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Cite This: Anal. Chem. XXXX, XXX, XXX−XXX

Electrochemical Detection of Gallic Acid-Capped Gold Nanoparticles Using a Multiwalled Carbon Nanotube-Reduced Graphene Oxide Nanocomposite Electrode Hashwin V. S. Ganesh, Bhargav R. Patel, Hamid Fini, Ari M. Chow, and Kagan Kerman* Department of Physical and Environmental Sciences, University of Toronto, Scarborough 1265 Military Trail, Toronto, ON M1C 1A4, Canada Downloaded via KEAN UNIV on July 18, 2019 at 01:24:40 (UTC). See https://pubs.acs.org/sharingguidelines for options on how to legitimately share published articles.

S Supporting Information *

ABSTRACT: Recently, a plethora of ecofriendly methods have been developed for the synthesis of AuNPs using a multitude of biogenic agents. Polyphenols from plants are particularly attractive for producing AuNPs because in addition to helping with the synthesis of AuNPs, the polyphenol capping of the NPs can be used as a platform for versatile applications. Polyphenol-capped AuNPs could also make the detection of AuNPs possible, should they be released into the environment. Because polyphenols are redox-active, they can be used as a probe to detect AuNPs using electrochemical techniques. In this work, we have developed an MWCNT-rGO nanocomposite electrode for the sensitive detection of AuNPs capped with gallic acid (GA, a green-tea-derived polyphenol) using differential pulse voltammetry (DPV). The reduction of gallic acid-capped AuNPs was used as the quantification signal, and the calibration curve displayed a detection limit of 2.57 pM. Using cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS), we have shown that the modification of the electrode surface with an MWCNT-rGO hybrid nanocomposite resulted in a 10-fold increase in current response leading to the sensitive detection of GA-AuNPs compared to unmodified electrodes. We have also demonstrated the applicability of the electrochemical sensor in detecting GA-AuNPs in various analytical matrixes such as human serum and natural creek water (Highland Creek, ON) with good recovery.

G

expensive synthetic chemical reducing agent.17 NPs synthesized using such methods have also been shown to be free of contamination and to be sustainable, less expensive, and suitable for mass production.17 AuNPs synthesized using ecofriendly methods offer various advantages compared to AuNPs produced using physicochemical methods. Although the synthesis of AuNPs is facile, the natural reducing agents can also act as capping and stabilizing agents, preventing aggregation or agglomeration of NPs. In some instances, NPs synthesized using such methods have been shown to have improved properties compared to the properties of chemically produced NPs. For example, silver nanoparticles (AgNPs) produced using biogenic methods have been shown to exhibit approximately 20-fold-higher antimicrobial activity when compared to chemically produced AgNPs.18 Polyphenols from plants, in the form of extracts as well as pure compounds, have been used to synthesize AuNPs. 19 Polyphenols are particularly attractive for producing AuNPs because in addition to helping with the synthesis of AuNPs, polyphenol capping of the NPs can be further utilized as a

old nanoparticles (AuNPs) have become a vital component of research and development in nanotechnology,1 electronics,2 and biomedical3 and materials science.4 Numerous methods have been developed for the synthesis of AuNPs.1 One of the most widely used methods for the preparation of AuNPs was developed by Brust and Schriffin5 in 1994, which involved creating organic-soluble alkanethiol-stabilized-AuNPs through a biphasic reduction protocol using tetraoctylammonium bromide (TOAB) as the phase-transfer reagent and sodium borohydride (NaBH4) as the reducing agent. Although the AuNPs synthesized using the above method have numerous advantages such as higher stability and the ability to be dried and redispersed in solution without aggregation, a major drawback is that they require the use of toxic chemicals such as toluene and generate hazardous byproducts as part of the synthesis process.5 Recently, a plethora of methods have been developed for the synthesis of ecofriendly AuNPs. AuNPs have been synthesized using a multitude of biogenic agents such as amino acids,6 citric acid,7 polyphenols,8 reducing sugars,9 saponin,10 terpenoids,11 carboxylic acids,12 ketones,13 flavonoids,14 heterocyclic compounds,15 and proteins.16 The synthesis of AuNPs is a bottom-up approach that involves using a reducing agent that is naturally available, as opposed to using an © XXXX American Chemical Society

Received: May 6, 2019 Accepted: June 28, 2019 Published: June 28, 2019 A

DOI: 10.1021/acs.analchem.9b02132 Anal. Chem. XXXX, XXX, XXX−XXX

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Analytical Chemistry platform for interesting applications.8 In this work, we have used gallic acid (GA, a green-tea-derived polyphenol) for the single-step synthesis of GA-capped AuNPs (GA-AuNPs) at room temperature. The GA-based synthesis of AuNPs has previously been reported by various groups.19,20 More interestingly, because polyphenols are redox-active, they can be used for the subsequent detection of GA-AuNPs. This is of particular relevance to the exponential increase in the application of nanomaterials in household items as well as the increase in the waste accumulation of nanomaterials. For example, Health Canada,21 the European Union22 (EU), and the U.S. Environmental Protection Agency23 (U.S. EPA) are paying close attention to the fate, transport, and health effects of NPs in the environment. Numerous research groups have reported extensively on the health and environmental toxicity effects of nanomaterials such as AuNPs.24−29 Therefore, the use of GA-AuNPs might not only serve as a green nanotechnology approach but could also help us detect the AuNPs, should they be released into the environment. Electrochemical techniques are particularly well suited for the detection and characterization of polyphenol-capped AuNPs, in this case, GA-AuNPs. Electrochemical sensors for the direct detection of AuNPs using the oxidation, reduction, and electrocatalytic signal of gold have previously been reported.30−32 Electrochemical techniques offer several advantages, including high sensitivity, high selectivity, rapid measurements, cost-effectiveness, and ease of use. The modification of electrode surfaces with nanomaterials,33 polymer films,34,35 nanoparticles,36,37 nanocomposites,38−40 and self-assembled monolayers41−43 (SAMs) is a popular approach to improving the performance of electrochemical sensors. For example, Wang et al.44 have reported carbon nanotube (CNT)-modified glassy carbon electrodes (GCEs) for the detection of phenolic compounds. NP-modified electrodes have also been developed to detect various analytes. For instance, a GCE modified with AuNPs has been reported for the nonenzymatic detection of glucose.45 Nanocomposite electrodes that incorporate metallic nanoparticles with nanomaterials such as reduced graphene oxide (rGO) and multiwalled carbon nanotubes (MWCNTs) have also been reported for various sensing applications.46−49 Nanomaterialmodified electrodes are able to offer improved performance for reasons such as the improvement in the electron transfer kinetics that occurs at the interface of these electrodes, increases in the electrochemically active surface area, electrocatalytic effects facilitated by the nanomaterials, and changes in the surface geometry.33,37,50,51 In this article, we have successfully developed MWCNTrGO nanocomposite-modified GCEs and deployed them for the electrochemical detection of GA-AuNPs using differential pulse voltammetry (DPV). Furthermore, we have also demonstrated the applicability of this sensor for detecting GA-AuNPs in relevant real sample matrixes such as in human serum and Highland Creek water (Scarborough, ON).

to 4.0−7.0 with KOH. All solutions were prepared with ultrapure Milli-Q water (dH2O) with a resistivity of 18.3 MΩ cm. Synthesis and Characterization of Gallic Acid-Capped Gold Nanoparticles (GA-AuNPs). GA-AuNPs were synthesized at room temperature by adding HAuCl4·3H2O solution (16.66 mM stock) to GA (10 mM) in a 10 mL total reaction volume under a blanket of nitrogen gas with vigorous stirring and protection from light. The GA stock solution (100 mM) was kept in amber vials and deoxygenated by purging with nitrogen gas to suppress the auto-oxidation of GA. Depending on the volume of HAuCl4·3H2O stock solution added, colored GA-AuNPs solutions of various sizes were obtained (Supporting Information Figure S1). Following this, the GA-AuNPs were centrifuged at 2000 rcf for 10 min and supernatant was discarded to remove unreacted GA. This centrifugation step was repeated once more using dH2O. The GA-AuNPs pellets were resuspended with dH2O at the required concentrations and subsequently characterized using UV−vis spectrophotometry, transmission electron microscopy (TEM), Fouriertransform infrared spectroscopy (FT-IR), and Fourier-transform Raman spectroscopy (FT-RAMAN). GA-AuNP sizes were estimated from TEM images using ImageJ software and further confirmed using theoretical calculations. UV−Vis Spectrophotometry. UV-vis was carried out using a Synergy H1 microplate reader (Biotek, Mississauga, ON). An aliquot (100 μL) of each GA-AuNP sample was placed in a 96well plate, and a UV−vis scan was performed from 300 to 750 nm at a resolution of 5 nm. The instrument was operated with Gen5 2.0 software (Biotek, Mississauga, ON). TEM. An aliquot of a GA-AuNP sample was spotted onto a UV-activated carbon-coated copper mesh grid (Electron Microscopy Sciences, Hatfield, PA) for 15 min and blot dried. The samples were then imaged on a Hitachi H-7500 transmission electron microscope (Hitachi Ltd., Tokyo, Japan). FT-IR. Data were obtained using an α-p FT-IR spectrometer (Bruker Corp., Billerica, MA), equipped with a deuterated lanthanum α-alanine-doped triglycine sulfate (DLaTGS) detector. GA and GA-AuNPs were mixed with dry potassium bromide (KBr) powder and pelleted, and FT-IR spectra were obtained in transmission mode. FT-IR spectra were obtained at a resolution of 2 cm−1 at a scan rate of 16, between 4000 and 375 cm−1. FT-IR spectra were baseline corrected and smoothed once with an average of 17 data points. The instrument was operated with Opus 6.5 software (Bruker Corp., Billerica, MA). FT-Raman. Spectra were obtained using a multiRAM FTRaman spectrometer (Bruker Corp., Billerica, MA) equipped with a Nd:YAG laser operating at 1064 nm with a maximum laser power of 1 W and a liquid-nitrogen-cooled highsensitivity Ge diode detector. An aliquot of GA-AuNPs was placed in a cuvette with a reflective back mirror, and FTRaman spectra were obtained at a resolution of 1 cm−1 with 16 scans over a Raman shift range of 3600−100 cm−1. A laser output power of 0.15 W was used, which was low enough to prevent possible laser-induced sample damage while providing a high signal-to-noise ratio. The instrument was operated with multiRAM software (Bruker Corp., Billerica, MA). Preparation and Characterization of MWCNT-rGO Nanocomposite-Modified GCE. MWCNT-rGO-GCE electrodes were constructed by drop-casting a MWCNT-GO solution (0.2 mg/mL) onto the GCE surface, followed by UV

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EXPERIMENTAL SECTION Materials. Gallic acid (GA) monohydrate, HAuCl4·3H2O, potassium ferrocyanide, potassium ferricyanide, multiwalled carbon nanotubes (diameter 110−170 nm, length 5−9 nm, 90+% purity), graphene oxide (GO), and human serum were purchased from Sigma-Aldrich (Oakville, ON) and used under ethical guidelines. Phosphate buffer solution (PBS) was prepared from H3PO4 (0.1 M); the pH range was adjusted B

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Figure 1. (A) UV−vis absorption spectra of GA-AuNPs synthesized at various concentrations of HAuCl4·3H2O: (a) 167, (b) 250, (c) 333, (d) 375, (e) 500, (f) 537, and (g) 666 μM at pH 7. (B) TEM images of various sizes of GA-AuNPs: (A) 60−70 nm and (B) 54 nm (c, d) 50 nm with scale bars indicating 200 nm. (C) FT-Raman and (D) FT-IR spectra of gallic acid solid crystals (blue) and a GA-AuNPs solution (orange). FTRaman spectra were obtained at a resolution of 1 cm−1 with 16 scans over a Raman shift range of 3600−100 cm−1

surface at −1.2 V for 2 min. The electrode surfaces were subsequently air-dried under a N2 atmosphere and imaged using a Hitachi S530 scanning electron microscope (Hitachi Ltd., Tokyo, Japan). TEM. A MWCNT-rGO nanocomposite sheet was gently placed on a UV-activated carbon-coated copper mesh grid (Electron Microscopy Sciences, Hatfield, PA) for 10 min and blot dried. The samples were then imaged on a Hitachi H-7500 transmission electron microscope (Hitachi Ltd., Tokyo, Japan).

irradiation for 20 min to result in a direct UV-mediated reduction of GO onto the surface. Following this, further electrochemical reduction of MWCNT-rGO was carried out at −1.2 V for 10 min at pH 3. This resulted in stable MWCNTrGO nanocomposite sheet formation on the GCE surface. Characterization of the MWCNT-rGO-modified GCE was carried out using cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), scanning electron microscopy (SEM), and transmission electron microscopy (TEM). Electrochemical measurements were performed with a μAutolab PGSTAT 128N (EcoChemie, Utrecht, The Netherlands) potentiostat/galvanostat controlled by NOVA 2.1.2 software. All electrochemical measurements were carried out with a conventional three-electrode cell at room temperature. A platinum wire counter electrode and a Ag/AgCl reference electrode were used in connection with a modified GCE as the working electrode. Electrochemical impedance spectroscopy (EIS) measurements were performed in 2.5 mM [Fe(CN)6]3−/4− prepared in 0.1 M KCl over a frequency range of 0.1 Hz to 10 kHz with 0.02 V amplitude (rms). CV measurements were performed in 2.5 mM [Fe(CN)6]3−/4− prepared in 0.1 M KCl at a scan rate of 50 mV/s. A Metrohm Titrando pH meter (model 888) was used for pH measurements. SEM. Measurements on the modified electrode surfaces were performed by directly electrodepositing gold (from a 4 mM HAuCl3 solution) onto the nanocomposite electrode

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RESULTS AND DISCUSSION GA-AuNPs were synthesized using a facile single-pot synthesis method at room temperature. The characterization of GAAuNPs using UV−vis (Figure 1A) and TEM (Figure 1B) revealed that a range of AuNP sizes were obtained depending on the concentration of HAuCl4·3H2O solution added during the synthesis (Supporting Information Figure S1). From the UV−vis spectra, it was observed that the size of the GA-AuNPs decreased as the concentration of HAuCl4·3H2O solution increased. At lower concentrations (Figure 1A, a−c), the UV− vis spectra exhibited a red shift and a broader peak, confirming that the AuNPs were larger in size. Previously, UV−vis studies have shown that as the size of AuNPs increased, they exhibited a red shift,52 and as the size of AuNPs decreased, they exhibited a blue shift.53 C

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Figure 2. Differential pulse voltammograms of GA-AuNPs (85 pM) at MWCNT-rGO-GCE at (a) pH 4, (b) pH 5, (c) pH 6, and (d) pH 7. DPV measurements were performed in 100 mM PBS at an amplitude of 25 mV and a step potential of 5 mV. Scans were carried out from +0.6 V to −0.2 V vs Ag/AgCl.

crystals were absent in the GA-AuNPs solution, further indicating that the majority of the GA present on the surface of the AuNPs was present in its oxidized form.56 Finally, as previously reported by Manjumeena et al.,57 the peak at 1635 cm−1 in the GA-AuNPs IR spectrum confirmed that the GAAuNPs synthesized using GA were stable under these experimental conditions. Following the characterization of GA-AuNPs, electrochemical studies were carried out for the sensitive detection of GA-AuNPs. First, pH studies were carried out to determine the ideal pH for the detection of GA-AuNPs at a GCE. This was done by performing DPV of GA-AuNPs in buffer solutions at pH values of 4−7 (Figure 2). As shown in Figure 2, as the pH of the medium increased, the peak reduction potential shifted toward less-positive values, indicating the involvement of protons in the electroreduction processes. A plot of peak cathodic potential (Ep) vs pH (Supporting Information Figure S2A) showed a linear relationship with pH and shifted to negative potentials, which is in agreement with the theoretical slope of 0.059 V/pH (where m and n are the numbers of protons and electrons, respectively). According to this result, it can be concluded that the reduction process of GA-AuNPs has a slope of 0.065 V/pH and shows that the redox reaction for GA-AuNPs contains equal m and n values. As Au(III) was reduced to form AuNPs, GA was oxidized to its quinonic form. The proposed process for the detection of GA-AuNPs is attributed to the reduction of the oxidized form of GA, as shown in Scheme 1. As shown in Figure 2, GA-AuNPs (in PBS) at pH 5 gave the highest response in terms of the cathodic peak current obtained

TEM analysis revealed that the GA-AuNPs had a similar size distribution (representative images shown in Figure 1B). For example, at a low HAuCl4·3H2O concentration of 250 μM (sample b), gold nanourchins were obtained that ranged in size from 60 to 70 (Figure 1B, a). At higher HAuCl4·3H2O concentrations of 500 μM (sample e) and 537 μM (sample f), smaller spherical GA-AuNPs were obtained, with average sizes of 54 nm (Figure 1B, b) and 50 nm (Figure 1B, c), respectively. In particular, at a concentration of 537 μM (sample f), a monodisperse solution of spherical GA-AuNPs (50 nm) was obtained with a sharp absorption peak at 535 nm and therefore was chosen as the ideal concentration for the synthesis of GA-AuNPs for further characterization and electrochemical studies. Nanoparticles sizes were determined from the UV−vis and TEM data as reported by Liu et al.54 To confirm that the AuNPs were capped with GA, FTRaman and FT-IR analyses were carried out. FT-Raman is an especially attractive technique for confirming the adsorption of chemical species onto metallic nanoparticle surfaces because they undergo a surface-enhanced Raman scattering (SERS) effect. The Raman band of the chemical species is amplified by several orders of magnitude when adsorbed onto metallic NPs.55 As shown in the Raman spectra in Figure 1C, a SERS effect was observed when GA-AuNPs were subjected to Raman analysis, suggesting that the AuNPs were capped with GA. FTIR analysis was also carried out on solid crystals of GA and dry GA-AuNPs. As shown in Figure 1D, the IR spectrum of GAAuNPs showed an intense, broad band at around 3300 cm−1 (O−H stretching). This broad band can also be attributed to the intermolecular hydrogen-bonded network of phenolic groups on the surface of GA-AuNPs. The binding of the carboxylic moiety of GA to the NP surface was confirmed by the disappearance of the broad band between 2920 and 3080 cm−1, which is characteristic of O−H stretching vibrations of carboxylic groups. Furthermore, the CO stretching frequency observed at 1695 cm−1 in the IR spectrum of GAAuNPs was observed following the nanoparticle synthesis reaction, suggesting that GA is present in its oxidized form on the surface of AuNPs. Moreover, the peaks at 1612, 1538, 1467, and 1335 cm−1 (corresponding to aromatic CC and phenolic C−O vibrations) seen in the IR spectrum of GA solid

Scheme 1. Proposed Electrochemical Reduction of the Oxidized Form of Gallic Acid

D

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Figure 3. (A) SEM image of the MWCNT-rGO-modified GCE electrode. The scale bar represents 5 μm. (B) TEM image of the MWCNT-rGOmodified GCE electrode. The scale bar represents 1 μm.

(Supporting Information Figure S2B) and the peak shape; therefore, pH 5 was chosen as optimal for the detection of GAAuNPs using DPV. To improve the sensitivity of the detecting electrode, the GCE was then modified with an MWCNT-rGO nanocomposite. This was performed using a combination of physical58 (UV irradiation) and electrochemical reduction59 as outlined in the methods section. The MWCNT-rGOmodified GCE was subsequently characterized using SEM and TEM (Figure 3). SEM and TEM images revealed that the MWCNTs were incorporated at various orientations within the nanocomposite structure, resulting in a structurally stable nanocomposite sheet. In addition, MWCNT-rGO-GCE electrodes were also characterized using electrochemical methods such as CV (Figure 4A) and EIS (Figure 4B). CV studies showed an increase in the Faradaic responses for the MWCNT-rGO-GCE compared to bare GCE, suggesting that the electrode modification using the nanocomposite layer resulted in an increase in electron transfer characteristics. Two different equivalent circuits were used to simulate the EIS behavior of the bare and modified electrodes. Circuit I, which was used for bare GCE and MWCNT-rGO-modified GCE, is similar to the Randles equivalent circuit, whereas the ideal elements for double-layer capacitance and Warburg elements are replaced by R1 and the solution resistance is shown by RS. Values for RS and Q1 are within a reasonable range, and the N value is close to 0.5, which is the ideal value for a Warburg element. The values for Qdl and R1, which are of more interest, are quite different for bare GCE and MWCNTrGO-modified GCE. The Q1 value for MWCNT-rGOmodified GCE is almost 10 times that of the bare GCE. Considering eq 1, which applies to ideal capacitors, this difference could be attributed to a very large microscopic area for the nanocomposite-modified electrode C = εε0A /d

Figure 4. (A) Cyclic voltammograms of bare GCE, MWCNT-GCE, rGO-GCE, and MWCNT+rGO-GCE at a scan rate of 50 mV/s in 2.5 mM [Fe(CN)6]3−/4− with 0.1 M KCl. (B) Nyquist plots of bare GCE, MWCNT-GCE, rGO-GCE and MWCNT-rGO-GCE in 2.5 mM [Fe(CN)6]3−/4− with 0.1 M KCl over a frequency range from 0.1 Hz to 100 kHz.

structure and also the presence of transition-metal ions in CNTs.62 On the other hand, MWCNT-only modified GCE and rGO-only modified electrodes exhibit more complicated behavior in EIS. The electrode behavior was simulated using a circuit with three electrical pathways instead of the two pathways that are used in a Randles equivalent circuit, as the simplest one for this purpose. Although Qdl still represents the double-layer capacitance, the other two pathways were not easy to interpret in terms of attributing the EIS behavior of the electrodes to the physical picture of the surface. This complex behavior was attributed to the fact that these two surfaces may behave like a porous medium which creates many different pathways for electrons to reach the electroactive species in the

(1)

where C is the capacitance, ε is the dielectric constant of the material, ε0 is the vacuum permittivity, A is the area of the electrode, and d is the distance between the electrodes. The observations are in good agreement with our knowledge of CNTs and rGO.60,61 The value for R1 is 10-fold smaller for MWCNT-rGO modified GCE, which suggests a smaller charge transfer resistance compared to that of the bare GCE. These two observations were also in agreement with our CV results, where the amount of charge for MWCTN-rGO-modified GCE is much larger and ΔEpeak is smaller than that for bare GCE. These results were attributed to the larger area for microscopic E

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Figure 5. EIS-based studies for the surface modifications using Nyquist and Bode plots with the proposed equivalent circuits to fit the experimental data for (A) bare GCE, (B) rGO-modified GCE, (C) MWCNT-modified GCE, and (D) rGO-MWCNT-modified-GCE.

the real and imaginary impedance values, we can express the following

solution by penetrating the porous medium from the solid bed of the electrode. We hypothesized that the relatively large resistances are due to partial contact between the particles or nanotubes in those media, which can be imagined by considering eq 2, which applies to ideal resistors R = ρl/A

C(ω) = C(ω) − jC″(ω)

(3)

C′(ω) = −Z″(ω)/ω |Z(ω)|2

(4)

C″(ω) = Z′(ω)/ω |Z(ω)|2

(5)

(2)

where R is the resistance, ρ is the resistivity, l is the length, and A is the cross-sectional area of the resistor. Obviously, a smaller contact surface (A) will generate a larger resistance. This could be the case for the contact points of MWCNTs and rGO in the respective electrodes. We imagine that the combination of MWCNTs and rGO produces a uniform and better-packed medium compared to a medium which shows one pathway with a large surface area. To investigate these phenomena in detail, other microscopic tools should be employed, which is out of the scope of this work. It should also be noted that the relatively large ΔEpeak values for MWCNT-only modified GCE and rGO-only modified GCE are also in agreement with our EIS observations, as shown in Figure 4A. The values of the various equivalent circuit elements following EIS fitting are also summarized in Supporting Information Table S1. To render a better understanding of the results, impedance data was analyzed in terms of the complex capacitance.63 Using

where Z′(ω) and Z′′(ω) are the real and imaginary components of the complex impedance Zω and ω is the angular frequency ω = 2πf. At low frequencies, C′(ω) corresponds to the capacitance of the electrode material and C′′(ω) corresponds to the dissipation of energy by irreversible processes that lead to hysteresis.64 Data in terms of a Bode modulus plot is shown in Supporting Information Figure S5, showing the same trend as the Nyquist C′′ vs C′ plot. Supporting Information Figure 5S shows circular path behavior for all of the modified GCEs (MWCNTs only, rGO only, and MWCNTs-rGO) as well as the bare GCE, which is a general feature of the dissipative capacitive electrode response. Diameters of circles are increased because of increased capacitance values for bare GCE and then modified GCEs with rGO, MWCNTs, and MWCNTs-rGO. Data in Supporting Information Figure S5 F

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Figure 6. Differential pulse voltammograms of MWCNT-rGO-modified GCEs in 0.1 M PBS (pH 5) at various concentrations of GA-AuNPs ranging from 0 to 329 pM. DPV measurements were performed at an amplitude of 25 mV and at a step potential of 5 mV. Concentrations of GAAuNPs were plotted against the cathodic peak current (Ipc) signals to generate a calibration curve.

illustrates the gain in capacitance obtained from the different modifiers on the GCE for rGO, MWCNTs, and MWCNTsrGO in comparison to bare GCE, with values of C, evaluated at 0.1 Hz, obtained as 0.40 mF/cm2 for bare GCE, 0.55 mF/cm2 for MWCNTs only, 0.84 mF/cm2 for rGO only, and 3.05 mF/ cm2 for MWCNTs-rGO. On the basis of these results, the C for MWCNT-rGO-GCE is approximately 8 times the values of the bare GCE, 1.5 times the values of GCE/MWCNTs, and 2 times the values of rGO-GCE, which is in good agreement with those obtained from EIS. MWCNT-rGO-modified GCEs were then deployed for the detection of GA-AuNPs. As shown in Supporting Information Figure S3B, MWCNT-rGO modification of GCE resulted in an approximately 4-fold increase in current response at the working electrode compared to the bare GCE. More interestingly, with the nanocomposite modification, the detectable concentration of GA-AuNPs was significantly improved. A calibration curve was constructed for the determination of GA-AuNPs using the MWCNT-rGO-GCE electrode (Figure 6). Control studies were also performed using citrate-AuNPs as shown in Supporting Information Figure 3A. The calibration curve was linear up to a GA-AuNPs concentration of 329 pM with a detection limit of 2.57 pM (LOD = 3σblank/m, where m is the slope of the calibration curve). The calibration curve consisted of two linear segments, from 29 to 125 pM and another linear segment from 145 to 329 pM with differences in the slopes for the calibration curve with low and high concentrations of the analytes. At lower analyte concentrations, because of a high number of active sites, the slope of the first linear segment of the calibration is high. However, at higher analyte concentrations, because of decreasing active sites, the slope of the second linear segment of the calibration curve decreased. To examine the applicability of MWCNT-rGO-modified GCEs at detecting GA-AuNPs in real-life samples, the performance of the nanocomposite electrodes was tested in various matrixes such as human serum and creek water (Highland Creek, Scarborough, ON). Studies with these real samples were performed strictly under the ethical guidelines and code of the University of Toronto. As shown in Table 1, MWCNT-rGO-GCE was successfully applied for the detection of GA-AuNPs in both analytical matrixes tested. Recovery was calculated using eq 6:

Table 1. Detection of GA-AuNPs in Human Serum and Creek Water Samples (n = 3)a sample creek water

human serum

spiked (pM)

detected ± SD (pM)

45 125 230 300 60 125 220 250

42.5 126.8 229.6 300.6 55.8 123.6 207.1 234.3

± ± ± ± ± ± ± ±

recovery (%)

1.8 2.5 3.8 6.2 2.0 3.2 4.2 4.4

94.4 101.4 99.8 100.2 92.8 98.9 94.2 93.7

“Spiked” is the amount of standard solution that was added to the sample. “Detected” is the amount of spiked analyte that was determined from the spiked solution.

a

% recovery =

Cspiked sample − Cunspiked sample Cadded

× 100

(6)

The repeatability of the MWCNT-rGO-modified GCE electrodes was investigated using DPV for repetitive (n = 10) measurements of GA-AuNPs (Supporting Information Figure S6). The relative standard deviation was calculated to be 1.03%. The reproducibility of the modified electrodes was studied using five independent GCEs prepared with the same nanocomposite composition, and the corresponding cathodic peak currents were obtained. The relative standard deviation of the modified electrodes was calculated to be 2.16% (n = 5). The long-term stability of the modified electrodes was also investigated by recording the differential pulse voltammograms of the modified GCEs (n = 5) for a period of 1 month. During this time, the modified GCEs were stored at room temperature and their responses were recorded twice a week. The Ipc signals were retained at 98.3%, suggesting that the modified GCEs demonstrated excellent stability.

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CONCLUSIONS We have demonstrated a highly sensitive electrochemical technique to detect GA-AuNPs using MWCNT-rGO hybrid nanocomposite GCEs. We have also demonstrated that the developed sensor has a detection limit of 2.57 pM for the detection of GA-AuNPs. With the exponential increase in applications of nanomaterials in daily used products, the development of sensitive analytical methods to detect them is G

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(project no. 35272), a Discovery Grant (project no. 3655) from the Natural Sciences and Engineering Research Council of Canada (NSERC), and the Canada Foundation for Innovation (project no. 35272). We thank Prof. Meissam Noroozifar (Department of Physical and Environmental Sciences, University of Toronto at Scarborough) for his valuable guidance with complex capacitance analysis and Dr. Bob Temkin (Centre for the Neurobiology of Stress, University of Toronto at Scarborough) for his technical assistance with electron microscopy.

of imminent concern to reduce public health and environmental issues related to acute and/or chronic exposure to nanomaterials. Polyphenol-coated AuNPs are finding novel applications in fields such as nanobiotechnology and environmental technology. For example, polyphenol-capped AuNPs synthesized using Taxus baccata extract have been used as a potent anticancer agent.65 Fazal et al.66 have reported polyphenol-capped AuNPs synthesized from the cocoa plant for photothermal cancer therapy. Our developed hybrid nanocomposite electrochemical sensor can be deployed to detect such polyphenol-capped AuNPs in biologically relevant matrixes. The toxicity of engineered nanoparticles that are deployed for environmental remediation efforts is also becoming an area of emerging concern.28 For example, catechin-capped AuNPs have been developed for the detection of lead in environmental water samples.67 We envision that our developed electrochemical approach can aid in the detection of such polyphenol-capped AuNPs in biological and environmental samples. This work forms the basis of an ongoing project in our laboratory for the future development of electrochemical sensors for the detection of other metallic NPs such as silver (AgNPs), iron (FeNPs), and cobalt (CoNPs) that are capped with redox-active polyphenols.

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ASSOCIATED CONTENT

* Supporting Information S

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.analchem.9b02132. Image of GA-AuNPs synthesis; plot of cathodic peak potential vs pH; plot of peak current response vs pH; DPVs of citrate-capped AuNPs (control) and gallic acid; DPVs of GA-AuNPs at bare and nanocompositemodified electrodes; DPVs of gallic acid and GAAuNPs at the nanocomposite electrode; equivalent circuit elements following EIS fitting; Bode−Bode plots; DPVs of repeatability studies; and the electrochemical mechanism of gallic acid (PDF)

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REFERENCES

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AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. Tel: +1 416 287 7249. ORCID

Kagan Kerman: 0000-0003-3511-8570 Author Contributions

H.V.S.G. and K.K. conceived the experiments. H.V.S.G. performed all experiments and analyzed all data. B.R.P. jointly performed UV−vis and FT-IR experiments. H.F. performed the fitting and interpretation of EIS data. H.V.S.G, A.M.C., and K.K wrote the manuscript. Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS H.V.S.G. gratefully acknowledges a Ph.D. award from Queen Elizabeth II Graduate Scholarships in Science and Technology (QEII-GSST). B.R.P. gratefully acknowledges a Ph.D. award from Ontario Graduate Scholarship (OGS). K.K. acknowledges financial support from the Canada Research Chair Tier-2 award for “Bioelectrochemistry of Proteins” (project no. 950231116), the Ontario Ministry of Research and Innovation H

DOI: 10.1021/acs.analchem.9b02132 Anal. Chem. XXXX, XXX, XXX−XXX

Article

Analytical Chemistry

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DOI: 10.1021/acs.analchem.9b02132 Anal. Chem. XXXX, XXX, XXX−XXX