Nitrogen and Phosphorus Co-Doped Carbon Nanodots as a Novel

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Nitrogen and Phosphorus Co-doped Carbon Nanodots as a Novel Fluorescent Probe for Highly Sensitive Detection of Fe3+ in Human Serum and Living Cells Bingfang Shi, Yubin Su, Liang-Liang Zhang, Mengjiao Huang, Rongjun Liu, and Shulin Zhao ACS Appl. Mater. Interfaces, Just Accepted Manuscript • DOI: 10.1021/acsami.6b01325 • Publication Date (Web): 25 Mar 2016 Downloaded from http://pubs.acs.org on March 26, 2016

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ACS Applied Materials & Interfaces

Nitrogen and Phosphorus Co-doped Carbon Nanodots as a Novel Fluorescent Probe for Highly Sensitive Detection of Fe3+ in Human Serum and Living Cells

Bingfang Shi,a,b Yubin Su,a Liangliang Zhang,a,* Mengjiao Huang,a Rongjun Liua and Shulin Zhaoa,*

a

State Key Laboratory for the Chemistry and Molecular Engineering of Medicinal Resources, School of Chemistry and Pharmacy, Guangxi Normal University, 15 Yucai Road, Guilin 541004, China

b

Key Laboratory of Regional Ecological Environment Analysis and Pollution Control of West Guangxi, College of Chemistry and Environmental Engineering, Baise University, 21 Zhongshan Road, Baise, 533000, China

ABSTRACT: Chemical doping with heteroatoms can effectively modulate physicochemical and photochemical properties of carbon dots (CDs). However, the development of multi heteroatoms co-doped carbon nanodots is still in its early stage. In this work, a facile hydrothermal synthesis strategy was applied to synthesize multi heteroatoms (nitrogen and phosphorus) co-doped carbon nanodots (N,P-CDs) using glucose as carbon source, and ammonia, phosphoric acid as dopant, respectively. Compared with CDs, the multi heteroatoms doped CDs resulted in dramatic improvement in the electronic characteristics and surface chemical activities. Therefore, the N,P-CDs prepared as described above exhibited a strong blue emission and a sensitive response to Fe3+. The N,P-CDs based fluorescent sensor was then applied to sensitively determine Fe3+ with a 1

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detection limit of 1.8 nM. Notably, the prepared N,P-CDs possessed negligible cytotoxicity, excellent biocompatibility, and high photostability. It was also applied for label-free detection of Fe3+ in complex biological samples and the fluorescence imaging of intracellular Fe3+, which indicated its potential applications in clinical diagnosis and other biologically related study. KEYWORDS: Nitrogen, phosphorus, doped carbon nanodots, cell imaging, Fe3+ detection

1. INTRODUCTION The level of Fe3+ is a significant factor for evaluating the water quality. Moreover, Fe3+, an essential trace element in plants and animals, performs crucial roles in cellular metabolism, enzyme catalysis, oxygen transport in haemoglobin and as a cofactor in enzyme-based reactions.1−5 Fe3+ overload and deficiency can disturb the cellular homeostasis, and result in various diseases, such as anemia, arthritis, intelligence decline, heart failure, diabetes and cancer.6,7 Therefore, detecting Fe3+ is vital for the early identification and diagnosis of these diseases.8 Conventional qualitative and quantitative strategies for Fe3+detection, such as spectrophotometry,9 atomic absorption spectrometry,10 inductively coupled plasma mass spectrometry11 and voltammetry,12 required complex instrumentation and tedious sample preparation procedures, which limited their prosperous applications.8 In recent years, many chemosensors based on organic dyes,13 semiconductor quantum dots,14 fluorescent metal nanoclusters15 and fluorescent metal organic frameworks16 are particularly attractive due to their admirable selectivity. However, specialized synthetic skills and complicated purification procedures are needed to prepare materials mentioned above.17 Additionally, most of these chemosensors are greatly limited in their ability to detect Fe3+ in living cells, because they are poisonous, biologically incompatible, and water-insoluble. Thus, it becomes increasingly important and even urgent to create novel probes possessing the good photostability, biocompatibility, and sensitivity enough to detect Fe3+ in living cells. 2


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Recently, tremendous interest has focused on carbon dots (CDs) because of their low toxicity, good photostability, excellent biocompatibility, cell membrane permeability and tunable surface functionalities.18−20 Consequently, CDs have been widely used in fluorescent bioimaging and biosensing.21−25 However, CDs have a low quantum yield (QY), which limits their applications. Their band gap and electronic density could be effectively modulated using chemical doping with heteroatoms, which would enhance the QY of CDs and are critically required in biological imaging.26 In the past few years, the design and development of fluorescence sensors based on the doped CDs for the determination of Fe3+ is becoming an important project because of their simplicity, easy monitoring, rapid response and high sensitivity.8,17,26–29 For example, Li et al. achieved highly selective detection of Fe3+ (with a detection limit of 4.2 nM) by synthesizing sulfur-doped graphene quantum dots (S-GQDs) using one-step electrolysis.8 Zhang et al. proposed a Fe3+ assay with a 2.5 nM detection limit using nitrogen-doped carbon quantum dots (N-CDs) based fluorescence sensor, which was then applied for the fluorescence imaging of intracellular Fe3+.17 Indeed, most of the fluorescent Fe3+ sensors are signaled by fluorescence quenching originating from the special coordinate interaction between Fe3+ and electron-rich sites on the surface. Therefore, preparing electron-rich CDs that are an electron donor and are prone to binding with Fe3+ is still imminently desired in the detection of intracellular Fe3+. It is well-known that nitrogen is a general dopant in the preparation of CDs because not only its atomize size is comparable to carbon, but also it can bond strongly with the carbon atoms due to its five valence electrons.30 What’s more, C-Dots become n-type after doping with N atoms.31 As for phosphorous, when it behaves as an n-type donor, it can cause a substitutional defect in sp3-bonded diamond thin films.32 Thus, the electronic characteristics of C-Dots could be altered by doping with nitrogen and phosphorus atoms in order to create more active sites, which could produce new and 3



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unanticipated properties. More recently, nitrogen and phosphorus have been reported used as heteroatoms to prepare the doped CDs.33–35 Although these new approaches have shown success in synthesizing carbon nanomaterials co-doped with nitrogen and phosphorus, they inevitably require rigorous synthesis temperature (heating rate 1.5 oC/min to 180 oC,33 900 oC,34 and control temperature to avoid any potential explosion35). Herein, we proposed a facile and high-output hydrothermal approach for synthesizing nitrogen and phosphorus co-doped carbon dots (N,P-CDs) that use glucose as carbon source, and ammonia, phosphoric acid as dopant, respectively (Scheme 1). The prepared N,P-CDs exhibited sufficient water solubility and excellent fluorescence properties; they can also resist light illumination and extreme pH. Moreover, a label-free chemosensor based on the as-prepared N,P-CDs have been developed and showed a sensitive response to Fe3+ in the concentration range of 5−100 nM with the detection limit of 1.8 nM. Significantly, the as-prepared N,P-CDs possessed negligible cytotoxicity, excellent biocompatibility. This proposed chemosensor based N,P-CDs was further applied for detection of Fe3+ in human serum and intracellular Fe3+ imaging.

2. EXPERIMENTAL DETAILS 2.1. Reagents and materials. Glucose, ammonia, phosphoric acid, FeCl3, CuCl2, CaCl2, CoCl2, Zn(NO3)2, AgNO3, CdCl2, Mg2SO4, NaCl, MnCl2, and CrCl3 were purchased from Aladdin Chemistry Co. Ltd. (Shanghai, China). Other chemical solvents and reagents were of analytical grade and did not require additional purification before use. A Milli-Q plus 185 equip from Millipore (Bedford, MA) was emploied to purify the water. 2.2. Characterization. The apparatus for the N,P-CDs characterization are same with that used in previous work.36 4


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2.3. Synthesis of the N,P-CDs. A hydrothermal method was used to prepare the N,P-CDs. Briefly, glucose (1.0 g), ammonia (2 mL), and phosphoric acid (2 mL) were mixed with 2 mL ultrapure water, after which the mixture was transferred into a 50 mL Teon-lined stainless steel autoclave. The mixture was then heated at 160 °C for 5 h under vigorous stirring. After being cooled to room temperature naturally, the resulting dark brown mixture was centrifuged for 15 min at 12,000 rpm to dislodge non-fluorescent deposit, and then dialyzed for 48 h in a dialysis membrane (1000 MWCO, Shanghai Green Bird Science & Technology Development Co., China). Under vacuum condition, the prepared N,P-CDs were then dried for 48 h at room temperature and stored at 4°C for later use. 2.4. Detection of Fe3+ using N,P-CDs. The detection of Fe3+ was conducted in Tris-HCl (10 mM, pH =7.0) buffer solution at room temperature. Typically, 10 µL N,P-CDs (10.0 µg mL-1) were first dispersed into 80 µL Tris-HCl buffer, and then different concentrations of Fe3+ were added. The resulting solution was shaken to ensure complete mixture and incubated for 35 min before recording the fluorescence emission spectra at 437 nm. For comparison, the same procedure was also performed on blank experiments. All experiments were conducted in triplicate, and the standard deviation was plotted as the error bar. To evaluate the feasibility of the proposed method for analysis of Fe3+ in complex biological samples, the chemosensors based on N,P-CDs was challenged by human serum obtained from the No. 5 People’s Hospital (Guangxi, China). After human serum samples were centrifuged and removed the protein deposit, the supernatant was diluted 100-fold with Tris-HCl buffer (10 mM, pH=7.0). Thus, 10 µL N,P-CDs (10.0 µg mL-1) and 10 µL different concentration of Fe3+ were dispersed in 80 µL diluted human serum. After the resulting solution incubated for 35 min at room temperature, the fluorescence emission spectra at 437 nm were recorded. 5



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2.5. Cytotoxicity investigation. T24 cells were seeded in a 96-well plate that contained Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal bovine serum (FBS, invitrogen). The cells were cultured first for 24 h in a humidified atmosphere containing 5% CO2 at 37 oC. Then, different concentrations (0, 5, 10.0, 30.0, 90.0, and 270.0 µg mL-1) of the N,P-CDs solution were added into each well, followed by incubation for 24 h. PBS buffer was used to wash samples three times, after which every cell well received fresh culture medium containing 20 µL MTT (5 mg mL-1), followed by an additional 4 h incubation period. The culture medium was then removed, and the obtained mixtures were dissolved in 100 µL DMSO and shaken for 10 min. The optical density (OD) of the mixture was measured at 450 nm with a Microplate Reader Model. The cell viability was estimated according to the following equation:

Cell viability (%) =

OD OD

treated

× 100%

control

Where ODcontrol was obtained in the absence of N,P-CDs, and ODtreated was obtained in the presence of N-CNs. 2.6. Fluorescence imaging of living cells. The T24 cells in the DMEM supplemented with 10 % fetal bovine serum were added into each 35 mm glass culture dishes. The cells were cultured for 24 h in an incubator (37 oC, 5% CO2). Then, 40 µL N,P-CDs solution (10.0 µg mL-1) was added into the cultured mixture and followed by another 24 h culturing period. Before imaging, the N,P-CDs loaded cells were rinsed three times, and incubated with 1 mL PBS buffer (0.2 M) containing different concentration of Fe3+ at pH=7.0 for 20 min. Images of the cells were immediately captured at ambient temperature on a confocal laser scanning microscopy.

3. RESULTS AND DISCUSSION 6


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3.1. Characterization of the N,P-CDs. The morphology and structure of the N,P-CDs were characterized through images obtained via transmission electron microscopy (TEM) and shown in Figure 1. As displayed in Figure 1a, the prepared N,P-CDs were well monodispersed. In the selected area, the size of N,P-CDs were distributed mainly in a narrow range of 2.2 to 3.5 nm, and almost 63% of N,P-CDs are concentrated at an average size of 3.0 nm (Figure 1b), a result comparable to N-CDs as previously reported (around 3.0 nm)37 and smaller than that of reported N,P-CDs (average 8.1 ± 2.7 nm).33 High resolution TEM (HRTEM) imaging (inset of Figure 1a) showed that the prepared N,P-CDs displayed a highly crystalline structure, with a 0.20 nm lattice parameter, which was similar to graphitic (sp2) carbon’s (102) diffraction plane.38 Fourier transform infrared spectroscopy (FTIR) spectrum was performed to investigate functional groups on the surface of the as-prepared N,P-CDs. Figure S1 shows that the stretching vibration of O-H was assigned to the absorption band at 3441 cm−1, and N-H was assigned to the 2989 cm−1 absorption band.39,40 Sharp peaks at 1383 cm−1 and 1058 cm−1 resulted from the bending vibration of N-H and the starching vibrations of C-N, respectively,41,42 which indicated the doping of N atoms into the N,P-CDs. Simultaneously, the stretching vibration of PO43– resulted in the peak at 531 cm−1, which confirms the presence of P atoms in the N,P-CDs. The results of FTIR indicate that the N,P-CDs are rich in hydroxyl, carbonyl, amino and phosphoric acid groups on their surfaces. These functional groups improved the hydrophilicity and stability of the N,P-CDs in an aqueous system. X-Ray photoelectron spectra (XPS) were carried out to further identify the surface functional state and components of the N,P-CDs. The XPS survey spectrum showed that C1s, N1s, P2s, P2p, and O1s signals appeared at 285.0, 398.5, 188.8, 132.9 and 530.5 eV, respectively (Figure 2a). In addition, composition analysis of the N,P-CDs showed the following elemental composition: carbon 7



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(62.0%), nitrogen (6.8%), phosphorus (5.2%) and oxygen (26.0%). The doping percentages of N and P are higher than that of previously reported N,P-CDs.33 The high resolution C1s spectrum of N,P-CDs was well-fitted into C-C/C-N/C-P bonds (285.1 eV), C=C (284.5 eV), C–O (286.0 eV), and O–C=O (288.0 eV) (Figure 2b),34 which was consistent well with the results of the FTIR results. Here, the peak located at 285.1 eV reflected bonding structure in the C–N/C-P bonds that differed from typical bonds and which originated from the doping of N and P atoms. Two main peaks in the N1s spectrum indicate the binding energy at 399.3 eV for the N-H bond and at 401.1 eV for C–N/N–N/N–P bond (Figure 2c).43 These results further confirmed that N atoms had been presented in N,P-CDs. The P2p XPS spectrum of N,P-CDs was able to be decomposed into peaks at 133.0 eV and 133.8 eV (Figure 2d) indicating bonds between P–O and P–C/P–N.44 These results illustrated PO43– was indeed covalently bonded to the N,P-CDs, which was also in good agreement with FTIR results. All results of XPS and FTIR spectra clearly confirmed that N and P atoms had perfectly doped in the CDs. It was deduced that the electronic properties and surface chemical reactivities of N,P-CDs may be improved because of the introduction of N and P atoms. 3.2. Optical properties of the N,P-CDs. Optical properties of the prepared N,P-CDs were examined under different excitation wavelengths for fluorescence behavior and confirmed via observation of the UV−vis absorption spectrum. As illustrated in Figure S2, the prepared N,P-CD solutions showed an intense absorption band at 276 nm, and which was suggested to result from π–π* transitions of C=C, C=N or N=P groups.33 Additionally, the N,P-CDs solution displayed a broad UV−vis absorption at 300 nm, which was ascribed to the n−π* transition of C=O.8 A bright blue luminescence of N,P-CDs was observed under excitation at 365 nm (Figure 3, inset). Additionally, the emission wavelength of N,P-CDs was red-shifted with the excitation wavelength increasing from 316 nm to 406 nm (Figure 3), as in the case of most of the carbon 8


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nanomaterials.45−47 The excitation-dependent fluorescence behaviour of N,P-CDs may be resulted from their different sizes and surface states.48,49 In addition, the QY of as-prepared N,P-CDs was measured to be 30% using quinine sulfate as the reference (QY=54%, λex=336 nm), which was higher than other doped CDs.8,28 Thus, it was competent for acting as a highly sensitive chemosensor. To investigate the stability of the N,P-CDs, the effects of pH, ionic strengths, and UV exposure on their fluorescence intensities were measured. As presented in Figure S3, the N,P-CDs exhibited stable fluorescent in the range of pH 4 to 9. Figure S4 shows how ionic strength affected N,P-CD stability in Tris-HCl buffer solution (10 mM, pH=7.0) with varying NaCl concentrations. The fluorescent intensity changed only slightly, even in concentrations of NaCl up to 5.0 M, which was result from no ionization of the groups located on the N,P-CDs surface.17 Furthermore, the fluorescence intensity had little change after continuous Xe lamp (365 nm) irradiation for 40 min (Figure S5), which suggested that the N,P-CDs possessed excellent photostability. Taking the results above, it was confirmed that the N,P-CDs had great potential for sensing applications under physiological conditions. 3.3. Fluorescence response of N,P-CDs to Fe3+. Given the special covalent binding between Fe3+ and amino as well as phosphoric acid groups,50 exploration was begun into the feasibility of the N,P-CDs’ fluorescence response to Fe3+. As shown in Figure S6, an obviously decrease in the fluorescence intensity of N,P-CDs at 437 nm were observed in the presence of Fe3+, which revealed that N,P-CDs do respond to Fe3+. Inspired by this, we deduced that Fe3+ had coordinated with amino and phosphoric acid groups on the surface of the N,P-CDs, and then the electrons in the excited state S1 of the N,P-CDs transfer to the half-filled 3d orbits of Fe3+, resulting in fluorescence quenching because of the nonradiative electron/hole recombination annihilation (Figure 4).51 9



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To get in sight into the mechanism of fluorescence quenching of N,P-CDs in the presence of Fe3+, UV–vis absorption spectra and fluorescence lifetime experiments were performed. As shown in Figure S7, the absorption peaks of the N,P-CDs gradually shifted to a longer wavelength when Fe3+was present, which was attributable to the formation of stable metal complexes because of strong chemical bonding between N,P-CDs and Fe3+. Figure S8 shows that the fluorescence lifetime of the N,P-CDs decreased from 2.95 ns (τ1=2.38, τ2=0.57) to 2.51 ns (τ1=2.22, τ2=0.29) after the addition of Fe3+. The change of fluorescence lifetime indicated a dynamics quenching occurred.8,52 3.4. Detection of Fe3+ using N,P-CDs as probes. In order to obtain the best performance for Fe3+ assay, the effects of pH and reaction time were optimized. As shown in Figure S9, the optimal pH value and reaction time were found to be 7.0 and 35 min, respectively. Fig. 5a showed the fluorescence intensity of N,P-CD at 437 nm decreased gradually with increasing concentrations of Fe3+. The change of fluorescence intensity of the N,P-CDs upon adding different Fe3+ concentrations in the range from 5 nM to 800 nM were shown in Figure S10a, which confirmed Fe3+ sensitivity in N,P-CD fluorescence. Figure S10b indicates that the N,P-CDs’ fluorescent quenching value (∆F=F0−F, F0 and F represent the fluorescence intensities of N,P-CD at 437 nm in the absence and presence of Fe3+, respectively) gradually increased as Fe3+concentrations increased. Moreover, the ∆F value indicated that the linear relationship was good in relation to Fe3+ concentration ranging from 5 nM to 100 nM (Figure 5b), and the equation for linear regression is ∆F=0.1058C+3.0475 with a correlation coefficient of 0.9958, where C is the concentration of Fe3+ (nM). The detection limit was estimated to be 1.8 nM (according to a signal-to-noise ratio of 3), which was much lower than most of those obtained by other fluorescent probes for Fe3+ (Table 1). Inspired by ultra-sensitivity of the proposed chemosensor based N,P-CDs for Fe3+ assay, the selectivity and competition experiments were performed. As illustrated in Figure 6, except for Fe3+ 10


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and Cu2+, no obvious change of the fluorescent quenching values (F/F0) of N,P-CDs was observed upon adding different metal ions including Cd2+, Pb2+, Zn2+, Ag+, Ca2+, Cr3+, Mg2+ and Na+. The result also indicated that Cu2+ strongly influenced the performance of Fe3+ detection, which may be resulted from the greater binding affinity and quicker chelating kinetics between N/O functional groups of N,P-CDs and Cu2+ as well as Fe3+ than other metal ions mentioned above.61,62 Besides, the redox potentials of Fe3+/Fe2+ and Cu2+/Cu+ are in the middle of the conduction and valence bands of N,P-CDs, so photoinduced electrons on N,P-CDs surface can, as a result, transfer to the complexed Fe3+ and Cu2+, leading to fluorescence quenching.6,63 Fortunately, the effect of Cu2+ can be circumvented by using triethanolamine (TEA) as chelating agents. Indeed, the coexistence of Cu2+ with the N,P-CDs–Fe3+ mixture in the presence of TEA does not affect Fe3+ detection (Figure S11). As clearly shown by these results, the proposed chemosensor based on N,P-CDs was highly selective for Fe3+ detection and could meet the selective requirements for bioimaging applications. 3.5. Detection of Fe3+ in human serum samples. Given the efficiency of Fe3+ in N,P-CD fluorescence quenching, the proposed N,P-CD based chemosensor was applied for the detection of Fe3+ in complex biological samples. Human serum samples were spiked with standard solutions with different concentrations of Fe3+ and measured by the proposed method. Table S1 showed that the recoveries were obtained in the range of 88.4–102.4%, and the relative standard deviation (RSD, n=6) was less than 3.2%, which indicated the high analytical precision of the current methods. All observations further approved the reliability and feasibility of developed N,P-CDs based chemosensor for monitoring Fe3+ in biological samples. 3.6. Intracellular imaging of Fe3+. In order to explore the further biological applications, the cytotoxicity of N,P-CDs were evaluated using T24 cells through MTT assays. Figure 7 showed that the cell viability were estimated to be greater than 89% upon addition of N,P-CDs with a high dose 11



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of 270 µg/mL for 24 h. High cell viability indicated the low toxicity and superior biocompatibility of the N,P-CDs, which make a favorable case for sue of N,P-CDs in detecting Fe3+ in living cells. Consequently, confocal fluorescence images were conducted to demonstrate further the usefulness of N,P-CDs in intracellular Fe3+ imaging for T24 cells. As displayed in Figure 8, a intracellular region exhibited a noticeable blue emission (Figure 8a). As expected, the fluorescence brightness of T24 cells becomes weaker as introduction of exogenous Fe3+ into the N,P-CDs-treated T24 cells. Furthermore, no significant changes in cell morphology confirmed the low cytotoxicity of the N,P-CDs. All observations indicated that the proposed chemosensor could be applied for effectively semiquantitative imaging Fe3+ in live cells.

4. CONCLUSION In summary, hydrothermal reaction can be effective in preparing doped CDs with different surface groups. The as-prepared N,P-CDs, which were synthesized through a facile hydrothermal approach, were remarkably water-soluble and excellently stable even under extreme pH, ionic strengths, and light illumination. Moreover, the electronic properties and surface chemical reactivities of N,P-CDs had been substantially improved due to doping of N and P atoms, which was indeed beneficial for chemically binding between N,P-CDs and Fe3+. The N,P-CDs emitting strong blue fluorescence were used as a novel chemosensor for the highly sensitive detection of Fe3+ with detection limit as low as 1.8 nM. The results for monitoring Fe3+ in complex biological samples, and fluorescence images for measuring intracellular Fe3+ indicated that the low cytotoxicity and excellent biocompatibility of N,P-CDs was promising as an efficient platform for clinical diagnosis.

■ ASSOCIATED CONTENT Supporting Information 12


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The Supporting Information is available free of charge on the ACS Publications website at DOI: ××.××××/acsami.×××××××. Figures S1−S11, including FT-IR spectra of the as-prepared N,P-CDs, UV−vis absorption and fluorescent emission spectra of the N,P-CDs, effect of pH on the fluorescence intensity of the N,P-CDs, effect of ionic strengths on the fluorescence intensity of the N,P-CDs, the fluorescence intensity of the N,P-CDs under 365 nm UV light at different time, the fluorescence spectra of the N,P-CDs in presence of Fe3+, the UV-vis spectra of as-prepared N,P-CDs in presence of Fe3+, fluorescence decays of the N,P-CDs in the absence and presence of Fe3+, effect of reaction time and pH on the analysis of Fe3+ using N,P-CDs, fluorescence spectra of N,P-CDs with different concentration of Fe3+, F/F0 values of the chemosensor based the N,P-CDs in the presence of Fe3+, Fe3+ and Cu2+, as well as Fe3+, Cu2+ and TEA. Table S1 (results for the determination of Fe3+ in human serum samples).

■ AUTHOR INFORMATION Corresponding Author *E-mail: [email protected]. *E-mail: [email protected]. Notes The authors declare no competing financial interest.

■ ACKNOWLEDGMENTS Financial support from the Natural Science Foundation of China (Nos. 21405025, 21575031), Natural

Science

Foundations

of

Guangxi

Province

(Nos.

13


2014GXNSFBA118047,


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2015GXNSFDA139006), IRT1225, CMEMR2013-A12, and Guangxi Pharmaceutical Industry Talent Highland Project (1417) is gratefully acknowledged.

■ REFERENCES (1) Hentze, M. W.; Muckenthaler, M. U.; Galy, B.; Camaschella, C. Two to Tango: Regulation of Mammalian Iron Metabolism. Cell 2010, 142, 24−38. (2) Wu, J. S.; Liu, W. M.; Ge, J. C.; Zhang, H. Y.; Wang, P. F. New Sensing Mechanisms for Design of Fluorescent Chemosensors Emerging in Recent Years. Chem. Soc. Rev. 2011, 40, 3483−3495. (3) Eisenstein, R. S. Iron Regulatory Proteins and the Molecular Control of Mammalian Iron Metabolism. Annu. Rev. Nutr. 2000, 20, 627−662. (4) D'Autreaux, B.; Tucker, N. P.; Dixon, R.; Spiro, S. A Non-Haem Iron Centre in the Transcription Factor NorR Senses Nitric Oxide. Nature 2005, 437, 769−772. (5) Rouault, T. A. The Role of Iron Regulatory Proteins in Mammalian Iron Homeostasis and Disease. Nat. Chem. Biol. 2006, 2, 406−414. (6) Zhang, S.; Li, J.; Zeng, M.; Xu, J.; Wang, X.; Hu, W. Polymer Nanodots of Graphitic Carbon Nitride as Effective Fluorescent Probes for the Detection of Fe3+ and Cu2+ Ions. Nanoscale 2014, 6, 4157−4162. (7) Narayanaswamy, N.; Govindaraju, T. Aldazine-based Colorimetric Sensors for Cu2+ and Fe3+. Sensor Actuat B-Chem. 2012, 161, 304−310. (8) Li, S.; Li, Y.; Cao, J.; Zhu, J.; Fan, L.; Li, X. Sulfur-Doped Graphene Quantum Dots as a Novel Fluorescent Probe for Highly Selective and Sensitive Detection of Fe3+. Anal. Chem. 2014, 86, 10201−10207. 14


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Figure Legends

Scheme 1.

Schematic illustrations for the synthesis of N,P-CDs and Fe3+ detection.

Figure 1.

TEM image (a) and the diameter distribution (b) of N,P-CDs.

Figure 2.

XPS survey spectrum (a), C1s XPS spectrum (b), N1s XPS spectrum (c), and P2p XPS spectrum (d) of the as-prepared N,P-CDs.

Figure 3.

Fluorescence emission spectra for N,P-CDs in aqueous solution. Inset: the photographs of N,P-CDs under visible light (left) and a UV beam of 365 nm (right).

Figure 4.

Fluorescence quenching mechanism of Fe3+ on the N,P-CDs.

Figure 5.

(a) Fluorescence spectra of N,P-CDs (10.0 µg mL-1) with different concentration of Fe3+ (from top to bottom: 0, 0.005, 0.01, 0.02, 0.05, 0.08, 0.1, 0.2, 0.8, 10.0, 20.0, 80.0, 100.0, 320.0 and 500.0 µM). (b) The linear calibration plot for ∆F versus different concentration of Fe3+.

Figure 6.

F/F0 values of the N,P-CDs based chemosensor in the absence (black column) and presence (red column) of different metal ions (the concentration of metal ions were 100 µM).

Figure 7. Figure 8.

The viability of T24 cells in different N,P-CD concentrations. Confocal fluorescence images of T24 cells after incubation with N,P-CDs (a), N,P-CDs+500.0 µM Fe3+ in pH 7.4 PBS buffer (b). From left to right: fluorescent image excited with a 405 nm laser and bright field, respectively. The scale bar is 20 µm.

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Scheme 1

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Figure 1

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Figure 2

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Figure 3

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Figure 4

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Figure 5

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Figure 6

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Figure 7

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Figure 8

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Table 1. Comparison of different fluorescent probes for Fe3+ detection Fluorescent probes Aminoantipyrine Graphene oxides nanosheets Pyrazoline derivative Rhodamine B Schiff-base Rhodamine B derivative-functionalized graphene quantum dots Anthracene-appended amino acids 2,5-diphenylfuran and 8-hydroxyquinoline Phosphonic acid-functionalized fluorine derivatives Sulfur-doped graphene quantum dots Nitrogen-doped carbon dots N and S doped carbon dots

Detection limit (µM) 0.211 17.9 1.4 0.11; 1.6

Ref. 53 54 55 56

0.02

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10 0.97 0.02; 0.01 0.0042 0.0025 0.80

57 58 59 8 17 60 This work

Nitrogen and phosphorus co-doped carbon nanodots

0.0018

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TOC Graphic

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Nitrogen and Phosphorus Co-Doped Carbon Nanodots as a Novel

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