Article pubs.acs.org/JPCC
Optical Properties of 2‑Methacryloyloxyethyl PhosphorylcholineProtected Au4 Nanoclusters and Their Fluorescence Sensing of C‑Reactive Protein Junya Yoshimoto,† Arunee Sangsuwan,† Issey Osaka,‡ Kazuko Yamashita,† Yasuhiko Iwasaki,† Mitsuru Inada,§ Ryuichi Arakawa,† and Hideya Kawasaki*,† †
Faculty of Chemistry, Materials and Bioengineering, and §Faculty of Engineering Science, Kansai University, 3-3-35 Yamate-cho, Suita 564-8680, Japan ‡ Center for Nano Materials and Technology, Advanced Institute of Science and Technology, 1-1 Asahidai, Nomi-shi, Ishikawa 923-1292, Japan S Supporting Information *
ABSTRACT: We present the solution synthesis of thiolated 2-methacryloyloxyethyl phosphorylcholine (MPC)-protected Au nanoclusters (NCs). This watersoluble lipid-mimetic MPC was first used for the size focusing synthesis of thiolate (SR)-protected Aun(SR)m NCs. Au25(MPC)18 and Au4(MPC)4 NCs are selectively synthesized, without the need for electrophoretic or chromatographic isolation of size mixed products, by including ethanol or not in the solvent. The Au4(MPC)4 NCs emit at yellow wavelengths (580−600 nm) with a quantum yield (3.6%) and an average lifetime of 1.5 μs. Also for the first time, we report Creactive protein (CRP) sensing using Au NCs, with a detection limit (5 nM) low enough for the clinical diagnosis of inflammation. This is based on the quenching effect of specific CRP−MPC interactions on the fluorescence of the Au4(MPC)4 NCs.
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INTRODUCTION The unique physiochemical properties of thiolate (SR)protected Au nanoclusters (Aun(SR)m NCs) have attracted substantial research interest, leading to applications in optics, catalysis, chemical sensing, and magnetism.1−9 These properties vary with the protecting ligand chosen as well as with the size of the NCs (i.e., the number of Au atoms in the cluster core). Indeed, the electronic and optical properties of the NCs depend on their size and on the atomic number of the constituent element, while the surface ligand species affect the surface chemistry of the clusters, notably their solubility and solution-phase properties.1−9 The solution synthesis of atomically precise Au NCs is therefore desirable in order to optimize their physiochemical properties for particular applications.1−13 Size focusing methods, based on etching Au NCs using excess thiol ligands, allow the one-pot-for-one-size synthesis of atomically precise Aun(SR)m NCs without the need for the electrophoretic or chromatographic isolation of size mixed products.9,14−16 This approach has widely been applied for the synthesis of Aun(SR)m NCs with organo-soluble ligands such as phenylethanethiol.9 The literature is sparse however on the size-focusing synthesis of water-soluble Aun(SR)m NCs,15,17−20 despite their recognized biomedical potential, notably as antimicrobial agents, for cancer therapy, and for bioimaging and biosensing.7,21−23· Crucial for such applications is the avoidance of nonspecific interactions with biomolecules (e.g., proteins) in biological © 2015 American Chemical Society
matrices, which affect the stability of Aun(SR)m NCs in human plasma. However, blood contains many proteins rich in SH and NH groups that readily degrade Au NCs, as reported recently for Au25(SR)18 NCs.24 The present study was therefore motivated by the resistance to protein adsorption, cell attachment, and biofilm formation of 2-methacryloyloxyethyl phosphorylcholine (MPC).25−27 We present the solution synthesis of lipid-mimetic MPC-protected Au NCs, Au25(MPC)18 and Au4 (MPC)4, using the size focusing approach. The Au4(MPC)4 NCs emit in the yellow optical region with a quantum yield (3.6%) and an average lifetime of 1.5 μs. Furthermore, specific interactions between MPC and Creactive protein (CRP) quench the fluorescence of the Au4(MPC)4 NCs, affording sensitive (5 nM detection limit) CRP sensing.
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EXPERIMENTAL SECTION Materials. All the chemicals were used as received without further purification. Tetrachloroauric (III) acid (HAuCl4·3H2O, 99.9% purity), tetraoctylammonium bromide (TOAB, > 98%), sodium borohydride (NaBH4, 99.99%), diisopropylamine (98%), 1,6-hexanedithiol, chloroform (99.0%), ethanol Received: April 24, 2015 Revised: May 31, 2015 Published: June 2, 2015 14319
DOI: 10.1021/acs.jpcc.5b03934 J. Phys. Chem. C 2015, 119, 14319−14325
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Figure 1. (a) UV−vis absorption spectrum and (b) positive-mode electrospray ionization mass spectrum of Au25(MPC)18 nanoclusters aged in water−ethanol for 12 and 10 h, respectively.
(HPLC grade), acetone (>99.5%), human serum albumin (HSA), 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES), and 2-morpholinoethanesulfonic acid monohydrate (MES) were purchased from Wako Chemicals, Japan. MPC was purchased from NOF Co., Japan. Human plasma CRP was purchased from Sigma-Aldrich Co. LLC. Pure water (resistivity 18.2 MΩ·cm) was obtained using a Barnstead NANOpure DIwater system. Human fresh blood was drawn from healthy volunteers and mixed with a 1/9 volume of 3.8% sodium citrate. The citrated whole blood was immediately centrifuged at 2800 rpm for 10 min to obtain platelet-poor plasma (PPP). The experiment with human plasma was approved by the Kansai University ethics committee (No. 010) Synthesis of Thiolated MPC. We synthesized thiolated MPC according to the literature,28 that is, a thiol−ene reaction was used to functionalize one of the mercapto groups in 1,6hexanedithiol with MPC. Specifically, MPC (14.76 g, 50 mmol) and 1,6-hexanedithiol (15.03 g, 100 mmol) were dissolved in a 200 mL round-bottom flask containing 100 mL of degassed chloroform. Di-isopropylamine (278.8 μL, 2.0 mmol) was added, and the mixture was stirred for 22 h at room temperature. After precipitation in acetone, the mixture was placed for 2 h in a vacuum desiccator to eliminate the residual acetone, dissolved in water, and finally lyophilized. Synthesis of Water-Soluble Au NCs. For the Au25(MPC)18 NCs, HAuCl4·3H2O (41 mg, 0.1 mmol), TOAB (63 mg, 0.165 mmol), and ethanol (10 mL) were mixed in a 50 mL three-neck flask. After stirring for 20 min at 300 rpm, MPC (446 mg, 1.0 mmol) in 5.0 mL of water (Au/ MPC = 1:10) was injected into the solution using a syringe. After stirring for 60 min at 300 rpm, NaBH4 (38 mg, 1.0 mmol, Au/NaBH4 = 1:10) was freshly dissolved in cold water (5 mL) and then rapidly poured into the reaction solution under vigorous stirring, which was maintained at 200 rpm for overnight (10−12 h). The reaction mixture was centrifuged for 5 min at 6000 rpm to remove insoluble side products, and the supernatant was dried in a vacuum rotary evaporator at room temperature. The dried, black, oil-like clusters were precipitated by adding acetone (∼15 mL), washed with fresh acetone, and then collected by centrifugation. The precipitate was washed three times with a mixture of acetone and ethanol (9:1 v/v), and then a different acetone−ethanol mixture (6:4 v/
v) was added to extract pure Au25(MPC)18 NCs from the precipitate, which were then dried in vacuum. For the Au4(MPC)4, NCs, HAuCl4·3H2O (41 mg, 0.1 mmol) and water (10 mL) were mixed in a 50 mL three-neck flask. After stirring for 20 min at 300 rpm, MPC (446 mg, 1.0 mmol) in 5.0 mL of water (Au/MPC = 1:10) was injected into the solution using a syringe. After stirring for 60 min at 300 rpm, NaBH4 (38 mg, 1.0 mmol, Au/NaBH4 = 1:10) was freshly dissolved in cold water (5 mL) and then rapidly poured into the reaction solution under vigorous stirring, which was maintained at 200 rpm for 1 week. The byproducts were then removed by centrifugation (10000 rpm, 10 min) and the supernatant was dialyzed through a membrane with a molecular weight cutoff of 3.5 kDa. Finally, the Au4(MPC)4 NCs were precipitated by adding acetone to the solution, and dried in vacuum. Fluorescence Sensing of CRP. Au4(MPC)4 NCs were dissolved in HEPES (0.1 M, pH 7.4), and the concentration was adjusted to 20 mg/mL. The Au4(MPC)4 NC solution (10 μL) was mixed with 250 μL of 2 mM CaCl2 in HEPES (pH 7.4), and 230 μL of 0.1 M MES buffer (pH 5.5) in a plastic tube. Appropriate amounts of 2800 nM CRP/HEPES buffer were then added to the tube, and the resulting mixtures were submitted to fluorescence analyses. Characterization. Ultraviolet−visible (absorption) and fluorescence (excitation and emission) spectra were recorded using JASCO V-670 and FP-6300 instruments, respectively. The absolute quantum yield of the fluorescence in the Au NC solution was measured using a Hitachi F-7000 spectrophotometer with an integrating sphere and a GG395 cutoff filter. The time-dependent photoluminescence of the Au NC solution was measured using an intensified charge coupled device (iCCD) camera. The excitation source was a 266 nm Nd:YAG laser (Ultra, Quantel, France) delivering 8 ns pulses at 20 Hz. A monochromator, (Spectrograph 300i, Acton, Massachusetts, USA) was attached to the iCCD camera, (PIMAX, Princeton, USA). X-ray photoelectron spectroscopy (XPS) measurements were performed on a Quantera SXM device (Physical Electronics, Inc., USA) using monochromatic Al Kα radiation (1486.7 eV). The base pressure was approximately 2 × 10−8 Torr, and the binding energies were referenced to the hydrocarbon C 1s signal at 284.7 eV to compensate for charging effects. 14320
DOI: 10.1021/acs.jpcc.5b03934 J. Phys. Chem. C 2015, 119, 14319−14325
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Figure 2. (a) UV−visible spectrum and (b) positive-mode electrospray ionization mass spectrum of as-synthesized Au4(MPC)4 nanoclusters aged in water for 10 h.
Figure 3. High-resolution positive mode MALDI FT-ICR mass spectrum of Au4(MPC)4 NCs aged in water for 10 h.
Figure 4. (a) Thermogravimetric trace of the Au4(MPC)4 nanoclusters recorded under nitrogen flow after 10 h reaction. (b) Au 4f X-ray photoelectron spectrum of Au4(MPC)4 nanoclusters.
H2O/MeOH; sample flow rate, 3−5 μL/min; capillary temperature, 200 °C; spray voltage, 3.9 kV. Matrix-assisted laser desorption/ionization Fourier transform ion cyclotron resonance mass spectrometry (MALDI FT-ICR MS) was carried out on a SolariX 9.4T device (Bruker Daltonics Inc., Billerica, MA). Positive ions were generated from a MALDI source. Typical MALDI conditions were used, namely, time domain, 1 M; acquisitions, 3; mass-to-charge (m/ z) scan range, 1000−4000; time-of-flight, 1.5 ms; plate offset,
Thermogravimetric analysis (TGA) was carried out using a Thermo plus EVO device (Rigaku, Japan) at a heating rate of 10 °C/min under nitrogen flow. Nanoelectrospray ionization mass spectrometry (ESI-MS) was conducted in positive mode on MPC−Au NC solutions (∼1 mg/mL) using an Exactive Plus Orbitrap instrument (Thermo Scientific). A dynamic nanospray probe attachment was used and the spray tip was made from a fused silica capillary. The following settings were used: solvent, 4:1 (v/v) 14321
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Figure 5. (a) Fluorescence spectra following excitation at 254 nm (orange line) and 350 nm (red line, intensity ×10), of Au4(MPC)4 nanoclusters. Photographs of the corresponding Au4(MPC)4-nanocluster aqueous solutions are inset. (b) Luminescence decay profile of the Au4(MPC)4 nanoclusters fit to a lifetime of 1.5 μs.
that the prominent peak at m/z = 2586.1 should be assigned to [Au25(MPC)18 + 5H]5+, confirming the molecular formula of the NCs as Au25(MPC)18. With pure water used instead of the water−ethanol mixture, in contrast with Figure 1a, the UV−visible spectrum of the prepared product shows an absorption peak at 370 nm with a shoulder at 335 nm (Figure 2a). This peak appears after 7 h size focusing and increases in intensity thereafter (Figure S2a in the SI). The wavelength of this absorption (370 nm) is close to those reported in the literature for small Au8−15 and Au5 NCs (370−420 and 330 nm, respectively).29,30 The most prominent peak in the ESI-MS data obtained for these NCs (Figure 2b) appears at m/z = 1283.3. The 0.5 Da spacing of the isotope peaks indicates that the ions carry a positive charge of 2, and good agreement between simulated and experimental isotope patterns is achieved with the elemental formula [Au4(MPC)4 + 2H]2+ (Figure S3 in the SI). This assignment is confirmed by the high-resolution MALDI FT-ICR MS data in Figure 3, in which the intense peak at m/z = 2565.53057 corresponds within 0.001 Da to the value expected for [Au4(MPC)4 + H]+. The other peaks observed in the MALDI mass spectrum (see Figure 3 for assignments) stem from the harder ionization process used (compared with ESI). No peaks assignable to larger Au NCs (e.g., Au25(MPC)18) are visible. The molecular formula of these NCs is also confirmed by thermogravimetric analysis (Figure 4a). Indeed, the 70.3% weight loss measured for the two-step thermal decomposition of the MPC ligands is approximately close to the value (69.3%) expected for the decomposition of Au4(MPC)4 NCs. We also conducted TGA analysis of the MPC ligand itself, in which the one-step weight loss of MPC ligand at less than 500 °C was observed (Figure S4 in the SI). The second weight loss of Au4(MPC)4 at more than 500 °C is attributed to the enhanced thermal stability of the ligand via the sulfur−gold bonding of Au4(MPC)4 NCs. The first weight loss step is likely related to the elimination of the phosphorylcholine group, −CO2C2H4PO4C2H4N(CH3)3, since such elimination of the phosphorylcholine group was also observed in MALDI mass spectrum of Au4(MPC)4 (Figure 3). It has been reported that stepwise fragmentation of the semiring staple motifs on the surface of Au25(SR)18 NCs can lead to the detection of smaller units, including Au21(SR)14 and cyclic [Au(SR)]4 in mass spectra.31,32 However, it is unlikely
100 V; deflector plate voltage, 200 V; laser power, 40%; laser shots, 500; frequency, 200 Hz; laser diameter, small; front and back trap-plate voltages, 0.4 and 0.5 V, respectively; analyzer entrance, −2.0 V; side kick, 0.0 V; side kick offset, −0.5 V; sweep excitation power, 15.0%.
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RESULTS AND DISCUSSION Synthesis and Characterization. With size-focusing methods, the choice of solvent governs the size of the resulting
Figure 6. UV−visible absorption at different times of Au4(MPC)4 NCs in human blood plasma.
NCs; in this case, a mixed solvent (water−ethanol at 1:1 v/v) or pure water led to the synthesis of Au25(MPC)18 or Au4(MPC)4 NCs, respectively. For the Au25(MPC)18 NCs, size focusing is evidenced by the emergence during synthesis of a characteristic Au25(SR)18 absorption and photoluminescence peak after 5 h (Figure S1 in the Supporting Information (SI)). Four distinct absorption peaks at 400, 450, 550, 670, and 780 nm are observed in the UV−vis spectrum of the product (Figure 1a) after 12 h reaction in the mixed solvent (water−ethanol at 1:1 v/v), which are consistent with the characteristic peaks of Au25(SR)18 NCs. The well-defined absorption spectrum suggests that the purity of the Au25(MPC)18 NCs in the raw product is already high, obviating the need for electrophoretic or chromatographic isolation. The 0.2 Da spacing of the isotope peaks in the ESI-MS data (Figure 1b) indicates that the ions carry a positive charge of five, and 14322
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Figure 7. (a) Fluorescence response to 330 nm excitation of Au4(MPC)4 nanoclusters in the presence of different concentrations of C-reactive protein (CRP). (b) Quenching effects of CRP (red closed circles) and human serum albumin (HSA, blue closed circles) on Au4(MPC)4 nanoclusters. (F and F0 are respectively the fluorescences measured in the presence and absence of the proteins.).
Human Blood Plasma Stability. The time-dependent UV−visible experiments on glutathione(GSH)- and bovine serum albumin(BSA)-protected Au NCs in human blood plasma of Zhang et al. show surface plasmon resonance (SPR) absorption within 1 h, from the large Au nanoparticles with 5− 30 nm via the NC agglomeration.24 However, the excellent biocompatibility of MPC, in particular its very weak interaction with blood plasma, motivated similar experiments on the Au4(MPC)4 NCs. Indeed, no signs of agglomeration (viz. SPR absorption) are observed in the UV−vis spectra recorded for Au4(MPC)4 NCs (Figure 6). Even after 72 h, most of Au4(MPC)4 NCs are still present in human plasma, as evidenced by the persistence of the peak at 370 nm, although its decreasing intensity suggests that the NCs degrade gradually (Figure 6). This nonaggregation is the effect of the biocompatible MPC ligands on the surface of the Au NCs, which prevent undesired protein interactions in human plasma.25−27 Fluorescence Sensing of CRP. C-reactive protein is a prototypic acute-phase protein in humans and an important biomarker,36 with serum levels increasing by up to 1000-fold within 24 h of severe inflammation. Each of CRP’s covalently linked subunits has a calcium-dependent binding site for phosphorylcholine (PC),37 allowing the concentration and activation dynamics of human CRP to be studied via PCfunctionalized cell-membrane-mimetic interfaces on plasmonic sensors. The PC side-chain of MPC can play the same role; recently indeed, label-free detection of CRP was demonstrated using MPC-protected Au nanoparticles on the basis of the change in color of Au nanoparticle solutions upon (CRPinduced) aggregation.28 Here, we exploit the fluorescence quenching of Au4(MPC)4 NCs in the presence of CRP/Ca2+ to the same end. Figure 7 shows fluorescence spectra of Au4(MPC)4 NC solutions obtained 60 min after adding different concentrations of CRP. The peak intensity (at 585 nm) decreases with increasing CRP concentrations; the fluorescence quenching (F0/F) is linearly proportional to the CRP concentration in the 5−50 nM range (Figure 7b inset, F/ F0 = 1.013 + 0.00538 × [CRP], R2 = 0.992), where F and F0 are the fluorescence intensities in the presence and absence of CRP, respectively. The method is highly sensitive, with a detection limit of 5 nM (∼6 mg/L), lower than the high-risk threshold for cardiovascular diseases (>10 mg/L CRP).38 The quenching mechanism is not clear at present, but is presumably related to the specific binding to CRP of the PC group of the
that these Au4(MPC)4 NCs are fragmentation products of the ionization process, since they are detected both by ESI- and MALDI-MS. Furthermore, the UV−visible spectra of the Au4(MPC)4 and Au25(MPC)18 NCs differ substantially (Figure 2a vs Figure 1a). An explanation consistent with the experimental data is that the Au4(MPC)4 NCs are actually present in aqueous solution and arise from the etching of larger MPC-protected NCs in the presence of excess free MPC. The stable complexes for Au(I) have been reported to be tetranuclear triphosphines.33,34 The Au valence state in Au4(MPC)4 NCs was investigated by XPS. The XPS extended spectrum showed the presence of carbon (C), oxygen (O), nitrogen (N), gold (Au), sulfur (S), and phosphorus (P), which contain the elements of MPC ligands of Au4(MPC)4 NCs (Figure S5 in the SI). The Au 4f peak appears at 84.4 eV, which is close to the peak of Au(I) at 84.7 eV (Figure 4b).35 This suggests that the oxidation state of Au in Au4(MPC)4 is Au(I). Fluorescence Properties. In aqueous media, the Au25(MPC)18 NCs produce fluorescence in the near-infrared (NIR) region with less than 0.3% quantum yield (Figure S1b in the SI), which is consistent with previous reports on the fluorescence of Au25(SR)18 NCs.10 For the Au4(MPC)4 NCs, however, strong yellow (585 nm) fluorescence is evidenced in Figure 5a along with weaker red/NIR emission (>650 nm). Photoluminescence at 585 nm leads to excitation maxima at 262 and 327 nm (Figure S6 in the SI). The emission maximum appears at the same wavelength following excitation at 280− 405 nm (Figure S7 in the SI), indicating real fluorescence from a single type of Au NC. However, the relative intensities of the yellow and red−NIR emissions depend on the excitation wavelength, the former dominating following 254 nm excitation (see the orange fluorescence line and the corresponding photograph in Figure 5a), while dual emission is observed for 350 nm excitation (red line and corresponding photograph in Figure 5a). The quantum yield measured for the yellow emission (585 nm) following 330 nm excitation is 3.6%, indicating that the Au4(MPC)4 NCs are highly luminescent. Furthermore, the large Stokes shift and the 1.5 μs lifetime of the corresponding luminescence decay profile (Figure 5b) are consistent with the presence of Au(I) in the Au4(MPC)4 NCs. This emission is therefore attributed to ligand-to-metal charge transfer from the sulfur atoms in the thiolates to the Au atoms (predominantly in the +1 oxidation state) and subsequent radiative relaxation.35 14323
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(4) Lu, Y.; Chen, W. Sub-nanometre sized metal clusters: From synthetic challenges to the unique property discoveries. Chem. Soc. Rev. 2012, 41, 3594−3623. (5) Maity, P.; Xie, S.; Yamauchi, M.; Tsukuda, T. Stabilized gold clusters: From isolation toward controlled synthesis. Nanoscale 2012, 4, 4027−4037. (6) Udayabhaskararao, T.; Pradeep, T. New protocols for the synthesis of stable Ag and Au nanocluster molecules. J. Phys. Chem. Lett. 2013, 4, 1553−1564. (7) Luo, Z.; Zheng, K.; Xie, J. Engineering ultrasmall water-soluble gold and silver nanoclusters for biomedical applications. Chem. Commun. 2014, 50, 5143−5155. (8) Kurashige, W.; Niihori, Y.; Sharma, S.; Negishi, Y. Recent progress in the functionalization methods of thiolate-protected gold clusters. J. Phys. Chem. Lett. 2014, 5, 4134−4142. (9) Jin, R. Atomically precise metal nanoclusters: Stable sizes and optical properties. Nanoscale 2015, 7, 1549−1565. (10) Negishi, Y.; Nobusada, K.; Tsukuda, T. Glutathione-protected gold clusters revisited: Bridging the gap between gold(I)-thiolate complexes and thiolate-protected gold nanocrystals. J. Am. Chem. Soc. 2005, 127, 5261−5270. (11) Jadzinsky, P. D.; Calero, G.; Ackerson, C. J.; Bushnell, D. A.; Kornberg, R. D. Structure of a thiol monolayer-protected gold nanoparticle at 1.1 Å resolution. Science 2007, 318, 430−433. (12) Heaven, M. W.; Dass, A.; White, P. S.; Holt, K. M.; Murray, R. W. Crystal structure of the gold nanoparticle [N(C8H17)4][Au25(SCH2CH2Ph)18]. J. Am. Chem. Soc. 2008, 130, 3754−3755. (13) Zhu, M.; Aikens, C. M.; Hollander, F. J.; Schatz, G. C.; Jin, R. Correlating the crystal structure of a thiol-protected Au25 cluster and optical properties. J. Am. Chem. Soc. 2008, 130, 5883−5885. (14) Zhu, M.; Lanni, E.; Garg, N.; Bier, M. E.; Jin, R. Kinetically controlled, high-yield synthesis of Au25 clusters. J. Am. Chem. Soc. 2008, 130, 1138−1139. (15) Shichibu, Y.; Negishi, Y.; Tsunoyama, H.; Kanehara, M.; Teranishi, T.; Tsukuda, T. Extremely high stability of glutathionateprotected Au25 clusters against core etching. Small 2007, 3, 835−839. (16) Jin, R.; Qian, H.; Wu, Z.; Zhu, Y.; Zhu, M.; Mohanty, A.; Garg, N. Size focusing: A methodology for synthesizing atomically precise gold nanoclusters. J. Phys. Chem. Lett. 2010, 1, 2903−2910. (17) Qian, H.; Zhu, M.; Lanni, E.; Zhu, Y.; Bier, M. E.; Jin, R. Conversion of polydisperse Au nanoparticles into monodisperse Au25 nanorods and nanospheres. J. Phys. Chem. C 2009, 113, 17599−17603. (18) Wu, Z.; Suhan, J.; Jin, R. One-pot synthesis of atomically monodisperse, thiol-functionalized Au25 nanoclusters. J. Mater. Chem. 2009, 19, 622−626. (19) Kumar, S.; Jin, R. Water-soluble Au25(capt)18 nanoclusters: Synthesis, thermal stability, and optical properties. Nanoscale 2012, 4, 4222−4227. (20) Yu, Y.; Luo, Z.; Yu, Y.; Lee, J. Y.; Xie, J. Observation of cluster size growth in CO-directed synthesis of Au25(SR)18 nanoclusters. ACS Nano 2012, 6, 7920−7927. (21) Sun, J.; Jin, Y. Fluorescent gold nanoclusters: Recent progress and sensing applications. J. Mater. Chem. C 2014, 2, 8000−8011. (22) Kawasaki, H.; Kumar, S.; Li, G.; Zeng, C.; Kauffman, D.; Yoshimoto, J.; Iwasaki, Y.; Jin, R. Generation of singlet oxygen by photoexcited Au25(SR)18 clusters. Chem. Mater. 2014, 26, 2777−2788. (23) Zhang, X. D.; Luo, Z.; Chen, J.; Song, S.; Yuan, X.; Shen, X.; Wang, W.; Sun, Y.; Gao, K.; Zhang, L.; et al. Ultrasmall glutathioneprotected gold nanoclusters as next generation radiotherapy sensitizers with high tumor uptake and high renal clearance. Sci. Rep. 2015, 5, 8669. (24) Zhang, X. D.; Wu, D.; Shen, X.; Liu, P. X.; Fan, F. U.; Fan, D. J. In vivo renal clearance, biodistribution, toxicity of gold nanoclusters. Biomaterials 2012, 33, 4628−4638. (25) Ishihar, K.; Ziats, N. P.; Tierney, B. P.; Nakabayashi, N.; Anderson, J. M. Protein adsorption from human plasma is reduced on phospholipid polymers. J. Biomed. Mater. Res., Part A 1991, 25, 1397− 1407.
Au4(MPC)4 NCs. For the sake of comparison, we also recorded the fluorescence spectra of Au4(MPC)4 NCs for 60 min after adding different concentrations of HSA, the most abundant protein in human blood plasma (Figure S8 in the SI). In this case, only limited fluorescence quenching is observed because the interactions between Au4(MPC)4 and HSA are weak. Figure 7b emphasizes the much greater quenching effect of CRP.
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CONCLUSION This paper reports the first successful use of water-soluble, lipid-mimetic MPC ligands for the size focusing synthesis of Aun(SR)m NCs. Au4(MPC)4 or Au25(MPC)18 NCs are readily obtained by using respectively water or water−ethanol as solvent. The Au4(MPC)4 fluoresce strongly at 585 nm in aqueous media, with a quantum yield (3.6%) and an average lifetime of 1.5 μs. The fluorescence of the Au4(MPC)4 is however quenched in the presence of CRP, via specific interactions between the latter and the MPC ligand. A CRP sensing method is proposed on this basis, whose detection limit (5 nM) is below the risk threshold for cardiovascular diseases. This is the first time CRP sensing was performed using Au clusters.
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ASSOCIATED CONTENT
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AUTHOR INFORMATION
S Supporting Information *
Time evolution of the UV−visible and the fluorescence spectra of crude MPC−Au NCs, experimental and simulated electrospray ionization mass spectrum of Au 4 (MPC) 4 NCs, thermogravimetric trace of the MPC ligand itself, XPS spectra of Au 4 (MPC) 4 NCs, photoluminescence spectrum of Au4(MPC)4, and fluorescence response of Au4(MPC)4 NCs in the presence of different concentrations of human serum albumin. The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/ acs.jpcc.5b03934. Corresponding Author
*E-mail:
[email protected]. Notes
The authors declare no competing financial interest.
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ACKNOWLEDGMENTS We thank Prof. Umeda and Prof. Y. Nishiyama at Kansai University for the measurements of the quantum yields of the MPC-Au NCs. This work was supported by JSPS KAKENHI Grant No. (15H03520, 15H03526, and 26107719), and also by Hitachi Metals Materials Science Foundation 2014 and a MEXT-supported Strategic Research Foundation Grant-aided Project for Private Universities, 2010−2014. The work of MS measurements was partly supported by Nanotechnology Platform Program of the Ministry of Education, Culture, Sports, Science and Technology (MEXT), Japan.
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REFERENCES
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DOI: 10.1021/acs.jpcc.5b03934 J. Phys. Chem. C 2015, 119, 14319−14325
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DOI: 10.1021/acs.jpcc.5b03934 J. Phys. Chem. C 2015, 119, 14319−14325
