-Cu(II)based fluorescence turn-on probe [28], 4Amino nicotinic acid mediated synthesis of gold nanoparticles [29], Electrochemically controlling oxygen functional groups in grapheme oxide for the optimization in the electro-catalytic oxidation of L-methionine [30], modified carbon paste electrode [31]. However, these electrodes have several drawbacks including tedious preparation. In the present work, we focused on synthesis and catalytic activity of Ag-Au bimetal nanoparticles (BNPs) system. The Ag-Au BNPs modified glassy carbon electrode was investigated for the electrochemical oxidation of L-Met using cyclic voltammetry, linear sweep voltammetry and chronoamperometry. The present modified electrode shows good results for the determination of L-Met. 2. Experimental
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(CV), linear sweep voltammetry (LSV) and chronoamperometry were performed with a CHI604C electrochemical workstation (CH Instruments, USA). 2.3. Synthesis of Ag-Au BNPs Bimetal colloidal solution of Ag-Au was synthesized using reducing and stabilizing agent by chemical reduction method. 50 mL of sterile double distilled water containing 0.0158 g of AgNO3 (0.00005 mol) was added and refluxed using oil path for 1 h at 90–100 °C. Then 5 mL of 1% Trisodium citrate (TSC) was slowly added to the reaction mixture. Initially the visible change of the solution from colourless to greenish yellow colour shows the Ag nanoparticle formed. After 30 min of boiling 0.0172 g of HAuCl4 (0.00004 mol) was added dropwise directly to the reaction mixture which is in the round bottom flask and heating was continued with 1.30 h. The reaction mixtures colour changed to dark blue and after completion to dark pink colour solution which indicates the formation of Ag-Au BNPs.
2.1. Reagents and materials 2.4. Preparation of modified electrode Silver nitrate (AgNO3), gold(III) chloride (HAuCl4), trisodium citrate, L-methionine and 5% Nafion-117 were purchased from Sigma-Aldrich and were used without further purification. All aqueous solutions were prepared with deionized (DI) water. Double distilled water was used throughout the studies. All other reagents were of analytical grade. The 0.1 mol L−1 phosphate buffer solutions (PBS), which was made from Na2HPO4, NaH2PO4, and H3PO4 was employed as a supporting electrolyte in the electrochemical measurements.
2.2. Characterization Optical absorbance studies were recorded on a SHIMADZU dualbeam spectrometer (Model UV-1650 PC) operated at a resolution of 1 nm. The morphologies were characterized using a field emission scanning electron microscopy (Hitachi FE-SEMSU6600) and transmission electron microscopy (TEM, JEOL2000). XRD were acquired on an X'Pert X-ray diffraction spectrometer (Philips, USA). The particle size was calculated by using X-ray line broadening technique by employing DebyeScherrer equation. t¼
0:9 λ βcos θ
where, t = particle diameter (in Å), λ = wavelength of the radiation (in Å), β = full-width at half maximum (FWHM, in radians) and θ is the Bragg diffraction angle (in degrees). XPS measurements were performed with Omicron nanotechnology, GMBH, Germany XM1000monochromator with Al Kα radiation of 1483 eV operated at 300 W (20 mA emission current, 15 kV) and a base pressure of 5 × 10−5 mbar. The survey scan was performed with a step size of 0.5 eV along with 50 eV as the pass energy. The high-resolution scan was done with 0.03 eV as the step size and 20 eV as the pass energy with three sweep segments. Zeta-potential measurements were carried out using a Malvern Zetasizer Ver. 6.32 in disposable capillary cells with a volume of 750 mL. The samples were equilibrated for 2 h at 25 °C and measurements of each sample were repeated three times. The zeta-potential distributions were calculated applying standard Smoluchowski fitting. Interpretation was performed by determining the mean values of the resulting peak. If samples having multiple peaks, average mean value, weighted by the peak integrals, were calculated. Dynamic light scattering (DLS) was used to measure the hydrodynamic diameter and the particle size distribution of the as-synthesized colloidal sols. Measurements were carried out by a Malvern Zetasizer Ver. 6.32 working at a fixed angle of 173°. Synthesized sols were properly diluted with water and poured in a polystyrene cuvette before measurement. Data obtained were analyzed using Zetasizer software. Cyclic voltammetry
Electrochemical activity of Ag-Au BNPs was measured in a conventional three electrode cell. A platinum wire served as a counter electrode, Ag/AgCl as a reference electrode and glassy carbon (GC) coated with Ag-Au BNPs as a working electrode. Prior to modification, the glassy carbon electrode (GCE) was polished with 1, 0.3 and 0.05 μm alumina slurry, rinsed thoroughly with ultra pure water between each polishing step, then washed successively with 1:1 nitric acid, acetone and ultra pure water in an ultrasonic bath and dried in air. To fabricate working electrode, 0.5 mL BNPs were mixed with 1 mL of ethanol and sonicated for 30 min. 20 μL of this solution was pipetted onto the surface of GC electrode and dried for 15 min. While ethanol was evaporated, the electrode was coated with a layer of the Ag-Au BNPs. 10 μL of 5% Nafion117 solution was then placed onto the GC electrode and dried for 15 min to form a membrane on the top. The Nafion membrane facilitates Ag-Au BNPs to stick on the electrode surface and works as a proton exchange membrane. Finally, the obtained modified electrode was activated by several successive scans with a scan rate of 50 mV s−1 in phosphate buffer solution (pH 7.4) until a steady voltammogram was obtained. The modified electrode was stored at 4 °C when not in use. 3. Results and discussion 3.1. Characterization of Ag-Au core-shell BNPs 3.1.1. UV–Vis analysis Fig. 1 shows the UV–Vis absorption spectra for the Ag-Au core-shell BNPs, Ag and Au nanoparticles (NPs), respectively. The Ag-Au core-shell BNPs show a characteristic Plasmon peak at 540 nm which is broad and red shifted compared with the characteristic peaks of pure Au and pure Ag NPs. The pure Au and Ag NPs show a characteristic peak at around 530 nm and 410 nm (Fig. 1a and b), respectively. The results accord with previous results [32]. The core-shell formation is concluded from the fact that the optical absorption spectrum shows only one plasmon band [33]. When the solution contains only Ag or Au NPs, absorption peak around 400 or 520 nm should be observed. But Fig. 1c shows there is no such absorption peaks. So the results suggested that there is the Ag-Au core-shell formation. 3.1.2. Zeta potential and dynamic light scattering Zeta potential (ZP) values reveal details about the surface charge and stability of the synthesized Ag-Au core-shell BNPs. The ZP value was found to be _26.6 mV for particles with an average size 40 nm (Fig. 2a). This can be attributed to extremely high surface energies of nanoparticle suspensions. The stability of the Ag-Au core-shell BNPs was tested after repeated intervals (1, 3 and 6 months) using UV–Vis
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the Ag-Au core-shell BNPs was measured by DLS. DLS measures the scattering intensity based on Rayleigh scattering [34]. DLS gives the average particle size of the synthesized Ag-Au core-shell BNPs and found to be 40 nm as shown in Fig. 2b. Polydispersity index of the Ag-Au core-shell BNPs was found to be 0.539, which indicates that the BNPs are generally polydispersed in nature. 3.1.3. XRD analysis Fig. 3 shows the XRD patterns of the prepared Ag-Au (1:1) core-shell BNPs. The peaks were characterized by (111), (200), (220), (311) and (222) peaks corresponding to 2θ values of 38.2, 44.2, 64.6, 77.4, and 81.6 can be assigned to the face-centered cubic (fcc) lattice planes of respectively. By using the Scherrer equation [35], the average particles size is calculate and found to be of 25 nm. Here, the (200) reflection of Ag-Au core-shell BNPs was used to calculate the crystallite size. Note that the diffraction peaks of the Ag-Au core-shell BNPs are much broader compared with those of the monometal Ag and Au NPs owing to the fact that the BNPs are smaller than the monometal NPs. Fig. 1. UV–Vis absorption spectra of (a) Ag NPs, (b) Au NPs and (c) Ag-Au core-shell BNPs.
and ZP measurements and no significant shift either in absorption peak or ZP value was observed. This implies that Ag-Au core-shell BNPs prepared using this protocol can be stored for relatively large periods without compromising their stability. Dynamic light scattering (DLS) is a technique, which determines polydispersity, hydrodynamic sizes and aggregation of particles in suspension. The hydrodynamic diameter of
3.1.4. FE-SEM/EDX analysis FE-SEM image (Fig. 4a) provides more detailed information involving in the formation of Ag-Au core-shell BNPs as spherical morphology. It can be clearly seen that the BNPs are distributed uniformly and having average size of 30 nm in diameter. To further demonstrate the Ag-Au core-shell BNPs, their EDX was also analyzed as shown in Fig. 4b, it is obvious that the elements Ag and Au were present. 3.1.5. TEM and HR-TEM studies Fig. 5a and b illustrate TEM images and size distributions of the AgAu core-shell BNPs prepared by chemical reduction method. One can see that the Ag-Au core-shell BNPs are clearly separated, which shows that the Ag-Au core-shell BNPs exhibit the in homogeneous morphology and showed that the central portions of the particles appeared to be lighter than their edges, denoting the formation of core-shell structures. The HR-TEM of an individual Ag and Au NPs in Ag-Au BNPs and its fast Fourier transform (FFT) image (inset) are presented in Fig. 5c and Fig. 5d, different growth directions are present within an Ag-Au coreshell like structure. The multiple lattice fringes with an interplanar spacing of 0.235 and 0.234 nm were observed, all of which is consistent with the interplanar distance of Ag and Au (111) planes fcc Ag and Au. The electron diffraction pattern indicates that the crystal structure is fcc (Fig. 5e). The size distribution histogram and the BNPs occupied
Fig. 2. Representative (a) zeta potential and (b) dynamic light scattering for Ag-Au coreshell BNPs.
Fig. 3. XRD pattern of Ag-Au core-shell BNPs.
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respectively. The XPS result indicates the presence of Ag and Au atoms in the core-shell BNPs. 3.2. Applications of Ag-Au core-shell BNPs 3.2.1. Electrochemisty of Ag-Au/GC modified electrodes We have examined the electrocatalytic activity of Ag-Au on GCE towards L-Met electrooxidation in buffer solution (pH 7.00) using cyclic voltammetry (Fig. 8). We obtained higher oxidation current with less positive potential for L-Met at pH 7.00. Bare GCE shows an oxidation wave at 0.756 V for L-Met (curve a). On the other hand, the Ag/GCE modified GCE exhibits an oxidation wave for L-Met at 0.764 (curve b). A comparison of the cyclic voltammograms of L-Met at bare GCE (curve b), Au/GCE (curve 8c) demonstrates the oxidation peak potential of L-Met at Ag-Au/GCE occurs at a potential about 119 mV less positive than at a bare GCE (potential shift from 0.932 to 0.637 V). The clear voltammetric signal with higher oxidation current and less positive potential for L-Met at Ag-Au/GCE was attributed to the oxidation of L-Met catalyzed by Ag-Au/GCE. We obtained higher oxidation current with less positive potential for L-Met at pH 7.0. L-Met containing \\SH group selectively bind to the nanorods and the other end containing \\NH2 and\\COOH are free for further interaction form zwitterions at its electrostatic point and form end to end self assembly. Thiol group selectively binds to the nanorods and\\COOH group forming zwitterions at a particular pH [36,37]. The same mechanism follows the oxidation of L-Met on Ag-Au core-shell BNPs. The oxidation peak current of L-Met on Ag-Au core-shell BNPs is higher than that of Ag/GCE and Au/GCE, which might be attributed to the synergistic effect of Ag and Au NPs present in the BNPs.
Fig. 4. (a) FE-SEM image and (b) EDX of Ag-Au core-shell BNPs.
surface volume are given in Fig. 5f and Fig. 5g. The colloid is composed of Ag-Au core-shell BNPs and the average particle size is 30 nm. The energy dispersive X-ray analysis (EDX) (Fig. 5h) confirms that the nanoparticles observed in the TEM images consist of Ag and Au atoms.
3.1.6. AFM morphology studies The shape of Ag-Au core-shell BNPs was then characterized by the AFM. The Ag-Au bimetal colloid for AFM experiments were prepared by depositing a drop of the colloid containing Ag-Au core-shell BNPs onto mica flakes and letting them dry completely at room temperature. Fig. 6 shows the noncontact AFM image of the Ag-Au core-shell BNPs. Two dimensional (Fig. 6a) and three dimensional (Fig. 6b) AFM image of Ag-Au core-shell BNPs indicated that nearly spherical structure. Each particle appears to be an agglomeration of several smaller particles. The diameter of each particle was estimated by examining the height profile or bearing ratio mapped by the AFM tip. The particles size data are shown in Fig. 6c, it is in good agreement with the measurements that done on the similar surfaces by TEM. The particle aggregation is still small and the majority of the particles are well separated.
3.1.7. XPS studies XPS can provide with further information regarding the chemical state of the Ag-Au core-shell BNPs. Fig. 7a presents XPS survey scan of the Ag-Au sample and high resolution XPS shows the Ag3d (Fig. 7b) and Au4f (Fig. 7c) spectral regions. The high resolution XPS from 368 to 372 eV (Fig. 7b) shows the binding energies of Ag3d5/2 at 368 eV and Ag3d3/2 at 374.29 eV and the splitting of the 3d doublet (6 eV). These binding energies reveal that Ag is present in the metallic state. Au4f regions for Ag-Au can be fitted into two sets of spin-orbit doublets. In Fig. 7c, Au4f7/2 and Au4f5/2 lines appears at 83.93 and 87.79 eV,
3.2.2. Effect of scan rate The cyclic voltammograms of Ag-Au core-shell BNPs modified GC electrode at various scan rates were also investigated (Fig. 9a). The effect of different scan rates (ν ranging from 25 to 150 mV s−1) on the current response of L-Met (1.0 × 10−4 mol L−1) on Ag-Au/GCE in phosphate buffer (pH 7.0) was studied and their corresponding expansion spectrum shown in Fig. 9b, a plot of current (ipa) versus square root of scan rate (ν1/2) gave a straight line relationship (Fig. 9c) and a plot of potential versus scan rate give a straight line (Fig. 9d). This revealed that the linearity of the relationship was realized up to a scan rate of 150 mV s−1. This also indicated that the charge transfer was under diffusion control. A good linear relationship was found for the oxidation peak currents and potentials at different scan rates. The oxidation peak currents increased linearly with the linear regression equations as ipa (μA) = 0.34527 (mV) + 0.00915, r2 = 0.995 suggesting that the reaction is a diffusion-controlled electrode reaction. The surface concentration of the Ag-Au/GCE can be estimated by using Brown-Anson model [38]. The peak current is related to the surface concentration of electroactive species, by following equation: Ip ¼
n2 F 2 AI ν 4RT
where, n is the number of electron involved in the reaction, F is the Faraday constant (96.485C mol−1), A is the surface geometrical area of the electrode (0.07 cm2), I* is the surface concentration of the electrode (mol cm−2), ν is the scan rate (50 mV s− 1), R is the gas constant (8.314 J mol−1 K−1) and T is the absolute temperature (298 K). From the slope of peak current versus scan rate (Fig. 9c) the surface concentration of the Ag-Au BNPs modified electrode has been calculated 3.19 × 10−9 mol cm−2. 3.2.3. Effect of pH on L-met oxidation It well known, the electrochemical behaviour of L-Met is dependent on the pH value of the aqueous solution [20,39]. The effect of pH value on the electro oxidation of L-Met at the surface of Ag-Au/GCE was
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Fig. 5. Representative (a, b) TEM images, (c, d) HR-TEM images (inset figure FFT), (e) corresponding SAED pattern, (f) distribution histogram, (g) surface plot and (h) corresponding EDX of Ag-Au core-shell BNPs.
investigated by using of different phosphate buffer solutions (pH 3.00– 11.00). The current response of L-Met at Ag-Au/GCE increases from pH 3.00 to 7.00 and then a decrease is noted at pH values higher than 7.00 (not shown). The electrooxidation of L-Met at a GCE occurs in two steps, corresponding to the formation of sulfoxide and sulfone species [40], involving the adsorption and the protonation/deprotonation of the thiol group, followed by electrochemical oxidation. Pingarron's group have reported that at Au modified carbon paste electrodes preadsorption on the Au surface occurs followed by the oxide-catalyzed oxidation to the sulfone occurs where the mechanism has a pH dependent
response involving 4 electrons and 4 protons [20]. In our case in Ag-Au/ GCE due to the negligible adsorption, L-Met undergoes a one step oxidation reaction with formation of L-Met sulfone (Scheme 1). 3.2.4. Electrochemical impedance spectroscopy (EIS) EIS is another powerful tool for studying the interface properties of surface-modified electrodes. The Nyquist plot of the EIS includes a semicircular portion and a linear portion. The semicircle portion at high frequencies corresponds to electron transfer limited process, and linear portion at lower frequencies corresponds to diffusion process [41].
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Fig. 7. XPS of Ag-Au coresheel BNPs: (a) survey scan, (b) high resolution scans for Ag3d and (c) Au4f. Fig. 6. Topography AFM images of Ag-Au core-shell BNPs: (a) 2D, (b) 3D and (c) height profile.
Fig. 10 shows the EIS curves of the bare and modified GCEs. The values of charge-transfer resistance (RCT) for bare GCE, Ag/GCE, Au/GCE, and AgAu/GCE modified electrodes have been estimated as 256, 238, 213 and 179 Ω cm−2, respectively. The RCT value of Ag-Au/GCE modified GCE is much lower than that of the other modified GCEs. Fig. 10 presents the envoy impedance spectrum of the bare GC electrode (curve a), as GCE was modified with Ag NPs (curve b), the RCT significantly decreased
compared to that of bare GCE. This indicates that the NPs formed an interpenetrating network in favor of diffusion probes and interfacial electron transfers. RCT continued to decrease with modifications with Au and Ag-Au BNPs, respectively (curves c and d). This result shows AgAu core-shell BNPs provides better electron transfer interface between electrode surface and electrolyte solution and also between electro active sites of immobilized L-Met and electrode. Compared to monometal NPs, bimetal Ag-Au core-shell BNPs is an ideal platform for sensing LMet and acts as better biosensors.
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Scheme 1. Representative electrooxidation of L-methionine.
positive potential shift in the oxidation peak potential upon each increment of 50 μM. The oxidation currents had a linear relationship with the concentration of L-Met and the linear regression equation is I (μA) = 1.241 + 0.00108 (μM), with a correlation coefficient of r2 = 0.995 (Fig. 11b). Fig. 8. Cyclic voltammograms of phosphate buffer solution (pH 7.0) at scan rate of 50 mV s−1 with (a) bare GCE, (b) Ag/GCE, (c) Au/GCE and (d) Ag-Au/GCE.
3.2.5. Linear sweep voltammetry obtained for L-Met Fig. 11a shows the linear sweep voltammograms (LSVs) obtained for in the concentration range of 50–500 μM at the Ag-Au/GCE modified electrode. The oxidation current of L-Met increased with a slight
L-Met
3.2.6. Amperometric detection of L-Met We have also carried out the amperometric method to examine the sensitivity of Ag-Au/GCE towards the detection of L-Met individually. Fig. 12 shows the amperometric i–t curve for L-Met with different concentrations using the modified electrode in a stirred 0.1 M PBS at the fixed potential of 0.63 V. It is seen that the current response increases slightly for initial addition of 50 μM and increases significantly for the further addition of 50 μM L-Met with a time interval of 100 s. The
Fig. 9. (a) Cyclic voltammograms of 0.5 mM L-Met at Ag-Au/GCE at different scan rate in 0.1 M phosphate buffer solution (pH 7.0): (i–vi) 25, 50, 75, 100, 125 and 150 mV s−1, respectively, (b) corresponding expand of potential window spectrum for different scan rate, (c) variation of the electrocatalytic currents versus the square root of scan rate and (d) variation of the potential versus the square root of scan rate.
M. Murugavelu, B. Karthikeyan / Materials Science and Engineering C 70 (2017) 656–664
Fig. 10. Electrochemical impedance spectra recorded at 0.1 M phosphate buffer solution (pH 7.0) with (a) bare GCE, (b) Ag/GCE, (c) Au/GCE and (d) Ag-Au/GCE.
calibration curve of Fig. 12 depicts the linear relationship between the amperometric current and the concentration of L-Met from 50 to 500 μM at Ag-Au/GCE.
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Fig. 12. Amperometric responses of the Ag-Au/GCE after the subsequent addition of L-Met solution in a 0.1 M PBS (pH 7.0) at 0.63 V.
3.2.7. Interference studies The practical application of Ag-Au/GCE was tested by measuring the concentration of L-Met in human blood serum samples. It is very important to determine the concentration of L-Met in the presence high concentration of Glutamine (Glu) because its concentration is generally higher than L-Met in human fluids such as blood serum, plasma and urine. Fig. 13 shows the LSVs obtained for 5 mM and 0.5 mM Glu of each L-Cystine (L-Cys) and L-Met at Ag-Au BNPs modified GCE in 1 M PBS (pH 7.0). It can be seen from Fig. 13 that a very clear voltammetric signal was found for L-Met at 0.75 (±0.03) V even in the presence of 10fold higher concentration of Glu and an equal concentrations of L-Cys. This result reveals that the present modified electrode could be applied to determine of L-Met even in the presence of high concentration of Glu. 3.2.8. Reproducibility and stability of the Ag-Au/GCE In order to investigate the stability of the Ag-Au/GCE modified electrode by CV in PBS (pH 7.0) containing 0.5 mM of L-Met; 25 continuous
Fig. 11. (a) LSVs obtained for L-Met in the concentrations ranging from 50 to 500 mM. LMet was added in steps of 50 mM each at the Ag-Au/GCE modified electrode in PBS (pH 7.0) and (b) represent linearity curve at different concentrations.
Fig. 13. LSVs obtained for 5 mM of Glu, and 0.5 mM each L-Cys and L-Met at Ag-Au/GCE in 0.1 M PBS (pH 7.0).
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project research F. No. 37-33/2010. One of the authors M. Murugavelu acknowledges UGC Networking Resource Centre, University of Hyderabad for providing the characterization facility and is thankful to Dr. Tushar Jana, School of Chemistry, University of Hyderabad for the laboratory facility.
References
Fig. 14. Cyclic voltammetric stability of Ag-Au/GCE modified GCE in PBS containing 0.5 mM of L-Met at a scan rate of 50 mV s−1.
cycles were carried out at a scan rate of 50 mV s−1. The results show only 1.71% loss from the initial determination value (Fig. 14). This result indicates that Ag-Au/GCE modified electrode has a good stability, reproducibility and does not undergo surface fouling. The storage stability of the sensors was investigated over a period of 2 weeks by storing the AgAu/GCE in 0.1 M PBS (pH 7.0) at room temperature. No obvious changes were observed during the first week and the response maintained up to 96.0%. This result showed that the stability of the Ag-Au/GCE was satisfactory. Thus it has good stability, excellent reproducibility and stability and may be mainly ascribed to the noble metals Ag and Au in the modified electrode. 4. Conclusion In this report, the Ag-Au core-shell BNPs have been synthesized and characterized using various methods. Ag-Au core-shell BNPs modified GC electrode (Ag-Au/GCE) was fabricated by drop casting method. The as-prepared electrode exhibited a high catalytic efficiency to the oxidation of L-Met due to the high surface area of the Ag-Au core-shell structure. Ag-Au/GCE shows good characteristics, such as a large determination range (50 μM – 1 mM), low detection limit (30 μM), under the optimum conditions. The constructed Ag-Au/GCE was established for the sensing of L-Met with robust stability and good reproducibility. Acknowledgements The author (BK) thanks University Grants Commission (UGC), New Delhi, for the granting financial support through a major research
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