Sensitive arginine sensing based on inner filter effect of Au nanoparticles on the fluorescence of CdTe quantum dots

Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 173 (2017) 105–113

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Sensitive arginine sensing based on inner filter effect of Au nanoparticles on the fluorescence of CdTe quantum dots Haijian Liu a,1, Ming Li a,1, Linye Jiang a, Feng Shen b, Yufeng Hu c,⁎, Xueqin Ren a,⁎ a b c

Department of Environmental Science and Engineering, College of Resources and Environmental Sciences, China Agricultural University, Beijing 100193, China Agro-Environmental Protection Institute, the Ministry of Agriculture, Tianjin 300191, China School of Food and Environment, Dalian University of Technology, Panjin 124221, China

a r t i c l e

i n f o

Article history: Received 3 May 2016 Received in revised form 29 August 2016 Accepted 29 August 2016 Available online 31 August 2016 Keywords: Arginine CdTe quantum dots Fluorescence Gold nanoparticles Inner filter effect

a b s t r a c t Arginine plays an important role in many biological functions, whose detection is very significant. Herein, a sensitive, simple and cost-effective fluorescent method for the detection of arginine has been developed based on the inner filter effect (IFE) of citrate-stabilized gold nanoparticles (AuNPs) on the fluorescence of thioglycolic acid-capped CdTe quantum dots (QDs). When citrate-stabilized AuNPs were mixed with thioglycolic acid-capped CdTe QDs, the fluorescence of CdTe QDs was significantly quenched by AuNPs via the IFE. With the presence of arginine, arginine could induce the aggregation and corresponding absorption spectra change of AuNPs, which then IFE-decreased fluorescence could gradually recover with increasing amounts of arginine, achieving fluorescence “turn on” sensing for arginine. The detection mechanism is clearly illustrated and various experimental conditions were also optimized. Under the optimum conditions, a decent linear relationship was obtained in the range from 16 to 121 μg L−1 and the limit of detection was 5.6 μg L−1. And satisfactory results were achieved in arginine analysis using arginine injection, compound amino acid injection, even blood plasma as samples. Therefore, the present assay showed various merits, such as simplicity, low cost, high sensitivity and selectivity, making it promising for sensing arginine in biological samples. © 2016 Published by Elsevier B.V.

1. Introduction Arginine is the only one with a guanidine group of the natural amino acid, which has the strongest alkaline and highest isoelectric point [1]. Arginine plays an important role in many biological mechanisms [2–4], for example, which is an antioxidant and regulates the levels of hormone, has immune ability, and helps to maintain blood pressure [5]. Therefore, the sensitive detection of arginine is of great importance. A variety of methods for the detection of arginine have been reported, for example, high-performance liquid chromatography, electrochemical methods, flow injection analysis, liquid chromatography–tandem mass spectroscopy, molecularly imprinted technology [6–10] and fluorescence spectrometry [11–14]. Although these analytical methods have high sensitivity and selectivity, most of them have drawbacks that include expensive equipment, the need for professional operators, complicated laboratory procedures, which restrict their practical applications. Wu's group reported a new Zn(II)–terpyridine complex functions as a reporter for the detection of arginine via FRET [12]. Recently, he reported a new

⁎ Corresponding authors. E-mail addresses: [email protected] (Y. Hu), [email protected] (X. Ren). 1 Haijian Liu and Ming Li contributed equally to this work.

http://dx.doi.org/10.1016/j.saa.2016.08.057 1386-1425/© 2016 Published by Elsevier B.V.

rhodamine–thiourea/Al3 + complex sensor for the fast visual detection of arginine in aqueous media [11]. Ding's group demonstrated the use of the ternary system containing the cationic fluorophore, anionic surfactant, and Cu2 + as a sensitive and selective sensor to arginine [13]. More recently, Ding and co-authors reported a new ternary sensor system based on fluorophore/SDS/Cu2 + by using pyrene-modified cationic fluorophore containing two imidazolium groups for sensitive detection of arginine over different concentration range [14]. Unfortunately, these methods were involved extensive organic framework or required complex procedure in synthesis and purification for their uses as molecular reorganization. Therefore, there is a great demand to develop a rapid, simple, inexpensive, and sensitive method for arginine determination. With the rapid development of nanotechnology, gold nanoparticles (AuNPs) have attracted extensive attention because of their unique physical, chemical, and optical properties [15]. Various morphologies of AuNPs show different colors; monodisperse gold nanoparticles are wine red, so the color of the solution would be transformed into purple or blue after aggregation. By taking advantage of this distinct color change, AuNPs modified with various ligands have been widely applied for the detection of many analytes, such as metal ions, small molecules, and proteins [16,17]. Rawat and co-workers demonstrated the use of ascorbic acid, Trp, hydroxyl amine, quercetin and 4-amino nicotinic acid assembled gold nanoparticles as a colorimetric sensor for visual

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detection of multiple amino acids [18,19]. However, the above probe preparations involve high cost and complex modification largely limits their application. Quantum dots (QDs) have superior luminescent properties [20,21], for example, high fluorescence quantum yield, excellent photostability, and size- and composition-tunable emission wavelength [22–27]. Therefore, QDs have been widely applied in the fluorescent sensing of analytes as ideal probes. For example, Hamid Reza Rajabi's group has employed them as ion and small molecular probes [42–44]. In addition, many biosensors based on the fluorescence resonance energy transfer (FRET) between QDs as donors and AuNPs as acceptors have been developed, while they also have been investigated in bioanalysis [28– 30]. Several research groups have designed various strategies for detecting analytes based on the FRET [38,45,46]. The fluorescence of QDs can be quenched by AuNPs through both the energy-transfer process and the inner filter effect (IFE). The formation of FRET assemblies is complicated, and it is necessary to keep a certain distance between the donor and receptor. However, the inner filter effect (IFE) refers to the absorption of the excitation or emission light of fluorophores by absorbers in the detection system. The IFE would occur effectively when the absorption spectra of the absorber possess complementary overlaps with the fluorescence excitation or emission spectra of fluorophores. Correspondingly, the linking between the absorbers and the lumiphore is not required in IFE detection systems. Consequently, controlling the IFE process is simple and relatively low cost. IFE has been used as an efficient strategy for the design and development of novel sensors by converting the analytical absorption signals into fluorescence signals, which has been proven to commendably enhance the sensitivity and selectivity compared to other fluorescence quenching methods [47,48]. Whereas, this method for arginine detection based on the IFE of citrate-stabilized AuNPs on the fluorescence of water-soluble CdTe QDs capped with thioglycolic acid (TGA) has not been reported. Therefore, we establish the strategy of a simple, sensitive, turn-on fluorescent

technique for the detection of arginine by taking advantage of the principle. Both AuNPs and QDs require no special modification, which offers considerable flexibility and simplicity. Because of the intensive absorption of AuNPs, the fluorescence of CdTe QDs is quenched in the presence of AuNPs. Arginine can easily interact with the carboxyl groups of citrate-capped AuNPs through electrostatic and hydrogen-bonding interactions under the appropriate pH conditions because of the guanidine group of arginine. As a result, the AuNPs aggregate and the solution color changes from red to blue. The absorption of AuNPs decreases. Meanwhile, the decreased fluorescence of CdTe QDs is recovered, based on which arginine can be easily detected. The principle of this method is illustrated in Scheme 1. The proposed biosensor demonstrates a simple, sensitive, and selective method for arginine determination. 2. Experimental 2.1. Materials Cadmium nitrate tetrahydrate, sodium tellurite, sodium borohydride (NaBH4), and amino acids were purchased from Beijing J&K Co., Ltd. (Beijing, China). Thioglycolic acid (TGA) was purchased from TCI (Shanghai) Development Co., Ltd. (Shanghai, China). Chloroauric acid (HAuCl4) and trisodium citrate dehydrate were obtained from Sinopharm Chemical Reagent (Beijing, China). The arginine working solution was obtained via dilution. Arginine hydrochloride injection was purchased from Shanghai Xinyi Jinzhu Pharmaceutical Co., Ltd. (Shanghai, China). Compound amino acid injection was purchased from SinoSwed Pharmaceutical Corp. Ltd. (Jiangsu, China). The pH of the solution was adjusted using Britton–Robinson (BR) buffer. All of the reagents were purchased as analytical grade and were used without further purification. Water was purified using a Milli-Q system (Millipore, Bedford, MA, USA).

Scheme 1. Schematic of the fluorescence change mechanism for the detection of arginine based on the IFE of citrate-capped AuNPs on CdTe QDs.

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2.2. Instrumentation The absorption spectra were recorded on a UV-2102 UV–vis spectrophotometer (Unico Instrument Co., Ltd. Shanghai, China). The fluorescence spectra were acquired on a F-7000 fluorescence spectrophotometer (Hitachi Ltd., Tokyo, Japan) at the excitation wavelength of 365 nm, with both the exciting and emission slits set at 5 nm. Transmission electron microscopy (TEM) images of the nanoparticles were acquired on an H-7500 (Hitachi Ltd., Tokyo, Japan). 2.3. Preparation of Water-soluble TGA-CdTe QDs TGA-capped CdTe QDs were prepared according to the procedure described previously, with some slight modification [31]. Briefly, 0.0617 g of Cd(NO3)2 was first added to 50 mL of water in a threeneck flask, and 18 μL of thioglycolic acid (TGA) was added while stirring. Then, the pH of the solution was adjusted to 10 with 1 M NaOH solution, and the solution was diluted to 200 mL. Then, 0.08 g of NaBH4 and 0.0088 g of Na2TeO3 were added successively to the above solution. The mixture was then refluxed at 100 °C for 1 h to obtain the required TGA-capped CdTe QDs solution. The crude CdTe QDs solution was washed with equal isopropanol and centrifuged to remove excess precursors. The centrifugal purification operation was repeated three times. Finally the QDs deposit was dissolved in water. The concentration of as-prepared CdTe QDs was approximately 2.7 × 10−5 mol L−1, as determined by the excitonic absorption peak value and the extinction coefficient per mole (ε = 10,043 (D)2.12) of CdTe nanoparticles [32]. 2.4. Synthesis of Citrate-capped AuNPs In a typical procedure, all glassware was soaked with freshly prepared aqua regia for 24 h, rinsed with distilled water, and oven dried prior to use. AuNPs were synthesized by Frens' method, as reported previously [33]. Briefly, 50 mL of 1 mM HAuCl4 was added to a 100-mL round-bottom flask equipped with a condenser and heated to boiling. Then, 5 mL of 38.8 mM sodium citrate was rapidly injected into the above solution under vigorous stirring, resulting in a color change of the mixture solution from light gray to wine red. Boiling continued for 10 min; then, the heating device was removed, and stirring was continued for an additional 15 min. After the solution cooled down to room temperature, it was filtered through a 0.22 μm Millipore membrane filter and then the prepared AuNP solution was stored in a 4 °C refrigerator prior to use. The concentration of the solution of AuNPs was calculated to be approximately 4.68 × 10−9 mol L−1 based on Beer's law using UV–vis spectroscopy [34]. 2.5. Fluorescence Detection of Arginine In order to verify the inner filter effect of Au nanoparticles on the fluorescence of CdTe quantum dots, we made the following experiment,

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1 mL of CdTe QDs (2.7 × 10−5 mol L−1) was mixed with 2 mL of AuNPs with various concentrations. Then, the emission spectra were recorded at an excitation of 365 nm. The fluorescence data were analyzed by plotting the fluorescence spectra. The detection of arginine steps is as follows, 500 μL of AuNP solution and 500 μL of BR buffer were mixed with 500 μL of arginine solutions of various concentrations, and the mixture was incubated at room temperature for 15 min; then, 1 mL of CdTe QDs (2.7 × 10− 5 mol L− 1) was added to the above solution. Afterwards, the fluorescence emission spectra were recorded at an excitation of 365 nm. The calibration curve for arginine was established according to the fluorescence enhancement efficiency, which was determined as (F − F0)/F0, where F0 and F are the maximum emission intensity of the system in the absence and presence of arginine, respectively. The selectivity for arginine was studied by adding other amino acids instead of arginine in the same manner. 3. Results and Discussion 3.1. Characterization of AuNPs and TGA-CdTe QDs The preparation of AuNPs was carried out following Frens' citrate reduction method [35]. As shown in curve a of Fig. 1A, the citrate-stabilized AuNP colloid solution has a strong characteristic plasma absorption at 522 nm, and the colloidal solution is red. The average particle size of AuNPs was 13 nm, as determined by transmission electron microscopy, and the AuNPs were well dispersed in aqueous solution, as shown in Fig. 2A. A water-soluble one-pot hydrothermal process was employed for TGA-CdTe QDs synthesis, and the size and the optical properties of the QDs could be tuned by changing the heating time. The size of TGA-CdTe QDs was difficult to characterize using TEM because of their small dimensions and their tendency to aggregate when drying on a copper grid [36]. Fig. 1B shows the TGA-CdTe QD absorption spectrum, according to the empirical fitting function [32]:     D ¼ 9:8127  10−7  λ3 − 1:7147  10−3  λ2 þ 1:0064λ−194:84ðnmÞ

ð1Þ

where D (nm) is the particle size of the TGA-CdTe QDs, and λ is the wavelength of the first excitonic absorption peak of the corresponding QDs. Using this formula, we calculate the average particle size of TGACdTe QDs as 2.3 nm. The molar concentration of TGA-CdTe QDs can be calculated using the Lambert–Beer law: A ¼ εcl

ð2Þ

where A is the value of the first excited absorption peak of TGA-CdTe QDs, c (mol L−1) is the molar concentration of the corresponding QDs, and l (cm) is the path length (cm) of the radiation beam and was set at 1.0 cm here. According to the literature [25], ε can be obtained by

Fig. 1. (A) Absorption spectra of AuNPs (a) and fluorescence emission spectra of TGA-CdTe QDs (b) (B) absorption spectra of TGA-CdTe QDs.

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Fig. 2. (A) TEM image of AuNPs. (B) TEM image of AuNPs after addition of arginine.

the formula:

From Fig. 3, it can be seen that the fluorescence intensity of TGACdTe QDs was quenched gradually with increasing concentration of AuNPs. The Stern–Volmer equation was also obtained (Fig. S3, Supporting Information). The quenching constant Ksv value is determined to be 2.49 × 106 M−1. The citrate-stabilized AuNPs in aqueous solution are stabilized against aggregation because of the negative citrate ions [36]. However, TGA-CdTe QDs also have a negative charge because of the ionization of the -COOH group in TGA (pKa = 3.53) at pH 9.6 [37]. Because both AuNPs and TGA-CdTe QDs possess negative charges, there is electrostatic repulsion between them. However, FRET is a common mechanism arising from the donor-receptor assembly structure, which can be formed by electrostatic interactions between positively charged quantum dots and negatively charged AuNPs. The distance

between the donor QDs and receptor AuNPs could be shortened because of the electrostatic interactions, leading to the quenched fluorescence intensity of QDs [38]. Under a pH of 5.0–7.0, i.e., weak acid or neutral conditions, a FRET donor-acceptor assembly could be established between TGA-modified QDs and citrate ligand-capped AuNPs by hydrogen bond interaction [36]. At pH 9.6, the negatively charged CdTe QDs and the negatively charged AuNPs will not form FRET through electrostatic interactions or hydrogen bonding. Instead, with the coexistence of TGA-QDs and AuNPs, the absorption spectra of the AuNPs did not show significant changes (curve a and b in Fig. 5A), indicating that AuNPs and QDs did not form complexes and energy transfer did not occur. Therefore, the decrease in the fluorescence intensity of TGA-CdTe QDs should be attributed to the IFE of AuNPs on the fluorescence of TGA-CdTe QDs. The formation of a FRET system is complex and difficult to control [39]. Compared with this, the IFE system is simpler. In addition, the fluorescence lifetime of CdTe QDs was hardly changed in the presence of AuNPs. This result further suggested that the quenched fluorescence of CdTe QDs should be attributed to IFE between CdTe QDs and AuNPs (Fig. S4 Supporting information). With increasing AuNP concentration, the absorbance of the AuNP is enhanced, resulting in a decrease in the fluorescence emission of TGA-CdTe QDs. Therefore, the emission intensity of TGA-CdTe QDs can be adjusted by the absorbance of AuNPs via IFE in a simple approach. When the concentration of AuNPs was 2.34 × 10−9 mol L−1, the fluorescence intensity of 2.7 × 10− 5 mol L− 1 TGA-CdTe QDs was quenched by approximately 90%. This effective quenching is attributed to the high extinction coefficient of AuNPs, which displays one advantage of using metal nanoparticles as the absorber compared with conventional chromophores.

Fig. 3. The fluorescence emission spectra of TGA-CdTe QDs in the presence of AuNPs. The concentration of AuNPs is 0, 1.17 × 10–9, 1.3 × 10–9, 1.46 × 10–9, 1.67 × 10–9, 1.8 × 10–9, 1.95 × 10–9, 2.34 × 10–9, 2.93 × 10–9, 3.9 × 10–9, and 4.68 × 10–9 mol L−1, respectively. The concentration of TGA-CdTe is 2.7 × 10–5 mol L−1; pH 9.6.

Fig. 4. Absorption spectra of 2.34 × 10−9 mol L−1 AuNPs in the presence of arginine at various concentrations. The concentration of arginine in the sample are 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000 and 1200 μg L−1.

ε ¼ 10043ðDÞ2:12 :

ð3Þ

The molar concentration of TGA-CdTe QDs calculated using this equation was approximately 2.7 × 10−5 mol L−1. In addition, as shown in curve b of Fig. 1A, the TGA-CdTe QDs show a maximum fluorescence emission at 550 nm, which is similar to the maximum absorption of AuNPs (520 nm). It is clear that the absorption spectrum of AuNPs overlaps well with the fluorescence emission spectrum of TGA-CdTe QDs from Fig. 1A. Thus, when the two types of materials coexist, the fluorescence intensity of TGA-CdTe QDs can be greatly quenched or decreased. 3.2. IFE Leading to the Fluorescence Changes of TGA-CdTe QDs

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Fig. 5. (A) Absorption spectra: (a) AuNPs; (b) AuNPs + CdTe QDs; (c) AuNPs + arginine; (d) AuNPs + arginine + CdTe QDs; (e) CdTe QDs; (f) CdTe QDs + arginine. (B) Fluorescence spectra: (a) CdTe QDs; (b) CdTe QDs + arginine; (c) AuNPs + arginine + CdTe QDs; (d) AuNPs + CdTe QDs. TGA-CdTe QDs, 2.7 × 10−5 mol L−1; AuNPs, 2.34 × 10−9 mol L−1; arginine, 500 μg L−1.

3.3. Interactions Between Arginine and AuNPs In Fig. 4, we can see that arginine can cause changes in the absorption spectra of AuNPs. With increasing concentration of arginine, the characteristic surface plasmon absorption peak of AuNPs at 522 nm decreased gradually; at the same time, new absorption peaks appeared at long wavelength (approximately 650 nm) and increased gradually, resulting in a gradual color change of AuNP solution from red wine to purple (or gray). The corresponding color change of AuNPs driven by arginine has been used for the colorimetric detecting of arginine, but this colorimetric technique generally displays a lower sensitivity than that of the fluorescence detecting assays. As is well known, the surface plasmon absorption of AuNPs is closely related to the distance between particles. When the distance between the particles decreases, the first absorption peak is weakened because of the electronic coupling between the AuNPs, resulting in a red shift of the absorption peak [39]. As shown in Scheme 1, arginine with positively charged guanidine could bind to the surface of citrate-stabilized AuNPs through electrostatic attraction, and arginine molecules could be combined together through hydrogen-bonding interactions, eventually leading to AuNP aggregation [40]. Arginine-induced aggregation of AuNPs was further confirmed by TEM. As shown in Fig. 2, the AuNPs were in a monodisperse state (Fig. 2A) in the absence of arginine. However, after the addition of arginine they were aggregated quickly (Fig. 2B). These results were consistent with the change in the absorption spectra (Fig. 4). Therefore,

the AuNP system can regulate the fluorescence emission intensity of TGA-CdTe QDs via IFE, which is a new theory for the development of a fluorescence detection method for arginine. More importantly, the fluorescent sensor composed of CdTe QDs and AuNPs has various advantages, such as enhanced signals, and lower detection limit. 3.4. IFE-based TGA-CdTe QDs Fluorescence Response to Arginine Concentration From the above analysis, it is clear that arginine can cause the aggregation of AuNPs and the fluorescence intensity of TGA-CdTe QDs can be quenched by AuNPs. Thus, it is reasonable to expect that the IFE of AuNPs on TGA-CdTe QDs could be influenced by arginine, which aggregates the AuNPs and modulates the IFE process. To verify the principle for arginine detection, the effect of arginine on the absorption and fluorescence spectra of AuNPs-CdTe QDs was investigated. As shown in Fig. 5, the absorption spectra and fluorescence spectra of TGA-CdTe QDs (curve e in Fig. 5A and curve a in Fig. 5B, respectively) were consistent with those of the mixture of arginine and TGA-CdTe QDs (curve f in Fig. 5A and curve b in Fig. 5B), which indicates that there is no interaction between QDs and arginine. The absorption spectra of AuNPs remained almost unchanged in the presence of TGA-CdTe QDs (curves a and b in Fig. 5A), indicating that there was no interaction between AuNPs and TGA-CdTe QDs. Therefore, the arginine-induced changes in the absorption spectrum of AuNPs were identical with or without the presence of TGA-CdTe QDs (curves c and d in Fig. 5A), indicating that AuNP aggregation is caused by arginine. When TGA-CdTe QDs were mixed with AuNPs, the fluorescence intensity of TGA-CdTe QDs was quenched by the IFE of AuNPs (curve d in Fig. 5B). However, the quenched fluorescence of TGA-CdTe QDs was restored in the presence of arginine (curve c in Fig. 5B). Meanwhile, the TGA-CdTe QD fluorescence spectral shape was not changed obviously in the presence of arginine and AuNPs, which indicated that the increase in fluorescence intensity came from TGA-CdTe QDs rather than any other newly formed fluorescence material. Thus, the recovery of the observed fluorescence intensity was due to the interaction between the arginine and AuNPs, which then affected the fluorescence emission of TGA-CdTe QDs. Arginine could reduce the absorption of AuNPs at 522 nm, which reduced the IFE of AuNPs on TGA-CdTe QDs. Subsequently, the fluorescence of TGA-CdTe QDs was recovered. Based on this principle, we developed a new, sensitive IFE-based fluorescence method for determination of arginine. 3.5. Optimization of Analysis Conditions

Fig. 6. The influence of different pH on (F − F0) / F0 for the detection of 31 μg L−1 arginine. AuNPs, 2.34 × 10−9 mol L−1; CdTe QDs, 2.7 × 10−5 mol L−1.

In this work, the proposed method for arginine detection is based on the analyte-induced decrease in the absorbance of the AuNPs, then recovery of the fluorescence of CdTe QDs. Therefore, the analysis

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Fig. 7. Time-dependent absorption spectra (A) and time-A522 variation curve (B) of AuNPs (2.34 × 10−9 mol L−1) in the presence of arginine (500 μg L−1).

conditions, such as media pH, AuNP concentration, and interaction time, have a considerable influence on the performance of the proposed arginine assay. The pH is an important factor in this work. On the one hand, the fluorescence emission intensity of TGA-CdTe QDs was influenced by the pH of the environment [32]. On the other hand, as the most alkaline amino acid, arginine has the highest pI, at approximately 10.8 [41]. The pH affects the electrical properties of arginine, which then affects the

Fig. 8. Fluorescence response to arginine of a solution containing 2.7 × 10−5 mol L−1 CdTe QDs to arginine in the presence of various concentration of AuNPs. a; 2.34 × 10−9 mol L−1 b; 1.95 × 10−9 mol L−1 c; 2.93 × 10−9 mol L−1.

interaction between arginine and AuNPs. When the pH is less than the isoelectric point of arginine, the positively charged guanidine group of arginine can connect to the surface of citrate-capped AuNPs through electrostatic and hydrogen-bonding interactions [29]. However, if the pH is much lower, it not only affects the fluorescence intensity of TGA-CdTe QDs, but also increases the interference of other amino acids, affecting the selectivity of arginine. Therefore, we investigated the effect of pH from 9.4 to 10.8 on the detection of arginine. As Fig. 6 shows, when the media pH is 9.6, the value of (F − F0)/F0 was the maximum. Thus, we chose 9.6 as the optimal pH for further experiments. The reaction time of arginine-AuNP binding and AuNP–CdTe QD interactions were also investigated. After adding arginine to the AuNP solution, the absorption spectrum and arginine-AuNP binding time was studied by recording the spectra every 1 min. As Fig. 7 shows, the absorption spectrum showed almost no change after 15 min. Therefore, the interaction between arginine and AuNPs was completed within 20 min. Therefore, we chose 15 min as the incubation time for arginine–AuNP binding. However, the fluorescence intensity of CdTe QDs was immediately quenched after the addition of CdTe QDs to the AuNP solution. Therefore, it can be directly measured without incubation after the addition of CdTe QDs into the above arginine-AuNP mixture. The fluorescence response towards arginine was easily affected by the concentration of AuNPs in the system. As shown in Fig. 8, when the concentration of AuNPs was 2.34 × 10−9 mol L−1, both the sensitivity and linear range in the experiment were the best. With increasing concentration of AuNPs, the fluorescence of CdTe QDs was quenched more effectively and the background fluorescence was also decreased, but the fluorescence response was weak for low concentrations of arginine (Fig. S5B Supporting information). Meanwhile, the assay sensitivity was reduced. However, when the concentration of AuNPs was lower, the fluorescence response with low concentrations of arginine improved, but the sensitivity of the method was influenced and the linear

Fig. 9. Fluorescence emission spectra of QDs-AuNPs system in the presence of various concentrations of arginine (A) and linear plot of (F-F0)/F0 versus the concentration of arginine (B) for concentrations of 16, 31, 46, 61, 76, 91, 106 and 121 μg L−1, respectively. TGA-CdTe QDs; 2.7 × 10−5 mol L−1 AuNPs; 2.34 × 10−9 mol L−1.

H. Liu et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 173 (2017) 105–113 Table 1 Comparison of LODs of different arginine detecting methods.

Table 2 Determination results of arginine in spiked samples. LOD (μg

Method Reversed-phase liquid chromatography NMR spectroscopy Molecularly imprinted technology NPs-based UV–visible spectroscopy NPs-based UV–visible spectroscopy AuNPs-based colorimetry Fluorescence spectroscopy Fluorescence spectroscopy Fluorescence spectroscopy Fluorescence spectroscopy AuNPs–CdTe IFE-based fluorescence spectroscopy

Linear range

L−1)

0.00001–0.00015 0.48–1.74 μM – 0.1–100 μM 2.5–1250 μM 1–100 μM 0.08–13 μM – 0–120 μM 0–35 μM 0–100 μM 16–121 μg L−1

111

5400.2 0.015 6.97 174.2 69.7 2.6 400.7 29.6 0.9 5.6

Added (μg

Found (μg

Ref.

Sample L−1)

L−1)

Recovery ± RSD (%) (n = 3)

Precision (%)

Accuracy (%)

[49]

1 2 3

18.75 39.61 61.13

93.75 ± 2.47 99.02 ± 1.62 101.88 ± 2.75

2.47 1.62 2.75

−6.2 −0.9 1.8

[50] [8] [18] [51] [40] [12] [11] [13] [14] This work

range was narrow because of the high background fluorescence (Fig. S5A Supporting information).

3.6. Sensitivity and Selectivity Under the optimum conditions, the analytical parameters of the developed method for arginine detection were investigated. As shown in Fig. 9, the fluorescence intensity of the system enhanced gradually with increasing arginine concentration. Correspondingly, the UV–vis absorbance spectra of the AuNPs at 520 nm gradually decreased (Fig. 2). The ratio between the increased fluorescence intensity (F − F0) and F0 was linearly related to the arginine concentration in the linear range from 16 to 121 μg L− 1 (Fig. 9A), which could be expressed as (F − F0)/F0 = 0.08345 C arginine −0.46527 with a correlation coefficient of 0.9943 (Fig. 9B). The detection limit (3σ/slope) was estimated to be 5.6 μg L−1 (where σ is the relative standard deviation of a blank solution, n = 11). Compared with other methods (Table 1), the detection limit of this method is quite competitive. To explore the specific detection of arginine using the developed method, the effects of the other 19 amino acids were examined. In addition, the interferences of major metal ions and anions were investigated for determination of arginine. We selected Na+, Mg2+, K+, Cu2+, Ag+, Fe2+, Co2+, Zn2+, Ni2+, and Cl− as the interference ions in this experiment. The selectivity of the fluorescence assay to arginine compared with other metal ions was shown in Fig. S2 (Supporting information). The fluorescence spectra of AuNP-QDs in the presence of arginine and the fluorescence spectra of AuNP-QDs in the presence of other amino acids, metal ions and anions are shown in Fig. 10A, and the corresponding responses are shown in Fig. 10B. It can be seen that most of the other amino acids did not interfere with the detection of arginine. The amino

20 40 60

acids are negatively charged under the acidity of the experiment and do not induce AuNP aggregation because of electrostatic repulsion; therefore, they also do not cause any change in the fluorescence intensity. Although the concentrations of interfering substances were 100-fold higher than that of arginine, the fluorescence intensity of TGA-QDs had no obvious change (Fig. S2 Supporting information). Thus, the proposed method exhibits a highly selective fluorescence response to arginine. 3.7. Detection of Arginine in Real Samples To verify the practicality of this method for the detection of arginine, the fluorescence response for spiked arginine samples are shown in Table 2. Arginine hydrochloride injection (sample 1 in Table 3) and compound amino acid injection (sample 2 in Table 3) were used as real samples, and the detection results are shown in Table 3. The arginine injection recovery was 100.42%, with a relative standard deviation of 2.61%. The compound amino acid injection recovery was 98.84%, with a relative standard deviation of 1.77%. This assay was also applied to detect arginine in blood plasma. The results are presented in Table 4. The blood plasma samples were collected from Peking University Third Hospital. The collected samples were diluted to 50 times and then spiked with different concentrations of Arginine (10, 50 and 100 μg L−1). The spiked samples were analyzed separately by the aforementioned procedure for detection of arginine. These results indicate that the proposed fluorescence analysis is accurate, reliable, and can be used in practical applications. 4. Conclusions In this work, a sensitive turn-on fluorescent detection analysis for arginine was developed using the inner filter effect of AuNPs on CdTe QDs. Upon addition of CdTe QDs to AuNPs, the fluorescence intensity of CdTe QDs was quenched because of the IFE of AuNPs. When the pH was lower than the pI of arginine, the arginine was positively charged. When arginine was incubated with AuNPs, it induced the aggregation of AuNPs because of electrostatic and hydrogen-bonding interactions. It also decreased their characteristic surface plasmon absorption, which then led to the recovery of the IFE-modified decrease in fluorescence

Fig. 10. Fluorescence emission spectra (A) and the corresponding (F-F0)/F0 (B) of AuNPs-QDs in the presence of arginine or other amino acids. Concentrations: AuNPs, 2.34 × 10−9 mol L−1; QDs, 2.7 × 10−5 mol L−1; arginine, 46 μg L−1; the other amino acids, 1000 μg L−1; Na+, Mg2+, Cl−, 1000 μg L−1; pH 9.6.

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Table 3 Determination results of arginine in arginine hydrochloride injection and compound amino acid injection. Sample

Certified concentration (μg L−1)

Found (μg L−1)

Recovery ± RSD (%) (n = 3)

Precision (%)

Accuracy (%)

1 2

24 32

24.10 31.63

100.42 ± 2.61 98.84 ± 1.77

2.61 1.77

0.4 −1.1

Table 4 Determination results of arginine in blood plasma samples. Blood plasma sample

Added (μg Found (μg Recovery ± RSD L−1) (%) (n = 3) L−1)

Precision Accuracy (%) (%)

1 2 3

10 50 100

1.78 2.34 1.46

8.41 46.78 104.57

84.1 ± 1.78 93.56 ± 2.34 104.57 ± 1.46

−15.9 −6.4 4.5

Accuracy was calculated from (found concentration − known concentration) / known concentration × 100 in the Tables 2, 3, 4.

intensity of CdTe QDs. This proposed method showed high precision and sensitivity, with a detection limit as low as 5.6 μg L− 1. Because AuNPs and QDs require no additional modification or label, the method is simple and rapid. Moreover, this IFE-based fluorescent arginine assay method is more sensitive than absorbance-based methods using AuNPs as colorimetric probes. Therefore, the proposed method is a promising approach for detection of arginine in real samples.

Acknowledgments This work was supported by the Chinese National Scientific Foundation (21375146).

Appendix A. Supplementary data Supplementary data to this article can be found online at http://dx. doi.org/10.1016/j.saa.2016.08.057.

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