Biosensors and Bioelectronics 65 (2015) 397–403
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Methionine-directed fabrication of gold nanoclusters with yellow fluorescent emission for Cu2 þ sensing Hao-Hua Deng a,b, Ling-Na Zhang a,b, Shao-Bin He a,b, Ai-Lin Liu a,b, Guang-Wen Li a, Xin-Hua Lin a,b, Xing-Hua Xia c, Wei Chen a,b,n a
Department of Pharmaceutical Analysis, Fujian Medical University, Fuzhou 350004, China Nano Medical Technology Research Institute, Fujian Medical University, Fuzhou 350004, China c State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210093, China b
art ic l e i nf o
a b s t r a c t
Article history: Received 21 July 2014 Received in revised form 4 September 2014 Accepted 30 October 2014 Available online 4 November 2014
In the past few years, fluorescent gold nanoclusters (AuNCs) have gained much attention in many areas of physics, chemistry, materials science, and biosciences due to their unique physical, electrical, and optical properties. Herein, we reported for the first time the synthesis of water soluble, monodispersed AuNCs by using methionine both as a reductant and a stabilizer. The synthetic process is green and simple, and the resulting AuNCs capped by methionine (Met-AuNCs) would be biocompatible with bioorganisms. UV–visible absorption, photoluminescence, X-ray photoelectron spectroscopy (XPS), and Fourier transform infrared (FTIR) were carried out to demonstrate the chemical composition and optical properties of the as-prepared Met-AuNCs. The Met-AuNCs possess many attractive features including intense yellow fluorescence (emission maximum at 530 nm), a long fluorescence lifetime (181 ns and 1651 ns), high colloidal stability (pH-, temperature-, salt- and time-stability), and a large Stoke's shift (110 nm), holding great promise as late-model analytical tools for life science and environmental studies. Moreover, the assynthesized Met-AuNCs can serve as an efficient fluorescent probe for selective detection of Cu2 þ by fluorescence quenching. The limit of detection for Cu2 þ was determined to be 7.9 nM and linear response over the Cu2 þ concentrations range from 50 nM to 8 μM. Furthermore, the new-constructed probe allows simple and rapid detection of the concentrations of Cu2 þ in soil, with results demonstrating its great feasibility for the determination of copper in real samples. & 2014 Elsevier B.V. All rights reserved.
Keywords: Methionine Gold nanoclusters Fluorescence Copper ion Quenching
1. Introduction Metal nanoclusters, consisting of a few to roughly a hundred atoms, have become one of the important kinds of nanomaterials being extensively pursued in current nanoscience research (Liu et al., 2013b; Shang et al., 2011c). They bridge the evolution of properties of materials between isolated atoms and those of the bulk. Their excellent optical, electrical, and physical properties are very promising for applications in single-molecule optoelectronics, catalysis, biological imaging/labeling, and sensing (Chen et al., 2012a, 2012b; Han and Wang 2011; Han et al., 2012; Hyotanishi et al., 2011; Lee et al., 2005; Liu et al., 2013a, 2011; Myers et al., 2011; Nair et al., 2013; Shang et al., 2011a). Among various metal nanoclusters, considerable efforts have been dedicated to the n Corresponding author at: Fujian Medical University, Department of Pharmaceutical Analysis, 88 Jiaotong Road, Fuzhou 350004, Fujian, China. Fax: þ86 591 22862016. E-mail address: [email protected] (W. Chen).
http://dx.doi.org/10.1016/j.bios.2014.10.071 0956-5663/& 2014 Elsevier B.V. All rights reserved.
research of gold nanoclusters (AuNCs) because of their chemical stability and elegant optical properties. Over the past few years, biological molecules (such as protein, peptide, nuclear acid, and amino acid) have been widely used as eco-friendly biotemplates for metal nanocrystals synthesis (Yang et al., 2011; Zhou et al., 2009). The biomolecule-mediated nanoclusters show lower toxicity and higher biocompatibility to organisms, holding great promise as late-model analytical tools for life science and environmental studies. Quite a number of attempts have been made in this respect. Xie et al. first reported a green method for preparing highly red-emitting Au25 clusters using bovine serum albumin (BSA) as biotemplate (Xie et al., 2009). Inspired by this work, researchers found many different proteins including pepsin, lysozyme, trypsin, horseradish peroxidase, and transferrin could also serve as efficient bioscaffolds for fabricating AuNCs (Kawasaki et al., 2011a, 2011b; Le Guevel et al., 2011; Wei et al., 2010; Wen et al., 2011). Pradeep et al. synthesized two fluorescent AuNCs (namely Au25 and Au8) from mercaptosuccinic acid-stabilized gold nanoparticles by etching with a natural tripeptide, glutathione (GSH) (Muhammed et al., 2008).
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The same group also introduced a novel way for the preparation of Au23 clusters by core etching of GSH-protected Au25 clusters (Muhammed et al., 2009). Liu et al. established a method to synthesize water-soluble and red-emitting AuNCs using single-stranded DNA (ss-DNA) as a host and dimethylamine borane (DMBA) as a reductant (Li et al., 2012). Chen et al. introduced a top-down etching strategy to prepare “magic” Au8 clusters from gold particles with the help of biomolecules (protein, peptide, DNA, or amino acid) under sonication in water (Zhou et al., 2009). Later, the same group also prepared bluish green-emitting Au10 clusters through a simple manner with histidine as both reducing and stabilizing molecule (Yang et al., 2011). More recently, Yang et al. synthesized blue-emitting AuNCs by HAuCl4 and N2H2 H2O (reducing agent) in the presence of lysine as a template (Yang et al., 2013). Although considerable progress has been made, finding new biomolecules for the formation of AuNCs is still of great importance. In this work, we reported for the first time a green synthetic process of AuNCs by using methionine as both reducing and stabilizing ligands. Compared to the previously reported strategies for the synthesis of metal nanocluster, the approach presented here is simple, time-saving, and does not require toxic reductants (e.g., NaBH4). The resulting AuNCs capped/stabilized by methionine (Met-AuNCs) exhibit intense yellow fluorescence maximized at 530 nm. Moreover, the Met-AuNCs were found to be a sensitive fluorescent “turn-off” sensor for the selective monitoring of Cu2 þ .
2. Experimental 2.1. Materials All chemicals and solvents were of at least analytical grade and commercially available. HAuCl4 4H2O, methionine, NH3 H2O, and Rhodamine 6G were obtained from Aladdin Reagent Company (Shanghai, China). H2SO4, C2H5OH, and NaOH were brought from Sinopharm Chemical Reagent Co. Ltd. (Shanghai, China). Deionized water was used throughout all experiments. 2.2. Preparation of Met-AuNCs All glassware used in the following procedures was cleaned in a bath of freshly prepared solution of HNO3–HCl (1:3, v/v), rinsed thoroughly in water and dried in air prior to use. AuNCs were synthesized in a blending manner. Typically, methionine (4 mL, 0.1 M) and NaOH (0.6 mL, 0.5 M) were added to an aqueous solution of HAuCl4 (0.4 mL, 20 mg/mL). A fast fading of the light yellow color of the mixture solution was observed, indicating the formation of Au(III)-methionine complexes due to the interaction of gold ions with the sulfur of methionine. The mixture solution was incubated at 37 °C for 6 h, obtaining a pale yellow solution. After that an aqueous solution of H2SO4 (0.5 mL, 1 M) was added into the mixture to precipitate AuNCs. After centrifuged at 6000 rpm for 2 min, the AuNCs were collected and rinsed by H2SO4 (5 mL, 0.1 M) for three times. Then the AuNCs were dissolved in a solution of NH3 H2O (1.4%) and incubated at 70 °C for 30 min. The resulting solution of AuNCs was stored in the dark at 4 °C for later use. 2.3. Characterization The UV–vis absorption spectrum and the photoluminescence spectrum were measured by UV-2450 UV–vis spectrophotometer (Shimadzu, Japan) and Cary Eclipse fluorescence spectrophotometer (Agilent, USA), respectively. Time decay measurements were conducted on a F900 time-correlated single photo
counting fluorescence lifetime spectrometer (Edinburgh Analytical Instruments, UK). X-ray photoelectron spectroscopy (XPS) was performed on an ESCALAB 250 XI electron spectrometer (Thermo, USA) using monochromatic Al Kα radiation (1486.6 eV) for analysis of the surface composition and chemical states of the product. The Fourier transform infrared spectroscopy (FTIR) was measured at the wavenumber ranging from 400 to 4000 cm 1 using a Nicolet Avatar 360 FTIR spectrophotometer. All measurements were performed at room temperature under ambient conditions. 2.4. Met-AuNCs-based sensor for Cu2 þ For Cu2 þ sensing, 0.04 mL as-prepared Met-AuNCs were added into 0.36 mL phosphate buffer (10 mM, pH ¼ 7) containing different concentrations of Cu2 þ . The mixture was incubated at room temperature for 1 min. The photoluminescence spectra were measured by Cary Eclipse fluorescence spectrophotometer. 2.5. Analysis of soil sample Real samples were analyzed to estimate the analytical performance of this new sensor. Acidic digestion of soil sample was performed according to EPA method 3050B. A soil sample (0.01 g) was weighed in an Erlenmeyer flask, and 10 mL of HNO3 1:1 (v/v) was added. The solution was heated on a hot plate to ∼95 °C without boiling, and this temperature was maintained for 15 min. After cooling to less than 70 °C, 5 mL of concentrated HNO3 was added and the sample was refluxed for 30 min at ∼95 °C without boiling. This step was repeated for a second time. The sample was then evaporated to 5 mL without boiling. After cooling to less than 70 °C, 2 mL water was added followed by the slow addition of 10 mL of H2O2 (30%). Care must be taken to ensure that losses do not occur due to excessively vigorous effervescence caused by rapidly adding the strong oxidizer, H2O2. The solution was then heated until effervescence subsided. After cooling to less than 70 °C, 5 mL of concentrated HCl and 10 mL water were added and the sample was refluxed for 15 min without boiling. After cooling to room temperature, the sample was filtered and diluted to 100 mL using water (Chen et al., 2009). The obtained aqueous soil sample (1 mL) was further diluted with 49 mL water and was adjusted to pH ¼ 7 with 5 M NaOH. Afterward, 0.04 mL as-prepared Met-AuNCs were added into 0.36 mL sample solution. The mixture was incubated at room temperature for 1 min before measuring.
3. Results and discussion 3.1. Synthesis of Met-AuNCs Fig. 1A depicted the synthetic procedures of Met-AuNCs with intense yellow fluorescent emission. By incubating methionine and HAuCl4 in NaOH solution at 37 °C for 6 h, a pale yellow solution was obtained. The crude product was then purified through an acid precipitation process. After purification, the precipitate was dissolved by NH3 H2O and incubated at 70 °C for 30 min to obtain the resulting Met-AuNCs for subsequent characterization and application. The Met-AuNCs solution emits intense yellow fluorescence (Fig. 1B) under UV light irradiation. In contrast, neither pure methionine nor HAuCl4 exhibits photoluminescence under the equal conditions, demonstrating that the observed significant fluorescence originates from the gold core of the Met-AuNCs. Photographs of the solutions mentioned before in room light were also showed in Fig. 1B.
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Fig. 1. (A) Schematics of the formation of Met-AuNCs. (B) Photographs of methionine solution in room light (a) and UV light at 365 nm (d), HAuCl4 solution in room light (b) and UV light at 365 nm (e), and Met-AuNCs solution in room light (c) and UV light at 365 nm (f).
Additional experiments were carried out to optimize the synthesis of brightly luminescent AuNCs. The synthetic procedure consists of two steps: (i) slow reducing process to form relatively weak fluorescent AuNCs, and (ii) fast ripening process to generate intense fluorescent AuNCs. Regarding the first step, experiments involving the influence of the concentration of methionine, the concentration of NaOH, and the reaction time were performed. It can be seen from Fig. S1, the fluorescence intensity of the product was the most intense when the methionine concentration was increased to 0.1 M and diminished if the methionine concentration increased further. The influence of the concentration of NaOH on the fluorescence intensity of AuNCs was revealed in Fig. S2. It is clear that a concentration of 0.5 M gives the highest intensity of the product. Under the optimal conditions, the first-step reaction proceeded to completion within 6 h (Fig. S3). As for the second step, the influence of the concentration of NH3 H2O, ripening temperature, and time was taken into consideration. From Fig. S4, it can be seen that the photoluminescence intensity of the MetAuNCs increased when the concentration of NH3 H2O increased from 0.07% to 1.4% and no distinct change was observed if the concentration increased further. The highest fluorescence intensity was achieved when the ripening process was performed at 70 °C for 30 min (Figs. S5 and S6). Under the optimal conditions, the Met-AuNCs obtained in the second-step show fluorescence about 3 times higher than those obtained in the first-step. Although the exact mechanism of the ripening process is not yet clear, there is
almost no change of the wavelength of maximum peaks in the excitation and emission spectra before and after the ripening process (Fig. S7), demonstrating that the emitters from the two steps are the same components. 3.2. Characterization of Met-AuNCs Fig. 2A displays the absorption spectrum of the as-prepared Met-AuNCs. The product missed the specific surface plasmon resonance (SPR) peak (around 520 nm) of larger gold nanocrystals. And instead, it showed a weak but noticeable absorption band centered on 330 nm. In comparison, the absorption spectra of pure methionine and HAuCl4 were also included in Fig. 2A and no peaks at 330 nm were observed in the two samples. By fluorescence spectrophotometry (excited at 420 nm), the Met-AuNCs solution exhibited strong photoluminescence around 530 nm (Fig. 2B). Besides, neither pure methionine nor HAuCl4 exhibited obvious luminescence under the equal conditions. Thus, the observed strong fluorescence should stem from the gold core of the resulting AuNCs. The emission spectrum shows unchanged over a wide excitation wavelength range of 360–460 nm (Fig. S8), indicating that the emission of AuNCs should be from the relaxed states rather than scattering effects (Shang et al., 2011b). It is worth mentioning that, contrast to semiconductor quantum dots (QDs), the AuNCs demonstrated here show a clear excitation band, which would be propitious to Förster resonance energy transfer
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Fig. 2. (A) UV–vis absorption spectra of the aqueous Met-AuNCs, HAuCl4, and methionine. (B) Photoluminescence excitation and emission spectra of Met-AuNCs, and emission spectra of methionine and HAuCl4 were taken as controls. (C) Au(4f) XPS spectrum of Met-AuNCs. (D) S(2p) XPS spectrum of Met-AuNCs.
(FRET) applications. The luminescence quantum yield (QY) of MetAuNCs was 2.8%, calculated with Rhodamine 6G (QY ¼0.95 in ethanol) as the reference, which is nearly two magnitude orders higher than those AuNCs prepared by using NaBH4 as a reductant (Negishi et al., 2004). Additionally, the dominant fluorescence lifetime of the Met-AuNCs was about 181 ns (44.06%) and 1651 ns (55.94%) (Fig. S9), providing a neat opportunity for fluorescence lifetime imaging in future (Shang et al., 2011a). Transmission electron microscopy (TEM) was used to confirm the structure of as-synthesized AuNCs, which revealed an average diameter of about 2.5 nm for the Met-AuNCs (Fig. S10). Electronic structure of the Met-AuNCs sample was conducted by X-ray photoelectron spectroscopy (XPS). The Au(4f) XPS spectrum (Fig. 2C) manifested the binding energies of Au(4f5/2) and Au(4f7/2) located at 88.1 and 84.2 eV, respectively, which explicitly suggests that the electronic structures of Au in Met-AuNCs are Au (0) and Au(I), coexisting (Shang et al., 2011a, 2011b; Zhou et al., 2010). According to reports, the Au(I) surrounded on the surface of AuNCs core plays a vital role in the formation of highly fluorescent AuNCs (Wang et al., 2013). After further reduction of the MetAuNCs with NaBH4, the fluorescence of Met-AuNCs showed a 77% decrease and the corresponding absorption in visible range displayed a slight increase (Fig. S11). The S(2p) XPS spectrum showed a dominant peak located at 162.6 eV, which is attributed to the binding of sulfur atoms of methionine to the gold surface (Fig. 2D). In addition, C(1S), O(1S), and N(1S) core-level photoemission spectra shown in Fig. S12 derive from the protecting ligand, making clear the AuNCs are capped by methionine. Infrared measurement of Met-AuNCs (Fig. S13) showed a suppression of the -C-S stretching vibration in the region of 790–830 cm 1 but
nearly no change of νasNH3 þ (1630 cm 1) and νasCOO (1530 cm 1) (Cao and Fischer, 2002; Vujacic et al., 2009), indicating that S-atom but not ‒NH2 and ‒COOH groups from the methionine molecule is involved in the formation of coordinative bonds in the Met-AuNCs. The functional groups (‒NH2 and ‒ COOH) from methionine on the surface of nanoclusters may favor further modification of the AuNCs. 3.3. Stability of fluorescent Met-AuNCs High stability of fluorescent nanomaterials is an important ingredient to evaluate the possibility of their practical applications. As shown in Fig. 3A, the curve which represents the fluorescence intensity of Met-AuNCs at 530 nm under various pH values from 2 to 12 fluctuates slightly, demonstrating that the Met-AuNCs have a relative high pH-stability. Apart from pH stability, temperature stability is another indispensable factor to ensure that the fluorescent nanomaterials are practical for applications. As shown in Fig. 3B, the curve which depicts the fluorescence intensity of AuNCs suffered a small change when temperature varied from 20 to 70 °C, implying that the Met-AuNCs have a high temperaturestability. Moreover, the variation in fluorescent intensity of the Met-AuNCs exposed to different concentrations of NaCl was examined. The results revealed in Fig. 3C show that the fluorescent intensity of the AuNCs decreased with the increasing concentration of NaCl. However, the fluorescent intensity of as-prepared Met-AuNCs preserved 50% of the initial intensity even in high ionic strength (1 M NaCl). Moreover, there was a small quenching of the fluorescent intensity (only a 6.8% decrease) and no change in the position of the emission peak for Met-AuNCs after storing in
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Fig. 3. The fluorescence stability of Met-AuNCs under (A) various pH values, (B) various temperatures, and (C) different concentrations of NaCl. (D) The fluorescence spectra of as-prepared Met-AuNCs before and after storing in dark at 4 °C for two months.
the dark at 4 °C for two months (Fig. 3D). This demonstrates that the as-prepared Met-AuNCs are particularly suitable for long-term use. It should be noted that the ultra-high stability of the MetAuNCs comes from two crucial factors: protective shell formed by methionine around AuNCs and stabilization effect of Au (I) surrounded on the surface of AuNCs core (Wang et al., 2013). Due to their ultra-high stabilities, the as-synthesized Met-AuNCs may broaden the application of fluorescent nanomaterials. 3.4. Fluorescent sensing of Cu2 þ using Met-AuNCs With improved features in terms of simplicity, selectivity, sensitivity, and miniaturizability, metal nanoclusters (e.g., Au and AgNCs or Au/AgNCs) have been used to construct fluorescent sensors for the detection of specific analytes of chemical as well as biological origin (Yuan et al., 2013). In this study, we found that Cu2 þ could selectively quench the fluorescence of Met-AuNCs, which can be used as a “turn off” probe for Cu2 þ . In order to verify whether the quenching fluorescence of Met-AuNCs was due to the coordination of Cu2 þ with methionine, which could impede Au–S charge transfer, or the fluorescence quenching by metal–metal interaction, ethylenediaminetetraacetate (EDTA) was used as chelator in competition with Met-AuNCs for Cu2 þ . It can be seen from Fig. 4, the fluorescence intensity of Met-AuNCs, after quenching by Cu2 þ , could effectively restore (94% of its initial value) when EDTA was introduced, which indicated that the interaction between Cu2 þ and Met-AuNCs was indeed through coordination.
Fig. 4. (A) Fluorescence regeneration results for quenching by Cu2 þ upon the addition of EDTA. (a) Met-AuNCs, (b) Met-AuNCs in presence of 10 μM Cu2 þ , and (c) Met-AuNCs containing 10 μM Cu2 þ after the addition of 20 μM EDTA. (B) The corresponding photographs.
Considering the appreciable changes in fluorescent properties of Met-AuNCs toward Cu2 þ , the potential of establishing a novel fluorescent probe for Cu2 þ determination was assessed. The kinetic behaviors of fluorescence of Met-AuNCs in the presence of Cu2 þ were monitored and the results show that the interaction between Cu2 þ and Met-AuNCs could be fulfilled within few seconds to quench the fluorescence (Fig. S14). With the concentrations of Cu2 þ increasing, the fluorescence intensity of Met-AuNCs gradually declined (Fig. 5A and B). There is a linear relationship between fluorescence quenching and the concentrations of Cu2 þ within a range from 50 nM to 8 μM (Fig. 5B inset). The limit of
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Fig. 5. (A) Emission spectra of Met-AuNCs in the presence of varying concentrations of Cu2 þ . a–k: 0, 0.05, 0.1, 0.2, 0.5, 1, 2, 4, 6, 8, 10 μM. (B) The plot of the fluorescence intensity (I) of Met-AuNCs recorded at 530 nm versus the concentration of Cu2 þ . Inset: linear relationship between the logarithm of I and the concentration of Cu2 þ . (C) Fluorescence quenching effect [(I0 I)/I0] of the Met-AuNCs at 530 nm incubated with a concentration of 6 μM different tested cations. Samples marked with 1–18 corresponding to Cu2 þ , Co2 þ , Ni2 þ , Mn2 þ , Ca2 þ , Mg2 þ , Ba2 þ , Cd2 þ , Pb2 þ , Zn2 þ , Fe2 þ , Fe3 þ , Al3 þ , Cr3 þ , Ag þ , Na þ , K þ , and NH4 þ , respectively. (D) Fluorescence quenching effect [(I0 I)/I0] of the Met-AuNCs at 530 nm incubated with a concentration of 6 μM different tested anions. Samples marked with 1–18 corresponding to Cu2 þ , SCN , F , Cl , Br , I , ClO4 , BrO3 , IO3 , SO32 , S2O32 , S2O82 , NO2 , NO3 , Ac , CO32 , H2PO4 , and EDTA2 , respectively.
Table 1 Comparison of the proposed method with other reported approaches for the detection of Cu2 þ based on fluorescent nanoclusters. Probe Mercaptoundecanoic acid-AuNCs BSA-AuNCs DNA-AgNCs Azobenzene modified poly(acrylic acid)-AgNCs DNA-AgNCs Glutathione-AuNCs DNA-Cu/Ag NCs Dithiothreitol-AuNCs Met-AuNCs
Linear range (M) 8
6
1.0 10 –1.0 10 5.0 10 7–1.0 10 4 0–1.0 10 5 5.0 10 6–1.5 10 4 5.0 10 6–1.25 10 4 1.0 10 7–6.25 10 6 5.0 10 9–2.0 10 7 5.0 10 6–5.0 10 5 5.0 10 8–8.0 10 6
detection (LOD) was 7.9 nM (3s) and the relative standard deviation (RSD) was 1.4% for the determination of 1 μM Cu2 þ (n¼ 9). The fluorescence quenching constant (Ksv) was calculated to be 5.5 105 M 1 by linear regression of the plot based on the Stern– Volmer equation, I0/I ¼1 þKsv[Q], where I0 and I are the fluorescence intensity of the Met-AuNCs at 530 nm in the absence and presence of Cu2 þ , and [Q] is the concentration of Cu2 þ . This approach was comparable and even more efficient for the detection of Cu2 þ compared to other nanoclusters-based fluorescence methods previously reported (Table 1). To test whether this quenching behavior by Cu2 þ is selective, the fluorescence of MetAuNCs in the presence of other ions was then investigated. It can
LOD (M) 8
8.7 10 3.0 10 7 1.0 10 8 5.0 10 6 5.0 10 7 8.6 10 8 2.7 10 9 8.0 10 8 7.9 10 9
Time
Ref.
– – 12 min 2 min – 3 min 30 min – 1 min
Guo et al., 2012 Cao et al., 2013 Zhang and Ye, 2011 Liu et al., 2012b Liu et al., 2012a Zhang et al., 2013 Su et al., 2010 Ding et al., 2014 This work
be seen from Fig. 5C and D, all the tested cations and anions showed nearly negligible change in the fluorescence intensity of Met-AuNCs. Fluorescence quenching of Met-AuNCs was also observed in the presence of Hg2 þ or S2 , and the tolerable limit was found to be 0.5 and 1 μM, respectively. However, the concentrations of Hg2 þ in most environmental samples are well below those of Cu2 þ , and S2 is difficult to coexist in the presence of Cu2 þ in the water (Huang et al., 2010). Therefore, Hg2 þ and S2 should not interfere with the detection of Cu2 þ in most of the practical conditions. All these results demonstrate that the rapid and green “turn off” sensor holds great promise for the determination of Cu2 þ in environmental samples. In order to validate the feasibility
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of our new-constructed method, we applied it to detect the concentrations of Cu2 þ in soil sample. Analytical results (Table S1) showed that the recoveries ranged from 92.1% to 107% in the spiked samples with the relative standard deviation (RSD) of 0.4– 2.5%, revealing high accuracy and good precision.
4. Conclusion In summary, we have demonstrated the methionine-directed synthesis of fluorescent AuNCs. The resulting AuNCs capped by methionine would be biocompatible with bioorganisms and easily functionalized with target ligands via the carboxylic or amino groups. They exhibit excellent properties, such as intense yellow fluorescence, long fluorescence lifetime, high colloidal stability, and large Stoke's shift. Combined with their photoluminescence properties, the nanoclusters have a wide prospect in the construction of light-emitting diodes, chemosensors, and biological labeling/imaging systems. As a demonstration, the Met-AuNCs were used as rapid, sensitive and selective “turn off” nanoprobes for Cu2 þ detection.
Acknowledgments We acknowledge the financial support of the National Natural Science Foundation of China (21175023), the Program for New Century Excellent Talents in University (NCET-12-0618), the Natural Science Foundation of Fujian Province (2012J06019), and the Medical Elite Cultivation Program of Fujian, PR China (2013-ZQNZD-25).
Appendix A. Supplementary material Supplementary data associated with this article can be found in the online version at http://dx.doi.org/10.1016/j.bios.2014.10.071.
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Methionine-directed fabrication of gold nanoclusters with yellow fluorescent emission for Cu2 þ sensing Hao-Hua Deng a,b, Ling-Na Zhang a,b, Shao-Bin He a,b, Ai-Lin Liu a,b, Guang-Wen Li a, Xin-Hua Lin a,b, Xing-Hua Xia c, Wei Chen a,b,n a
Department of Pharmaceutical Analysis, Fujian Medical University, Fuzhou 350004, China Nano Medical Technology Research Institute, Fujian Medical University, Fuzhou 350004, China c State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210093, China b
art ic l e i nf o
a b s t r a c t
Article history: Received 21 July 2014 Received in revised form 4 September 2014 Accepted 30 October 2014 Available online 4 November 2014
In the past few years, fluorescent gold nanoclusters (AuNCs) have gained much attention in many areas of physics, chemistry, materials science, and biosciences due to their unique physical, electrical, and optical properties. Herein, we reported for the first time the synthesis of water soluble, monodispersed AuNCs by using methionine both as a reductant and a stabilizer. The synthetic process is green and simple, and the resulting AuNCs capped by methionine (Met-AuNCs) would be biocompatible with bioorganisms. UV–visible absorption, photoluminescence, X-ray photoelectron spectroscopy (XPS), and Fourier transform infrared (FTIR) were carried out to demonstrate the chemical composition and optical properties of the as-prepared Met-AuNCs. The Met-AuNCs possess many attractive features including intense yellow fluorescence (emission maximum at 530 nm), a long fluorescence lifetime (181 ns and 1651 ns), high colloidal stability (pH-, temperature-, salt- and time-stability), and a large Stoke's shift (110 nm), holding great promise as late-model analytical tools for life science and environmental studies. Moreover, the assynthesized Met-AuNCs can serve as an efficient fluorescent probe for selective detection of Cu2 þ by fluorescence quenching. The limit of detection for Cu2 þ was determined to be 7.9 nM and linear response over the Cu2 þ concentrations range from 50 nM to 8 μM. Furthermore, the new-constructed probe allows simple and rapid detection of the concentrations of Cu2 þ in soil, with results demonstrating its great feasibility for the determination of copper in real samples. & 2014 Elsevier B.V. All rights reserved.
Keywords: Methionine Gold nanoclusters Fluorescence Copper ion Quenching
1. Introduction Metal nanoclusters, consisting of a few to roughly a hundred atoms, have become one of the important kinds of nanomaterials being extensively pursued in current nanoscience research (Liu et al., 2013b; Shang et al., 2011c). They bridge the evolution of properties of materials between isolated atoms and those of the bulk. Their excellent optical, electrical, and physical properties are very promising for applications in single-molecule optoelectronics, catalysis, biological imaging/labeling, and sensing (Chen et al., 2012a, 2012b; Han and Wang 2011; Han et al., 2012; Hyotanishi et al., 2011; Lee et al., 2005; Liu et al., 2013a, 2011; Myers et al., 2011; Nair et al., 2013; Shang et al., 2011a). Among various metal nanoclusters, considerable efforts have been dedicated to the n Corresponding author at: Fujian Medical University, Department of Pharmaceutical Analysis, 88 Jiaotong Road, Fuzhou 350004, Fujian, China. Fax: þ86 591 22862016. E-mail address: [email protected] (W. Chen).
http://dx.doi.org/10.1016/j.bios.2014.10.071 0956-5663/& 2014 Elsevier B.V. All rights reserved.
research of gold nanoclusters (AuNCs) because of their chemical stability and elegant optical properties. Over the past few years, biological molecules (such as protein, peptide, nuclear acid, and amino acid) have been widely used as eco-friendly biotemplates for metal nanocrystals synthesis (Yang et al., 2011; Zhou et al., 2009). The biomolecule-mediated nanoclusters show lower toxicity and higher biocompatibility to organisms, holding great promise as late-model analytical tools for life science and environmental studies. Quite a number of attempts have been made in this respect. Xie et al. first reported a green method for preparing highly red-emitting Au25 clusters using bovine serum albumin (BSA) as biotemplate (Xie et al., 2009). Inspired by this work, researchers found many different proteins including pepsin, lysozyme, trypsin, horseradish peroxidase, and transferrin could also serve as efficient bioscaffolds for fabricating AuNCs (Kawasaki et al., 2011a, 2011b; Le Guevel et al., 2011; Wei et al., 2010; Wen et al., 2011). Pradeep et al. synthesized two fluorescent AuNCs (namely Au25 and Au8) from mercaptosuccinic acid-stabilized gold nanoparticles by etching with a natural tripeptide, glutathione (GSH) (Muhammed et al., 2008).
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The same group also introduced a novel way for the preparation of Au23 clusters by core etching of GSH-protected Au25 clusters (Muhammed et al., 2009). Liu et al. established a method to synthesize water-soluble and red-emitting AuNCs using single-stranded DNA (ss-DNA) as a host and dimethylamine borane (DMBA) as a reductant (Li et al., 2012). Chen et al. introduced a top-down etching strategy to prepare “magic” Au8 clusters from gold particles with the help of biomolecules (protein, peptide, DNA, or amino acid) under sonication in water (Zhou et al., 2009). Later, the same group also prepared bluish green-emitting Au10 clusters through a simple manner with histidine as both reducing and stabilizing molecule (Yang et al., 2011). More recently, Yang et al. synthesized blue-emitting AuNCs by HAuCl4 and N2H2 H2O (reducing agent) in the presence of lysine as a template (Yang et al., 2013). Although considerable progress has been made, finding new biomolecules for the formation of AuNCs is still of great importance. In this work, we reported for the first time a green synthetic process of AuNCs by using methionine as both reducing and stabilizing ligands. Compared to the previously reported strategies for the synthesis of metal nanocluster, the approach presented here is simple, time-saving, and does not require toxic reductants (e.g., NaBH4). The resulting AuNCs capped/stabilized by methionine (Met-AuNCs) exhibit intense yellow fluorescence maximized at 530 nm. Moreover, the Met-AuNCs were found to be a sensitive fluorescent “turn-off” sensor for the selective monitoring of Cu2 þ .
2. Experimental 2.1. Materials All chemicals and solvents were of at least analytical grade and commercially available. HAuCl4 4H2O, methionine, NH3 H2O, and Rhodamine 6G were obtained from Aladdin Reagent Company (Shanghai, China). H2SO4, C2H5OH, and NaOH were brought from Sinopharm Chemical Reagent Co. Ltd. (Shanghai, China). Deionized water was used throughout all experiments. 2.2. Preparation of Met-AuNCs All glassware used in the following procedures was cleaned in a bath of freshly prepared solution of HNO3–HCl (1:3, v/v), rinsed thoroughly in water and dried in air prior to use. AuNCs were synthesized in a blending manner. Typically, methionine (4 mL, 0.1 M) and NaOH (0.6 mL, 0.5 M) were added to an aqueous solution of HAuCl4 (0.4 mL, 20 mg/mL). A fast fading of the light yellow color of the mixture solution was observed, indicating the formation of Au(III)-methionine complexes due to the interaction of gold ions with the sulfur of methionine. The mixture solution was incubated at 37 °C for 6 h, obtaining a pale yellow solution. After that an aqueous solution of H2SO4 (0.5 mL, 1 M) was added into the mixture to precipitate AuNCs. After centrifuged at 6000 rpm for 2 min, the AuNCs were collected and rinsed by H2SO4 (5 mL, 0.1 M) for three times. Then the AuNCs were dissolved in a solution of NH3 H2O (1.4%) and incubated at 70 °C for 30 min. The resulting solution of AuNCs was stored in the dark at 4 °C for later use. 2.3. Characterization The UV–vis absorption spectrum and the photoluminescence spectrum were measured by UV-2450 UV–vis spectrophotometer (Shimadzu, Japan) and Cary Eclipse fluorescence spectrophotometer (Agilent, USA), respectively. Time decay measurements were conducted on a F900 time-correlated single photo
counting fluorescence lifetime spectrometer (Edinburgh Analytical Instruments, UK). X-ray photoelectron spectroscopy (XPS) was performed on an ESCALAB 250 XI electron spectrometer (Thermo, USA) using monochromatic Al Kα radiation (1486.6 eV) for analysis of the surface composition and chemical states of the product. The Fourier transform infrared spectroscopy (FTIR) was measured at the wavenumber ranging from 400 to 4000 cm 1 using a Nicolet Avatar 360 FTIR spectrophotometer. All measurements were performed at room temperature under ambient conditions. 2.4. Met-AuNCs-based sensor for Cu2 þ For Cu2 þ sensing, 0.04 mL as-prepared Met-AuNCs were added into 0.36 mL phosphate buffer (10 mM, pH ¼ 7) containing different concentrations of Cu2 þ . The mixture was incubated at room temperature for 1 min. The photoluminescence spectra were measured by Cary Eclipse fluorescence spectrophotometer. 2.5. Analysis of soil sample Real samples were analyzed to estimate the analytical performance of this new sensor. Acidic digestion of soil sample was performed according to EPA method 3050B. A soil sample (0.01 g) was weighed in an Erlenmeyer flask, and 10 mL of HNO3 1:1 (v/v) was added. The solution was heated on a hot plate to ∼95 °C without boiling, and this temperature was maintained for 15 min. After cooling to less than 70 °C, 5 mL of concentrated HNO3 was added and the sample was refluxed for 30 min at ∼95 °C without boiling. This step was repeated for a second time. The sample was then evaporated to 5 mL without boiling. After cooling to less than 70 °C, 2 mL water was added followed by the slow addition of 10 mL of H2O2 (30%). Care must be taken to ensure that losses do not occur due to excessively vigorous effervescence caused by rapidly adding the strong oxidizer, H2O2. The solution was then heated until effervescence subsided. After cooling to less than 70 °C, 5 mL of concentrated HCl and 10 mL water were added and the sample was refluxed for 15 min without boiling. After cooling to room temperature, the sample was filtered and diluted to 100 mL using water (Chen et al., 2009). The obtained aqueous soil sample (1 mL) was further diluted with 49 mL water and was adjusted to pH ¼ 7 with 5 M NaOH. Afterward, 0.04 mL as-prepared Met-AuNCs were added into 0.36 mL sample solution. The mixture was incubated at room temperature for 1 min before measuring.
3. Results and discussion 3.1. Synthesis of Met-AuNCs Fig. 1A depicted the synthetic procedures of Met-AuNCs with intense yellow fluorescent emission. By incubating methionine and HAuCl4 in NaOH solution at 37 °C for 6 h, a pale yellow solution was obtained. The crude product was then purified through an acid precipitation process. After purification, the precipitate was dissolved by NH3 H2O and incubated at 70 °C for 30 min to obtain the resulting Met-AuNCs for subsequent characterization and application. The Met-AuNCs solution emits intense yellow fluorescence (Fig. 1B) under UV light irradiation. In contrast, neither pure methionine nor HAuCl4 exhibits photoluminescence under the equal conditions, demonstrating that the observed significant fluorescence originates from the gold core of the Met-AuNCs. Photographs of the solutions mentioned before in room light were also showed in Fig. 1B.
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Fig. 1. (A) Schematics of the formation of Met-AuNCs. (B) Photographs of methionine solution in room light (a) and UV light at 365 nm (d), HAuCl4 solution in room light (b) and UV light at 365 nm (e), and Met-AuNCs solution in room light (c) and UV light at 365 nm (f).
Additional experiments were carried out to optimize the synthesis of brightly luminescent AuNCs. The synthetic procedure consists of two steps: (i) slow reducing process to form relatively weak fluorescent AuNCs, and (ii) fast ripening process to generate intense fluorescent AuNCs. Regarding the first step, experiments involving the influence of the concentration of methionine, the concentration of NaOH, and the reaction time were performed. It can be seen from Fig. S1, the fluorescence intensity of the product was the most intense when the methionine concentration was increased to 0.1 M and diminished if the methionine concentration increased further. The influence of the concentration of NaOH on the fluorescence intensity of AuNCs was revealed in Fig. S2. It is clear that a concentration of 0.5 M gives the highest intensity of the product. Under the optimal conditions, the first-step reaction proceeded to completion within 6 h (Fig. S3). As for the second step, the influence of the concentration of NH3 H2O, ripening temperature, and time was taken into consideration. From Fig. S4, it can be seen that the photoluminescence intensity of the MetAuNCs increased when the concentration of NH3 H2O increased from 0.07% to 1.4% and no distinct change was observed if the concentration increased further. The highest fluorescence intensity was achieved when the ripening process was performed at 70 °C for 30 min (Figs. S5 and S6). Under the optimal conditions, the Met-AuNCs obtained in the second-step show fluorescence about 3 times higher than those obtained in the first-step. Although the exact mechanism of the ripening process is not yet clear, there is
almost no change of the wavelength of maximum peaks in the excitation and emission spectra before and after the ripening process (Fig. S7), demonstrating that the emitters from the two steps are the same components. 3.2. Characterization of Met-AuNCs Fig. 2A displays the absorption spectrum of the as-prepared Met-AuNCs. The product missed the specific surface plasmon resonance (SPR) peak (around 520 nm) of larger gold nanocrystals. And instead, it showed a weak but noticeable absorption band centered on 330 nm. In comparison, the absorption spectra of pure methionine and HAuCl4 were also included in Fig. 2A and no peaks at 330 nm were observed in the two samples. By fluorescence spectrophotometry (excited at 420 nm), the Met-AuNCs solution exhibited strong photoluminescence around 530 nm (Fig. 2B). Besides, neither pure methionine nor HAuCl4 exhibited obvious luminescence under the equal conditions. Thus, the observed strong fluorescence should stem from the gold core of the resulting AuNCs. The emission spectrum shows unchanged over a wide excitation wavelength range of 360–460 nm (Fig. S8), indicating that the emission of AuNCs should be from the relaxed states rather than scattering effects (Shang et al., 2011b). It is worth mentioning that, contrast to semiconductor quantum dots (QDs), the AuNCs demonstrated here show a clear excitation band, which would be propitious to Förster resonance energy transfer
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Fig. 2. (A) UV–vis absorption spectra of the aqueous Met-AuNCs, HAuCl4, and methionine. (B) Photoluminescence excitation and emission spectra of Met-AuNCs, and emission spectra of methionine and HAuCl4 were taken as controls. (C) Au(4f) XPS spectrum of Met-AuNCs. (D) S(2p) XPS spectrum of Met-AuNCs.
(FRET) applications. The luminescence quantum yield (QY) of MetAuNCs was 2.8%, calculated with Rhodamine 6G (QY ¼0.95 in ethanol) as the reference, which is nearly two magnitude orders higher than those AuNCs prepared by using NaBH4 as a reductant (Negishi et al., 2004). Additionally, the dominant fluorescence lifetime of the Met-AuNCs was about 181 ns (44.06%) and 1651 ns (55.94%) (Fig. S9), providing a neat opportunity for fluorescence lifetime imaging in future (Shang et al., 2011a). Transmission electron microscopy (TEM) was used to confirm the structure of as-synthesized AuNCs, which revealed an average diameter of about 2.5 nm for the Met-AuNCs (Fig. S10). Electronic structure of the Met-AuNCs sample was conducted by X-ray photoelectron spectroscopy (XPS). The Au(4f) XPS spectrum (Fig. 2C) manifested the binding energies of Au(4f5/2) and Au(4f7/2) located at 88.1 and 84.2 eV, respectively, which explicitly suggests that the electronic structures of Au in Met-AuNCs are Au (0) and Au(I), coexisting (Shang et al., 2011a, 2011b; Zhou et al., 2010). According to reports, the Au(I) surrounded on the surface of AuNCs core plays a vital role in the formation of highly fluorescent AuNCs (Wang et al., 2013). After further reduction of the MetAuNCs with NaBH4, the fluorescence of Met-AuNCs showed a 77% decrease and the corresponding absorption in visible range displayed a slight increase (Fig. S11). The S(2p) XPS spectrum showed a dominant peak located at 162.6 eV, which is attributed to the binding of sulfur atoms of methionine to the gold surface (Fig. 2D). In addition, C(1S), O(1S), and N(1S) core-level photoemission spectra shown in Fig. S12 derive from the protecting ligand, making clear the AuNCs are capped by methionine. Infrared measurement of Met-AuNCs (Fig. S13) showed a suppression of the -C-S stretching vibration in the region of 790–830 cm 1 but
nearly no change of νasNH3 þ (1630 cm 1) and νasCOO (1530 cm 1) (Cao and Fischer, 2002; Vujacic et al., 2009), indicating that S-atom but not ‒NH2 and ‒COOH groups from the methionine molecule is involved in the formation of coordinative bonds in the Met-AuNCs. The functional groups (‒NH2 and ‒ COOH) from methionine on the surface of nanoclusters may favor further modification of the AuNCs. 3.3. Stability of fluorescent Met-AuNCs High stability of fluorescent nanomaterials is an important ingredient to evaluate the possibility of their practical applications. As shown in Fig. 3A, the curve which represents the fluorescence intensity of Met-AuNCs at 530 nm under various pH values from 2 to 12 fluctuates slightly, demonstrating that the Met-AuNCs have a relative high pH-stability. Apart from pH stability, temperature stability is another indispensable factor to ensure that the fluorescent nanomaterials are practical for applications. As shown in Fig. 3B, the curve which depicts the fluorescence intensity of AuNCs suffered a small change when temperature varied from 20 to 70 °C, implying that the Met-AuNCs have a high temperaturestability. Moreover, the variation in fluorescent intensity of the Met-AuNCs exposed to different concentrations of NaCl was examined. The results revealed in Fig. 3C show that the fluorescent intensity of the AuNCs decreased with the increasing concentration of NaCl. However, the fluorescent intensity of as-prepared Met-AuNCs preserved 50% of the initial intensity even in high ionic strength (1 M NaCl). Moreover, there was a small quenching of the fluorescent intensity (only a 6.8% decrease) and no change in the position of the emission peak for Met-AuNCs after storing in
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Fig. 3. The fluorescence stability of Met-AuNCs under (A) various pH values, (B) various temperatures, and (C) different concentrations of NaCl. (D) The fluorescence spectra of as-prepared Met-AuNCs before and after storing in dark at 4 °C for two months.
the dark at 4 °C for two months (Fig. 3D). This demonstrates that the as-prepared Met-AuNCs are particularly suitable for long-term use. It should be noted that the ultra-high stability of the MetAuNCs comes from two crucial factors: protective shell formed by methionine around AuNCs and stabilization effect of Au (I) surrounded on the surface of AuNCs core (Wang et al., 2013). Due to their ultra-high stabilities, the as-synthesized Met-AuNCs may broaden the application of fluorescent nanomaterials. 3.4. Fluorescent sensing of Cu2 þ using Met-AuNCs With improved features in terms of simplicity, selectivity, sensitivity, and miniaturizability, metal nanoclusters (e.g., Au and AgNCs or Au/AgNCs) have been used to construct fluorescent sensors for the detection of specific analytes of chemical as well as biological origin (Yuan et al., 2013). In this study, we found that Cu2 þ could selectively quench the fluorescence of Met-AuNCs, which can be used as a “turn off” probe for Cu2 þ . In order to verify whether the quenching fluorescence of Met-AuNCs was due to the coordination of Cu2 þ with methionine, which could impede Au–S charge transfer, or the fluorescence quenching by metal–metal interaction, ethylenediaminetetraacetate (EDTA) was used as chelator in competition with Met-AuNCs for Cu2 þ . It can be seen from Fig. 4, the fluorescence intensity of Met-AuNCs, after quenching by Cu2 þ , could effectively restore (94% of its initial value) when EDTA was introduced, which indicated that the interaction between Cu2 þ and Met-AuNCs was indeed through coordination.
Fig. 4. (A) Fluorescence regeneration results for quenching by Cu2 þ upon the addition of EDTA. (a) Met-AuNCs, (b) Met-AuNCs in presence of 10 μM Cu2 þ , and (c) Met-AuNCs containing 10 μM Cu2 þ after the addition of 20 μM EDTA. (B) The corresponding photographs.
Considering the appreciable changes in fluorescent properties of Met-AuNCs toward Cu2 þ , the potential of establishing a novel fluorescent probe for Cu2 þ determination was assessed. The kinetic behaviors of fluorescence of Met-AuNCs in the presence of Cu2 þ were monitored and the results show that the interaction between Cu2 þ and Met-AuNCs could be fulfilled within few seconds to quench the fluorescence (Fig. S14). With the concentrations of Cu2 þ increasing, the fluorescence intensity of Met-AuNCs gradually declined (Fig. 5A and B). There is a linear relationship between fluorescence quenching and the concentrations of Cu2 þ within a range from 50 nM to 8 μM (Fig. 5B inset). The limit of
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Fig. 5. (A) Emission spectra of Met-AuNCs in the presence of varying concentrations of Cu2 þ . a–k: 0, 0.05, 0.1, 0.2, 0.5, 1, 2, 4, 6, 8, 10 μM. (B) The plot of the fluorescence intensity (I) of Met-AuNCs recorded at 530 nm versus the concentration of Cu2 þ . Inset: linear relationship between the logarithm of I and the concentration of Cu2 þ . (C) Fluorescence quenching effect [(I0 I)/I0] of the Met-AuNCs at 530 nm incubated with a concentration of 6 μM different tested cations. Samples marked with 1–18 corresponding to Cu2 þ , Co2 þ , Ni2 þ , Mn2 þ , Ca2 þ , Mg2 þ , Ba2 þ , Cd2 þ , Pb2 þ , Zn2 þ , Fe2 þ , Fe3 þ , Al3 þ , Cr3 þ , Ag þ , Na þ , K þ , and NH4 þ , respectively. (D) Fluorescence quenching effect [(I0 I)/I0] of the Met-AuNCs at 530 nm incubated with a concentration of 6 μM different tested anions. Samples marked with 1–18 corresponding to Cu2 þ , SCN , F , Cl , Br , I , ClO4 , BrO3 , IO3 , SO32 , S2O32 , S2O82 , NO2 , NO3 , Ac , CO32 , H2PO4 , and EDTA2 , respectively.
Table 1 Comparison of the proposed method with other reported approaches for the detection of Cu2 þ based on fluorescent nanoclusters. Probe Mercaptoundecanoic acid-AuNCs BSA-AuNCs DNA-AgNCs Azobenzene modified poly(acrylic acid)-AgNCs DNA-AgNCs Glutathione-AuNCs DNA-Cu/Ag NCs Dithiothreitol-AuNCs Met-AuNCs
Linear range (M) 8
6
1.0 10 –1.0 10 5.0 10 7–1.0 10 4 0–1.0 10 5 5.0 10 6–1.5 10 4 5.0 10 6–1.25 10 4 1.0 10 7–6.25 10 6 5.0 10 9–2.0 10 7 5.0 10 6–5.0 10 5 5.0 10 8–8.0 10 6
detection (LOD) was 7.9 nM (3s) and the relative standard deviation (RSD) was 1.4% for the determination of 1 μM Cu2 þ (n¼ 9). The fluorescence quenching constant (Ksv) was calculated to be 5.5 105 M 1 by linear regression of the plot based on the Stern– Volmer equation, I0/I ¼1 þKsv[Q], where I0 and I are the fluorescence intensity of the Met-AuNCs at 530 nm in the absence and presence of Cu2 þ , and [Q] is the concentration of Cu2 þ . This approach was comparable and even more efficient for the detection of Cu2 þ compared to other nanoclusters-based fluorescence methods previously reported (Table 1). To test whether this quenching behavior by Cu2 þ is selective, the fluorescence of MetAuNCs in the presence of other ions was then investigated. It can
LOD (M) 8
8.7 10 3.0 10 7 1.0 10 8 5.0 10 6 5.0 10 7 8.6 10 8 2.7 10 9 8.0 10 8 7.9 10 9
Time
Ref.
– – 12 min 2 min – 3 min 30 min – 1 min
Guo et al., 2012 Cao et al., 2013 Zhang and Ye, 2011 Liu et al., 2012b Liu et al., 2012a Zhang et al., 2013 Su et al., 2010 Ding et al., 2014 This work
be seen from Fig. 5C and D, all the tested cations and anions showed nearly negligible change in the fluorescence intensity of Met-AuNCs. Fluorescence quenching of Met-AuNCs was also observed in the presence of Hg2 þ or S2 , and the tolerable limit was found to be 0.5 and 1 μM, respectively. However, the concentrations of Hg2 þ in most environmental samples are well below those of Cu2 þ , and S2 is difficult to coexist in the presence of Cu2 þ in the water (Huang et al., 2010). Therefore, Hg2 þ and S2 should not interfere with the detection of Cu2 þ in most of the practical conditions. All these results demonstrate that the rapid and green “turn off” sensor holds great promise for the determination of Cu2 þ in environmental samples. In order to validate the feasibility
H.-H. Deng et al. / Biosensors and Bioelectronics 65 (2015) 397–403
of our new-constructed method, we applied it to detect the concentrations of Cu2 þ in soil sample. Analytical results (Table S1) showed that the recoveries ranged from 92.1% to 107% in the spiked samples with the relative standard deviation (RSD) of 0.4– 2.5%, revealing high accuracy and good precision.
4. Conclusion In summary, we have demonstrated the methionine-directed synthesis of fluorescent AuNCs. The resulting AuNCs capped by methionine would be biocompatible with bioorganisms and easily functionalized with target ligands via the carboxylic or amino groups. They exhibit excellent properties, such as intense yellow fluorescence, long fluorescence lifetime, high colloidal stability, and large Stoke's shift. Combined with their photoluminescence properties, the nanoclusters have a wide prospect in the construction of light-emitting diodes, chemosensors, and biological labeling/imaging systems. As a demonstration, the Met-AuNCs were used as rapid, sensitive and selective “turn off” nanoprobes for Cu2 þ detection.
Acknowledgments We acknowledge the financial support of the National Natural Science Foundation of China (21175023), the Program for New Century Excellent Talents in University (NCET-12-0618), the Natural Science Foundation of Fujian Province (2012J06019), and the Medical Elite Cultivation Program of Fujian, PR China (2013-ZQNZD-25).
Appendix A. Supplementary material Supplementary data associated with this article can be found in the online version at http://dx.doi.org/10.1016/j.bios.2014.10.071.
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