Oxidation at the Core–Ligand Interface of Au Lipoic Acid Nanoclusters

Aug 14, 2014 - Multivalent binding of the lipoic acid ligands on Au core drastically reduces the ... core charge states, ligand polarity, and ligand m...
0 downloads 0 Views 2MB Size
Download PDF
Article pubs.acs.org/JPCC

Oxidation at the Core−Ligand Interface of Au Lipoic Acid Nanoclusters That Enhances the Near-IR Luminescence Jie Jiang, Cecil V. Conroy, Maksim M. Kvetny, Gabriel J. Lake, Jonathan W. Padelford, Tarushee Ahuja, and Gangli Wang* Department of Chemistry, Georgia State University, Atlanta, Georgia 30302, United States S Supporting Information *

ABSTRACT: A new surface oxidation mechanism that enhances the luminescence of Au nanoclusters is discovered in Au nanoclusters synthesized with disulfide lipoic acid. The quantum efficiency increased from 1 to 2% up to 10% upon the sulfur oxidation at the Au−ligand interface. Relatively low quantum efficiency has been a bottleneck barrier to exploit the appealing near-infrared luminescence from molecular-like gold nanoclusters for broader and effective applications. Combined IR, XPS, and NMR analysis reveals that the outer sulfur atoms on ca. half of the lipoic acid ligands were partially oxidized accompanying the luminescence enhancement. Opposite to those from the widely studied monothiolate Au nanoclusters, the quantum efficiency increases at lower pH. The observation is explained by the electron density changes at the core−ligand interfaces under the ligand dielectric layer. Multivalent binding of the lipoic acid ligands on Au core drastically reduces the exchange or addition of monothiols into ligand monolayer. The improved resistance to excess thiols is significant because the stability of Au nanoclusters is a critical concern in their applications in thiol-rich physiological environment. The fundamental materials and chemistry insights suggest promising routes to further enhance the near-IR luminescence and chemical stability that are critical factors in biomedical and sensing applications.

■

INTRODUCTION Molecular-like metal nanoclusters (NCs) are fundamentally interesting in that they can be characterized by molecular compositions not just by the size and shape. Analogous to the broad interest in various nanomaterials that are at the transition regime of small molecules and bulk materials, these metal NCs bridge the transition from small molecules to larger nanomaterials. The revolutionary progress in the recent structural determination of Au1−3 and Ag4,5 NCs lay the foundation for possible correlations with their unique electrochemical, optical, magnetic, catalytic, and other physicochemical properties.6−9 Understanding of the photoluminescence from Au NCs stabilized by various ligands remains elusive.8,10 Several features make the luminescence from these Au NCs technically significant: near-infrared emission with drastically reduced interferences, nontoxic components, less photobleachable and photoblinking, large Stokes shift, and long lifetime for lifetime imaging. Because of the highly advantageous near-IR spectrum range and the high molar absorptivity of Au materials by nature, research efforts have been focused on improving the relatively low quantum efficiency (QE) which is the key limit for their effective applications, i.e., in bioimaging and sensing. Unlike the energetics elucidated by UV−vis absorption and electrochemistry in which experiments and theories have found decent agreements,9 improvements in the emission properties, especially the QE, are primarily achieved via empirical approaches.11−14 The energy states corresponding to the © 2014 American Chemical Society

near-IR emission can be attributed to hybridized atomic orbitals of Au and S and thus referred to as surface states. Note the emission is less dependent on the overall energetics of the Au NCs, which is known to vary depending on the number of Au atoms in the core, the number of ligands, and their binding structures.15−17 While no predictive theory is available for further enhancement, it has been reported that core size, core charge states, ligand polarity, and ligand molecular structures are the intrinsic parameters of Au NCs affecting their emission QE.11−17 Albeit a comparison of the definitive QE values is complicated by different standards employed (the large Stokes shift makes it a challenge for the correction of both excitation and emission),14,18−21 the QE reported so far are sufficiently strong to be competitive for real applications, which make the aforementioned advantages from various Au NCs relevant for applications and superior over the counterparts such as small molecule dyes or semiconductor quantum dots. Redox activities and relevant optical transitions of Au-thiolate NCs have been primarily discussed in nonaqueous solvents.6,22 From the widely studied Au25PET18 NCs (PET as 2phenylethanethiolate), the photoluminescence was shown to intensify upon the chemical oxidation of the Au cores.11 Electrogenerated chemiluminescence was also observed and Received: June 17, 2014 Revised: August 8, 2014 Published: August 14, 2014 20680

dx.doi.org/10.1021/jp5060217 | J. Phys. Chem. C 2014, 118, 20680−20687

The Journal of Physical Chemistry C

Article

Scheme 1. Oxidation of the Lipoic Acid Ligands on Au22 NCs

Figure 1. Transitions of (a) absorbance and (b) luminescence during room temperature synthesis. The spectrum features at each step, particularly the changes of the band gap (absorbance approaching zero), were enlarged in the inset in panel a. The emission intensity at 740 nm was plotted versus time in the panel b inset.

(NaBH4, ≥99%), sodium hydroxide (NaOH, ≥97%), deuterium oxide (D2O, ≥99.9%), and 3,3′-diethylthiatricarbocyanine iodide (DTTC, ≥99%) were purchased from Sigma-Aldrich and used as received. Potassium phosphate monobasic (KH 2 PO 4 , ≥99.5%) and potassium phosphate dibasic (K2HPO4, ≥99.0%) were purchased from Fluka. Phosphate buffer solution (10 mM) was prepared by mixing different volumes of K2HPO4 (10 mM) and KH2PO4 (10 mM) solution. In all preparations, nanopure water (>18 MΩ·cm) from a Barnstead system was used. Measurements. UV−vis absorbance spectra (UV−vis) were recorded with a Shimadzu UV-1700 spectrophotometer. Fluorescence spectra were measured with a Horiba Jobin-Yvon Fluorolog 311 spectrometer equipped with PMT visible and InGaAs near IR detectors. A Hoya Y-44 440 nm-long-pass filter was used to collect emission spectra. QE was calculated from the combined detector responses with the corresponding absorbance values around 0.05 at the wavelength of excitation (400 nm). DTTC in methanol was used as standard (QE = 21%).31 MALDI mass spectra were collected with an ABI 4800 TOF-TOF analyzer with R-cyano-4-hydroxycinnamic acid (CHCA) as matrix. Infrared spectra were acquired on a PerkinElmer Spectrum 100 FT-IR spectrometer. NMR spectra (1D, 2D COSY, and HSQC) were collected with a Bruker NMR 400 MHz spectrometer. XPS analysis was conducted with a SSX-100 X-ray photoelectron spectrometer using an Al KR X-ray source (1486.6 eV). All XPS spectra were calibrated with respect to the C 1s peak at 284.7 eV. The XPS spectra were fitted with symmetric Gauss−Lorentz sum function (Gaussian−Lorentzian ratio at 0.8) after Shirley background subtraction. Synthesis of Au Nanoclusters Stabilized by Lipoic Acid at Room and Elevated Temperatures. The synthesis followed reported procedure with some modifications.13 Lipoic

interpreted based on the voltammetric and optical properties.23 In aqueous environment, the large dielectric constant of water limits the resolution of charging current and thus excludes many interesting voltammetric features such as the quantized double-layer charging. Accordingly, the redox activities in aqueous system, beyond surface self-assembly type Au− monothiolate interactions, are rarely explored. Our group has been interested in the multidentate dithiol/dithiolate core− ligand binding.24−26 The multivalent binding from individual ligands is entropy favored. Accordingly, the chemical stability of ligand monolayer should be improved by the introduction of dithiolates over monothiolates.27,28 This feature is believed significant for their applications in thiol-rich physiological environments. Furthermore, the structural constraint imposed on possible formation of the thiol-bridging motif (RS−Au−SR) will affect the optical and electrochemical properties as reported previously.24−26 Lipoic acid (LA) was employed in this report to create highly luminescent Au−LA NCs. Illustrated in Scheme 1, binding to Au will open the five-atom disulfide ring and form 1,3 dithiolate−Au interactions, which align with our earlier studies of 1,2 and 1,4 dithiol/dithiolate Au NCs.26,29 Our interest in lipoic acid also resides in its widespread medical and nutritional functions involving its redox activities in thiol−disulfide conversions.30 Surprisingly, the QE was found to enhance up to 10-fold, or to 10% through the purification process, accompanied by partial oxidation of sulfur. The QE increases at lower pH. The trend is opposite compared with those in monothiolate Au NCs, which is successfully explained by the elucidated surface oxidation mechanism.

■

EXPERIMENTAL SECTION Chemicals. Tetrachloroauric acid trihydrate (HAuCl4· 3H2O, >99.99%), lipoic acid (LA, ≥99%), sodium borohydride 20681

dx.doi.org/10.1021/jp5060217 | J. Phys. Chem. C 2014, 118, 20680−20687

The Journal of Physical Chemistry C

Article

acid (0.15 mmol) was first dissolved in nanopure water at slightly basic pH adjusted with NaOH for better solubility. A HAuCl4 solution was then added to reach a final Au:LA mole ratio of 1:3 under vigorous stirring. The final solution pH was adjusted to 11.0. The Au(III)−disulfide reaction was stirred over ca. 4 h. After the completion of absorbance transitions, NaBH4 (2 equiv) was added at either room temperature or heated at 65 °C. The reaction proceeded over a period of time determined by changes in luminescence. The crude products were filtered to remove any insoluble precipitates, concentrated by removal of solvent via evaporator, and followed by two different purification methods. In the first route, the synthesized NCs were subject to dialysis using regenerated cellulose dialysis tube in nanopure water, which was recharged every few hours over a period of 3−4 days. In the other route, the synthesized NCs were dissolved in 2 mL of fresh nanopure water and thrice ultracentrifuged at 15 000 rpm for 30 min. Similar molecular weight cutoff membranes (3.0K for dialysis and 3.5K for ultracentrifuge) were employed in the two purification routes. Further, the NCs from the same synthetic batch were split into two portions, each purified with one route to compare the two purification methods. The final products were collected after the removal of solvent under vacuum at room temperature.

Figure 2. Impacts of purification routes on the optical features. The absorption (left) was normalized at 250 nm to 1. The luminescence intensity (right) was normalized by the absorbance at the excitation wavelength of 400 nm. The absorbance value at 400 nm was ca. 0.05 for each sample to ensure negligible self-absorption and quenching. The break on wavelength axis separates the responses from two different detectors.

the luminescence QE at ca. 0.6% was much lower than the 10% QE from our results explained in the following sections. A series of lipoic acid derivatives have been employed to synthesize near-IR luminescent Au NCs with no composition elucidated.20,21 Well-defined AuXSY patterns were observed under negative mode by MALDI-MS analysis shown in Figure SI-2. At low m/ z range, a strong peak at m/z 1831.2 matches the Au8S8− fragment (also confirmed by the theoretical isotopic patterns). At higher m/z range, the peak at 5307.8 m/z matches Au25S12−, with a series of peaks corresponding to loss and gain of Au2S/ AuS units distributed on either side. With the increase in laser intensity, the Au8S8− intensity decreases while the m/z peaks around Au25S12− increase. The transition suggests that albeit all are fragments, the Au8S8− better correlates with the original Au NCs while those around 5K m/z reflect subsequent fragmentations and recombinations. Similar AuXSY patterns were also detected from purified Au−LA NCs further heated for 8 h in aqueous solution at 65 °C. The results from different synthetic routes are consistent and suggest the favorable compositions, which reveal good thermostability of these Au− LA NCs. Although molecular ions have not been captured due to the fragmentation and recombination that are known problematic for MALDI analysis, the observed AuXSY patterns are insightful in the elucidation of a possible composition. Unlike the well-known Au25 NCs that have a Au13 core,2,3,34 Au8 structure have been reported as the inner core of Au20, Au22, and Au24 NCs.32,35−39 In reference to the absorption features, and the general trend that optical band gap increases as the nanocluster size decreases, we propose the Au−LA NCs being Au22 instead of the well-studied Au25 NCs that have a 1.33 eV optical band gap.40 We also propose there are 12 lipoic acid ligands per nanocluster based on the results discussed next. Drastically Enhanced Luminescence QE through Purification. A unique feature was discovered in the Au−LA NCs system not observed previously: the QE of the Au NCs was found to differ drastically depending on the purification process. Shown in Figure 2, from the same synthetic batch of Au−LA NCs that had an initial QE at ca. 1% prior to purification (w/o purification), the luminescence QE was found to increase drastically through multiday dialysis, up to 10%. In

■

RESULTS AND DISCUSSION The absorbance and luminescence features at key synthetic steps are summarized in Figure 1. Lipoic acid itself displays an absorption band at ca. 330 nm. After the addition of HAuCl4, this band diminished gradually and stabilized after ca. 4 h, monitored by the absorbance at the 300−400 nm range. The absorption transition corresponds to the Au(III)−disulfide reaction that disrupts the five-atom disulfide ring, accompanied by solution color changes from light yellow to colorless. Upon further reduction by NaBH4, the solution immediately turned dark orange. The optical band gap extended from 3.1 to 1.9 eV (400 to 630 nm, Figure 1a inset). This is characteristic for the conversion of small Au−SR complexes (also referred as Au− thiolate oligo-polymers) to Au NCs. A weak absorption band at ca. 505 nm could be resolved from the purified samples (inset of Figure 1a and Figure 2). Prior to reduction by NaBH4, the photoluminescence in the near-IR range was negligible. The emission intensified gradually during the reduction process. It took ca. 16 h to reach maximum emission intensity at room temperature followed by a decrease at i.e. 20 h, while about 3 h to peak when heated at 65 °C as shown in Figure SI-1. At higher temperatures (i.e., 80 °C) during synthesis, the Au−LA NCs appeared to be unstable and the emission intensity was lower. The controlled heating apparently facilitated the luminescence increase during the synthesis and induced slightly higher QE prior to purification. The 505 nm absorption band and absorption band gap (1.9 eV) are in agreement with the Au22 NCs stabilized by either glutathione32,33 or lipoic acid13 in recent reports. There are also differences in absorption and/or luminescence properties compared to other Au22 NCs. It is known that NCs with the same Au core size but different ligand compositions could display different optical spectra. For example, glutathionestabilized Au22 NCs, such as Au22(SG)16, Au22(SG)17, and Au22(SG)18, have similar band gap but different absorption peaks at 560, 540, and 515 nm, respectively.32,33 Nienhaus and co-workers reported protein adsorption on Au NCs stabilized by lipoic acid with comparable absorption features.13 However, 20682

dx.doi.org/10.1021/jp5060217 | J. Phys. Chem. C 2014, 118, 20680−20687

The Journal of Physical Chemistry C

Article

and thiosulfonate (R−SO2−SR at 1340 and 1150 cm−1) were not observed and ruled out. IR results suggested that some sulfur atoms were oxidized during dialysis. Correspondingly, those oxidation peaks were not present or very weak from the centrifuged samples that were promptly purified with less exposure to air (O2). While lipoic acid is known oxidizable forming various S−Ox species as shown in Figure SI-6,42 oxidation of sulfur bonded to Au has not been reported in the extensive studies of various Au− monothiolate NCs and the recent studies employing lipoic acid. We propose that some lipoic acid ligands might have one of the two sulfur atoms only weakly bonded to Au after synthesis and thus prone to oxidation. If both sulfur atoms on one lipoic acid were oxidized, the ligand would dissociate from Au surface and be removed by dialysis. To actively control and expedite the oxidation process and to attest the proposed mechanism, the centrifuged Au−LA NCs were treated with a strong oxidant H2O2. The spectrum is included in Figure 3d. The emerging peaks at 1045 and 1187 cm−1 are consistent with those observed from dialyzed Au NCs. The results strongly support the hypothesis that these peaks arise from the oxidized sulfur species. Meanwhile, a new peak at ca. 1125 cm−1 and a broader one at 1190 cm−1 can also be resolved, which match those from alkyl−sulfate species. Apparently H2O2 did induce the spectrum features but generated more complex products due to more aggressive oxidation. For reference, lipoic acid itself was titrated with H2O2 shown in the Supporting Information (Figure SI-6). Those peaks at 1045 and 1190 cm−1, and the weak shoulders around 1125 and 1306 cm−1, were found to increase with the addition of H2O2. The features are consistent with those from Au−LA NCs and support the proposed oxidization mechanism. A general concern in the synthesis of dithiol/dithiolate Au NCs is whether one of the sulfur is reduced into thiol and not bound to Au during synthesis. The S−H stretching at ca. 2500−2600 cm−1 range was not observed from the purified Au−LA NCs. The IR spectrum of the synthetic mixture prior to purification, included in Figure SI-7, displayed significant background around 1000 cm−1 due to synthetic byproducts (i.e., boronic species, excess ligands) that limited the resolution of any oxidized sulfur signals. Elimination of the broad background signals around 1000 cm−1 further affirmed the efficacy of both purification routes. Other major IR peaks of the Au−LA NCs and the pH dependence, such as the alkyl stretching at 2932/2855 cm−1, the carbonyl stretching at 1690 cm−1, and asymmetric/symmetric carboxylate stretching at 1560/1405 cm−1, etc., are explained following the IR spectra collected at pH 4.0 and pH 7.4 shown in Figure SI-7. Charge States of Gold and Sulfur by XPS Analysis. The binding energies (BE) of Au and S were analyzed by X-ray photoelectron spectroscopy (XPS) shown in Figure 4. The Au 4f7/2 band was found at 84.5 and 84.3 eV from dialyzed and centrifuged samples, respectively, which lie between Au(0) (83.9 eV) and Au(I)−thiolates (85−86 eV). The results are consistent with monothiolate stabilized Au NCs and confirms the presence of Au(0) core in both NCs. A mild positive shift of 0.2 eV BE could correspond to the oxidation effects on the overall Au core charge states through the dialysis process. The oxidized sulfur species are evident in Figure 4c,d. The S 2p3/2 peak at 162.6 eV is consistent with the reported ligand features (RS−Au) in various monothiolate Au NCs. New peaks from oxidized sulfurs were observed at higher binding energies. Specifically, the peak at 168.2 eV, matching the BE of R−SO3

comparison, a mild increase if any, from ca. 1 to 2%, was observed from the portion purified via ultracentrifuge that was complete within hours. The normalized emission spectrum showed negligible peak shift with a maximum at ca. 740 nm. Such drastic enhancement in QE upon dialysis has never been observed in other Au NCs to the best of our knowledge. Because both purification processes were confirmed effective by IR and 1H NMR analysis, the QE enhancement involves factors more than simple purification effects. The absorption features of the purified Au−LA NCs are also included in Figure 2. The normalized absorption spectrum from the dialyzed sample is between the centrifuged and the original ones. The optical band gap is 1.9 eV for all three samples as plotted in Figure SI-4. The absorption band at 2.5 eV (505 nm) is much weaker in the dialyzed spectrum. Similar absorption transition has been reported in the oxidation of anionic Au25 into neutral Au25 NCs.41 Because no other absorption bands were discernible and the band gap remained unchanged, the absorption changes suggest possible oxidation of the Au−LA NCs rather than core size changes. The same impact by purification was observed from different batches of Au NCs synthesized at room temperature and at higher temperatures. The excitation, emission, and absorbance features are included in Figure SI-5. The unexpected QE dependence on purification appears to be universal in Au−LA systems. The finding suggests a new route to further improve the QE of the near-IR luminescence that is highly favorable for biomedical imaging. Oxidized Sulfur Characterized by Infrared Spectroscopy. A series of IR spectra collected under comparable conditions are presented in Figure 3. Compared with the

Figure 3. FTIR spectra of (a) lipoic acid sodium salt (prepared from a LA−NaOH solution), (b) centrifuged Au−LA NCs, (c) dialyzed Au− LA NCs, and (d) centrifuged Au−LA NCs treated with 2 equiv of H2O2. Each spectrum was recorded from a drop-cast sample after being air-dried. The broad features around 3000 cm−1 are affected by hydrogen bonding from residue H2O.

spectra from lipoic acid itself and the centrifuged Au−LA NCs, two new distinct peaks at 1045 and 1187 cm−1 can be identified from the dialyzed sample. These peaks match well with the S O symmetric and asymmetric stretching bands in alkylsulfonic acid/sulfonate species, such as 2,3-dimercapto-1-propanesulfonic acid (DMPS), which have two peaks at 1042 cm−1 and 1179 cm−1 (R-SO3Na), respectively. In reference to the characteristic IR signals of other organosulfur species in the region from 600 to 1500 cm−1, sulfone (R−SO2−C at ca. 1300 and 1135 cm−1), sulfate (RSO4H at ca. 1250 and 1060 cm−1), 20683

dx.doi.org/10.1021/jp5060217 | J. Phys. Chem. C 2014, 118, 20680−20687

The Journal of Physical Chemistry C

Article

Figure 4. XPS analysis of Au 4f (a, b) and S 2p (c, d) bands. Panels a and c were taken from a dialyzed sample while panels b and d from a centrifuged sample. The S 2p peaks were deconvoluted based on characteristic BEs of organosulfur species (R−SO2/R−SO at 166.2 eV, R−SO3 at 168.2 eV). The fitting data are summarized in Table SI-1.

is 64 Hz on Au NCs, while that of free lipoic acid is 164 Hz (0.41 ppm) and that of dihydrolipoic acid (DHLA) is 42 Hz.44 Those splittings are too large for J coupling and suggest the two protons being magnetically nonequivalent in all species. Compared with DHLA splitting induced by the C-6 stereocenter, the larger splitting in Au−LA NCs suggests that, albeit one of the sulfur is oxidized, its interaction with the Au core is sufficiently strong to impose additional constraint that makes the two H-7s more magnetically different. The decrease in the splitting of the two H-7s from free lipoic acid to Au−LA NCs provides undoubted structural evidence for the disulfide bond breakage or the disruption of the tight five-atom dithiolane ring in Au−LA NCs as observed in UV−vis and IR spectrum transitions. Additional insights can be learned from the comparison of the chemical shifts of H-6 and H-8s on Au-LA NCs with those from DHLA, lipoic acid, and representative R−CHx−SOx compounds. The related chemical shifts are summarized in Tables SI-2−SI-4. First, the H-6 and H-8s on Au-LA NCs displayed 0.57 and 0.54 ppm downfield shift compared with free DHLA, which immediately ruled out the existence of unbound SH groups after the reductive synthesis. Second, the H-8s peak at 3.24 ppm is in the representative range of both R−CH2−SO3 (2.90−3.40 ppm) and R−CH2−S−Au (3.17 ppm in Au 25 (SCH 2 CH 2 Ph) 18 , 4 5 3.30−3.40 ppm in Au25(SG)1846). The downfield shift compared with H-8s on DHLA is also consistent with RCH2−SH to RCH2−S−Au conversion. Therefore, this H-8s peak could include both the oxidized ligands and those forming classic Au−S bonding. Third, the H-6 peak at 3.60 ppm is shifted upfield by 0.13 ppm compared with free LA and downfield by 0.57 ppm compared with DHLA. No oxidation of the inner S (on C-6) was observed based on the significant downfield shift compared to R2−CH−SO3 (ca. 3.00 ppm).47 In the HSQC spectrum, 1H resonances at 3.60 ppm (H-6) and 3.24 ppm (H-8) correlate with carbon signals at 47.9 and 49.1 ppm, respectively. The weak crossing signals indicate that both sulfurs are localized near Au and thus suffer significant broadening effects. The C6 peak is more broadened and barely discernible, indicating C8 being less restricted or weaker interactions with Au. Compared with the reported alkylsulfonate species R−CH2−SO3 at ca. 52.0 ppm and R2−CH−SO3 at ca. 60.0 ppm,47 such chemical shifts also suggest the oxidation at the outer sulfur on C-8 rather than the inner one on C-6. While the chemical shift of C-6 at 47.9 ppm is close to the reported CH carbon signals in other R2−CH−S−Au system such as Au−tiopronin NCs (48.0 ppm),48 the C-8 peak at 49.1 ppm is downfield shifted more significantly if compared with

species, is more prominent from dialyzed Au−LA NCs. The band at 166.2 eV from the centrifuged sample was attributed to the intermediate oxidized species such as R−SO and R−SO2 that were ultimately converted to sulfonate species through longer exposure to air/water, i.e., through dialysis.43 The intensity of this band varied from different batches, presumably due to the variations in their exposure to air/water from sample preparation to XPS data collection. Qualitatively, this band gradually diminishes, and the R−SO3 band intensifies with respect to the Au−SR band. About 25% of sulfur atoms, or half of the LA ligands if one of two sulfurs per ligand, was ultimately oxidized from these measurements. The XPS BE analysis supports the IR observation and provides key insights into the proposed oxidation mechanism. The sulfur oxidation is fundamentally different from earlier reports that primarily studied the redox activities of Au cores or charge effects on the peripheral/terminal groups in ligand monolayer. Chemical Bonding and Composition by NMR. The next question to answer is which one of the two S atoms on LA ligand is oxidized or whether the oxidization is selective. The NMR spectra are shown in Figure 5. For reference, the 1D proton NMR spectrum and the molecular structure of free lipoic acid are included in panel a. The chemical shifts at carbon 6−8 positions will be most informative for the elucidation of S bonding at the core−ligand interface and thus the focus of the following discussion. It is important to mention that due to more significant motion restriction and heterogeneity in chemical environment after cluster formation, NMR signals at the α-position from S (and β-position that is also closer to Au core, i.e., 5−8 positions) are more broadened and/or shifted compared to those positions further away along the ligand (i.e., 4−1 positions). Further, in the 1D spectrum of Au NCs, the absence of sharper peaks corresponding to unbound free lipoic acid confirms the efficacy of the purification. Sharper NMR features corresponding to smaller complexes or synthetic intermediates are plotted in Figure SI-8, which reflects less effective purifications and could be misinterpreted in the analysis of Au NCs. The peak assignments in the 1H NMR spectrum are elucidated together with 2D spectra analysis, including through-bonds 1H−1H correlation spectrum (COSY) and one-bond 1H−13C correlation spectrum (HSQC). In the COSY spectrum, the peak at 3.14 ppm is assigned to the protons on C-8 (H-8s), which showed cross-peaks with the adjacent H-7s. The H-6 peak appears at 3.60 ppm and couples with both H-7s and additionally with the H-5s peak. The crosstalk between the signals at 2.26 and 2.10 ppm further confirms the H-7s assignment. Interestingly, the splitting of the two H-7s 20684

dx.doi.org/10.1021/jp5060217 | J. Phys. Chem. C 2014, 118, 20680−20687

The Journal of Physical Chemistry C

Article

from classic point of view. Second and likely more important, the inner sulfur on C-6 might be less accessible than the outer one on C-8 depending on the bonding structures at the core− ligand interface. Given that ca. 1 out of 4 sulfur atoms being oxidized from XPS analysis, and combined with MS data and literature report of Au22SR18,32 we propose that there are 12 LA ligands per Au−LA NC. Among the 24 sulfur atoms, 6 being oxidized and the other 18 are responsible for direct bonding with Au. The proposed composition will therefore allow the construction of similar structures discovered in monothiolate Au NCs. Further studies, particularly with structural information, are required to confirm this hypothesis. Luminescence QE at Different pH: Dipoles Buried under the Ligand Monolayer versus at the Periphery. An opposite trend was observed in the pH dependence of luminescence QE from the oxidized Au−LA NCs compared with the monothiolate Au NCs that have attracted extensive research interest. As shown in Figure 6, the QE of Au−LA NCs

Figure 6. (a) pH dependence of emission QE of Au−LA NCs. Each color represents a series of pH adjustments. (b) Concentration effects on the QE pH dependence of the oxidized Au−LA NCs. The emission intensity was normalized by the corresponding absorption values at the excitation wavelength of 400 nm. The absorbance of the stock solution (concentration C0) was below 0.5. The solution pH was adjusted by either HCl or NaOH in respective measurements.

increased and plateaued when the solution pH decreased from 12.0 to 3.0−4.0. Protonation of the terminal carboxyl group will neutralize the ligand charges. Unlike the Au−LA NCs, the QE of the near-IR luminescence is known to decrease for carboxylic acid-terminated monothiolate Au NCs due to the decrease in monolayer polarity.19 The unique pH dependence is attributed to the oxidized SOx species beneath the ligand chain that serves as a dielectric layer as illustrated in Scheme 1 and NMR Figure 5a. An elimination of the terminal negative charges will effectively reduce the electron density surrounding the dielectric layer. Because the oxidized SOx is beneath the dielectric layer, a decrease in the electronic coupling with the peripherals will increase the oxidation induced polarization at the core−ligand interface. This is similar to the introduction of a positive charge at ligand terminal groups or the oxidation of Au core described in the literature.11,16 Electronic/dipole coupling across the dielectric layer is also supported by the impacts on the charging energy by charge/dipole intrusions into ligand monolayer in voltammetric studies.50 The argument is further supported by the much weaker pH dependence, if any, from the centrifuged Au−LA NCs that are less oxidized. The proposed scenario also explains the recent results in which PEGlation or zwitterion insertion on lipoic acid was found to enhance the emission.20,21 The hydrophilic headgroup and hydrophobic chain of lipoic acid render the Au−LA NCs excellent performance/stability in a wide pH range from 2.2 up to 11.0. The linear dependence in

Figure 5. NMR spectra of dialyzed Au−LA NCs in D2O (4.80 ppm as solvent peak): (a) comparison of free lipoic acid and Au−LA NCs; (b) COSY spectrum; (c) HSQC spectrum. The solution pH was adjusted to slightly basic with NaOH for better solubility.

other R−CH2−S−Au signals (34.0−39.0 ppm46,49). Therefore, the significant downfield shift of C-8 further attests the oxidation of sulfur on C-8. Figure SI-9 provides the 1H spectrum of both dialyzed and centrifuged (less oxidized) Au− LA NCs. Unfortunately, H-6 and H-8 signals could not be resolved presumably due to the sample heterogeneity. A very weak peak at 2.60 ppm could indicate the intermediate or partial oxidation. Two factors could have contributed to the selective oxidation. First, the higher electron density of the sulfur on C-8 is more favorable for the oxidation than the one on C-6 20685

dx.doi.org/10.1021/jp5060217 | J. Phys. Chem. C 2014, 118, 20680−20687

The Journal of Physical Chemistry C UV−vis absorbance with the Au−LA NCs concentration, and the overlapping normalized spectra, suggest negligible aggregation at each pH (Figures SI-10 and SI-11). Shown in Figure 6b, the luminescence QE clearly has two zones at high and low pH in a series of Au−LA NCs concentrations. The terminal carboxyl groups will be largely deprotonated at higher pH, i.e. 7.2 and 11.0, and protonated at lower pH 2.2 and 4.0. The pKa of lipoic acid is ca. 4.7, while the pKa of sulfonic acid (