Gold Nanoclusters@Ru(bpy)32+-Layered Double Hydroxide Ultrathin

Jul 10, 2015 - Fax/phone: +86 10 64411957. Abstract. Abstract Image. Herein, it is the first report that a cathodic electrochemiluminescence (ECL) res...
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Gold Nanoclusters@Ru(bpy)32+-Layered Double Hydroxide Ultrathin Film as a Cathodic Electrochemiluminescence Resonance Energy Transfer Probe Yingchang Yu,† Chao Lu,*,† and Meining Zhang‡ †

State Key Laboratory of Chemical Resource Engineering, Beijing University of Chemical Technology, Beijing 100029, China Department of Chemistry, Renmin University of China, Beijing 100872, China

‡

S Supporting Information *

ABSTRACT: Herein, it is the first report that a cathodic electrochemiluminescence (ECL) resonance energy transfer (ERET) system is fabricated by layer-by-layer (LBL) electrostatic assembly of CoAl layered double hydroxide (LDH) nanosheets with a mixture of blue BSA−gold nanoclusters (AuNCs) and Ru(bpy)32+ (denoted as AuNCs@Ru) on an Au electrode. The possible ECL mechanism indicates that the appearance of CoAl− LDH nanosheets generates a long-range stacking order of the AuNCs@Ru on an Au electrode, facilitating the occurrence of the ERET between BSA−AuNC donors and Ru(bpy)32+ acceptors on the as-prepared AuNCs@Ru−LDH ultrathin films (UTFs). Furthermore, it is observed that the cathodic ECL intensity can be quenched efficiently in the presence of 6-mercaptopurine (6MP) in a linear range of 2.5−100 nM with a detection limit of 1.0 nM. On the basis of these interesting phenomena, a facile cathodic ECL sensor has successfully distinguished 6-MP from other thiol-containing compounds (e.g., cysteine and glutathione) in human serum and urine samples. The proposed sensing scheme opens a way for employing the layered UTFs as a platform for the cathodic ECL of Ru(bpy)32+.

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using NaS2O8 as a coreagent. Similarly, Chen and his coauthors also reported that triethylamine can greatly improve the ECL emission of AuNCs.21 Hereafter, there has been a growing interest in red AuNC-based ECL due to their remarkable advantages, such as strong catalytic properties, low toxicity, easy labeling, and renewability.22−24 Obviously, red AuNCs with an emission peak at ∼620 nm are unsuitable donors for the cathodic ECL of Ru(bpy)32+. In principle, blue AuNCs with an emission peak at ∼450 nm are ideal donor candidates for the cathodic ECL of Ru(bpy)32+. However, blue AuNCs usually emit quite weak fluorescence with low fluorescent quantum yields.25,26 Nowadays, the aqueous solution of blue AuNCs has not yet been used as ERET donors for the cathodic ECL of Ru(bpy)32+. Therefore, it represents a great challenge to develop an efficient strategy to improve the ERET efficiency between blue AuNCs and Ru(bpy)32+. Layered double hydroxides (LDHs) are a family of layered materials with positively charged metal hydroxide layers and charge-balancing anions.27−29 Researchers demonstrated that highly efficient fluorescence resonance energy transfer (FRET) could occur between two kinds of fluorescence molecules in the

ince it has been discovered in 1966, tris(2,2′-bipyridine)ruthenium(II) (Ru(bpy)32+) as a powerful electrochemiluminescence (ECL) analytical reagent has been widely used in medical diagnostics, light-emitting devices, immunoassays, and food detections.1−3 The anodic ECL reactions of Ru(bpy)32+ are well documented in the literature.4−7 However, their extensive applications are limited because the oxygen evolution during the reaction can decrease anodic ECL intensities and inactivate work electrodes.8 Alternately, considerable effort has been devoted to the development of the cathodic ECL of Ru(bpy) 3 2+ for avoiding the disadvantages of anodic reactions.9−11 However, the cathodic ECL of Ru(bpy)32+ is hardly happened in aqueous solutions owing to the high instability of reaction intermediates.9 To solve these problems, scientists have aroused considerable interest in ECL resonance energy transfer (ERET) between suitable donors (e.g., luminol and quantum dots) and Ru(bpy)32+ acceptors.12−14 Unfortunately, traditional luminol is nonrenewable,15 and quantum dots are toxic.16 Therefore, it is highly essential to explore a novel ERET donor with renewability and excellent biocompatibility. Gold nanoclusters (AuNCs) have been extensively used as fluorescent probes for a variety of analytical applications and biological imaging owing to their excellent photostability, low toxicity, high emission rates, and good water-solubility.17−19 In 2011, Zhu et al.20 first observed ECL emission of red AuNCs © XXXX American Chemical Society

Received: June 11, 2015 Accepted: July 10, 2015

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DOI: 10.1021/acs.analchem.5b02208 Anal. Chem. XXXX, XXX, XXX−XXX

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tetrachloroaurate(III) trihydrate (HAuCl4·3H2O, Au ≥ 49%) and Na2S2O8 were obtained from J&K Scientific Ltd. (Shanghai, China). The polymer poly diallyldimethylammonium chloride (PDDA, 20%, w/v, MW ∼ 20 000) was purchased from Beijing HWRK Chem. Co. Ltd. (Beijing, China). NaOH, Al(NO3)3·9H2O, Mg(NO3)2·6H2O, Zn(NO 3)2 ·6H2O, Ni(NO 3)2 ·6H2O, Co(NO 3)2 ·6H2O, urea, HNO3, NaNO3, formamide, ascorbic acid, Na2HPO4, and NaH2PO4 were supplied by Beijing Chemical Reagent Company (Beijing, China). Phosphate buffer solution (PBS, pH = 7.5, 0.1 M) was prepared using 0.1 M Na2HPO4 and 0.1 M NaH2PO4. All solutions were prepared with deionized water (Milli Q, Millipore, Barnstead, CA). All reagents were of analytical grade and used without further purification. Preparation of BSA−AuNCs. BSA−AuNCs with a blue fluorescence emission at 450 nm were synthesized according to literature procedures.25 In brief, 5.0 mL HAuCl4 aqueous solution (10 mM) was added into 5.0 mL bovine serum albumin (BSA, 20 mg/mL) under vigorous stirring at 37 °C. Subsequently, ascorbic acid (5.0 μL, 0.35 mg/mL) was added dropwise. BSA−AuNCs could be obtained after the solution was kept at pH = 8 for 5 h under vigorous stirring. The asprepared BSA−AuNCs were purified by the dialysis membrane with a molecular weight cutoff of 2000 Da for 12 h, and then they were stored at 4 °C for further use. Preparation of AuNCs@Ru Solution. The AuNCs@Ru solution was produced by adding Ru(bpy)3Cl2 aqueous solution (0.05 M, 0.05 mL) into BSA−AuNC solution (15 mL) with stirring gently at room temperature until getting a uniform orange solution. Using the same procedures, the mixture solution of BSA and Ru(bpy)32+ was prepared by mixing 10 mg/mL BSA solution with Ru(bpy)3Cl2 (0.05 M, 0.05 mL) for the control experiments. Synthesis of CoAl−CO3 LDHs. We synthesized the CoAl− LDHs according to the method reported previously. 35 Co(NO3)2·6H2O (10 mM), Al(NO3)3·9H2O (5.0 mM), and urea (30 mM) were dissolved in 100 mL of deionized water, stirred vigorously and heated at 97 °C for 48 h. Afterward, the precipitate was centrifuged and washed thoroughly with deionized water until the pH value returned to 8.0. The CoAl−CO3 LDHs were obtained after drying at 60 °C for 24 h. The similar synthesis procedure as that of CoAl−CO3 LDHs was used to prepare ZnAl/MgAl/NiAl−CO3 LDHs, respectively. Synthesis of CoAl−LDH Nanosheets. CoAl−CO3 LDHs were exfoliated into the LDH nanosheet according to the method reported previously.35 Typically, 0.1 g of CoAl−CO3 LDHs were dissolved into 100 mL of water containing 0.005 M HNO3 and 1.5 M NaNO3. Under flowing nitrogen, the mixture was continuously agitated at ambient temperature for 48 h. The resulting CoAl−NO3 LDHs were centrifuged, washed, and vacuum-dried. Subsequently, 0.03 g of the as-prepared CoAl− NO3 LDHs were vigorously stirred for 48 h in 30 mL of formamide under N2 atmosphere at room temperature. A pink colloidal suspension of positively charged CoAl−LDH nanosheets was acquired, and it was used directly for the assembly of UTFs. Fabrication of the UTF-Modified Au Electrode. Before the fabrication, the Au electrode was polished sequentially with 0.3 and 0.5 μm alumina slurry and washed ultrasonically with ethanol and water. Then the Au electrode was rinsed electrochemically (0.1 M H2SO4; scan rate, 0.1 V/s) until a stable redox wave of H2SO4 was observed. We fabricated the

confined space of the interlayer galleries of LDHs.30,31 For example, pole (vinyl carbazole) (PVK) and tetraphenylethylene (TPE) molecules can be assembled in the interlayers of LDHs nanosheets to fabricate (TPE@PVK/LDH)n films, and thus the FRET efficiency between PVK and TPE was greatly improved within films owing to the orderly packing architecture of PVK and TPE in the confined space of LDHs.30 On the other hand, the chemiluminescence resonance energy transfer (CRET) efficiency between peroxynitrous acid donors and fluorescence dye acceptors could be improved remarkably owing to the efficient suppression of the intermolecular π−π stacking interactions among aromatic rings and the orderly packing of fluorescence molecules at the surface of the LDHs. As a result, the fluorescence lifetime and quantum yield of fluorescence dyes would increase when they were assembled on the surface of LDHs.32,33 The successful improvements of FRET and CRET in the LDH matrix inspirits us to construct an efficient ERET process by layer-by-layer (LBL) electrostatic assembly of CoAl−LDH nanosheets with some ECL molecules (i.e., ERET donors and acceptors). However, the ERET with LDH nanosheets have not been established yet. Among the family of LDHs, CoAl−LDHs are often used a new kind of high-performance electrode materials.34 In this study, we fabricated novel ECL ultrathin films (UTFs) on an Au electrode by LBL assembly of CoAl−LDH nanosheets with a mixture of blue BSA−AuNCs and Ru(bpy)32+ (denoted as AuNCs@Ru). The fabrication scheme of the UTFs was illuminated in Figure 1. The enhanced ECL signals could be

Figure 1. Schematic illustration of AuNCs@Ru−LDH UTFs for ECL resonance energy transfer between BSA−AuNCs and Ru(bpy)32+.

achieved by an efficient ERET between BSA−AuNCs and Ru(bpy)32+ on the as-prepared AuNCs@Ru−LDH UTFs. Furthermore, the selectivity and stability of the fabricated ECL sensor were examined by detecting 6-mercaptopurine (6MP). To the best of our knowledge, this is the first example of the cathodic ERET system to employ AuNCs as ECL donors, different from many reported ECL acceptors. The strategy indicated that the ERET in the LDH UTFs had a wide range of potential applications in molecular diagnostics.

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EXPERIMENTAL SECTION Chemicals and Materials. Bovine serum albumin (BSA), Ru(bpy)3Cl2 (Ru), 6-MP, cysteine, and glutathione were supplied by Sigma-Aldrich (St. Louis, MO). Hydrogen B

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Figure 2. (A) Thickness of the as-prepared CoAl−LDH nanosheets (inset, AFM image of the exfoliated CoAl−LDH nanosheets deposited on a fresh silicon substrate slice and Tyndall light scattering of CoAl−LDH nanosheets); (B) excitation (blue) and emission (red) spectra of BSA− AuNCs (inset, HRTEM image of BSA−AuNCs).

instrument. Mass spectra of the BSA−AuNCs and BSA were measured by a matrix assisted laser desorption ionization-timeof-flight (MALDI-TOF) mass spectrometer (Ultraflex, Bruker Daltonics, Germany). All the tests were carried out in positive mode. The fluorescence spectra were measured with a Hitachi F-7000 fluorescence spectrophotometer (Tokyo, Japan) with the excitation wavelength of 375 nm. The scanning rate was 1500 nm/min, and the excitation (or emission) slit was maintained at 5.0 nm. We used a Zetasizer DTS Nano (Malvern Instrument Ltd.) to measure the dynamic light scattering (DLS) at 25.0 ± 0.1 °C, and then particle size and zeta potential data were auto-calculated by the software. Electrochemical experiments were performed with a CHI 660E electrochemical analyzer (CHI). The ECL intensities were recorded by a BPCL luminescence analyzer (Institute of Biophysics, Chinese Academy of Science, Beijing, China) with the voltage of the photomultiplier tube (PMT) set at −900 V.

PDDA/(CoAl−LDH/AuNCs@Ru)n UTF-modified Au electrodes using the LBL electrostatic assembly. Briefly, the Au electrode was first immersed into the PDDA aqueous solution (1.0 mg/mL) for 10 min to own a surface rich in positive charge. Then the Au electrode was alternately dipped into the AuNCs@Ru solution and the colloidal CoAl−LDH nanosheets and washed thoroughly. After repeating the deposition of AuNCs@Ru and CoAl−LDH nanosheets for several times, the PDDA/(CoAl−LDH/AuNCs@Ru)n UTF-modified electrode was obtained. The PDDA/(CoAl−LDH/AuNCs) n and PDDA/(CoAl−LDH/BSA@Ru)n UTF-modified Au electrodes were prepared by displacing AuNCs@Ru solution with AuNCs or BSA@Ru solution. All the films were dried under nitrogen atmosphere. Electrochemical and ECL Experiments. In this work, we used a conventional three-electrode system. A modified Au electrode (3.0 mm in diameter) served as the working electrode, a platinum electrode was the auxiliary electrode, and a saturated Ag/AgCl electrode acted as the reference electrode. The electrochemical impedance spectroscopy (EIS) measurements were detected in a 5.0 mM Fe(CN)63−/4− solution under the frequency range from 0.01 Hz to 1000 kHz. The ECL reactions occurred in 0.1 M PBS (pH = 7.5) with 50 mM Na2S2O8 under the potential from 0 to −1.5 at the scanning rate of 100 mV/s. The appropriate amounts of 6-MP solutions were added into the electrolyte to form the solutions with a series of concentrations, and the selectivity and stability of the ERET probe were certified by detecting their ECL signals. Apparatus and Characterization. X-ray diffraction (XRD) patterns of CoAl−CO3 LDH, CoAl−NO3 LDH were recorded on a Rigaku D/max-2500VB2+/PC, 2θ scanning from 5° to 70°. Also the PDDA/(CoAl−LDH/AuNCs@Ru)n (n = 10, 20, 30) UTFs were scanned from 0.5° to 10°. The scanning electron microscope (SEM) was performed on a Hitachi S3500 SEM with an accelerating voltage of 20 kV. The morphology of the BSA−AuNCs was studied using high resolution transmission electron microscopy (HRTEM) (JEOL JEM-3010) with an accelerating voltage of 300 kV. Atomic force microscopy (AFM) in tapping mode was carried out on a NanoScope IIIa (Digital Instruments Co., Santa Barbara, CA)

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RESULTS AND DISCUSSION Characterization of CoAl−LDH Nanosheets and BSA− AuNCs. In this work, the LDH nanosheets were obtained by three steps: preparation of CoAl−CO3 LDHs, preparation of the CoAl−NO3 LDHs obtained from the CoAl−CO3 LDHs by the anion exchange method, and exfoliation of the CoAl−NO3 LDHs into the CoAl−LDH nanosheets. The XRD and SEM results of the CoAl−CO3 LDHs and CoAl−NO3 LDHs crystals were shown in Figures S1 and S2 in the Supporting Information. These samples were hexagonal platelets with a size of 5 μm. In comparison to the CoAl−CO3 LDHs, the CoAl−NO3 LDHs had a smaller 2θ angles with the interlayer spacing (d003).36 The AFM image of the CoAl−LDH nanosheets in the inset of Figure 2A showed that the thickness of the as-prepared LDH nanosheets was less than 1 nm, indicating the formation of unilamellar nanosheets.35 In addition, a clear Tyndall light scattering effect was observed from the transparent colloid solution, further demonstrating the existence of the LDH nanosheets (inset of Figure 2A).36 Figure 2B showed the excitation and emission spectra of the BSA−AuNCs, exhibiting an excitation maximum at 375 nm and an emission maximum at 450 nm, respectively. Additionally, the fluorescence emission intensity of the BSA−AuNCs at 450 nm C

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Figure 3. (A) XRD patterns for the PDDA/(CoAl−LDH/AuNCs@Ru)n UTFs with n = 10, 20, and 30; (B) EIS for the PDDA/(CoAl−LDH/ AuNCs@Ru)n UTF-modified Au electrode with different bilayer numbers (n) = 5, 10, 15, and 20.

Figure 4. (A) ECL curves of (a) the PDDA/(CoAl−LDH/AuNCs@Ru)10 UTF-modified Au electrode, (b) PDDA/(CoAl−LDH/AuNCs)10 UTFmodified Au electrode, (c) AuNCs@Ru-modified Au electrode, (d) AuNC-modified Au electrode, (e) a bare Au electrode, (f) PDDA/(CoAl− LDH/BSA@Ru)10 UTF-modified Au electrode, (g) Ru(bpy)32+-modified Au electrode, (h) CoAl−LDH-modified Au electrode; (B) ECL spectra of the PDDA/(CoAl−LDH/AuNCs)10 (blue) and the PDDA/(CoAl−LDH/AuNCs@Ru)10 (red) UTF-modified Au electrodes (inset, absorption spectrum of Ru(bpy)32+ (red) and ECL spectrum of BSA−AuNCs (blue).

stable self-assembly with the negative [email protected] Here, the multilayer PDDA/(CoAl−LDH/AuNCs@Ru)n UTFs were fabricated by alternately dipping the PDDA-modified Au electrode into a colloidal suspension of LDH nanosheets and an aqueous solution of AuNCs@Ru. The XRD patterns of PDDA/(CoAl−LDH/AuNCs@Ru)n showed that there was a linear increase in the intensity upon increasing bilayer number (n) up to 30 cycles (Figure 3A), indicating a long-range stacking order of the AuNCs@Ru on the Au electrode. In EIS diagrams, the semicircle portion at high frequencies is corresponding to the electron-transfer-limited process, while the linear part of lower frequencies represents the diffusionlimited electron-transfer process.37 Meanwhile, the increase of EIS signals could be used to monitor the LBL assembly processes.38 Figure 3B showed that the EIS signals grew linearly upon increasing bilayer number (n) up to 20 cycles, further demonstrating that the assembly of PDDA/(CoAl−LDH/ AuNCs@Ru)n UTFs was successful and the ECL reaction in

was much higher than that of BSA (Figure S3 in the Supporting Information), indicating the successful synthesis of the BSA− AuNCs.25 The HRTEM image of the BSA−AuNCs demonstrated that the particle size of the as-prepared BSA−AuNCs was around 2.0 nm (inset of Figure 2B), which was approximate with the DLS results (Figure S4 in the Supporting Information). Finally, the MALDI-TOF mass spectrometry of the BSA and the as-prepared BSA−AuNCs further confirmed the successful synthesis of the BSA−AuNCs (Figure S5 in the Supporting Information).26 Surface Morphology of PDDA/(CoAl−LDH/AuNCs@ Ru)n UTF-Modified Au Electrodes. Figure S6 in the Supporting Information showed the zeta potentials of the BSA−AuNCs and the AuNCs@Ru were negatively charged (−25.4 mV and −22.8 mV, respectively); however, the zeta potential of the CoAl−LDH nanosheets was positively charged (28.8 mV). The relatively high number of zeta potentials indicated that the positive LDH nanosheets could allow the D

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The DLS technique was used to monitor the sizes changes of the BSA−AuNCs containing 50 nM 6-MP, glutathione, and cysteine, respectively. The results (Table S1 in the Supporting Information) showed that the size of the BSA−AuNCs was bigger in the presence of 6-MP in comparison to those in the presence of glutathione and cysteine. In addition, after the BSA−AuNCs reacted with 6-MP, glutathione, and cysteine, respectively, they were used to fabricate the PDDA/(CoAl− LDH/AuNCs@Ru)10 UTFs. The results showed that the BSA−AuNCs after reacting with 6-MP could greatly quench the ECL intensity of the proposed system (Figure S12 in the Supporting Information). These results suggested that the great quenching effect of 6-MP on the ECL signals of the proposed system may be ascribed to the formation of bigger particles.45 Under the optimal conditions (Figures S7−S10 in the Supporting Information), it was apparent that the ECL quenching ratio (F0 − F1)/F0 was proportionate to the concentration of 6-MP at the range of 2.5−100 nM (Figure 5), where F0 is the ECL intensity in the absence of 6-MP and F1

the proposed system was controlled by two electron-transfer processes. Efficient ERET on PDDA/(CoAl−LDH/AuNCs@Ru)n UTF-Modified Au Electrodes. Under the optimum experimental conditions (Figures S7−S10 in the Supporting Information), the ECL behaviors of the PDDA/(CoAl− LDH/AuNCs@Ru)10 UTF-modified Au electrode were investigated in the range from 0 to −1.5 V vs Ag/AgCl in 0.1 M PBS (pH = 7.5) with 50 mM Na2S2O8 (Figure 4A). A remarkable stronger ECL signal at ∼−1.5 V in the cyclic voltammetry scanning was observed when the Au electrode was modified by the PDDA/(CoAl−LDH/AuNCs@Ru)10 UTFs. In comparison, a lower ECL intensity was found when the Ru(bpy)32+ was absent on the UTFs. In addition, a series of control experiments were carried out, including bare Au electrode, AuNC-modified Au electrode, LDH nanosheet-modified Au electrode, Ru(bpy)32+-modified Au electrode, AuNCs@Rumodified Au electrode, and PDDA/(CoAl−LDH/BSA@ Ru)10-modified electrode. The results showed that both AuNCs and AuNCs-Ru had weak ECL signals in the absence of the LDH nanosheets. More interestingly, the ECL intensities had no obvious changes when the Au electrode was modified only by Ru(bpy)32+, suggesting that Ru(bpy)32+ itself is not an ECL species in the proposed system.13 On the other hand, we found that the absorption spectrum of Ru(bpy)32+ could be well overlapped with the ECL spectrum of BSA−AuNCs (inset of Figure 4B), indicating that the ERET from BSA−AuNC donors to Ru(bpy)32+ acceptors could occur.39 To further illuminate this possibility, the ECL spectra of PDDA/(CoAl− LDH/AuNCs)10 UTFs and PDDA/(CoAl−LDH/AuNCs@ Ru)10 UTFs were compared using a series of optical filters (Figure 4B). Without the Ru(bpy)32+, there was an ECL emission at 475 nm on the PDDA/(CoAl−LDH/AuNCs)10 UTF-modified Au electrode. While the intensity of the peak at 475 nm was decreased, there appeared a new strong ECL emission peak at 620 nm on the PDDA/(CoAl−LDH/ AuNCs@Ru)10 UTF-modified Au electrode. In general, Ru(bpy)32+ has no ECL response in the negative potential on the Au electrode between 0 and −1.5 V and the cathodic ECL peak of Ru(bpy)32+ at 620 nm is derived from the ERET.13 Finally, it was calculated the ERET efficiency between BSA− AuNCs and Ru(bpy)32+ was about 13.9% in the absence of the CoAl−LDH nanosheets; while, the ERET efficiency between the BSA−AuNCs and Ru(bpy)32+ on the PDDA/(CoAl− LDH/AuNCs@Ru)n UTFs was about 49.6%.39 Therefore, we concluded that the enhanced ECL emissions in this work were attributed to the efficient ERET between the BSA−AuNCs and Ru(bpy)32+ on the PDDA/(CoAl−LDH/AuNCs@Ru)n UTFs. Applications of PDDA/(CoAl−LDH/AuNCs@Ru)n UTFs for 6-Mercaptopurine Probe. 6-Mercaptopurine (6-MP) is an antineoplastic chemotherapy drug with a thiol group.40 Various methods have contributed much to the determination of 6-MP.41−43 Recently, it is found that the fluorescence of BSA−AuNCs can be selectively quenched by 6-MP over other thiol-containing compounds, such as glutathione and cysteine.44 Here, in order to evaluate the applicability and reliability of the proposed ERET sensor, it was applied for detecting 6-MP in human serum and urine samples. In addition, the effects of typical commonly interferences present in human serum and urine samples were investigated to assess the selectivity of the developed methodology (Figure S11 in the Supporting Information). Interestingly, the proposed ECL method can distinguish 6-MP from glutathione and cysteine.

Figure 5. Calibration curve of 6-MP (nM).

is the ECL intensity in the presence of 6-MP. The detection limit is 1.0 nM (S/N = 3). In addition, the consecutive ECL signals of the PDDA/(CoAl−LDH/AuNCs@Ru)10 UTFmodified Au electrode for 10 cycles were shown in Figure S13 in the Supporting Information. The relative standard deviation (RSD) was 4.3%. Human serum and urine samples were obtained from healthy volunteers. After treated with acetonitrile, centrifuging, and filtering, these samples were spiked with 6-MP at different concentrations. The results were shown in Table 1. The recoveries are in the range of 95.0− 108.7%. Table 1. Determination of 6-MP in Spiked Real Samples samples

added (nM)

human serum 1

30.00 50.00 30.00 50.00 30.00 50.00 30.00 50.00

human serum 2 human urine 1 human urine 2

a

E

founda (nM)

recoverya (%)

± ± ± ± ± ± ± ±

95.0 ± 2.8 104.6 ± 2.3 106.0 ± 1.4 101.0 ± 3.3 108.7 ± 4.2 103.3 ± 4.7 105.1 ± 3.2 96.4 ± 3.5

28.50 52.32 31.79 50.46 32.60 51.63 31.52 48.22

0.83 1.14 0.42 1.65 1.25 2.36 0.97 1.74

All of the results were the mean of three determinations ± SD. DOI: 10.1021/acs.analchem.5b02208 Anal. Chem. XXXX, XXX, XXX−XXX

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CONCLUSIONS In conclusion, a novel AuNCs@Ru−LDH UTF was fabricated by integrating AuNCs@Ru with LDH nanosheets by the LBL assembly technique. The cathodic ECL behaviors of the asprepared AuNCs@Ru−LDH UTFs were investigated. A strong emission was observed through an efficient ERET from BSA− AuNC donors to Ru(bpy)32+ acceptors as a result of a longrange stacking order of the AuNCs@Ru on the Au electrode. The feasibility of the proposed ECL probe has been demonstrated by detecting 6-MP with good stability, reproducibility, and selectivity. The observed ECL enhancement in AuNCs@Ru−LDH UTFs opens a novel avenue to further study the cathodic ECL behaviors of Ru(bpy)32+ in the layered UTFs for sensitive detection of various biomolecules.

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REFERENCES

(1) Konz, T.; Alvarez, E. A.; Montes−Bayon, M.; Sanz−Medel, A. Anal. Chem. 2013, 85, 8334−8340. (2) Dong, Y.-P.; Gao, T.-T.; Zhou, Y.; Zhu, J.-J. Anal. Chem. 2014, 86, 11373−11379. (3) Kim, Y.; Kim, J. Anal. Chem. 2014, 86, 1654−1660. (4) Tang, X. F.; Zhao, D.; He, J. C.; Li, F. W.; Peng, J. X.; Zhang, M. N. Anal. Chem. 2013, 85, 1711−1718. (5) Zhang, X. W.; Chen, C. G.; Li, J.; Zhang, L. B.; Wang, E. K. Anal. Chem. 2013, 85, 5335−5339. (6) Qi, H. L.; Li, M.; Dong, M. M.; Ruan, S. P.; Gao, Q.; Zhang, C. X. Anal. Chem. 2014, 86, 1372−1379. (7) Long, Y.-M.; Bao, L.; Zhao, J.-Y.; Zhang, Z.-L.; Pang, D.-W. Anal. Chem. 2014, 86, 7224−7228. (8) Zu, Y. B.; Bard, A. J. Anal. Chem. 2000, 72, 3223−3232. (9) Gaillard, F.; Sung, Y.-E.; Bard, A. J. J. Phys. Chem. B 1999, 103, 667−674. (10) Parajuli, S.; Miao, W. J. Anal. Chem. 2013, 85, 8008−8015. (11) Hu, L. Z.; Li, H. J.; Zhu, S. Y.; Fan, L. S.; Shi, L. H.; Liu, X. Q.; Xu, G. B. Chem. Commun. 2007, 40, 4146−4148. (12) Li, M. Y.; Li, J.; Sun, L.; Zhang, X. L.; Jin, W. R. Electrochim. Acta 2012, 80, 171−179. (13) Wu, M.-S.; Shi, H.-W.; Xu, J.-J.; Chen, H.-Y. Chem. Commun. 2011, 47, 7752−7754. (14) Wu, M.-S.; Shi, H.-W.; He, L.-J.; Xu, J.-J.; Chen, H.-Y. Anal. Chem. 2012, 84, 4207−4213. (15) Li, L.; Li, M. Y.; Sun, Y. M.; Li, J.; Sun, L.; Zou, G. Z.; Zhang, X. L.; Jin, W. R. Chem. Commun. 2011, 47, 8292−8294. (16) Hoshino, A.; Fujioka, K.; Oku, T.; Suga, M.; Sasaki, Y. F.; Ohta, T.; Yasuhara, M.; Suzuki, K.; Yamamoto, K. Nano Lett. 2004, 4, 2163− 2169. (17) Shang, L.; Dong, S. J.; Nienhaus, G. U. Nano Today 2011, 6, 401−418. (18) Zhang, X.−D.; Luo, Z. T.; Chen, J.; Shen, Xiu.; Song, S. S.; Sun, Y. M.; Fan, S. J.; Fan, F. Y.; Leong, D. T.; Xie, J. P. Adv. Mater. 2014, 26, 4565−4568. (19) Chen, L.-Y.; Wang, C.-W.; Yuan, Z. Q.; Chang, H.-T. Anal. Chem. 2015, 87, 216−229. (20) Li, L. L.; Liu, H. Y.; Shen, Y. Y.; Zhang, J. R.; Zhu, J.-J. Anal. Chem. 2011, 83, 661−665. (21) Fang, Y.-M.; Song, J.; Li, J.; Wang, Y.-W.; Yang, H.-H.; Sun, J.-J.; Chen, G.-N. Chem. Commun. 2011, 47, 2369−2371. (22) Cheng, Y.; Lei, J. P.; Chen, Y. L.; Ju, H. X. Biosens. Bioelectron. 2014, 51, 431−436. (23) Chen, Y.; Shen, Y. Y.; Sun, D.; Zhang, H. Y.; Tian, D. B.; Zhang, J. R.; Zhu, J.-J. Chem. Commun. 2011, 47, 11733−11735. (24) Wu, Y. F.; Huang, J. H.; Zhou, T. Y.; Rong, M. C.; Jiang, Y. Q.; Chen, X. Analyst 2013, 138, 5563−5565. (25) Le Guével, X.; Hötzer, B.; Jung, G.; Hollemeyer, K.; Trouillet, V.; Schneider, M. J. Phys. Chem. C 2011, 115, 10955−10963. (26) Das, T.; Ghosh, P.; Shanavas, M. S.; Maity, A.; Mondal, S.; Purkayastha, P. Nanoscale 2012, 4, 6018−6024. (27) Wang, Q.; O’Hare, D. Chem. Rev. 2012, 112, 4124−4155. (28) Song, F.; Hu, X. L. Nat. Commun. 2014, 5, 5477. (29) Nicolosi, V.; Chhowalla, M.; Kanatzidis, M. G.; Strano, M. S.; Coleman, J. N. Science 2013, 340, 1420. (30) Li, Z.; Lu, J.; Qin, Y. M.; Li, S. D.; Qin, S. H. J. Mater. Chem. C 2013, 1, 5944−5952. (31) Shi, W. Y.; Fu, Y.; Li, Z. X.; Wei, M. Chem. Commun. 2015, 51, 711−713. (32) Wang, Z. H.; Teng, X.; Lu, C. Anal. Chem. 2013, 85, 2436− 2442. (33) Wang, Z. H.; Teng, X.; Lu, C. Anal. Chem. 2015, 87, 3412− 3418. (34) Huang, S.; Zhu, G.-N.; Zhang, C.; Tjiu, W. W.; Xia, Y.-Y.; Liu, T. X. ACS Appl. Mater. Interfaces 2012, 4, 2242−2249. (35) Liu, Z. P.; Ma, R. Z.; Osada, M.; Iyi, N.; Ebina, Y.; Takada, K.; Sasaki, T. J. Am. Chem. Soc. 2006, 128, 4872−4880.

ASSOCIATED CONTENT

S Supporting Information *

Power XRD patterns of (a) CoAl−CO3 LDHs and (b) CoAl− NO3 LDH; SEM images of (a) CoAl−CO3 LDHs and (b) CoAl−NO3 LDHs; fluorescence intensity (λem = 450 nm) of BSA−AuNCs, BSA, and blank (10 mg/mL); DLS measurements for (a) BSA and (b) blue BSA−AuNCs; MALDI MS data of BSA and blue BSA−AuNCs; zeta potentials of BSA− AuNCs, AuNCs@Ru, and CoAl−LDH nanosheets; relative ECL intensity of the PDDA/(LDH/AuNCs@Ru)n UTFmodified Au electrodes (LDHs are CoAl−LDH, ZnAl−LDH, MgAl−LDH, NiAl−LDH, respectively); relative ECL intensity of the PDDA/(CoAl−LDH/AuNCs@Ru)n UTF-modified Au electrodes as a function of n; relative ECL intensity of the PDDA/(CoAl−LDH/AuNCs@Ru)n UTF-modified Au electrodes as a function of pH value; CVs of the PDDA/(CoAl− LDH/AuNCs@Ru)n UTF-modified Au electrodes at various scan rates (a) 10, (b) 20, (c) 40, (d) 60, (e) 80, (f) 100 mV/s; inset, plots of peak current vs scan rate; selectivity study results of the proposed sensor for the analysis of 6-MP (100 nM), the concentration of each interference is 100 nM; relative ECL intensity of the PDDA/(CoAl−LDH/mixture−Ru)10 UTFmodified Au electrodes as a function of pH value; consecutive ECL signals of the PDDA/(CoAl−LDH/AuNCs−Ru)10 UTFmodified Au electrode; results of the particle size distribution. The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.analchem.5b02208.

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Article

AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. Fax/phone: +86 10 64411957. Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS This work was supported by National Basic Research Program of China (973 Program, Grant 2014CB932103), the National Natural Foundation of China (Grant 21375006), the 973 Program (Grant 2011CBA00503), and the Fundamental Research Funds for the Central Universities (Grants YS1406 and JD1311). We also thank Prof. Xue Duan, Beijing University of Chemical Technology, for his valuable discussion. F

DOI: 10.1021/acs.analchem.5b02208 Anal. Chem. XXXX, XXX, XXX−XXX

Article

Analytical Chemistry (36) Ma, R. Z.; Liu, Z. P.; Li, L.; Iyi, N.; Sasaki, T. J. Mater. Chem. 2006, 16, 3809−3813. (37) Ahirwal, G. K.; Mitra, C. K. Biosens. Bioelectron. 2010, 25, 2016− 2020. (38) Zhang, B.; Shi, S. X.; Shi, W. Y.; Sun, Z. Y.; Kong, X. G.; Wei, M.; Duan, X. Electrochim. Acta 2012, 67, 133−139. (39) Freeman, R.; Liu, X. Q.; Willner, I. J. Am. Chem. Soc. 2011, 133, 11597−11604. (40) Lennard, L.; Lilleyman, J. S.; Van Loon, J.; Weinshilboum, R. M. Lancet 1990, 336, 225−229. (41) Rosenfeld, J. M.; Taguchi, V. Y.; Hillcoat, B. L.; Kawai, M. Anal. Chem. 1977, 49, 725−727. (42) Cao, X.-N.; Lin, L.; Zhou, Y.-Y.; Shi, G.-Y.; Zhang, W.; Yamamoto, K.; Jin, L.-T. Talanta 2003, 60, 1063−1070. (43) Nikolić, K.; Velašević, K. Microchim. Acta 1990, I, 69−73. (44) Li, Z.; Wang, Y.; Ni, Y. N.; Kokot, S. Biosens. Bioelectron. 2015, 70, 246−253. (45) Han, B. Y.; Wang, E. K. Biosens. Bioelectron. 2011, 26, 2585− 2589.

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DOI: 10.1021/acs.analchem.5b02208 Anal. Chem. XXXX, XXX, XXX−XXX