Accepted Manuscript Title: An ultrasensitive method for the determination of melamine using cadmium telluride quantum dots as fluorescence probes Author: Xiafei Li Jin Li Huiyan Kuang Lei Feng Shoujun Yi Xiaodong Xia Haowen Huang Yong Chen Chunran Tang Yunlong Zeng PII: DOI: Reference:
S0003-2670(13)01244-0 http://dx.doi.org/doi:10.1016/j.aca.2013.09.042 ACA 232854
To appear in:
Analytica Chimica Acta
Received date: Revised date: Accepted date:
15-5-2013 14-9-2013 19-9-2013
Please cite this article as: X. Li, J. Li, H. Kuang, L. Feng, S. Yi, X. Xia, H. Huang, Y. Chen, C. Tang, Y. Zeng, An ultrasensitive method for the determination of melamine using cadmium telluride quantum dots as fluorescence probes, Analytica Chimica Acta (2013), http://dx.doi.org/10.1016/j.aca.2013.09.042 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
An ultrasensitive method for the determination of melamine using
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cadmium telluride quantum dots as fluorescence probes
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Xiafei Lia,b, Jin Lia,b, Huiyan Kuanga,b, Lei Fenga,b, Shoujun Yia,b, Xiaodong Xiaa,b,
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Haowen Huanga,b, Yong Chena, Chunran Tanga, Yunlong Zenga,b,c,*
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School of Chemistry and Chemical Engineering , Hunan University of Science and Technology, Xiangtan 411201,
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PR China
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b
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University of Science and Technology, Xiangtan 411201, PR China
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Key Laboratory of Theoretical Chemistry and Molecular Simulation of Ministry of Education of China, Hunan
State Key Laboratory for Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering,
Hunan University, Changsha 410082, PR China
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Abstract: An ultrasensitive and simple method for the determination of melamine was developed
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based on the fluorescence quenching of thioglycolic acid (TGA) capped CdTe quantum dots (QDs)
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at pH 11.0. In strong alkaline aqueous solution, the selectivity of the method has been greatly
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improved due to most heavy metal ions show no interference as they are in the precipitation form or in their anion form. Furthermore, CdTe quantum dots have higher quantum yields at higher pH. The method has a wider concentration range and lower detection limit. The influence factors on the determination of melamine were investigated and the optimum conditions were determined. Under
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optimum conditions, the fluorescence intensity change of TGA coated CdTe quantum dots was
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linearly proportional to melamine over a concentration range from 1.010-11 to 1.010-5 mol L-1
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with a correlation coefficient of 0.9943 and a detection limit of 5×10-12 mol L-1. The mechanism of
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fluorescence quenching of the QDs has been proposed based on the infrared spectroscopy
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information and electrophoresis experiments in presence of melamine under alkaline condition. The
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proposed method was employed to detect trace melamine in milk powder and pet feeds with
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satisfactory results.
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Keywords: Melamine; Ultrasensitive determination; High selectivity, Cadmium telluride quantum
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dots; Fluorescence probe.
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1. Introduction
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Melamine (1,3,5-triazine-2,4,6-triamine,C3H6N6) is a chemical compound used commonly for
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the production of melamine resins. As its high nitrogen level (66.6% by mass), melamine is illegally
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added to food, such as milk, cookies and animal feeds, which produces an incorrectly high reading
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in the measurement of protein content based on total nitrogen, resulting some serious melamine
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residue related problems [1]. Since the melamine contaminated animal feeds incident in USA in
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2007 and the accident of San Lu poisonous milk accident in China in 2008, the great importance of
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detecting melamine has been well recognized and the methods for melamine determination in
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various samples have been developed [2]. Most techniques for melamine assay such as GC/MS [3,4],
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HPLC [5,6] and CZE [7] require expensive apparatus and considerable time for identification and quantification. Hence, it is necessary to develop some simple, highly sensitive, selective method for the analysis of melamine in real samples. Recently, nanoparticles have attracted increasing attention in bioanalysis as their unique optical
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Technology, Xiangtan 411201, PR China. Tel.: (+86)13787423309
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E-mail address: [email protected]
Corresponding author at: School of Chemistry and Chemical Engineering, Hunan University of Science and
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and electronic properties. Several simple, fast, sensitive methods for the determination of melamine
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have been established [8−10] based on the change of visual colorimetric by the aggregation of some
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metal nanoparticles such as silver nanoparticles, gold nanoparticles. As a kind of typical
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nanomaterials, luminescence semiconductor quantum dots (QDs), such as CdS, CdSe, and CdTe,
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have attract great interest in recent years due to their quantum confinement effect and remarkable
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optical, electrical properties [11,12]. They have been widely used in biology, medicine, analytical
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chemistry, especially in sensors [13−15]. Recently, Gao [16] has developed a high sensitive method
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to detect melamine based on the energy transfer of CdTe quantum dots (CdTe@SiO2). This method
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has a wide linear range for melamine determination from 7.5 × 10−9 to 3.5 × 10−7 mol L-1 with a
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detection limit of 8.9 × 10−10 mol L-1. However, the methods for the determination of melamine was
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often at pH 5.0 − 8.0 [17−19]. There are no research reports about the detection of melamine in
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strong alkaline solution based on noble metal nanoparticles or quantum dots so far as we know. As
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known, heavy metal ions can strongly change the fluorescence intensity and ultraviolet-visible signal
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of nanoparticles (noble metal nanoparticles and quantum dots) [20−22], which will interfere with the
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target object analysis. But in strong alkaline solution, the selectivity and detection limit can be greatly improved as heavy metal ions are in their precipitate form or in their anion form at high pH. In this work, we have proposed an ultrasensitive method for the determination of melamine
using water-soluble TGA-CdTe QDs as the fluorescence probes at pH 11.0. Under alkaline
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conditions, CdTe QDs have high quantum yields( QYs), and heavy metal ions free from interference
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as they are in the form of precipitation or in their anion form. So the method showed high selectivity
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and sensitivity. This method was applied to detecting melamine in milk power and pet feeds with
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satisfactory results.
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2. Experimental
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2.1 Apparatus Fluorescence spectra were acquired on a FL 4500 Fluorescence Spectrometer (Hitachi, Japan).
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UV-vis absorption spectra were recorded on Lambda 35 spectrometer (Perkin-Elmer, USA). IR
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spectra were measured on a Spectrum One FT/IR spectrometer (Perkin–Elmer, USA). All optical
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measurements were carried out at room temperature.
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2.2 Reagent
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Tellurium dioxide (TeO2), sodium borohydride (NaBH4), cadmium nitrate (Cd(NO3)2),
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thioglycolic acid (TGA) and melamine were purchased from Shanghai Chemical Reagent Company
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(Shanghai, China). All other chemical reagents were of analytical-reagent grade and were used as
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received without further purification. Stock standard aqueous solution of melamine was prepared at
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a concentration of 0.01mol L-1, and other concentration of melamine was prepared by diluting the
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stock standard solution with double distilled water. The milk powder was purchased from the
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supermarket in Xiangtan. Double distilled water was used throughout all experiments.
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2.3 Preparation of water-soluble CdTe QDs The TGA-CdTe QDs, which with a molar ratio of TGA:Cd2+:Te2- was 3.9:1:0.5, was prepared
by using the reaction between Cd2+ and NaHTe solution at pH 12.0 with TGA as the stabilizing
reagent. Briefly, 0.08 g TeO2 powder was reduced by 0.05g NaBH4 in 5 ml water under a cold-water
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bath (about 15) for about 4 h, and then the aqueous solution was injected into 95 ml oxygen-free
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aqueous solution containing a certain amount of TGA and 1.0×10−3 mol L-1 of Cd(NO3)2 at pH 12.0.
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The resulting mixture was refluxed in boiling water bath at dark for 25 min under vigorously
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magnetic stirring. The TGA-CdTe QDs were obtained and stored in 4 at dark for use. The final
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concentration of CdTe QDs was approximately 5.0×10-3 mol L-1 (according to the Te2-
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concentration).
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2.4 Isolation and extraction of melamine
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According to the previous method reported by Li [17], the sample was prepared with some
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modifications and described as follows: First, 2.5 g sample was dissolved with 25 ml of 0.6 mol L-1
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trichloroacetic acid and mixed with a vortex for 5 min to precipitate protein in the sample matrix.
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And the mixture was centrifuged at 8000 r/min for 10 min. Then, the protein content in the
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supernatants was determined by Bicinchoninic Acid method [23]. After the protein in the filtrate was
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removed, the supernatants were transferred into a 25 ml beaker, adjusted to pH 7.0 with a small
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amount of 2 mol L-1 NaOH and filtered with a 0.22 μm filter. The obtained filtrate was the sample
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solution and was used in the following detection.
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2.5 Detection of melamine
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Melamine determination was carried out by successively adding 0.5 ml varying concentrations
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of melamine or sample solution into 4.5 ml 1×10-4 mol L-1 CdTe QDs solution (pH 11.0). The
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fluorescence emission of the solution was recorded at the peak wavelength (about 535 nm) with the excitation wavelength of 465 nm. All measurements were carried out under room temperature. 3. Results and discussion 3.1 Influence of pH
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The influence of pH on the luminescence quantum yield(QY) of CdTe QDs was investigated in
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different pH. The QY was measured and the results are recorded in Fig.1, by referring to Rhodamine
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6G (QY 95%) with the method described in Ref. [24]. As known, the photoluminescence of CdTe
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NCs was attributed to defect-related emission. The QYs of the NCs was strongly dependent on pH.
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As shown in Fig. 1, at lower pH, QYs of the QDs increased linearly with rise of pH and the QYs
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reaches a maximum (51.9 %) at the pH 11.0 and then decreased. This is because of the extremely
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small diameter, there are many trap sites on the surface of CdTe NCs, the more the trap sites, the
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lower the fluorescence efficiency. As known, TGA is a weak acid and the dissociation constants of
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TGA are pKa1 = 3.6 and pKa2 = 10.6 [25, 26]. With rise of pH, TGA2- (−SCH2COO−) increased, and
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more TGA2- coordinated with Cd2+ on the surface of the CNs. So the relative coverage rate of the
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particle surface with thiols was increased. Therefore, the trap sites on the CdTe surface will be
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decreased, and the fluorescence efficiency was improved. However, at higher pH(higher than 11.0),
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the relative fluorescence intensity of the QDs also tended to decrease. This phenomenon might be
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resulted from the formation of cadmium hydroxide product on the surface of QDs at higher pH [27].
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Insert Fig. 1
As seen from Fig.1, when the pH is less than 5.0, the QY of the QDs is very low, and the
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fluorescence intensity of the system is very weak. Therefore, the influence of pH on the performance of the method was investigated and the results were recorded in Table 1 at pH 5.0 to 12.0. From Table 1, one can notice that the method has a wider linear concentration range and lower detection limit at pH 11.0. The result shows the advantage of high sensitivity for melamine assay under strong
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alkaline condition. This is because high QY of the NCs is crucial for realizing a good sensitivity of
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the fluorescent sensors. The sensitivity for an analyte is determined at the signal-to-noise, which is
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largely dependent on the QYs of the fluorescent probes. Fluorescent probes with higher QYs can
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achieve the same luminescence intensity at lower concentrations relative to NCs with lower QYs,
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thus leading to a higher sensitivity.
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Quantum dots are always sensitive to metal ions. Hence, the influence of pH on the
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fluorescence intensity in the presence of some metal ions (1.0×10-6 mol L-1) was investigated (see
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Fig. S1). The fluorescence intensity of the QDs was changed obviously at pH 6.5 and almost was not
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changed at pH 11.0 in the presence of heavy metal ions(such as Fe3+, Cu2+, Hg2+, Pb2+ and Zn2+). In
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other words, heavy metal ions show interference at pH 6.5 and show no interference at pH 11.0.
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Presumably the heavy metal ions are in the form of precipitation or in their anion forms under strong
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at alkaline condition and free from interference.
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Melamine, a Lewis base that can accept a proton to form conjugate acid, may hydrolyze in an
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aqueous solution. Therefore, the media pH affects the form and stability of melamine. Bozzi [28] has
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studied the photocatalysis of melamine and pointed out that melamine remains stable in aqueous
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solution in detail. To examine the stability of melamine solution at pH 11.0, the experiments of fluorescence quenching of CdTe quantum solution by 1.0×10-7 mol L-1 melamine at pH 7.0 and pH 11.0 were carried out, respectively. There were no obvious differences in the quenched fluorescence intensity when the melamine solution was stored for 0 − 5.0 h at pH 11.0 or at pH 7.0, which
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indicated that melamine was stable and can be determined accurately in strong alkaline solution. So
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the following experiments were performed at pH 11.0.
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3.2 Influence of TGA molar ratio
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The influence of TGA molar ratio on the properties of fluorescence and UV–vis absorption of
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the QDs was investigated. Fig.2 depicts the fluorescence and UV–vis absorption spectra of
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TGA-CdTe QDs at different molar ratio of TGA and Cd2+. From Fig. 2 (a), one can see that the
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UV–vis absorption peaks are red shift with increasing TGA, which indicates that the size of the QDs
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increased with increasing the ratio of TGA. The diameters of as-prepared CdTe QDs were about 2.2,
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2.4, 2.5, 2.6, 2.8, 2.9 and 3.2 nm, respectively, corresponding to TGA ratio of 1.7, 2.4, 3.2, 3.9, 4.6,
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5.3 and 6.0, respectively, according to the calculation reported in literature [29]. It is obviously that
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the concentration of free cadmium ions decreased with the increasing TGA because of the formation
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of TGA-Cd2+ complex, leading to form larger diameter CdTe nanocrystals. From Fig. 2 (b), one can
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also see that the TGA-CdTe QDs generally have a broad emission ranging from 460 nm to 680 nm
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and the fluorescence spectra do not exhibit a tail on the right side, suggesting that the prepared QDs
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possess good fluorescence properties. All of the fluorescence spectra band are relatively narrow
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(with full width at half maximum of about 30~40 nm) and symmetric indicating that the prepared
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CdTe QDs are nearly mono-disperse and homogeneous.
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Insert Fig. 2
The size dependent sensitivity has been reported in photoluminescence, chemiluminescence
and electrochemiluminescence[30−33]. In this work, different sized CdTe QDs were used to study
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the influence of the size effect on the sensitivity and linear range for melamine assay, and the results
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were recorded in Table 2.
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Insert Table 2
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177 From Table 2, one can notice that the sensitivity of the method increased the maximum and
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then decreased with increasing size of the QDs. Furthermore, the linear concentration range reached
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the widest as the diameter of CdTe QDs is about 2.6 nm in the molar ratio of TGA: Cd2+: Te2- at 3.9:
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1: 0.5. This may be the most suitable size for melamine assay.
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3.3 Selection of the QDs concentration
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The fluorescence of the QDs can be quenched strongly by melamine. The fluorescence
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intensity of the solution changes very slightly, when a small amount of melamine was added to the
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solution with high concentration of CdTe QDs. The results indicated that the amount of QDs in the
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solution will greatly influence the detection sensitivity and linear range for melamine assay. So
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series of concentration of CdTe QDs was employed to investigate the effect of QDs concentration on
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the measurement(as shown in fig. S2). A wider concentration range and lower detection limit
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were obtained when 1.0×10-4 mol L-1 of the CdTe QDs solution was used. Consequently, this QDs
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concentration was chosen for the following experiments.
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3.4 Effect of reaction time
The influence of reaction time on the fluorescence intensity of the QDs in the presence of
melamine (1.0×10-8, 1.0×10-9, 1.0×10-10 mol L-1 respectively) was investigated. The results showed
that the fluorescence intensity of the QDs decreased rapidly when melamine was added to the QDs
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solution. After 5 min, the fluorescence intensity reached the lowest and kept unchanged more than
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50 min. So the reaction time was fixed at 10 min.
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3.5 Influence of coexisting substance
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To evaluate the detection selectivity, the influence of common coexisting ions in milk powder
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on the fluorescence quenching of the CdTe QDs in the presence of 1.0×10-7 mol L-1 melamine were
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investigated under optimum conditions. The tolerable concentration ratios relative to concentration
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levels usually found in milk for interference at the 5% level were over 1000 fold for Na+, K+, Cl-,
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NO3- and SO42-, 600 fold for Ca2+, Mg2+, PO43-, 200 fold for Fe3+, Cu2+, Zn2+, and 120 fold for Vc
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and Lactos. The results demonstrated that common metal ions had no interferences in detection of
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melamine. So, this method has the characteristic of high selectivity for the determination of
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melamine, as the heavy metal ions are in the form of precipitation or in their anion forms under
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strong at alkaline condition.
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3.6 Calibration curve and detection limit
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Under optimum conditions, a high linearity was observed (insert of Fig. 3) in the melamine
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concentration range from 1.0×10-11 to 1.0×10-5 mol L-1. The detection limit (LOD) is defined by the
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equation LOD = 3 S0/K, where S0 is the standard deviation of blank measurements (nine
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measurements, n = 9), K is the slope of calibration graph. The LOD is 5×10-12 mol L-1 and the
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relative standard deviation is 3.8 % for the determination of 1.0×10-8 mol L-1 melamine (n = 11).
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Insert Fig. 3
Comparison between our proposed method and some methods reported recently for the
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determination of melamine is presented in Table 3. The present method showed lower detection limit
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and wider linear range.
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Insert Table 3
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3.7 Applications The proposed method was employed to detect melamine in milk powder and pet feeds samples
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(Table 4) with satisfactory results. And the recovery studies were carried out on samples to which
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known amounts of melamine were added. The results are illustrated in Table 4, with satisfying
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recoveries of 97.3 − 105 %, suggesting that the proposed method was reliable and practical.
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The interaction between TGA and CdTe NCs was investigated by the FTIR spectrometry. As
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shown in Fig. S3 b, the absorption band due to S–H vibration observed at 2560 cm–1 disappeared
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and the characteristic absorption band of C=O vibration was shifted from 1700 cm–1 to 1556 cm–1,
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which strongly suggest that the thiol groups of TGA coordinated with Cd2+ ions on the NCs surface,
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and the hydrophilic hydroxyl groups faced outward, making NCs water-soluble. Generally, more than 15% of the atoms in a quantum dot are located on its surface [40], and the
luminescence of QDs is sensitive to their surface conditions. The molecular interactions with the surface of luminescence QDs would change their photophysical properties [41, 42]. As known, TGA
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exists in the form of anion and melamine is in the form of neutral molecular under alkaline condition.
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Therefore, TGA-CdTe QDs carry negative charge. As there were excess of cadmium ions being used
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in the preparation of TGA-CdTe QDs (as mentioned above, the molar ratio of TGA: Cd2+: Te2-: is 3.9:
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1: 0.5), the cadmium telluride crystals will carry positive charge as their surfaces attract these
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excessive ions in the solution. However, TGA-CdTe QDs carry negative charge owing to
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–
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melamine solution was added to this system, TGA on the nanoparticle surfaces was to be displaced
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by melamine to form melamine coated CdTe QDs in alkaline solution. The melamine coated CdTe
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QDs carry positive charge, as melamine is weak base (pKb= 9.0) and exists almost in neutral
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molecule at pH 11.0. Therefore, there are two opposite charge CdTe QDs in the presence of
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melamine. As a result, the fluorescence of QDs was quenched when the QDs aggregation occurred
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by electrostatic attraction of these two opposite charge nanoparticles. The mechanism can be
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described as following:
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SCH2COO–(TGA2-) coordinated with Cd2+ ions on the NCs surface. When the high concentration
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Insert Scheme 1
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The presumption was verified by IR information (Fig. S3) and electrophoresis (Fig.S4).
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Fig.S3 shows the infrared spectra of the TGA (Fig. S3a), TGA−capped CdTe QDs (Fig. S3b)
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and the QDs in the presence of melamine (Fig. S3c). The IR absorption broad bands (Fig. 4a) around, 3026.26– 3435.88 cm−1 (sυ O-H), 1386.58 cm−1 (mυ COO−), 1716.01 cm−1 (υ C=O), 1291.82 cm−1 (δ OH) indicate −COOH group. The peak at 2923.70 cm−1 (wυ C-H) represents -CH2 group. The
peak at 2566.14 cm−1 (wυ S-H) represents -SH group. There are coexisting IR absorption bands of
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-COOH, -CH2 and -SH on free TGA (see form Fig. S3a), whereas only bands -COO− (1386.21 cm−1,
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mυ COO−) and free -CH2 (2923.70 cm−1, wυ C-H) were observed on the surface of the TGA−capped
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CdTe QDs (seen from Fig. S3b). The disappearance of S-H group vibration 2566.14 cm−1 (wυ S-H)
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on the surface of functionalized CdTe QDs resulted from the covalent bonds between thiols and the
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Cd atom of the surface of CdTe QDs. Compared with TGA−capped CdTe QDs (Fig. S3b), the
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R-COO− (1576.60 cm−1) and –CH2 (2923.70 cm−1) disappeared in the presence of melamine(Fig.
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S3c), which indicated the loss of TGA capping ligand of CdTe QDs. On the other hand, the –NH2
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( 3136.08 – 3470.11 cm−1, sυ N-H ) and triazine ring (1551.26 cm−1) appear, which indicated the
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surface of CDTe QDs coated with melamine. The reaction was as following:
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[QDs-(SCH2COO2−)i]m- + j C3N6H6 → [QDs-(C3N6H6)j]n+ + i (SCH2COO)2-
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Figure S4 shows the electrophoresis of TGA-CdTe QDs and the melamine coated CdTe QDs,
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respectively. One can note that TGA-CdTe QDs move to the cathode (Fig. S4B) means them
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carrying a negative charge and that melamine-dTe QDs move to the anode electrode( Fig. S4 D)
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indicate the QDs with positive charge applying an electric field (100V). Hence, the mechanical of
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fluorescence quenched can first be the TGA on the surface of TGA- CdTe QDs with negative charge
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was placed by melamine to form positive charged melamine coated CdTe QDs. And then these two
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types of opposite charge nanoparticles aggregated by electrostatic attraction. As a result, the
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fluorescence of CdTe QDs quenched.
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as heavy metal ions in the form of precipitation or in their anion form. Hence, the method can be
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free from interference with metal ions in strong alkaline solution. Second, the lower concentration of
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the QDs can be used for the determination of melamine as the CdTe QDs have higher quantum yield.
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The method shows a wider linear concentration range and lower detection limit. Furthermore, the
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4. Conclusion
In summary, we have developed an ultrasensitive and simple method for the determination of
melamine using the QDs as fluorescence probes in strong alkaline media. The analysis of melamine at alkaline condition has advantages: First, the selectivity of the method has been greatly improved
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size dependent sensitivity appeared using QDs probe for melamine assay. The linear concentration
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range reached the widest with most sensitivity as the diameter of CdTe QDs is about 2.6 nm in the
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molar ratio of TGA: Cd2+: Te2- at 3.9: 1: 0.5. This may be the most suitable size for melamine assay.
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At last, the experiments revealed that melamine is quite steady and can be accurately detected in an
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alkaline aqueous solution. The method is simple, reliable and can be used in the analysis of practical
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samples.
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Acknowledgments
The authors gratefully acknowledge financial support from the Key Program of National
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Science Foundation of China (No.20936002), the Key Project of the National Eleventh-Five Year
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Research Program of China (2008BAI63B04), the State Key Laboratory for Chemo/Biosensing and
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Chemometrics Foundation (No.2008006), Scientific Research Fund of Hunan Provincial Education
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Department (No. 10K024) and the innovation Foundation of Graduate Student of Hunan University
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of Science and Technology (No. S090121, No. S120028).
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References
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[1] B.M. Kuehn, J. A. M. A. 301(2009) 473–475. [2] X. Zhu, S. Wang, Q. Liu, Q. Xu, S. Xu, H. Chen, J. Agric. Food Chem. 57(2009) 11075–11080. [3] X. Xu, Y. Ren, Y. Zhu, Z. Cai, J. Han, B. Huang, Y. Zhu, Analytica Chimica Acta 650(2009) 39–43. [4] X. Xia, S. Ding, X. Li, X. Gong, S. Zhang, H. Jiang, J. Li, J. Shen, Anal. Chim. Acta 651(2009) 196–200.
305
[5] A. Filazi, U.T. Sireli, H. Ekici, H.Y. Can, A. Karagoz, J. Dairy Sci. 95(2012) 602–608.
306
[6] V. Gopalakrishnan, R.S.J. John, Anal. Chim. Acta 665(2010) 227−230.
307
[7] N. Yan, L. Zhou, Z. Zhu, X. Chen, J. Agric. Food Chem. 57(2009) 807–811.
308
[8] Q. Cao, H. Zhao, Y. He, X. Li, L. Zeng, N. Ding, J. Wang, J. Yang, G. Wang, Biosens. Bioelectron. 25(2010)
14
Page 14 of 28
2680–2685.
310
[9] H. Ping, M. Zhang, H. Li, S. Li, Q. Chen, C. Sun, T. Zhang, Food Control. 23(2012) 191–197.
311
[10] D. Patra, T. H. Dhaddar, Talanta, 77(2009) 1549 –1554.
312
[11] W. Zhao, Y. Fung, Waisum O, M. P. L. Cheung, Anal Sci. Augu. 26(2010) 879 – 884.
313
[12] H. Zhang, Z. Zhou, B. Yang, J. Phys. Chem. B 107(2003) 8–13.
314
[13] M. Hou, J. Na, Anal. Bioanal. Chem. 397 (2010) 3589–3593.
315
[14] L. Guo, J. Zhong, J. Wu, F.F. Fu, G. Chen, X. Zheng, S. Lin, Talanta 82(2010) 1654–1658.
316
[15] Z. He, H. Zhu, P. Zhou, J. Fluoresc 22(2012) 193–199.
317
[16] F. Gao, Q. Ye, P. Cui, L. Zhang, J. Agric. Food Chem. 60(2012) 4550–4558.
318
[17] L. Li, B. Li, D. Cheng, L. Mao, Food Chem. 122(2010) 895–900.
319
[18] E.A.E Garber, J. Food Prot. 71(2008) 590–594.
320
[19] N. Ding, N. Yan, C. Ren, X. Chen, Anal. Chem. 82(2010) 5897–5899.
321
[20] C. Han, H. Li, Analyst 35(2010) 583–588.
322
[21] G.L. Wang, H.J. Jiao, X.Y. Zhu, Y.M. Dong, Z.J. Li, Talanta 93(2012) 398–403.
324 325 326
cr us
an
M
d
te
Ac ce p
323
ip t
309
[22] E.R. Goldman, I.L. Medintz, J.L. Whiteley, A. Hayhurst, A.R. Deschamps, M.E. Lassman, H. Mattoussi, J. A. C. S. 127(2005) 6744–6751.
[23] P.K. Smith, R.I. Krohn, G.T. Hermanson, A.K. Mallia, F.H. Gartner, M.D. Provenzano, E.K. Fujimoto, N.M. Goeke, B.J. Olson, D.C. Klenk, Anal. Biochem. 150(1985) 76–85.
327
[24] G. A. Crosby, J.N. Demas, J.Phys. Chem. 75(1971) 991–1024.
328
[25] Y. L. Wang, J. P. LU, Z. F. Tong, Bull. Mater. Sci. 33(2010) 543–546.
329
[26] D.P. Wrathall, R. M. Izatt, J.J. Christensen, J. Am. Chem. Soc. 86(1964) 4779–4783.
330
[27] M. Gao, S. Kirstein, H. Möwald, Andrey L. Rogach, A. Kornowski, A. Eychmuller, H. Weller, J. Phys. Chem. B
15
Page 15 of 28
102(1988) 8360–8363.
332
[28] A. Bozzi, M. Dhananjeyan, I. Guasaquillo, S. Parra, C. Pulgarin, C. Weins, J. Kiwi, J. Photochem. Photobio. A:
333
Chem. 162 (2004) 179–185.
334
[29] W.W. Yu, L. Qu, W. Guo, X. Peng, Chem Mater. 15(2003) 2854–2860.
335
[30] M. Li, Scott K. Cushing, Q. Wang, X. Shi, Lawrence A. Hornak, Z. Hong, N. Wu, J. Phys. Chem. Lett. 2(2011)
336
2125–2129.
337
[31] Y. Tang, W. Cheng, Langmuir, 29(2013)3125−3132.
338
[32] X. Liu, H. Ju, Anal. Chem. 80 (2008)5377–5382.
339
[33] C. Duan, H. Cui, Z. Zhang, B. Liu, J. Guo, W. Wang, J. Phys. Chem. C 111(2007)4561– 4566.
340
[34] L. Guo, J. Zhong, J. Wu, F. Fu, G. Chen, Y. Chen, X. Zheng, S. Lin, Analyst 136(2011) 1659–1663.
341
[35] S. Liu, J. Hu, H. Zhang, X. Su. Talanta, 101(2012) 368 – 373.
342
[36] J. Wu, F. Xu, K. Zhu, Z. Wang, Y. Wang, K. Zhao, X. Li, H. Jiang, S. Ding, Anal. Lett. 46(2013) 275 – 285.
343
[37] H. Zeng, R. Yang, Q. Wang, J. Li, L. Qu, Food Chem. 127(2011) 842–846.
344
[38] H. Lei, Y. Shen, L. Song, J. Yang, O.P. Chevallier, S.A. Haughey, H. Wang, Y. Sun; C. Elliott, Anal. Chim.
346 347 348 349
cr us an
M
d
te
Ac ce p
345
ip t
331
Acta. 665(2010) 84–90.
[39] J. Zhang, M. Wu, D. Chen, Z. Song, J. Food Comp. Anal. 24(2011) 1038– 1042. [40] A. Henglein, Chem. Rev. 89(1989):1861–1873. [41] Y. Chen, Z. Rosenzweig, Anal Chem. 74(2002):5132– 5138. [42] S. Banerjee, S. Kar, S. Santra, Chem Commun. (2008), 3037–3039.
350 351 352
16
Page 16 of 28
353
Figure captions
357
Fig. 1 Influence of pH on Quantum yield of CdTe QDs.
358
cr
355 356
ip t
354
Fig. 2 UV-vis absorption (a) and Fluorescence spectra (b) of TGA-CdTe QDs aqueous solution
360
(5.0×10-4 mol L-1), in different molar ratio of TGA: Cd2+: Te2-.
us
359
an
361
Fig.3 The fluorescence spectra of the TGA-CdTe QDs (5.0×10-5 mol L-1) in the presence of
363
melamine at various concentrations(from top to bottom): (a) 0, (b) 1.0×10-11, (c) 1×10-10, (d) 1×10-9,
364
(e) 1×10 -8, (f) 1×10-7, (g) 1×10-6, (h) 1×10-5 mol L-1. Insert: The calibration plots of melamine.
d te
366
Scheme 1 Mechanism of fluorescence quenching of the QDs in the presence of melamine.
367
Ac ce p
365
M
362
368 369 370 371 372 373 374
17
Page 17 of 28
375
Table 1
377
Influence of pH on the performances on the method (TGA-CdTe QDs: 5.0×10-5 mol L-1).
ip t
376
Linear range (μmol L-1)
Detection limits (μmol L-1)
5.0
0.004 − 2.5
0.002
6.0
0.002 − 5.5
7.0
0.0008 − 15
8.0
0.0065 − 8
9.1
0.0009 − 10
0.0001
10.0
0.0002 − 10
0.00004
0.00001 − 10
0.000005
0.00045 − 6
0.0001
380 381 382
te
d
M
an
0.0009
0.0004 0.0008
Ac ce p
12.0
us
pH
11.0
379
cr
378
383 384 385 386
18
Page 18 of 28
387
Table 2
389
Influence of diameter of the CdTe QDs on the performances on the method (TGA-CdTe QDs:
390
5.0×10-5 mol L-1).
ip t
388
Linear range (μmol L-1)
Detection limits (μmol L-1)
1.7
1.0×10-9 − 1.0×10-7
0.00080
2.4
1.0×10-9 − 1.0×10-6
3.2
1.0×10-9 − 1.0×10-5
3.9
394 395
an 0.00050
2.17 2.37
2.48
1.0×10-11 −1.0×10-5
0.000005
2.64
4.6
1.0×10-9 −1.0×10-5
0.00072
2.79
5.3
1.0×10-8 − 1.0×10-5
0.0030
2.88
1.0×10-7 − 1.0×10-5
0.015
3.16
te
d
M
0.00045
Ac ce p
393
Diameter of the QDs(nm)*
us
Molar ratio(TGA:Cd 2+)
6.0
392
cr
391
* established according to the calculation reported in literature [29].
396 397 398 399
19
Page 19 of 28
400
Table 3
401
Comparison of the performances of different methods.
0.396 − 0.793
Gas chromatography/mass spectrometry [3]
0.396 − 1.59
HPLC method [6]
Colorimetric method/AuNPs [11]
0.793
0.396 − 79.3
0.0792
0.001 − 10
4.0 − 170.0
M
Label-free silver nanoparticles [12]
0.6 – 1.6
0.0075 − 0.35
te
d
Label-free AuNPs [17]
CdS quantum dots as fluorescence probe [21]
0.002 − 50 0.0048 − 1.6
Ac ce p
Colorimetric method/AuNPs [22] FRET/fluoresce in-AuNPs[34]
0.1 − 4.0
CuInS2 QDs as fluorescence probe [35]
0.0793
7.93 − 634
an
Capillary zone electrophoresis [7]
FRET/CdTe@SiO2-AuNPs [19]
0.0396
us
Gas chromatography−mass spectrometric [2]
Detection limits (μmol L-1)
cr
Linear range (μmol L-1)
Methods
ip t
402
0.0008 2.32 0.2 0.00089 0.001 0.00064 0.001
0.01 − 10
0.005
icELISA with QDs-Ab2 [36]
0.069 − 0.90
0.031
FI-CL [37]
0.0015 − 0.64
0.0008
ELISA [38]
0.0006 − 0.0058
0.00002
Luminol-hydrogen peroxide chemiluminescence[39]
0.00002 – 0.002
0.000007
The present method
0.00001 − 10
0.000005
403 404 20
Page 20 of 28
406
Table 4
407
Determination of melamine in milk powder and pet feeds samples.
Added (μmol L-1)
Found (μmol L-1)
Milk powder 1
6.82
7.93
14.99
103.0
Milk powder 2
7.30
7.93
15.38
102.0
Milk powder 3
9.83
7.93
18.16
105.0
Milk powder 4
9.20
11.89
24.74
97.3
pet feeds 1
15.46
11.89
27.20
98.7
pet feeds 2
22.13
15.86
38.53
103.5
pet feeds 3
16.97
15.86
32.59
98.5
cr
an
M d
Recovery (%)
te
410
Background (μmol L-1)
Ac ce p
409
Sample
us
408
ip t
405
21
Page 21 of 28
OOCH2CS d C CS H2 C - OO
-
OO H 2C
OO
CdTe Cd 2 SCH2COO-
N H2 N
+
2
H2N
N N
N
Cd 2+ H
N
H2N
N
H Cd 2+ N
N
H2N
O-
2
OO CH
H NH2
Melamine
SC H
2 CO
H
N N
OCO
SCH2COO-
Cd2+
H SC
-
-
2+
Cd S C
N N H
N
NH 2
m-
N
NH2
H Cd 2 H + H N N H H N N N H NH2
Cd2+
an
n+
m-
n+
M
+
2+ i SCH2COO
Melamine coated CdTe QDs
Agglomerates
410
te
d
1 scheme
Ac ce p
412
N
N N
CdTe
Exc
411
H
+
n+
TGA coated CdTe QDs
m-
N 2 Cd
H H N 2
ip t
2+
CS
-
Cd 2+
N
cr
2 CS
n+
NH2 H N
NH2
us
2
CS
CH
m-
2C
CH
OO
SC H
O O -
-
SCH 2COO -
Exc
22
Page 22 of 28
412 Graphical Abstract
Melamine
SC H
2 CO
H NH2
H2N
N N
N
Cd 2+ H
H2N
O-
N
2
N
H2N
N
H Cd 2+ N
N
NH 2
m-
H
+
N
N N
N
NH2
H Cd 2 H + H N N H H N N N H NH2
Melamine coated CdTe QDs
an
Exc
m-
n+
n+
413 414
M
+
2+ i SCH2COO
CdTe
n+
TGA coated CdTe QDs
m-
N 2 Cd
Cd2+
N H
N
H H N 2
ip t
Cd2+
H
N N
N
cr
-
OO
2+
N
OCO
OO CH
+
SCH2COO-
-
-
O CO
2 H2 Cd S C H2 N CdTe Cd SCH2COO-
H SC
OOCH2CS d C CS H2 C - OO
S
2+
2C
-
Cd 2+
H N
NH2
us
2 CS
n+
NH2
2C
2
CS
CH
m-
H
CH
OO
SCH 2COO -
O O -
-
SC
Exc
415
Melamine takes place of the TGA on the surface of TGA-CdTe QDs with negative
416
charge to form melamine coated QDs changing the surface charge of the QDs, resulting
417
the fluorescence quenched as the QDs aggregation occurred by electrostatic attraction
419 420 421
d
te
Ac ce p
418
Agglomerates
of the two opposite charged nanocrystals.
23
Page 23 of 28
Highlights:
422
● An ultrasensitive and selective method for the determination of melamine was
423
developed at pH 11.0.
424
● The selectivity of the method was improved.
425
● The sensitivity of the method enhanced obviously as the CdTe QDs have higher QYs
426
at pH 11.
427
● The sensitivity and linear range for an analyze is size dependent using QDs PL
428
pbobes.
429
● Melamine takes the place of TGA resulting fluorescence quenched of QDs.
an
us
cr
ip t
421
M
430
Ac ce p
te
d
431
24
Page 24 of 28
Ac
ce
pt
ed
M
an
us
cr
i
Fig.1
Page 25 of 28
Ac
ce
pt
ed
M
an
us
cr
i
Fig.2a
Page 26 of 28
Ac
ce
pt
ed
M
an
us
cr
i
Fig.2b
Page 27 of 28
Ac
ce
pt
ed
M
an
us
cr
i
Fig.3
Page 28 of 28
S0003-2670(13)01244-0 http://dx.doi.org/doi:10.1016/j.aca.2013.09.042 ACA 232854
To appear in:
Analytica Chimica Acta
Received date: Revised date: Accepted date:
15-5-2013 14-9-2013 19-9-2013
Please cite this article as: X. Li, J. Li, H. Kuang, L. Feng, S. Yi, X. Xia, H. Huang, Y. Chen, C. Tang, Y. Zeng, An ultrasensitive method for the determination of melamine using cadmium telluride quantum dots as fluorescence probes, Analytica Chimica Acta (2013), http://dx.doi.org/10.1016/j.aca.2013.09.042 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
An ultrasensitive method for the determination of melamine using
2
cadmium telluride quantum dots as fluorescence probes
3
Xiafei Lia,b, Jin Lia,b, Huiyan Kuanga,b, Lei Fenga,b, Shoujun Yia,b, Xiaodong Xiaa,b,
4
Haowen Huanga,b, Yong Chena, Chunran Tanga, Yunlong Zenga,b,c,*
6
School of Chemistry and Chemical Engineering , Hunan University of Science and Technology, Xiangtan 411201,
cr
a
PR China
7
b
8
University of Science and Technology, Xiangtan 411201, PR China
9
c
an
Key Laboratory of Theoretical Chemistry and Molecular Simulation of Ministry of Education of China, Hunan
State Key Laboratory for Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering,
Hunan University, Changsha 410082, PR China
M
10
us
5
ip t
1
11
Abstract: An ultrasensitive and simple method for the determination of melamine was developed
13
based on the fluorescence quenching of thioglycolic acid (TGA) capped CdTe quantum dots (QDs)
14
at pH 11.0. In strong alkaline aqueous solution, the selectivity of the method has been greatly
16 17 18
te
Ac ce p
15
d
12
improved due to most heavy metal ions show no interference as they are in the precipitation form or in their anion form. Furthermore, CdTe quantum dots have higher quantum yields at higher pH. The method has a wider concentration range and lower detection limit. The influence factors on the determination of melamine were investigated and the optimum conditions were determined. Under
19
optimum conditions, the fluorescence intensity change of TGA coated CdTe quantum dots was
20
linearly proportional to melamine over a concentration range from 1.010-11 to 1.010-5 mol L-1
21
with a correlation coefficient of 0.9943 and a detection limit of 5×10-12 mol L-1. The mechanism of
22
fluorescence quenching of the QDs has been proposed based on the infrared spectroscopy
1
Page 1 of 28
information and electrophoresis experiments in presence of melamine under alkaline condition. The
24
proposed method was employed to detect trace melamine in milk powder and pet feeds with
25
satisfactory results.
26
Keywords: Melamine; Ultrasensitive determination; High selectivity, Cadmium telluride quantum
27
dots; Fluorescence probe.
28
1. Introduction
us
cr
ip t
23
Melamine (1,3,5-triazine-2,4,6-triamine,C3H6N6) is a chemical compound used commonly for
30
the production of melamine resins. As its high nitrogen level (66.6% by mass), melamine is illegally
31
added to food, such as milk, cookies and animal feeds, which produces an incorrectly high reading
32
in the measurement of protein content based on total nitrogen, resulting some serious melamine
33
residue related problems [1]. Since the melamine contaminated animal feeds incident in USA in
34
2007 and the accident of San Lu poisonous milk accident in China in 2008, the great importance of
35
detecting melamine has been well recognized and the methods for melamine determination in
36
various samples have been developed [2]. Most techniques for melamine assay such as GC/MS [3,4],
38 39 40
M
d
te
Ac ce p
37
an
29
HPLC [5,6] and CZE [7] require expensive apparatus and considerable time for identification and quantification. Hence, it is necessary to develop some simple, highly sensitive, selective method for the analysis of melamine in real samples. Recently, nanoparticles have attracted increasing attention in bioanalysis as their unique optical
41 42
*
43
Technology, Xiangtan 411201, PR China. Tel.: (+86)13787423309
44
E-mail address: [email protected]
Corresponding author at: School of Chemistry and Chemical Engineering, Hunan University of Science and
2
Page 2 of 28
and electronic properties. Several simple, fast, sensitive methods for the determination of melamine
46
have been established [8−10] based on the change of visual colorimetric by the aggregation of some
47
metal nanoparticles such as silver nanoparticles, gold nanoparticles. As a kind of typical
48
nanomaterials, luminescence semiconductor quantum dots (QDs), such as CdS, CdSe, and CdTe,
49
have attract great interest in recent years due to their quantum confinement effect and remarkable
50
optical, electrical properties [11,12]. They have been widely used in biology, medicine, analytical
51
chemistry, especially in sensors [13−15]. Recently, Gao [16] has developed a high sensitive method
52
to detect melamine based on the energy transfer of CdTe quantum dots (CdTe@SiO2). This method
53
has a wide linear range for melamine determination from 7.5 × 10−9 to 3.5 × 10−7 mol L-1 with a
54
detection limit of 8.9 × 10−10 mol L-1. However, the methods for the determination of melamine was
55
often at pH 5.0 − 8.0 [17−19]. There are no research reports about the detection of melamine in
56
strong alkaline solution based on noble metal nanoparticles or quantum dots so far as we know. As
57
known, heavy metal ions can strongly change the fluorescence intensity and ultraviolet-visible signal
58
of nanoparticles (noble metal nanoparticles and quantum dots) [20−22], which will interfere with the
60 61 62
cr
us
an
M
d
te
Ac ce p
59
ip t
45
target object analysis. But in strong alkaline solution, the selectivity and detection limit can be greatly improved as heavy metal ions are in their precipitate form or in their anion form at high pH. In this work, we have proposed an ultrasensitive method for the determination of melamine
using water-soluble TGA-CdTe QDs as the fluorescence probes at pH 11.0. Under alkaline
63
conditions, CdTe QDs have high quantum yields( QYs), and heavy metal ions free from interference
64
as they are in the form of precipitation or in their anion form. So the method showed high selectivity
65
and sensitivity. This method was applied to detecting melamine in milk power and pet feeds with
66
satisfactory results.
3
Page 3 of 28
67
2. Experimental
68
2.1 Apparatus Fluorescence spectra were acquired on a FL 4500 Fluorescence Spectrometer (Hitachi, Japan).
70
UV-vis absorption spectra were recorded on Lambda 35 spectrometer (Perkin-Elmer, USA). IR
71
spectra were measured on a Spectrum One FT/IR spectrometer (Perkin–Elmer, USA). All optical
72
measurements were carried out at room temperature.
73
2.2 Reagent
us
cr
ip t
69
Tellurium dioxide (TeO2), sodium borohydride (NaBH4), cadmium nitrate (Cd(NO3)2),
75
thioglycolic acid (TGA) and melamine were purchased from Shanghai Chemical Reagent Company
76
(Shanghai, China). All other chemical reagents were of analytical-reagent grade and were used as
77
received without further purification. Stock standard aqueous solution of melamine was prepared at
78
a concentration of 0.01mol L-1, and other concentration of melamine was prepared by diluting the
79
stock standard solution with double distilled water. The milk powder was purchased from the
80
supermarket in Xiangtan. Double distilled water was used throughout all experiments.
82 83 84
M
d
te
Ac ce p
81
an
74
2.3 Preparation of water-soluble CdTe QDs The TGA-CdTe QDs, which with a molar ratio of TGA:Cd2+:Te2- was 3.9:1:0.5, was prepared
by using the reaction between Cd2+ and NaHTe solution at pH 12.0 with TGA as the stabilizing
reagent. Briefly, 0.08 g TeO2 powder was reduced by 0.05g NaBH4 in 5 ml water under a cold-water
85
bath (about 15) for about 4 h, and then the aqueous solution was injected into 95 ml oxygen-free
86
aqueous solution containing a certain amount of TGA and 1.0×10−3 mol L-1 of Cd(NO3)2 at pH 12.0.
87
The resulting mixture was refluxed in boiling water bath at dark for 25 min under vigorously
88
magnetic stirring. The TGA-CdTe QDs were obtained and stored in 4 at dark for use. The final
4
Page 4 of 28
concentration of CdTe QDs was approximately 5.0×10-3 mol L-1 (according to the Te2-
90
concentration).
91
2.4 Isolation and extraction of melamine
ip t
89
According to the previous method reported by Li [17], the sample was prepared with some
93
modifications and described as follows: First, 2.5 g sample was dissolved with 25 ml of 0.6 mol L-1
94
trichloroacetic acid and mixed with a vortex for 5 min to precipitate protein in the sample matrix.
95
And the mixture was centrifuged at 8000 r/min for 10 min. Then, the protein content in the
96
supernatants was determined by Bicinchoninic Acid method [23]. After the protein in the filtrate was
97
removed, the supernatants were transferred into a 25 ml beaker, adjusted to pH 7.0 with a small
98
amount of 2 mol L-1 NaOH and filtered with a 0.22 μm filter. The obtained filtrate was the sample
99
solution and was used in the following detection.
us
an
M
d
2.5 Detection of melamine
te
100
cr
92
Melamine determination was carried out by successively adding 0.5 ml varying concentrations
102
of melamine or sample solution into 4.5 ml 1×10-4 mol L-1 CdTe QDs solution (pH 11.0). The
103 104 105 106
Ac ce p
101
fluorescence emission of the solution was recorded at the peak wavelength (about 535 nm) with the excitation wavelength of 465 nm. All measurements were carried out under room temperature. 3. Results and discussion 3.1 Influence of pH
107
The influence of pH on the luminescence quantum yield(QY) of CdTe QDs was investigated in
108
different pH. The QY was measured and the results are recorded in Fig.1, by referring to Rhodamine
109
6G (QY 95%) with the method described in Ref. [24]. As known, the photoluminescence of CdTe
110
NCs was attributed to defect-related emission. The QYs of the NCs was strongly dependent on pH.
5
Page 5 of 28
As shown in Fig. 1, at lower pH, QYs of the QDs increased linearly with rise of pH and the QYs
112
reaches a maximum (51.9 %) at the pH 11.0 and then decreased. This is because of the extremely
113
small diameter, there are many trap sites on the surface of CdTe NCs, the more the trap sites, the
114
lower the fluorescence efficiency. As known, TGA is a weak acid and the dissociation constants of
115
TGA are pKa1 = 3.6 and pKa2 = 10.6 [25, 26]. With rise of pH, TGA2- (−SCH2COO−) increased, and
116
more TGA2- coordinated with Cd2+ on the surface of the CNs. So the relative coverage rate of the
117
particle surface with thiols was increased. Therefore, the trap sites on the CdTe surface will be
118
decreased, and the fluorescence efficiency was improved. However, at higher pH(higher than 11.0),
119
the relative fluorescence intensity of the QDs also tended to decrease. This phenomenon might be
120
resulted from the formation of cadmium hydroxide product on the surface of QDs at higher pH [27].
M
an
us
cr
ip t
111
121
124 125 126 127 128
d te
123
Insert Fig. 1
As seen from Fig.1, when the pH is less than 5.0, the QY of the QDs is very low, and the
Ac ce p
122
fluorescence intensity of the system is very weak. Therefore, the influence of pH on the performance of the method was investigated and the results were recorded in Table 1 at pH 5.0 to 12.0. From Table 1, one can notice that the method has a wider linear concentration range and lower detection limit at pH 11.0. The result shows the advantage of high sensitivity for melamine assay under strong
129
alkaline condition. This is because high QY of the NCs is crucial for realizing a good sensitivity of
130
the fluorescent sensors. The sensitivity for an analyte is determined at the signal-to-noise, which is
131
largely dependent on the QYs of the fluorescent probes. Fluorescent probes with higher QYs can
132
achieve the same luminescence intensity at lower concentrations relative to NCs with lower QYs,
6
Page 6 of 28
133
thus leading to a higher sensitivity.
134 Insert Table 1
ip t
135 136
Quantum dots are always sensitive to metal ions. Hence, the influence of pH on the
138
fluorescence intensity in the presence of some metal ions (1.0×10-6 mol L-1) was investigated (see
139
Fig. S1). The fluorescence intensity of the QDs was changed obviously at pH 6.5 and almost was not
140
changed at pH 11.0 in the presence of heavy metal ions(such as Fe3+, Cu2+, Hg2+, Pb2+ and Zn2+). In
141
other words, heavy metal ions show interference at pH 6.5 and show no interference at pH 11.0.
142
Presumably the heavy metal ions are in the form of precipitation or in their anion forms under strong
143
at alkaline condition and free from interference.
M
an
us
cr
137
Melamine, a Lewis base that can accept a proton to form conjugate acid, may hydrolyze in an
145
aqueous solution. Therefore, the media pH affects the form and stability of melamine. Bozzi [28] has
146
studied the photocatalysis of melamine and pointed out that melamine remains stable in aqueous
148 149 150
te
Ac ce p
147
d
144
solution in detail. To examine the stability of melamine solution at pH 11.0, the experiments of fluorescence quenching of CdTe quantum solution by 1.0×10-7 mol L-1 melamine at pH 7.0 and pH 11.0 were carried out, respectively. There were no obvious differences in the quenched fluorescence intensity when the melamine solution was stored for 0 − 5.0 h at pH 11.0 or at pH 7.0, which
151
indicated that melamine was stable and can be determined accurately in strong alkaline solution. So
152
the following experiments were performed at pH 11.0.
153
3.2 Influence of TGA molar ratio
154
The influence of TGA molar ratio on the properties of fluorescence and UV–vis absorption of
7
Page 7 of 28
the QDs was investigated. Fig.2 depicts the fluorescence and UV–vis absorption spectra of
156
TGA-CdTe QDs at different molar ratio of TGA and Cd2+. From Fig. 2 (a), one can see that the
157
UV–vis absorption peaks are red shift with increasing TGA, which indicates that the size of the QDs
158
increased with increasing the ratio of TGA. The diameters of as-prepared CdTe QDs were about 2.2,
159
2.4, 2.5, 2.6, 2.8, 2.9 and 3.2 nm, respectively, corresponding to TGA ratio of 1.7, 2.4, 3.2, 3.9, 4.6,
160
5.3 and 6.0, respectively, according to the calculation reported in literature [29]. It is obviously that
161
the concentration of free cadmium ions decreased with the increasing TGA because of the formation
162
of TGA-Cd2+ complex, leading to form larger diameter CdTe nanocrystals. From Fig. 2 (b), one can
163
also see that the TGA-CdTe QDs generally have a broad emission ranging from 460 nm to 680 nm
164
and the fluorescence spectra do not exhibit a tail on the right side, suggesting that the prepared QDs
165
possess good fluorescence properties. All of the fluorescence spectra band are relatively narrow
166
(with full width at half maximum of about 30~40 nm) and symmetric indicating that the prepared
167
CdTe QDs are nearly mono-disperse and homogeneous.
169 170 171 172
cr
us
an
M
d
te
Ac ce p
168
ip t
155
Insert Fig. 2
The size dependent sensitivity has been reported in photoluminescence, chemiluminescence
and electrochemiluminescence[30−33]. In this work, different sized CdTe QDs were used to study
173
the influence of the size effect on the sensitivity and linear range for melamine assay, and the results
174
were recorded in Table 2.
175 176
Insert Table 2
8
Page 8 of 28
177 From Table 2, one can notice that the sensitivity of the method increased the maximum and
179
then decreased with increasing size of the QDs. Furthermore, the linear concentration range reached
180
the widest as the diameter of CdTe QDs is about 2.6 nm in the molar ratio of TGA: Cd2+: Te2- at 3.9:
181
1: 0.5. This may be the most suitable size for melamine assay.
182
3.3 Selection of the QDs concentration
us
cr
ip t
178
The fluorescence of the QDs can be quenched strongly by melamine. The fluorescence
184
intensity of the solution changes very slightly, when a small amount of melamine was added to the
185
solution with high concentration of CdTe QDs. The results indicated that the amount of QDs in the
186
solution will greatly influence the detection sensitivity and linear range for melamine assay. So
187
series of concentration of CdTe QDs was employed to investigate the effect of QDs concentration on
188
the measurement(as shown in fig. S2). A wider concentration range and lower detection limit
189
were obtained when 1.0×10-4 mol L-1 of the CdTe QDs solution was used. Consequently, this QDs
190
concentration was chosen for the following experiments.
192 193 194
M
d
te
Ac ce p
191
an
183
3.4 Effect of reaction time
The influence of reaction time on the fluorescence intensity of the QDs in the presence of
melamine (1.0×10-8, 1.0×10-9, 1.0×10-10 mol L-1 respectively) was investigated. The results showed
that the fluorescence intensity of the QDs decreased rapidly when melamine was added to the QDs
195
solution. After 5 min, the fluorescence intensity reached the lowest and kept unchanged more than
196
50 min. So the reaction time was fixed at 10 min.
197
3.5 Influence of coexisting substance
198
To evaluate the detection selectivity, the influence of common coexisting ions in milk powder
9
Page 9 of 28
on the fluorescence quenching of the CdTe QDs in the presence of 1.0×10-7 mol L-1 melamine were
200
investigated under optimum conditions. The tolerable concentration ratios relative to concentration
201
levels usually found in milk for interference at the 5% level were over 1000 fold for Na+, K+, Cl-,
202
NO3- and SO42-, 600 fold for Ca2+, Mg2+, PO43-, 200 fold for Fe3+, Cu2+, Zn2+, and 120 fold for Vc
203
and Lactos. The results demonstrated that common metal ions had no interferences in detection of
204
melamine. So, this method has the characteristic of high selectivity for the determination of
205
melamine, as the heavy metal ions are in the form of precipitation or in their anion forms under
206
strong at alkaline condition.
207
3.6 Calibration curve and detection limit
an
us
cr
ip t
199
Under optimum conditions, a high linearity was observed (insert of Fig. 3) in the melamine
209
concentration range from 1.0×10-11 to 1.0×10-5 mol L-1. The detection limit (LOD) is defined by the
210
equation LOD = 3 S0/K, where S0 is the standard deviation of blank measurements (nine
211
measurements, n = 9), K is the slope of calibration graph. The LOD is 5×10-12 mol L-1 and the
212
relative standard deviation is 3.8 % for the determination of 1.0×10-8 mol L-1 melamine (n = 11).
214 215 216
d
te
Ac ce p
213
M
208
Insert Fig. 3
Comparison between our proposed method and some methods reported recently for the
217
determination of melamine is presented in Table 3. The present method showed lower detection limit
218
and wider linear range.
219 220
Insert Table 3
10
Page 10 of 28
221 222
3.7 Applications The proposed method was employed to detect melamine in milk powder and pet feeds samples
224
(Table 4) with satisfactory results. And the recovery studies were carried out on samples to which
225
known amounts of melamine were added. The results are illustrated in Table 4, with satisfying
226
recoveries of 97.3 − 105 %, suggesting that the proposed method was reliable and practical.
227 Insert Table 4
an
228 229 3.8 Mechanism
M
230
us
cr
ip t
223
The interaction between TGA and CdTe NCs was investigated by the FTIR spectrometry. As
232
shown in Fig. S3 b, the absorption band due to S–H vibration observed at 2560 cm–1 disappeared
233
and the characteristic absorption band of C=O vibration was shifted from 1700 cm–1 to 1556 cm–1,
234
which strongly suggest that the thiol groups of TGA coordinated with Cd2+ ions on the NCs surface,
236 237 238
te
Ac ce p
235
d
231
and the hydrophilic hydroxyl groups faced outward, making NCs water-soluble. Generally, more than 15% of the atoms in a quantum dot are located on its surface [40], and the
luminescence of QDs is sensitive to their surface conditions. The molecular interactions with the surface of luminescence QDs would change their photophysical properties [41, 42]. As known, TGA
239
exists in the form of anion and melamine is in the form of neutral molecular under alkaline condition.
240
Therefore, TGA-CdTe QDs carry negative charge. As there were excess of cadmium ions being used
241
in the preparation of TGA-CdTe QDs (as mentioned above, the molar ratio of TGA: Cd2+: Te2-: is 3.9:
242
1: 0.5), the cadmium telluride crystals will carry positive charge as their surfaces attract these
11
Page 11 of 28
243
excessive ions in the solution. However, TGA-CdTe QDs carry negative charge owing to
244
–
245
melamine solution was added to this system, TGA on the nanoparticle surfaces was to be displaced
246
by melamine to form melamine coated CdTe QDs in alkaline solution. The melamine coated CdTe
247
QDs carry positive charge, as melamine is weak base (pKb= 9.0) and exists almost in neutral
248
molecule at pH 11.0. Therefore, there are two opposite charge CdTe QDs in the presence of
249
melamine. As a result, the fluorescence of QDs was quenched when the QDs aggregation occurred
250
by electrostatic attraction of these two opposite charge nanoparticles. The mechanism can be
251
described as following:
an
us
cr
ip t
SCH2COO–(TGA2-) coordinated with Cd2+ ions on the NCs surface. When the high concentration
M
252
te
254
Insert Scheme 1
d
253
The presumption was verified by IR information (Fig. S3) and electrophoresis (Fig.S4).
256
Fig.S3 shows the infrared spectra of the TGA (Fig. S3a), TGA−capped CdTe QDs (Fig. S3b)
257 258 259 260
Ac ce p
255
and the QDs in the presence of melamine (Fig. S3c). The IR absorption broad bands (Fig. 4a) around, 3026.26– 3435.88 cm−1 (sυ O-H), 1386.58 cm−1 (mυ COO−), 1716.01 cm−1 (υ C=O), 1291.82 cm−1 (δ OH) indicate −COOH group. The peak at 2923.70 cm−1 (wυ C-H) represents -CH2 group. The
peak at 2566.14 cm−1 (wυ S-H) represents -SH group. There are coexisting IR absorption bands of
261
-COOH, -CH2 and -SH on free TGA (see form Fig. S3a), whereas only bands -COO− (1386.21 cm−1,
262
mυ COO−) and free -CH2 (2923.70 cm−1, wυ C-H) were observed on the surface of the TGA−capped
263
CdTe QDs (seen from Fig. S3b). The disappearance of S-H group vibration 2566.14 cm−1 (wυ S-H)
264
on the surface of functionalized CdTe QDs resulted from the covalent bonds between thiols and the
12
Page 12 of 28
Cd atom of the surface of CdTe QDs. Compared with TGA−capped CdTe QDs (Fig. S3b), the
266
R-COO− (1576.60 cm−1) and –CH2 (2923.70 cm−1) disappeared in the presence of melamine(Fig.
267
S3c), which indicated the loss of TGA capping ligand of CdTe QDs. On the other hand, the –NH2
268
( 3136.08 – 3470.11 cm−1, sυ N-H ) and triazine ring (1551.26 cm−1) appear, which indicated the
269
surface of CDTe QDs coated with melamine. The reaction was as following:
cr
ip t
265
[QDs-(SCH2COO2−)i]m- + j C3N6H6 → [QDs-(C3N6H6)j]n+ + i (SCH2COO)2-
271
Figure S4 shows the electrophoresis of TGA-CdTe QDs and the melamine coated CdTe QDs,
272
respectively. One can note that TGA-CdTe QDs move to the cathode (Fig. S4B) means them
273
carrying a negative charge and that melamine-dTe QDs move to the anode electrode( Fig. S4 D)
274
indicate the QDs with positive charge applying an electric field (100V). Hence, the mechanical of
275
fluorescence quenched can first be the TGA on the surface of TGA- CdTe QDs with negative charge
276
was placed by melamine to form positive charged melamine coated CdTe QDs. And then these two
277
types of opposite charge nanoparticles aggregated by electrostatic attraction. As a result, the
278
fluorescence of CdTe QDs quenched.
282
Ac ce p
te
d
M
an
us
270
283
as heavy metal ions in the form of precipitation or in their anion form. Hence, the method can be
284
free from interference with metal ions in strong alkaline solution. Second, the lower concentration of
285
the QDs can be used for the determination of melamine as the CdTe QDs have higher quantum yield.
286
The method shows a wider linear concentration range and lower detection limit. Furthermore, the
279 280 281
4. Conclusion
In summary, we have developed an ultrasensitive and simple method for the determination of
melamine using the QDs as fluorescence probes in strong alkaline media. The analysis of melamine at alkaline condition has advantages: First, the selectivity of the method has been greatly improved
13
Page 13 of 28
size dependent sensitivity appeared using QDs probe for melamine assay. The linear concentration
288
range reached the widest with most sensitivity as the diameter of CdTe QDs is about 2.6 nm in the
289
molar ratio of TGA: Cd2+: Te2- at 3.9: 1: 0.5. This may be the most suitable size for melamine assay.
290
At last, the experiments revealed that melamine is quite steady and can be accurately detected in an
291
alkaline aqueous solution. The method is simple, reliable and can be used in the analysis of practical
292
samples.
cr us
293
ip t
287
Acknowledgments
The authors gratefully acknowledge financial support from the Key Program of National
295
Science Foundation of China (No.20936002), the Key Project of the National Eleventh-Five Year
296
Research Program of China (2008BAI63B04), the State Key Laboratory for Chemo/Biosensing and
297
Chemometrics Foundation (No.2008006), Scientific Research Fund of Hunan Provincial Education
298
Department (No. 10K024) and the innovation Foundation of Graduate Student of Hunan University
299
of Science and Technology (No. S090121, No. S120028).
300
References
302 303 304
M
d
te
Ac ce p
301
an
294
[1] B.M. Kuehn, J. A. M. A. 301(2009) 473–475. [2] X. Zhu, S. Wang, Q. Liu, Q. Xu, S. Xu, H. Chen, J. Agric. Food Chem. 57(2009) 11075–11080. [3] X. Xu, Y. Ren, Y. Zhu, Z. Cai, J. Han, B. Huang, Y. Zhu, Analytica Chimica Acta 650(2009) 39–43. [4] X. Xia, S. Ding, X. Li, X. Gong, S. Zhang, H. Jiang, J. Li, J. Shen, Anal. Chim. Acta 651(2009) 196–200.
305
[5] A. Filazi, U.T. Sireli, H. Ekici, H.Y. Can, A. Karagoz, J. Dairy Sci. 95(2012) 602–608.
306
[6] V. Gopalakrishnan, R.S.J. John, Anal. Chim. Acta 665(2010) 227−230.
307
[7] N. Yan, L. Zhou, Z. Zhu, X. Chen, J. Agric. Food Chem. 57(2009) 807–811.
308
[8] Q. Cao, H. Zhao, Y. He, X. Li, L. Zeng, N. Ding, J. Wang, J. Yang, G. Wang, Biosens. Bioelectron. 25(2010)
14
Page 14 of 28
2680–2685.
310
[9] H. Ping, M. Zhang, H. Li, S. Li, Q. Chen, C. Sun, T. Zhang, Food Control. 23(2012) 191–197.
311
[10] D. Patra, T. H. Dhaddar, Talanta, 77(2009) 1549 –1554.
312
[11] W. Zhao, Y. Fung, Waisum O, M. P. L. Cheung, Anal Sci. Augu. 26(2010) 879 – 884.
313
[12] H. Zhang, Z. Zhou, B. Yang, J. Phys. Chem. B 107(2003) 8–13.
314
[13] M. Hou, J. Na, Anal. Bioanal. Chem. 397 (2010) 3589–3593.
315
[14] L. Guo, J. Zhong, J. Wu, F.F. Fu, G. Chen, X. Zheng, S. Lin, Talanta 82(2010) 1654–1658.
316
[15] Z. He, H. Zhu, P. Zhou, J. Fluoresc 22(2012) 193–199.
317
[16] F. Gao, Q. Ye, P. Cui, L. Zhang, J. Agric. Food Chem. 60(2012) 4550–4558.
318
[17] L. Li, B. Li, D. Cheng, L. Mao, Food Chem. 122(2010) 895–900.
319
[18] E.A.E Garber, J. Food Prot. 71(2008) 590–594.
320
[19] N. Ding, N. Yan, C. Ren, X. Chen, Anal. Chem. 82(2010) 5897–5899.
321
[20] C. Han, H. Li, Analyst 35(2010) 583–588.
322
[21] G.L. Wang, H.J. Jiao, X.Y. Zhu, Y.M. Dong, Z.J. Li, Talanta 93(2012) 398–403.
324 325 326
cr us
an
M
d
te
Ac ce p
323
ip t
309
[22] E.R. Goldman, I.L. Medintz, J.L. Whiteley, A. Hayhurst, A.R. Deschamps, M.E. Lassman, H. Mattoussi, J. A. C. S. 127(2005) 6744–6751.
[23] P.K. Smith, R.I. Krohn, G.T. Hermanson, A.K. Mallia, F.H. Gartner, M.D. Provenzano, E.K. Fujimoto, N.M. Goeke, B.J. Olson, D.C. Klenk, Anal. Biochem. 150(1985) 76–85.
327
[24] G. A. Crosby, J.N. Demas, J.Phys. Chem. 75(1971) 991–1024.
328
[25] Y. L. Wang, J. P. LU, Z. F. Tong, Bull. Mater. Sci. 33(2010) 543–546.
329
[26] D.P. Wrathall, R. M. Izatt, J.J. Christensen, J. Am. Chem. Soc. 86(1964) 4779–4783.
330
[27] M. Gao, S. Kirstein, H. Möwald, Andrey L. Rogach, A. Kornowski, A. Eychmuller, H. Weller, J. Phys. Chem. B
15
Page 15 of 28
102(1988) 8360–8363.
332
[28] A. Bozzi, M. Dhananjeyan, I. Guasaquillo, S. Parra, C. Pulgarin, C. Weins, J. Kiwi, J. Photochem. Photobio. A:
333
Chem. 162 (2004) 179–185.
334
[29] W.W. Yu, L. Qu, W. Guo, X. Peng, Chem Mater. 15(2003) 2854–2860.
335
[30] M. Li, Scott K. Cushing, Q. Wang, X. Shi, Lawrence A. Hornak, Z. Hong, N. Wu, J. Phys. Chem. Lett. 2(2011)
336
2125–2129.
337
[31] Y. Tang, W. Cheng, Langmuir, 29(2013)3125−3132.
338
[32] X. Liu, H. Ju, Anal. Chem. 80 (2008)5377–5382.
339
[33] C. Duan, H. Cui, Z. Zhang, B. Liu, J. Guo, W. Wang, J. Phys. Chem. C 111(2007)4561– 4566.
340
[34] L. Guo, J. Zhong, J. Wu, F. Fu, G. Chen, Y. Chen, X. Zheng, S. Lin, Analyst 136(2011) 1659–1663.
341
[35] S. Liu, J. Hu, H. Zhang, X. Su. Talanta, 101(2012) 368 – 373.
342
[36] J. Wu, F. Xu, K. Zhu, Z. Wang, Y. Wang, K. Zhao, X. Li, H. Jiang, S. Ding, Anal. Lett. 46(2013) 275 – 285.
343
[37] H. Zeng, R. Yang, Q. Wang, J. Li, L. Qu, Food Chem. 127(2011) 842–846.
344
[38] H. Lei, Y. Shen, L. Song, J. Yang, O.P. Chevallier, S.A. Haughey, H. Wang, Y. Sun; C. Elliott, Anal. Chim.
346 347 348 349
cr us an
M
d
te
Ac ce p
345
ip t
331
Acta. 665(2010) 84–90.
[39] J. Zhang, M. Wu, D. Chen, Z. Song, J. Food Comp. Anal. 24(2011) 1038– 1042. [40] A. Henglein, Chem. Rev. 89(1989):1861–1873. [41] Y. Chen, Z. Rosenzweig, Anal Chem. 74(2002):5132– 5138. [42] S. Banerjee, S. Kar, S. Santra, Chem Commun. (2008), 3037–3039.
350 351 352
16
Page 16 of 28
353
Figure captions
357
Fig. 1 Influence of pH on Quantum yield of CdTe QDs.
358
cr
355 356
ip t
354
Fig. 2 UV-vis absorption (a) and Fluorescence spectra (b) of TGA-CdTe QDs aqueous solution
360
(5.0×10-4 mol L-1), in different molar ratio of TGA: Cd2+: Te2-.
us
359
an
361
Fig.3 The fluorescence spectra of the TGA-CdTe QDs (5.0×10-5 mol L-1) in the presence of
363
melamine at various concentrations(from top to bottom): (a) 0, (b) 1.0×10-11, (c) 1×10-10, (d) 1×10-9,
364
(e) 1×10 -8, (f) 1×10-7, (g) 1×10-6, (h) 1×10-5 mol L-1. Insert: The calibration plots of melamine.
d te
366
Scheme 1 Mechanism of fluorescence quenching of the QDs in the presence of melamine.
367
Ac ce p
365
M
362
368 369 370 371 372 373 374
17
Page 17 of 28
375
Table 1
377
Influence of pH on the performances on the method (TGA-CdTe QDs: 5.0×10-5 mol L-1).
ip t
376
Linear range (μmol L-1)
Detection limits (μmol L-1)
5.0
0.004 − 2.5
0.002
6.0
0.002 − 5.5
7.0
0.0008 − 15
8.0
0.0065 − 8
9.1
0.0009 − 10
0.0001
10.0
0.0002 − 10
0.00004
0.00001 − 10
0.000005
0.00045 − 6
0.0001
380 381 382
te
d
M
an
0.0009
0.0004 0.0008
Ac ce p
12.0
us
pH
11.0
379
cr
378
383 384 385 386
18
Page 18 of 28
387
Table 2
389
Influence of diameter of the CdTe QDs on the performances on the method (TGA-CdTe QDs:
390
5.0×10-5 mol L-1).
ip t
388
Linear range (μmol L-1)
Detection limits (μmol L-1)
1.7
1.0×10-9 − 1.0×10-7
0.00080
2.4
1.0×10-9 − 1.0×10-6
3.2
1.0×10-9 − 1.0×10-5
3.9
394 395
an 0.00050
2.17 2.37
2.48
1.0×10-11 −1.0×10-5
0.000005
2.64
4.6
1.0×10-9 −1.0×10-5
0.00072
2.79
5.3
1.0×10-8 − 1.0×10-5
0.0030
2.88
1.0×10-7 − 1.0×10-5
0.015
3.16
te
d
M
0.00045
Ac ce p
393
Diameter of the QDs(nm)*
us
Molar ratio(TGA:Cd 2+)
6.0
392
cr
391
* established according to the calculation reported in literature [29].
396 397 398 399
19
Page 19 of 28
400
Table 3
401
Comparison of the performances of different methods.
0.396 − 0.793
Gas chromatography/mass spectrometry [3]
0.396 − 1.59
HPLC method [6]
Colorimetric method/AuNPs [11]
0.793
0.396 − 79.3
0.0792
0.001 − 10
4.0 − 170.0
M
Label-free silver nanoparticles [12]
0.6 – 1.6
0.0075 − 0.35
te
d
Label-free AuNPs [17]
CdS quantum dots as fluorescence probe [21]
0.002 − 50 0.0048 − 1.6
Ac ce p
Colorimetric method/AuNPs [22] FRET/fluoresce in-AuNPs[34]
0.1 − 4.0
CuInS2 QDs as fluorescence probe [35]
0.0793
7.93 − 634
an
Capillary zone electrophoresis [7]
FRET/CdTe@SiO2-AuNPs [19]
0.0396
us
Gas chromatography−mass spectrometric [2]
Detection limits (μmol L-1)
cr
Linear range (μmol L-1)
Methods
ip t
402
0.0008 2.32 0.2 0.00089 0.001 0.00064 0.001
0.01 − 10
0.005
icELISA with QDs-Ab2 [36]
0.069 − 0.90
0.031
FI-CL [37]
0.0015 − 0.64
0.0008
ELISA [38]
0.0006 − 0.0058
0.00002
Luminol-hydrogen peroxide chemiluminescence[39]
0.00002 – 0.002
0.000007
The present method
0.00001 − 10
0.000005
403 404 20
Page 20 of 28
406
Table 4
407
Determination of melamine in milk powder and pet feeds samples.
Added (μmol L-1)
Found (μmol L-1)
Milk powder 1
6.82
7.93
14.99
103.0
Milk powder 2
7.30
7.93
15.38
102.0
Milk powder 3
9.83
7.93
18.16
105.0
Milk powder 4
9.20
11.89
24.74
97.3
pet feeds 1
15.46
11.89
27.20
98.7
pet feeds 2
22.13
15.86
38.53
103.5
pet feeds 3
16.97
15.86
32.59
98.5
cr
an
M d
Recovery (%)
te
410
Background (μmol L-1)
Ac ce p
409
Sample
us
408
ip t
405
21
Page 21 of 28
OOCH2CS d C CS H2 C - OO
-
OO H 2C
OO
CdTe Cd 2 SCH2COO-
N H2 N
+
2
H2N
N N
N
Cd 2+ H
N
H2N
N
H Cd 2+ N
N
H2N
O-
2
OO CH
H NH2
Melamine
SC H
2 CO
H
N N
OCO
SCH2COO-
Cd2+
H SC
-
-
2+
Cd S C
N N H
N
NH 2
m-
N
NH2
H Cd 2 H + H N N H H N N N H NH2
Cd2+
an
n+
m-
n+
M
+
2+ i SCH2COO
Melamine coated CdTe QDs
Agglomerates
410
te
d
1 scheme
Ac ce p
412
N
N N
CdTe
Exc
411
H
+
n+
TGA coated CdTe QDs
m-
N 2 Cd
H H N 2
ip t
2+
CS
-
Cd 2+
N
cr
2 CS
n+
NH2 H N
NH2
us
2
CS
CH
m-
2C
CH
OO
SC H
O O -
-
SCH 2COO -
Exc
22
Page 22 of 28
412 Graphical Abstract
Melamine
SC H
2 CO
H NH2
H2N
N N
N
Cd 2+ H
H2N
O-
N
2
N
H2N
N
H Cd 2+ N
N
NH 2
m-
H
+
N
N N
N
NH2
H Cd 2 H + H N N H H N N N H NH2
Melamine coated CdTe QDs
an
Exc
m-
n+
n+
413 414
M
+
2+ i SCH2COO
CdTe
n+
TGA coated CdTe QDs
m-
N 2 Cd
Cd2+
N H
N
H H N 2
ip t
Cd2+
H
N N
N
cr
-
OO
2+
N
OCO
OO CH
+
SCH2COO-
-
-
O CO
2 H2 Cd S C H2 N CdTe Cd SCH2COO-
H SC
OOCH2CS d C CS H2 C - OO
S
2+
2C
-
Cd 2+
H N
NH2
us
2 CS
n+
NH2
2C
2
CS
CH
m-
H
CH
OO
SCH 2COO -
O O -
-
SC
Exc
415
Melamine takes place of the TGA on the surface of TGA-CdTe QDs with negative
416
charge to form melamine coated QDs changing the surface charge of the QDs, resulting
417
the fluorescence quenched as the QDs aggregation occurred by electrostatic attraction
419 420 421
d
te
Ac ce p
418
Agglomerates
of the two opposite charged nanocrystals.
23
Page 23 of 28
Highlights:
422
● An ultrasensitive and selective method for the determination of melamine was
423
developed at pH 11.0.
424
● The selectivity of the method was improved.
425
● The sensitivity of the method enhanced obviously as the CdTe QDs have higher QYs
426
at pH 11.
427
● The sensitivity and linear range for an analyze is size dependent using QDs PL
428
pbobes.
429
● Melamine takes the place of TGA resulting fluorescence quenched of QDs.
an
us
cr
ip t
421
M
430
Ac ce p
te
d
431
24
Page 24 of 28
Ac
ce
pt
ed
M
an
us
cr
i
Fig.1
Page 25 of 28
Ac
ce
pt
ed
M
an
us
cr
i
Fig.2a
Page 26 of 28
Ac
ce
pt
ed
M
an
us
cr
i
Fig.2b
Page 27 of 28
Ac
ce
pt
ed
M
an
us
cr
i
Fig.3
Page 28 of 28
