InorganicâOrganic Hybrid 3D Redox ... - ACS Publications

Research Article pubs.acs.org/journal/ascecg

Inorganic−Organic Hybrid 3D Redox Nanoarchitecture for the Electrocatalytic Sensing of Thiols Bhaskar Manna and C. Retna Raj* Functional Materials and Electrochemistry Laboratory, Department of Chemistry, Indian Institute of Technology Kharagpur, Kharagpur−721302, West Bengal, India S Supporting Information *

ABSTRACT: We demonstrate the development of a novel redox active three-dimensional (3D) functional hybrid nanoarchitecture based on reduced graphene oxide (rGO) and sol−gel derived silicate network for the mediated electrocatalytic sensing of biological thiols. First, the thiol functionalized hybrid material was synthesized by the cross condensation reaction between graphene oxide (GO) and the in situ generated silanol derived from 3-(mercaptopropyl)trimethoxysilane (MPTS). The chemisorption of the hybrid material on Au electrode and the subsequent NaBH4 treatment yield the thiol-terminated 3D hybrid inorganic−organic self-assembly. The 3D hybrid assembly was further tailored with a redox active 4-methyl catechol (MCA) or catechol (CA) moiety by taking advantage of the bias-driven Michael addition reaction between the surface-confined thiol-terminated self-assembly and electrogenerated quinone. The Michael addition of quinone was quantitatively monitored by electrochemical quartz crystal microbalance (EQCM). The redox-tailored architecture displays reversible voltammetric response corresponding to the redox reaction of catechol/quinone redox couple with standard heterogeneous rate constant of 75.74 s−1 (for CA) and 71.43 s−1 (for MCA). The MCA-tailored 3D hybrid architecture efficiently mediates the oxidation of biological thiols (cysteine, homocysteine, and glutathione). However, the CA-tailored hybrid architecture favors the Michael addition of biological thiols, as evidenced by EQCM studies. The kinetics for the mediated electrocatalytic oxidation of biological thiols on MCA-tailored assembly was studied using a rotating disk electrode. The rate constant for the oxidation of cysteine is higher than those of the other two thiols. The MCA-tailored hybrid assembly is highly sensitive, and we could achieve a very low detection limit of 0.3 nM in flow injection analysis at the potential of 270 mV. The electrode is highly stable, and only 10% decrease in the initial amperometric current was observed after 7 days. KEYWORDS: Reduced graphene oxide, Inorganic−organic hybrid material, Michael addition, Biological thiols, Mediated oxidation, Electrocatalytic sensing

■

materials.10,11 The large surface area, 3D porous structure, enhanced electronic and ionic conductivity, and interface dominated properties favor efficient charge transport. The sol−gel process provides a versatile direct approach to produce a variety of hybrid materials with tunable functional properties for potential applications. Depending on the possible interactions connecting the inorganic and organic species, hybrid materials are generally classified into two categories, viz., class I and class II. Composite with weak interactions between the constituents such as van der Waals, hydrogen bonding, or weak electrostatic interactions is referred to as a class I hybrid. On the other hand, components of class II hybrid are interconnected by strong chemical interactions such as covalent bonds.12 Although the silica-based hybrid materials are being exploited for energy conversion and storage and electrochromic and corrosion protection applications, the majority of the works

INTRODUCTION Development of novel 3D functional hybrid materials is of significant interest for various applications including photovoltaics, electrocatalysis, bioanalysis, energy storage, etc.1−4 The properties of hybrid materials are not only based on the constituents but very often have new features depending on the nature of the constituents. The inorganic−organic hybrid materials offer attractive opportunities for many applications. The fascinating multifunctional nature of these hybrid materials makes them ideal for the fabrication of various functional devices ranging from biosensors to supercapacitors and batteries. Highly sensitive and stable amperometric biosensors for glucose, hydrogen peroxide, ammonia, etc., have been developed using sol−gel derived hybrid materials.4,5 Development of high performance energy storage devices using covalent and noncovalent inorganic−organic hybrid materials has also been demonstrated.6−9 The coexistence of electric and magnetic ordering and the paraelectric-to-ferroelectric phase transition have also been observed in functional hybrid © 2017 American Chemical Society

Received: July 25, 2017 Published: August 25, 2017 9412

DOI: 10.1021/acssuschemeng.7b02523 ACS Sustainable Chem. Eng. 2017, 5, 9412−9422

Research Article

ACS Sustainable Chemistry & Engineering Scheme 1. Schematic Representation of the Synthesis of GO-MPTS

hybrid assembly is highly sensitive, stable and it could detect the biological thiols at subnanomolar level.

are oriented toward electroanalytical applications as evidenced by the large number of publications.12,13 The sol−gel derived ceramic−carbon hybrid has interesting properties that can be tuned further according to the analytical requirements. The physically entrapped carbon inside the silicate network actually renders high mechanical stability and excellent electronic conductivity.4 One of the interesting properties of these hybrid material is their capability to encapsulate redox enzymes and retain the enzymatic activity into the 3D network for biosensing applications.14 Graphene is a one atom thick 2D honeycomb carbon network, the mother of all graphitic materials. The mechanical strength, carrier mobility, thermal stability, and flexibility make it a wonder material for electronic applications.15 Graphene oxide (GO), the oxidized form of graphene has variety of reactive oxygen functionalities.16 It can be used as potential materials for various applications ranging from biosensing to drug delivery owing to the presence of chemically tunable surface.16−18 Taking advantage of the oxygen functionalities, the covalent and noncovalent functionalization of small molecules and complex enzymes has been achieved.18 The covalent hybrid material based on sol−gel derived silicate and graphene is expected to have unique properties and would outperform the ceramic−carbon hybrid material. Matsuo et al. for the first time demonstrated the silylation of GO by octyltrichlorosilane and showed that the silylated GO is highly stable in ethanol.19 Ou et al. shown the grafting of hydroxylated octadecyltrichlorosilane and investigated the tribological performances.20 Meng et al. covalently grafted 3-azidopropyl functional moiety onto GO sheet using silicon alkoxide chemistry.21 Recently Li et al. covalently attached 3methacryloxypropyltrimethoxysilane to GO for oil spill cleanup.22 Our group is interested in the development of graphene-based functional hybrid material for electrochemical applications.8,17,23 Herein, we demonstrate the synthesis of inorganic−organic 3D hybrid material based on (3-mercaptopropyl)trimethoxysilane (MPTS) derived silicate network and GO and the bias-driven Michael addition on the hybrid selfassembly for mediated electrocatalytic sensing of biological thiols. The sensing approach involves the step-by-step buildup of the nanoarchitecture using MPTS and GO by taking advantage of the Michael addition reaction between thiolterminated 3D network and electrogenerated o-quinone. The

■

EXPERIMENTAL SECTION

Synthesis of GO-MPTS Hybrid Material. A modified Hummers method was used to synthesize GO from pristine graphite24 (Supporting Information). In a typical procedure for the synthesis of GO-MPTS hybrid material, GO (5 mg) was dispersed in water (10 mL) by sonication for ∼1 h. Then the GO dispersion was deaerated by purging argon gas and an aliquot of MPTS (100 μL) was injected into the GO dispersion in argon atmosphere at room temperature under constant stirring. HCl (100 μL of 0.1 M) was subsequently injected into the above mixture to initiate the hydrolysis and condensation of MPTS with GO. The stirring was continued for 16 h in an argon atmosphere. Then it was centrifuged and the residue was thoroughly washed with water and ethanol. The inorganic−organic hybrid material (GO-MPTS) was dried at 70 °C in vacuum overnight and used for further studies. Self-Assembling of GO-MPTS Hybrid Material on Au Electrode. Polycrystalline Au electrode was used for the selfassembling of GO-MPTS hybrid material. A well-polished and electrochemically cleaned polycrystalline Au electrode was soaked in GO-MPTS (1 mg/mL) dispersion in water for 10 min. The hybrid GO-MPTS self-assembles on the Au electrode through −SH groups by chemisorption. The self-assembly of GO-MPTS was repeatedly rinsed with water and kept in PBS before subjecting to electrochemical experiments. The real surface area of the polycrystalline Au electrode was determined from the charge consumed during the reduction of surface oxides.25 The real surface area of the electrode was used in the calculation of the surface coverage of self-assembled layer and sensitivity and in other cases. The oxygen functionalities of GO in the hybrid self-assembly on Au electrode were partially reduced by NaBH4 treatment. The GO-MPTS modified electrode was soaked in freshly prepared aqueous solution of NaBH4 (1 mM) for 10 min. Then, the electrode was washed with copious amount of water and stored in PBS before subjecting to electrochemical experiments. The hybrid assembly after NaBH4 treatment is referred to as rGO-MPTS. For FTIR, Raman, and TGA measurements, the GO-MPTS hybrid material in water was mixed with required NaBH4 and stirred at room temperature for 10 min. Bias-driven Michael Addition Reaction. The rGO-MPTS hybrid assembly was tailored with redox active CA or MCA by Michael addition reaction. Michael addition reaction was achieved electrochemically. The rGO-MPTS electrode was either biased at 0.5 V or subjected to potential cycling between −0.2 and 0.5 V in aqueous electrolyte containing CA or MCA. The electrochemically generated oquinone undergoes Michael addition reaction on the hybrid selfassembly and yields redox active hybrid nanoarchitecture. An optimum of 10 potential cycles and 1 mM of CA or MCA is required to achieve 9413


Research Article

ACS Sustainable Chemistry & Engineering

Figure 1. (A) FTIR and (B) XRD profiles of GO (a), GO-MPTS (b), rGO-MPTS (c).

epoxy symmetrical ring deformation vibration (1225 cm−1), and alkoxy C−O (1060 cm−1) (Figure 1A). GO-MPTS shows characteristic peaks for CH2 (asymmetric and symmetric stretching at 2925 and 2855 cm−1, respectively), S−H (2555 cm−1), Si−C (1254 cm−1), Si−O−C (1114 cm−1),27 Si−O−Si/ O−Si−O (asymmetric, symmetric stretching at 1030, 795 cm−1, respectively),28,29 C−S−H (bending mode at 885 cm−1),30 and C−S (688 cm−1). More importantly, observable decrease in the peak intensity at 3415 cm−1 was noticed and is ascribed to the cross condensation of the −OH groups of GO moiety with silanol −OH groups. The NaBH4 treatment of GO-MPTS, significantly alters the FTIR spectral features. The intensity of the peak corresponding to −CO moieties of GO at 1725 cm−1 significantly decreased, implying chemical reduction of −CO. The Raman spectra of GO and the hybrid material shows characteristic D and G bands at 1350 and 1593 cm−1, respectively (Figure S1). However, the intensity ratio (ID/IG) for the hybrid material is slightly higher than GO. Such increase can be accounted for presence of new sp3 carbons via C−O−Si linkage with the GO skeleton. When GO-MPTS is reduced to rGO-MPTS, both D and G bands are shifted to lower wavenumber and the ID/IG ratio of these bands increases suggesting the decrease in the average size of the sp2 domain in the carbon network.31 The XRD profile of GO shows characteristic diffraction at 2θ = 9.28° corresponding to (002) plane (Figure 1B). The covalent grafting of GO with MPTS significantly shifts the diffraction to lower angle, indicating that the covalent attachment of MPTS on the GO moiety increases the interlayer distance. The interlayer distance increases from 0.952 to 1.328 nm. In the case of rGO-MPTS, we observed broadening of the peak at lower angle and appearance of broad peak at higher angle presumably due to the partial loss of crystallinity. The height profile of the AFM image of GO shows that it has a thickness of ∼1.2 nm suggesting single layered GO (Figure S2). On the other hand, the GOMPTS hybrid material has a thickness of 5−6 nm. Such increase in the thickness of GO sheet can be ascribed to the covalent condensation of MPTS with GO and the possible interconnection of two GO sheets by silicate network. The TEM images show a wrinkled paper-like rough surface for the hybrid material (Figure S3). The energy dispersive spectral analysis shows characteristic signatures for C, O, S, and Si,

efficient Michael addition reaction on the hybrid self-assembly. The electrodes after Michael addition reaction were washed well with water and stored in PBS before subjecting to further experiments. Electrochemical Studies. All the voltammetric and amperometric measurements were carried out with a CHI643B electrochemical analyzer attached to a current booster (CH Instruments, Austin, TX). A two-compartment three-electrode cell with polycrystalline Au working, platinum wire auxiliary, and Ag/AgCl (3 M KCl) reference electrodes was used in the voltammetric and amperometric measurements. Electrochemical quartz crystal microbalance (EQCM) studies were performed with CHI400A computer controlled time-resolved EQCM (CHI, Austin, TX). AT-cut quartz crystals covered with Au deposited on a Cr layer on the crystals, having fundamental resonant frequency of 8 MHz, were used as the working electrode. The area of quartz crystal covered with Au was 0.196 cm2. The flow injection analysis experiments were performed with a CHI842B dual channel electrochemical detector (CH Instrument, Austin, TX, USA). The flow rate of the mobile phase (PBS of pH 7.2) was controlled by BAS LC peristaltic pump (PM-80) with Teflon tubing. An electrochemical cell with stainless steel auxiliary and dual Au working electrode (3 mm diameter) was used in the flow injection experiments. The thickness of the Teflon gasket was 0.002 in. The sample was delivered to the electrochemical cell at a flow rate of 0.5 mL min−1 from a reservoir. The hydrodynamic voltammetric experiments were performed with an Autolab potentiostat/galvanostat (302N) workstation.

■

RESULTS AND DISCUSSION Synthesis of GO-MPTS Hybrid Material and Characterization. Scheme 1 illustrates the synthesis of GO-MPTS hybrid. The synthetic method involves the hydrolysis of MPTS and cross-condensation with GO. Silicon alkoxides are known to undergo hydrolysis and condensation in aqueous solution of suitable pH to produce sol−gel silicate network.26 GO has large number of −OH functionalities and it can co-condense with the in situ generated silanol as shown in the scheme. It is interesting to note that the condensation can take place on both sides of GO, depending on the availability of −OH groups on the GO surface. Because large number of −OH groups are randomly present on the surface of GO sheet, condensation on both sides cannot be avoided. The cross-condensation reaction of silanol with GO yields 3D hybrid material GO-MPTS with large number of free thiol groups. The properties of this hybrid material are anticipated to be largely different from the sol−gel silicate or GO. The FTIR spectrum of GO shows characteristic signatures for the oxygen containing functionalities such as −OH (3415 cm−1), −CO (1725 cm−1), CC (1620 cm−1), 9414


Research Article


Figure 2. Deconvoluted C 1s (a) and Si 2p XPS (b) profiles of GO-MPTS.

Figure 3. (A) Self-assembly of GO-MPTS on Au electrode. (B) Cyclic voltammetric response of (a) GO-MPTS and (b) rGO-MPTS hybrid assembly on Au electrode in 0.1 M PBS (pH 7.2). Scan rate 100 mV s−1.

subjected to NaBH4 treatment. The 3D hybrid material has plenty of free thiol groups and some of these thiol groups chemisorb on the Au surface and form the self-assembly of GOMPTS (Figure 3A). The Raman spectra of GO-MPTS selfassembly on Au surface shows a characteristic band at 275 cm−1 corresponding to the Au−S vibration (Figure S7).37 The other bands at 357, 710, 796, 880, 990, 1110, and 1715 cm−1 correspond to C−C (bending), C−S (υstr), Si−O−Si/O−Si− O (υsym str), C−S−H (bending mode), Si−O−Si/O−Si−O (υasym str), Si−O−C (υstr), and CO, respectively. The D and G bands appear at 1350 and 1596 cm−1, respectively. The NaBH4 treatment of GO-MPTS self-assembly on the Au surface alters the Raman spectral features. The peak at 1715 cm−1 corresponding to −CO moieties significantly decreased implying chemical reduction of −CO. Figure 3B shows cyclic voltammograms of GO-MPTS and rGO-MPTS hybrid selfassembly on Au surface. It is interesting to note that the NaBH4 treatment of GO-MPTS significantly alters the capacitive properties. GO is known to be an insulator, and the capacitance is significantly less owing to the loss of conjugation. As the NaBH4 treatment partially removes the oxygen containing functionalities and restores the π-conjugation in the carbon network, it enhances the electronic conductivity and hence a significant increase (∼3.5 times) in the capacitance with respect to GO-MPTS was noticed. Moreover, the hybrid rGO- MPTS assembly favors facile diffusion of electrolyte and charge transport across the electrolyte−solution interface (Figure S8). The surface coverage (Γ) of the hybrid assembly on the

originating from the silicate and GO sheets of the hybrid material (Figure S3). The thermogravimetric analysis show characteristic mass change (Figure S4) and the careful analysis of the profile shows 43% grafting of MPTS network with GO. The surface survey XPS profile of GO-MPTS shows signatures for C, O, S, and Si (Figure S5) whereas that of GO shows signatures only for C and O (Figure S5). The deconvoluted C 1s XPS profile of GO (Figure S5) shows peaks centered at 284.45, 286.42, 287.1, 288.12, and 289 eV corresponding to the binding energy of CC/C−C, C−OH, C−O−C, CO, and O−CO functionalities, respectively.32,33 In the case of GOMPTS (Figure 2a), the new peaks observed at 283.8, 285.21, 285.63, and 286.79 eV correspond to C−Si, sp3C, C−S, and C−O−Si, respectively.32,34,35 As discussed earlier, the cross condensation of hydrolyzed MPTS with the GO moiety leads to the formation of C−O−Si linkage. The deconvoluted XPS profile of Si 2p (Figure 2b) shows peaks centered at 101.89 and 102.7 eV corresponding to Si−O−C and Si−O−Si/O−Si−O moieties, respectively36 confirming the formation of Si−O−C linkage. The BET analysis shows that the GO, GO-MPTS, and rGO-MPTS have the surface area of 153.28, 232.94, and 234.57 m2 g−1, respectively. The covalent attachment of MPTS with GO increases the surface area and significantly alters the pore size distribution (Figure S6). The hybrid material has the pore size distribution between 2.1 and 5 nm. Characterization of GO-MPTS Hybrid Assembly on Au Surface. As the main focus of our investigation is to explore the electrochemical applications, the GO-MPTS hybrid material was self-assembled on polycrystalline Au electrode and 9415


Research Article

ACS Sustainable Chemistry & Engineering Scheme 2. Development of GO-MPTS-Based Redox Nanoarchitecture

electrode surface was obtained from reductive desorption and was 2.03 ± 0.18 × 10−10 mol cm−2 (Figure S9). Bias-Driven Michael Addition. The hybrid 3D assembly on the Au electrode was further functionalized with redox active catechol moiety by taking advantage of the facile Michael addition reaction between thiol and quinone (Scheme 2). The GO-MPTS or rGO-MPTS self-assembly on the electrode surface has large number of free −SH groups (mercaptopropyl) and the Michael addition reaction was electrochemically achieved at pH 10 with the electrogenerated o-quinone. Although the exact surface pKa of −SH groups in the hybrid assembly is not known, they are deprotonated under our experimental conditions (vide infra). Note that the bulk pKa of the SH group of mercaptopropionic acid38 and mercaptopropanesulfonate39 lies between 9.9 and 10.3, and it is assumed that our self-assembly has a similar pKa. The o-quinone was generated electrochemically by the oxidation of CA or MCA (Figure 4), and the in situ generated quinone reacts with the thiolate groups at the electrode surface according to Scheme 2.

The electrode after Michael addition reaction shows welldefined reversible voltammetric response at 0.265 V characteristic of surface-confined redox species in neutral electrolyte in the absence of CA or MCA (inset of Figure 4). It is to be noted that MPTS or GO-MPTS or rGO-MPTS assembly shows such characteristic voltammetric response only after cycling the potential between −0.2 and 0.5 V or holding the potential at 0.5 V in electrolyte containing CA or MCA (Figure 4). In order to exclude the possibility of physical adsorption of CA or MCA on the electrode surface, a control experiment has been performed with an unmodified Au electrode. No characteristic voltammetric response was obtained for the unmodified electrode after potential cycling, implying that the voltammetric response obtained for the hybrid self-assembly is not due to the physical adsorption. Actually when the potential of the modified electrode is biased at 0.5 V or cycled between −0.2 to 0.5 V, o-quinone moiety is electrochemically generated and it undergoes Michael addition reaction with thiolate groups of the hybrid self-assembly on the electrode surface. The Michael addition reaction on the hybrid self-assembly depends on pH. The solution pH strongly controls the nature and yield of the Michael addition product. In the case of 4-substituted oquinones the S nucleophiles predominantly react via 1,6Michael addition at pH ≤ 7.40,41 However, 1,4-Michael addition preferentially occurs at pH ≥ 7 as the thiolate is a highly reactive nucleophile.34 In order to understand the role of pH, Michael addition reaction was performed at different pH ranging from 2 to 11. The Michael addition product was electrochemically characterized and it was noticed that the Γ of Michael addition product increases with increase in pH of the electrolyte and attains a maximum of (7.56 ± 0.24) × 10−12 mol cm−2 at pH 10 (Figure S10). As large Γ was obtained at pH 10, all further studies were performed with the redox hybrid nanoarchitecture obtained at this pH. The surface pKa of the −SH group of the hybrid assembly was obtained from the plot of Γ vs pH at which Michael addition reaction was performed and was found to be 9 which is close to the value of 9.9 obtained for mercaptopropanesulfonate.39 Quantitative Estimation of Michael Addition Product. In order to quantitatively estimate the Michael addition product on the electrode surface, EQCM studies were performed. The change in mass on the electrode during Michael addition reaction was monitored with EQCM by measuring the changes in the frequency of the quartz crystal (Supporting Information). The potential of the Au-coated quartz crystal modified with rGO-MPTS self-assembly was cycled between −0.2 to 0.5 V at a scan rate of 100 mV s−1 in pH 10 containing CA or MCA. As

Figure 4. Cyclic voltammogram obtained during the Michael addition reaction on the hybrid self-assembly of rGO-MPTS in 0.1 M bicarbonate buffer of pH 10 containing 1 mM MCA at a scan rate of 100 mV s−1. (inset) Voltammogram obtained for the hybrid assembly in PBS (pH 7.2) after Michael addition reaction. Sweep rate 100 mV s−1. 9416


Research Article


which increases the probability of covalent attachment via Michael addition reaction (Figure S11). Unlike CA, MCA has only one possible position for Michael addition at our experimental condition as one position is occupied with methyl group (Figure S11). The peak current linearly scales with sweep rate implying that the redox species is surface-confined to the electrode surface (Figure S12). The peak potential shifts negatively by ∼60 mV while changing the pH by one unit (Figure S13), suggesting the involvement of electrons and protons in 1:1 ratio. The heterogeneous electron transfer rate constant for the surface-confined redox species was calculated using Laviron’s approach42 and is listed in Table 1. The careful analysis of the electrochemical data presented in Table 1 reveals the vital role of GO layers of the inorganic−organic hybrid architecture. Although the MPTS self-assembly has 3D network, the Γ and ko are 2−3 times less than the hybrid architecture, strongly indicating that the honeycomb carbon framework favors the Michael addition reaction. The kinetics of the redox reaction on rGO-MPTS based architecture is highly facile with respect to the other architectures. It is understood that the covalent integration of the GO framework (i) exposes more number of −SH group for the Michael addition reaction and (ii) enhances the overall electrical conductivity of the sol− gel MPTS network. The 3D architecture and the enhanced electrical conductivity favor the free diffusion of electrolytes and CA/MCA for facile Michael addition reaction on the hybrid assembly. Electrocatalytic Sensing of Biological Thiols. The biological thiols such as CYS, HCYS, and GSH play crucial role in the biological system. They serve as antioxidant, inhibit cellular apoptosis, are involved in the formation of threedimensional complex protein structure and control the cellular redox states.43−45 Monitoring the concentration of these biological thiols is of great importance as their abnormal levels are related to cancer, aging, neurodegenerative, cardiovascular diseases, etc.46−49 Although several analytical methods have been developed in the past, the electrochemical methods received considerable interest. The electrochemical oxidation of these thiols on unmodified electrode requires large overpotential and several attempts have been made to decrease the overpotential.50−54 Achieving high sensitivity and a low detection limit is a challenging task in the development of electrochemical sensor for biological thiols. The electrochemical sensing of biological thiols can be achieved with catechol-like redox molecules in two different reaction pathways:55−57 (i) Michael addition reaction of electrogenerated quinone with thiol and (ii) mediated-electrocatalytic oxidation of thiol by electrogenerated quinone to produce the corresponding disulfide. The second pathway has advantage over the first as the catechol/quinone redox couple can be recycled for any number of times for the catalytic reaction. In the former pathway, the structural integrity of quinone/ catechol would be compromised during the Michael addition and the quinone used in the Michael addition reaction cannot

shown in Figure 5, significant decrease in the frequency was noticed while cycling the potential, implying the increase in the

Figure 5. Frequency−potential response obtained for rGO-MPTS hybrid assembly during the Michael addition reaction. The potential was cycled between −0.2 and 0.5 V in 0.1 M bicarbonate buffer of pH 10 containing MCA (1 mM) at a sweep rate of 100 mV s−1.

mass due to the Michael addition reaction of the electrochemically generated o-quinone moiety with thiol groups of the hybrid self-assembly. The change in the frequency is large in the first 2−3 cycles, and it does not significantly change after 10 cycles. The overall change in frequency after 10 cycles was 14.15 ± 0.41 Hz, corresponding to an increase in the mass of 98.37 ± 2.86 ng cm−2 on the electrode surface for MCA. In the case of CA, the mass change is slightly high (114.29 ± 2.02 ng cm−2) due to the availability of more sites for the nucleophilic attack (Figure S11). No such change in the frequency was observed for unmodified electrode implying that the change in the frequency noticed with the hybrid assembly is due to the chemical reaction of the electrochemically generated o-quinone moiety with the free thiolate groups. The number of CA and MCA molecules reacted with the thiolate groups on the electrode was calculated to be (63.74 ± 1.13) × 1013 and (48.52 ± 1.43) × 1013/cm2, respectively. Redox Electrochemistry of Hybrid Nanoarchitecture. The redox-tailored rGO-MPTS hybrid architecture displays well-defined voltammetric response corresponding to the redox reaction of catechol/quinone couple with ΔEp value of 22 ± 2 mV (inset of Figure 4). Such small ΔEp suggests fast electron transfer kinetics on the electrode surface. All the electrochemical characteristics of the Michael addition product obtained with MPTS, GO-MPTS, and rGO-MPTS assemblies are summarized in Table 1. It is interesting to note that the Γ of the CA-tailored architecture is higher than that of the MCAtailored architecture. The higher Γ value in case of CA is ascribed to the availability of more sites for nucleophilic attack

Table 1. Electrochemical Characteristics of Michael Addition Products Derived from CA and MCA CA

MCA

self-assembly

ΔEp mV

Γ (mol cm2) × 1012

ko, s−1

no. of molecules

ΔEp mV

Γ (mol cm2) × 1012

ko, s−1

no. of molecules

MPTS GO-MPTS rGO-MPTS

30 24 21

3.91 ± 0.31 6.55 ± 0.23 10.29 ± 0.19

35.17 53.25 75.74

2.38 × 1011 3.99 × 1011 6.27 × 1011

33 28 24

2.89 ± 0.47 4.83 ± 0.39 7.56 ± 0.24

33.28 49.76 71.43

1.76 × 1011 2.94 × 1011 4.61 × 1011

9417


Research Article


Figure 6. Cyclic voltammogram and the scheme illustrating the mediated electrocatalytic activity of rGO-MPTS-MCA hybrid redox nanoarchitecture toward CYS in 0.1 M PBS (pH 7.2). Scan rate 10 mV s−1.

be recycled back for further reaction as the reaction site would be permanently blocked. High sensitivity and low detection limit can be achieved in the second pathway, if quinone/ catechol moiety is confined on the electrode surface. Figure 6 displays the voltammetric response of MCA-tailored hybrid nanoarchitecture on Au electrode toward CYS. The dramatic increase in the anodic peak is ascribed to the mediated oxidation of CYS to the corresponding disulfide (Figure 6). The careful analysis of the voltammetric profile shows that though the oxidation peak appears at 270 mV, the onset potential for the oxidation is significantly less positive ( 1 Hz in the first cycle) with the CA-tailored architecture (Figure S16). Although earlier studies suggest the formation of minimal amount of biadduct with CA,58 our extensive EQCM study does not support the formation of such biadduct (Figure S14). Because the MCA-tailored self-assembly mediates the facile oxidation of CYS, all the amperometric sensing experiments have been performed with this self-assembly. Figure 8A is the amperometric response of the MCA-tailored architecture toward CYS. The potential of the electrode was held at 0.27 V and aliquots of CYS were injected at regular intervals. The sensitivity was obtained from the calibration plot and was 62.65 ± 0.82 nA μM−1 cm−2. The electrode has a linear response up to 90 μM with a response time of 11 s. The electrode could detect as low as 20 nM of CYS (S/N = 8) at the potential of 0.27 V in PBS (pH 7.2) (inset of Figure 8A). The analytical response is highly stable and the amperometric response does not change significantly with time. The operational stability of the electrode was investigated by polarizing the electrode at 0.27 V and injecting 20 μM of CYS (Figure S17). No significant change in the current was observed for long time. The electrode retains ∼90% of the amperometric current even after 7 days (Figure S17), supporting the long-term stability of the electrode. To further evaluate the performance of the 9419


ACS Sustainable Chemistry & Engineering obtained from the slope and intercept of the plot. The kcat for CYS is higher than the other two thiols (Figure S19, Table 2).

■

ACKNOWLEDGMENTS

■

REFERENCES

(1) Heo, J. H.; Im, S. H.; Noh, J. H.; Mandal, T. N.; Lim, C.-S.; Chang, J. A.; Lee, Y. H.; Kim, H.; Sarkar, A.; Nazeeruddin, M. K.; Gratzel, M.; Seok, S. I. Efficient Inorganic-Organic Hybrid Heterojunction Solar Cells Containing Perovskite Compound and Polymeric Hole Conductors. Nat. Photonics 2013, 7, 486−491. (2) Liu, B.; Yu, Z.-T.; Yang, J.; Hua, W.; Liu, Y.-Y.; Ma, J.-F. First Three-Dimensional Inorganic-Organic Hybrid Material Constructed From an “Inverted Keggin” Polyoxometalate and a Copper(I)-Organic Complex. Inorg. Chem. 2011, 50, 8967−8972. (3) Kumar, T. A.; Capua, E.; Tkachev, M.; Adler, S. N.; Naaman, R. Hybrid Organic-Inorganic Biosensor for Ammonia Operating under Harsh Physiological Conditions. Adv. Funct. Mater. 2014, 24, 5833− 5840. (4) Wang, B.; Li, B.; Deng, Q.; Dong, S. Amperometric Glucose Biosensor Based on Sol-Gel Organic-Inorganic Hybrid Material. Anal. Chem. 1998, 70, 3170−3174. (5) Chen, X.; Zhu, J.; Tian, R.; Yao, C. Bienzymatic Glucose Biosensor Based on Three Dimensional Macroporous Ionic Liquid Doped Sol−Gel Organic-Inorganic Composite. Sens. Actuators, B 2012, 163, 272−280. (6) El-Kady, M. F.; Ihns, M.; Li, M.; Hwang, J. Y.; Mousavi, M. F.; Chaney, L.; Lech, A. T.; Kaner, R. B. Engineering Three-Dimensional Hybrid Supercapacitors and Microsupercapacitors for High-Performance Integrated Energy Storage. Proc. Natl. Acad. Sci. U. S. A. 2015, 112, 4233−4238. (7) Aradilla, D.; Gao, F.; Lewes-Malandrakis, G.; Muller-Sebert, W.; Gentile, P.; Boniface, M.; Aldakov, D.; Iliev, B.; Schubert, T. J. S.; Nebel, C. E.; Bidan, G. Designing 3D Multihierarchical Heteronanostructures for High-Performance On-Chip Hybrid Supercapacitors: Poly(3,4-(ethylenedioxy)thiophene)-Coated Diamond/Silicon Nanowire Electrodes in an Aprotic Ionic Liquid. ACS Appl. Mater. Interfaces 2016, 8, 18069−18077. (8) Bag, S.; Raj, C. R. Layered Inorganic-Organic Hybrid Material Based on Reduced Graphene Oxide and α-Ni(OH)2 for High Performance Supercapacitor Electrodes. J. Mater. Chem. A 2014, 2, 17848−17856. (9) Reddy, A. L. M.; Gowda, S. R.; Shaijumon, M. M.; Ajayan, P. M. Hybrid Nanostructures for Energy Storage Applications. Adv. Mater. 2012, 24, 5045−5064. (10) Xu, G.-C.; Zhang, W.; Ma, X.-M.; Chen, Y.-H.; Zhang, L.; Cai, H.-L.; Wang, Z.-M.; Xiong, R.-G.; Gao, S. Coexistence of Magnetic and Electric Orderings in the Metal Formate Frameworks of [NH4][M(HCOO)3]. J. Am. Chem. Soc. 2011, 133, 14948−14951. (11) Pato-Doldan, B.; Gomez-Aguirre, L. C.; Bermudez-Garcia, J. M.; Sanchez-Andujar, M.; Fondado, A.; Mira, J.; Castro-Garcia, S.; SenarisRodriguez, M. A. Coexistence of Magnetic and Electrical Order in the New Perovskite-like (C3N2H5)[Mn(HCOO)3] Formate. RSC Adv. 2013, 3, 22404−22411. (12) Walcarius, A. Electrochemical Applications of Silica-Based Organic-Inorganic Hybrid Materials. Chem. Mater. 2001, 13, 3351− 3372. (13) Audebert, P.; Walcarius, A.; Gómez-Romero, P.; Sanchez, C. Electrochemistry of Sol-Gel Derived Hybrid Materials. Functional Hybrid Materials 2004, 173, 172. (14) Rabinovich, L.; Lev, O. Sol-Gel Derived Composite Ceramic Carbon Electrodes. Electroanalysis 2001, 13, 265−275 and references cited therein. (15) Allen, M. J.; Tung, V. C.; Kaner, R. B. Honeycomb Carbon: A Review of Graphene. Chem. Rev. 2010, 110, 132−145.

ASSOCIATED CONTENT

* Supporting Information S

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acssuschemeng.7b02523. Additional details on the reagents and materials, instrumentation, synthesis of GO, Sauerbrey equation, Raman spectra, AFM and TEM images, TGA, EDX, voltammogram for reductive desorption, XPS, pore size distribution plot, voltammetric and impedimetric response toward the Fe(CN)63−/4− redox couple, plot of surface coverage of Michael addition product vs pH, scheme illustrating the possible Michael addition reaction with CA and MCA, cyclic voltammograms of Michael addition product at different scan rate and at different pH values, scheme illustrating the possible biadduct formation, voltammetric response before and after the electrocatalytic sensing of biological thiols, EQCM response during sensing of thiols, amperometric response toward sensing of thiols, plots illustrating the stability, interference free sensing, hydrodynamic voltammograms, and table highlighting the superior performance of the hybrid architecture toward the sensing of biological thiols (PDF)

■

■

This work is supported by grants from Department of Science and Technology (DST) and Council of Scientific and Industrial Research (CSIR), New Delhi, India, and Indian Institute of Technology Kharagpur.

CONCLUSIONS We have demonstrated the development of novel inorganic− organic 3D hybrid assembly of rGO and MPTS for the electrocatalytic sensing of biological thiols. Redox tailoring of the hybrid assembly was achieved by taking advantage of the Michael addition reaction of rationally selected MCA with the surface-confined −SH groups under optimized potential. The 3D hybrid structure of the assembly favors facile Michael addition reaction of electrogenerated quinone. The covalently attached redox couple undergoes facile electron transfer kinetics. MCA-based architecture favors mediated oxidation whereas the CA-based assembly favors Michael addition of biological thiols. The rational selection of the redox mediator promotes the mediated electrocatalytic oxidation of biological thiols. Amperometric sensing of CYS at subnanomolar level has been achieved with the hybrid 3D redox nanoarchitecture. The EQCM technique has been employed to distinguish mediated oxidation of thiol from a Michael addition reaction. Our study shows a new approach for the fabrication of electrochemical sensors based on the hybrid rGO-MPTS 3D assembly. This hybrid assembly can be effectively used for the immobilization of redox enzymes to study the direct electrochemistry and for the development of amperometric biosensors. Our study further demonstrates that the MCA-tailored hybrid assembly is promising for the electrochemical sensing of biological thiols.

■

Research Article

AUTHOR INFORMATION

Corresponding Author

*Fax: +91 3222 282522. E-mail: [email protected]. ORCID

C. Retna Raj: 0000-0002-7956-0507 Notes

The authors declare no competing financial interest. 9420


Research Article

ACS Sustainable Chemistry & Engineering (16) Chen, D.; Feng, H.; Li, J. Graphene Oxide: Preparation, Functionalization, and Electrochemical Applications. Chem. Rev. 2012, 112, 6027−6053. (17) Manna, B.; Raj, C. R. Covalent Functionalization and Electrochemical Tuning of Reduced Graphene Oxide for the Bioelectrocatalytic Sensing of Serum Lactate. J. Mater. Chem. B 2016, 4, 4585−4593. (18) Georgakilas, V.; Otyepka, M.; Bourlinos, A. B.; Chandra, V.; Kim, N.; Kemp, K. C.; Hobza, P.; Zboril, R.; Kim, K. S. Functionalization of Graphene: Covalent and Non-Covalent Approaches, Derivatives and Applications. Chem. Rev. 2012, 112, 6156− 6214. (19) Matsuo, Y.; Tabata, T.; Fukunaga, T.; Fukutsuka, T.; Sugie, Y. Preparation and Characterization of Silylated Graphite Oxide. Carbon 2005, 43, 2875−2882. (20) Ou, J.; Wang, Y.; Wang, J.; Liu, S.; Li, Z.; Yang, S. Self-Assembly of Octadecyltrichlorosilane on Graphene Oxide and the Tribological Performances of the Resultant Film. J. Phys. Chem. C 2011, 115, 10080−10086. (21) Meng, D.; Sun, J.; Jiang, S.; Zeng, Y.; Li, Y.; Yan, S.; Geng, J.; Huang, Y. Grafting P3HT Brushes on GO Sheets: Distinctive Properties of the GO/P3HT Composites due to Different Grafting Approaches. J. Mater. Chem. 2012, 22, 21583−21591. (22) Li, H.; Liu, L.; Yang, F. Covalent Assembly of 3D Graphene/ Polypyrrole Foams for Oil Spill Cleanup. J. Mater. Chem. A 2013, 1, 3446−3453. (23) Dey, R. S.; Raj, C. R. Development of an Amperometric Cholesterol Biosensor Based on Graphene-Pt Nanoparticle Hybrid Material. J. Phys. Chem. C 2010, 114, 21427−21433. (24) Hummers, W. S.; Offeman, R. E. Preparation of Graphitic Oxide. J. Am. Chem. Soc. 1958, 80, 1339−1339. (25) Trasatti, S.; Petrii, O. A. Real Surface Area Measurements in Electrochemistry. Pure Appl. Chem. 1991, 63, 711−734. (26) Brinker, C. J. Hydrolysis and Condensation of Silicates: Effects on Structure. J. Non-Cryst. Solids 1988, 100, 31−50. (27) Scott, A.; Gray-Munro, J. E. The Surface Chemistry of 3Mercaptopropyltrimethoxysilane Films Deposited on Magnesium Alloy AZ91. Thin Solid Films 2009, 517, 6809−6816. (28) Ma, W.-S.; Li, J.; Deng, B.-J.; Zhao, X.-S. Preparation and Characterization of Long-Chain Alkyl Silane-Functionalized Graphene Film. J. Mater. Sci. 2013, 48, 156−161. (29) Finocchio, E.; Macis, E.; Raiteri, R.; Busca, G. Adsorption of Trimethoxysilane and of 3-Mercaptopropyltrimethoxysilane on Silica and on Silicon Wafers from Vapor Phase: An IR Study. Langmuir 2007, 23, 2505−2509. (30) Bandyopadhyay, S.; Dey, A. Convenient Detection of the Thiol Functional Group using H/D Isotope Sensitive Raman Spectroscopy. Analyst 2014, 139, 2118−2121. (31) Stankovich, S.; Dikin, D. A.; Piner, R. D.; Kohlhaas, K. A.; Kleinhammes, A.; Jia, Y.; Wu, Y.; Nguyen, S. T.; Ruoff, R. S. Synthesis of Graphene-Based Nanosheets via Chemical Reduction of Exfoliated Graphite Oxide. Carbon 2007, 45, 1558−1565. (32) Chen, W.; Yan, L.; Bangal, P. R. Chemical Reduction of Graphene Oxide to Graphene by Sulfur-Containing Compounds. J. Phys. Chem. C 2010, 114, 19885−19890. (33) Ma, H.-L.; Zhang, H.-B.; Hu, Q.-H.; Li, W.-J.; Jiang, Z.-G.; Yu, Z.-Z.; Dasari, A. Functionalization and Reduction of Graphene Oxide with p-Phenylene Diamine for Electrically Conductive and Thermally Stable Polystyrene Composites. ACS Appl. Mater. Interfaces 2012, 4, 1948−1953. (34) Nemanick, E. J.; Hurley, P. T.; Webb, L. J.; Knapp, D. W.; Michalak, D. J.; Brunschwig, B. S.; Lewis, N. S. Chemical and Electrical Passivation of Single-Crystal Silicon(100) Surfaces through a TwoStep Chlorination/Alkylation Process. J. Phys. Chem. B 2006, 110, 14770−14778. (35) Li, Z.; Wang, R.; Young, R. J.; Deng, L.; Yang, F.; Hao, L.; Jiao, W.; Liu, W. Control of the Functionality of Graphene Oxide for its Application in Epoxy Nanocomposites. Polymer 2013, 54, 6437−6446.

(36) Zhang, W.; Gu, H.; Li, Z.; Zhu, Y.; Li, Y.; Zhang, G.; Zhang, F.; Fan, X. General Acid and Base Bifunctional Graphene Oxide for Cooperative Catalysis. J. Mater. Chem. A 2014, 2, 10239−10243. (37) Behera, S.; Sampath, S.; Raj, C. R. Electrochemical Functionalization of a Gold Electrode with Redox-Active SelfAssembled Monolayer for Electroanalytical Application. J. Phys. Chem. C 2008, 112, 3734−3740. (38) Danehy, J. P.; Parameswaran, K. N. Acidic Dissociation Constants of Thiols. J. Chem. Eng. Data 1968, 13, 386−389. (39) Suwandaratne, N.; Hu, J.; Siriwardana, K.; Gadogbe, M.; Zhang, D. Evaluation of Thiol Raman Activities and pKa Values Using Internally Referenced Raman Based pH Titration. Anal. Chem. 2016, 88, 3624−3631. (40) Chavdarian, C. G.; Castagnoli, N., Jr. Synthesis, Redox Characteristics, and in Vitro Norepinephrine Uptake Inhibiting Properties of 2-(2-Mercapto-4,5-dihydroxyphenyl)ethylamine (6-Mercaptodopamine). J. Med. Chem. 1979, 22, 1317−1322. (41) Xu, R.; Huang, X.; Kramer, K. J.; Hawley, M. D. Characterization of Products from the Reactions of Dopamine Quinone with NAcetylcysteine. Bioorg. Chem. 1996, 24, 110−126. (42) Laviron, E. General Expression of the Linear Potential Sweep Voltammogram in the Case of Diffusionless Electrochemical Systems. J. Electroanal. Chem. Interfacial Electrochem. 1979, 101, 19−28. (43) Pompella, A.; Visvikis, A.; Paolicchi, A.; Tata, V. D.; Casini, A. F. The Changing Faces of Glutathione, a Cellular Protagonist. Biochem. Pharmacol. 2003, 66, 1499−1503. (44) Chae, H. Z.; Uhm, T. B.; Rhee, S. G. Dimerization of Thiolspecific Antioxidant and the Essential Role of Cysteine 47. Proc. Natl. Acad. Sci. U. S. A. 1994, 91, 7022−7026. (45) Sato, N.; Iwata, S.; Nakamura, K.; Hori, T.; Mori, K.; Yodoi, J. Thiol-Mediated Redox Regulation of Apoptosis. Possible Roles of Cellular Thiols other than Glutathione in T Cell Apoptosis. J. Immunol. 1995, 154, 3194−3203. (46) Lu, S. C. Regulation of Glutathione Synthesis. Mol. Aspects Med. 2009, 30, 42−59. (47) Townsend, D. M.; Tew, K. D.; Tapiero, H. The Importance of Glutathione in Human Disease. Biomed. Pharmacother. 2003, 57, 145− 155. (48) Barnham, K. J.; Masters, C. L.; Bush, A. I. Neurodegenerative Diseases and Oxidative Stress. Nat. Rev. Drug Discovery 2004, 3, 205− 214. (49) Refsum, H.; Ueland, P. M.; Nygard, O.; Vollset, S. E. Homocysteine and Cardiovascular Disease. Annu. Rev. Med. 1998, 49, 31−62. (50) Lee, P. T.; Goncalves, L. M.; Compton, R. G. Electrochemical Determination of Free and Total Glutathione in Human Saliva Samples. Sens. Actuators, B 2015, 221, 962−968. (51) Lin, Q.; Lin, C.; Wu, H.; Batchelor-McAuley, C.; Compton, R. G. Catalytic Single-Particle Nano-Impacts: Theory and Experiment. Poly(vinylferrocene)-Modified Graphene Nanoplatelet Mediated LCysteine Oxidation. J. Phys. Chem. C 2016, 120, 20216−20223. (52) Gong, K.; Zhu, X.; Zhao, R.; Xiong, S.; Mao, L.; Chen, C. Rational Attachment of Synthetic Triptycene Orthoquinone onto Carbon Nanotubes for Electrocatalysis and Sensitive Detection of Thiols. Anal. Chem. 2005, 77, 8158−8165. (53) Hosseini, H.; Ahmar, H.; Dehghani, A.; Bagheri, A.; Tadjarodi, A.; Fakhari, A. R. A Novel Electrochemical Sensor Based on MetalOrganic Framework for Electro-Catalytic Oxidation of L-Cysteine. Biosens. Bioelectron. 2013, 42, 426−429. (54) Devasurendra, A. M.; Zhu, T.; Kirchhoff, J. R. Detection of Thiols by o-Quinone Nanocomposite Modified Electrodes. Electroanalysis 2016, 28, 2972−2978. (55) Bucur, M. P.; Bucur, B.; Radulescu, C. M.; Covaci, O. I.; Radu, G. L. L-Cysteine Determination Based on Tyrosinase Amperometric Biosensors without Interferences from Thiolic Compounds. Anal. Lett. 2010, 43, 2440−2455. (56) Lee, P. T.; Thomson, J. E.; Karina, A.; Salter, C.; Johnston, C.; Davies, S. G.; Compton, R. G. Selective Electrochemical Determi9421


Research Article

ACS Sustainable Chemistry & Engineering nation of Cysteine with a Cyclotricatechylene Modified Carbon Electrode. Analyst 2015, 140, 236−242. (57) Lee, P. T.; Compton, R. G. Selective Electrochemical Detection of Thiol Biomarkers in Saliva using Multiwalled Carbon Nanotube Screen-Printed Electrodes. Sens. Actuators, B 2015, 209, 983−988. (58) Huang, X.; Xu, R.; Hawley, M. D.; Hopkins, T. L.; Kramer, K. J. Electrochemical Oxidation of N-Acyldopamines and Regioselective Reactions of Their Quinones with N-Acetylcysteine and Thiourea. Arch. Biochem. Biophys. 1998, 352, 19−30.

9422


InorganicâOrganic Hybrid 3D Redox ... - ACS Publications

Recommend Documents

InorganicâOrganic Hybrid 3D Redox ... - ACS Publications