Olga Burchak

Caroline Keller

Gérard Lapertot

Mathieu Salaün

Julien Danet

Yani Chen

Nedjma Bendiab

Brigitte Pépin-Donat

Christian Lomba

Jérôme Faure-Vincent

Anthony Vignon

David Aradilla

Peter Reiss

Pascale Chenevier
email: pascale.chenevier@cea.fr

Scalable chemical synthesis of doped silicon nanowires for energy applications †

A versatile, low-cost and easily scalable synthesis method is presented for producing silicon nanowires (SiNWs) as a pure powder. It applies air-stable diphenylsilane as a Si source and gold nanoparticles as a catalyst and takes place in a sealed reactor at 420 °C ( pressure <10 bar). Micron-sized NaCl particles, acting as a sacrificial support for the catalyst particles during NW growth, can simply be removed with water during purification. This process gives access to SiNWs of precisely controlled diameters in the range of 10 ± 3 nm with a high production yield per reactor volume (1 mg cm -3 ). The reaction was scaled up to 500 mg of SiNWs without altering the morphology or diameter. Adding diphenylphosphine results in SiNW n-type doping as confirmed by ESR spectroscopy and EDX analyses. The measured SiNW doping level closely follows the initial dopant concentration. Doping induces both an increase in diameter and a sharp increase of electrical conductivity for P concentrations >0.4%. When used in symmetric supercapacitor devices, 1% P-doped SiNWs exhibit an areal capacity of 0.25 mF cm -2 and retention of 80% of the initial capacitance after one million cycles, demonstrating excellent cycling stability of the SiNW electrodes in the presence of organic electrolytes.

Introduction

Elemental silicon, the central building block of microelectronics, also shows outstanding characteristics for energy conversion and energy harvesting applications. 1 The most wellknown example is the use of crystalline silicon in photovoltaic panels, which dominates solar cell technology. 2,3 For Li-ion batteries, silicon has attracted a lot of interest as a constituent of anode materials 4 due to its high Earth abundance, low discharge potential, very high theoretical charge capacity (>4000 mA h g -1 ), 5 and its abilityafter nanostructurationto stand mechanical strains due to swelling/deswelling in lithiation/delithiation. 6 Among the different nanostructured morphologies (0D nanoparticles, 1D nanowires, 2D nanosheets), silicon nanowires (SiNWs) are of particular interest for use in energy storage or energy conversion devices. First of all, they possess a very high aspect ratio (generally around 100 to 1000) favoring efficient charge transport as well as a good processability from dispersions or slurries facilitating e.g., the preparation of electrode materials. Furthermore, SiNWs exhibit a huge surface-to-volume ratio and their electrical conductivity can be increased via doping. The former features are particularly beneficial for their application as anodes in Li-ion batteries, 7 while the latter are of special interest for supercapacitors 8 and thermoelectrics. 9 SiNW growth was initially reported more than 60 years ago 10 and the potential application of SiNWs in electronics boosted research on controlled growth methods in the 1990s. 11,12 The control of n-and p-type doping is not only important for electronic devices 13,14 or optoelectronic p-n junctions, 15,16 it is also of great interest for SiNW based devices for energy storage or conversion. Thermoelectrics, 9 photoelectrocatalysis 17 and supercapacitors 18 are illustrative examples of applications requiring high electrical conductivity. Phosphorus-doped SiNWs also proved to lower the lithiation potential of Si in Li-ion batteries. 19,20 In general, n-type doping will be preferred as it leads to higher conductivity in Si than p-type doping. 21 For all these reasons, the development of scalable and cost-efficient synthesis methods giving access to large quantities of doped SiNWs is highly desirable. Current methods do not satisfy this demand. The high chemical versatility of Si opened very different ways for producing SiNWs in bottom-up and top-down approaches. 22 To date, the three most popular methods are metal assisted catalytic etching (MACE), chemical vapor deposition (CVD) and solution growth. 8,23 MACE relies on the catalytic Si dissolution by HF solution in contact with metal particles deposited on the surface of bulk Si. This method yields SiNW arrays 24,25 or SiNW decoration around Si grains 26 at low temperature (50 °C). The SiNWs are intrinsically doped if the bulk Si source is doped, i.e. when doped Si wafers are etched. The main drawback of this method is its high cost due to the use of silicon wafers. CVD and solution growth, on the other hand, both rely on crystallization of elemental Si obtained by cracking a molecular Si source. CVD growth is performed at 650 °C without catalysts 27 or at 400-500 °C with metal catalysts based on the vapor-liquid-solid (VLS) mechanism. Boron or phosphorus dopants added in the process are incorporated into the SiNWs during growth. 18,22 The safe handling of highly reactive and sensitive SiH 4 or SiCl 4 used as gaseous sources of Si is one challenge of this method. Furthermore, as in the case of MACE, CVD growth takes place on 2D substrates, and therefore the maximum SiNW surface area density is a factor intrinsically limiting the quantity, which can be produced in one reaction. Chemical methods, characterized by the use of liquid (or solid) Si precursors and solvents, can be divided into two classes: the first one taking place at ambient or low pressure 28,29 and the second implying high pressure conditions and solvents (e.g., toluene, hexane) in their supercritical state. 30,31 In both cases, liquid organosilanes replace SiH 4 as safer silicon sources while only supercritical growth has been reported to give access to larger quantities (0.1-1.0 g). However, reactions of organosilanes in supercritical medium are complex 32,33 due to simultaneous transformations of the silicon and organic moieties and interactions with the solvent, which makes controlled doping a challenging task. Moreover, the control of the NW diameter, in particular in the range of 10-50 nm, remains to be demonstrated.

In the present work, we present a robust and easily scalable method for the solvent-free chemical synthesis of thin (10 ± 3 nm) SiNWs using diphenylsilane as a Si source and small (1-2 nm) gold nanoparticles (AuNP) as a catalyst in a sealed reactor at 420 °C and with a pressure <10 bar. The catalyst nanoparticles are immobilized on micron-sized NaCl powder, which acts as a sacrificial 3D substrate easily removable by washing with water after NW growth. Pure SiNWs are obtained at a high production yield of 1 mg cm -3 of reactor volume and with a 70% chemical yield. N-type doping of the SiNWs is achieved by adding diphenylphosphine at concentrations of 0.025 to 1.5% as the dopant source. Using Raman and ESR spectroscopy as well as TGA, ICP-AES and EDX measurements, we demonstrate successful volume doping of the SiNWs. Doping results in a jump of electrical conductivity for dopant concentrations exceeding 0.4% and also induces an increase of the NW diameter. The obtained materials are used as nano-structured electrodes in supercapacitors, resulting in excellent areal capacity, power density and cycling stability, in particular in the case of 1% P-doped NWs.

Experimental section

Materials and reagents

Diphenylphosphine, propylene carbonate (PC), tetrabutylammonium tetrafluoroborate (TBABF 4 ) and carboxymethylcellulose sodium (CMC) were purchased from Sigma Aldrich. Gold tetrachloroauric acid, tetraoctylammonium bromide (TOAB), diphenylsilane, sodium chloride, carbon black Super-P (TIMCAL) and copper foil (thickness of 0.25 mm) were purchased from Alfa Aesar.

AuNP growth

Following Brust's method 34 optimized by Hostetler, 35 gold tetrachloroauric acid HAuCl 4 •3H 2 O dissolved in water is transferred to toluene using TOAB, then reduced with NaBH 4 and stabilized with DDT (DDT/Au molar ratio 2 : 1), leading to 1.5-2 nm diameter AuNPs quantitatively.

SiNW growth

SiNWs are obtained from a solution of diphenylphosphine PPh 2 H in diphenylsilane SiPh 2 H 2 at a molar ratio of 0.025% to 1.5%. AuNPs in solution in dichloromethane (5 mg mL -1 ) are drop-dried on a finely ground NaCl powder (2 mg AuNPs on 1 g NaCl) in a warm mortar at 80 °C. The PPh 2 H/SiPh 2 H 2 solution (0.6 mL, 3.2 mmol of diphenylsilane) and the powder are sealed in a 30 mL glass tube (16 mm outer diameter, 1.5 mm thickness) under vacuum. The glass reactor is heated at 420 °C for 4 h in a preheated tubular furnace. At the end of the reaction, one end of the glass tube is pulled out of the oven for 15 minutes to allow for local condensation of tetraphenylsilane, and then the glass tube is cooled to room temperature. Care must be taken while opening because of the pressure in the glass tube (2-3 bars). The black micropowder is washed with water, acetone and dichloromethane to recover 30-35 mg of pure SiNWs (35% yield in Si conversion). For EDX analysis and electrical conductivity measurements, SiNWs were prepared as pellets: dry SiNW powder was (6 to 20 mg) finely ground and pressed for 10 minutes under a 2t pressure in a 5 mm die in a SPECAC hydraulic press.

Preparation of electrodes

SiNWs were employed as active materials. CMC was used as a binder and Super P as a conducting matrix. The formulation used in this study consisted of 50% wt of SiNWs, 25% wt CMC and 25% wt conductive carbon (Super P), respectively, using a total mass of 222 mg. Firstly, the corresponding amounts were weighed using a Sartorius micro-balance into a Techni-dome enclosure. Subsequently, the components were transferred into a planetary ball mill system (Fritsch model), made of silicon nitride milling tools, and filled with Milli-Q distilled water (2 mL) at 700 rpm for 3 h. After that, the obtained slurry for the electrode's induction was prepared by the doctor blade method using a Smart coater to obtain a homogeneous and uniform coating onto copper foils. Finally, the electrodes were dried under vacuum at 90 °C overnight before cutting them into the required dimensions.

Elaboration of supercapacitor devices

SiNW-based supercapacitors were built from CR2032 coin cell devices (MSK-160D, MTI, USA) using a symmetric configuration. CR2032 devices were composed of coin cell cases, wave springs, stainless steel (SS) spacers, two separators (Whatman glass fiber paper) and two identical electrodes of SiNWs (mass loading of 2 ± 0.5 mg per electrode) on round copper substrates (1.76 cm 2 , diameter 15 mm), which were in contact with stainless steel spacers as a current collector. The gold catalyst was not removed from the SiNW material, as gold was shown not to interfere with the electrochemical behavior of supercapacitors in previous studies based on CVD-grown SiNWs. 36,37 Before assembling, the separator was soaked in a PC solution containing 0.5 M TBABF 4 (a total volume of 220 µl). 36

Electrochemical performances of supercapacitor devices

The electrochemical properties of the full devices were evaluated using CV curves, galvanostatic charge-discharge (GCD) cycles and electrochemical impedance spectra (EIS) using a multichannel VMP3 potentiostat/galvanostat with Ec-Lab software (Biologic, France) at a cell voltage of 2 V. CV curves were recorded for scan rates ranging from 0.01 to 40 V s -1 . GCD cycles were recorded at various current densities (0.14-4.53 mA cm -2 ), and EIS tests (Nyquist and Bode plots) were performed using a frequency range between 400 kHz and 10 mHz with an ac perturbation voltage of 10 mV at open circuit potential. Areal capacitance (AC, µF cm -2 ) was calculated using GCD cycles and CV curves using the following equations: AC = I/ A(dV/dt ) where I is the discharge current, A is the surface of the electrode and dV/dt corresponds to the slope of the discharging curve, and AC = iυ where i is the average capacitive current density ([Δj: ( j oxj red )/2]) in the middle of the cell voltage (1 V) and υ is the scan rate. The energy (E, mJ cm -2 ) and power (P, mW cm -2 ) density values were calculated by using E = 0.5 ACΔV 2 and P = E/t d , where ΔV is the cell voltage and t d is the discharge time. The coulombic efficiency (η) was determined along η = t d /t c where t c is the charge time and the cycling stability was measured by applying 2 V at a current density of 0.57 mA cm -2 . The electrochemical results reported in this study were analyzed from an average of three samples to ensure a good reproducibility. Before running any electrochemical experiment, the cells were equilibrated overnight. All measurements were carried out at room temperature in an argon-filled glovebox with oxygen and water levels less than 1 ppm.

Electron microscopy

SiNWs were imaged by scanning electron microscopy (SEM) on a Zeiss Ultra 55 microscope at an accelerating voltage of 5 kV.

Samples were prepared by drop casting a suspension of SiNWs in dichloromethane on a Si wafer substrate. Diameter histograms were recorded by measuring the diameter of all SiNW sections in each 0.5 × 1 µm 2 zone of SEM images using ImageJ (>500 counts per sample). SiNW film electrodes for supercapacitors were observed under the same conditions with a tilted angle of 45°.

Energy-dispersive X-ray spectroscopy (EDX) spectra were recorded on SiNW pellets (compacity >80%) in the same microscope equipped with an EDX probe (acceleration voltage 5 kV) and calibrated with standards of pure SiC, MgO and GaP.

Transmission electron microscopy (TEM) images were acquired on a Jeol JEM-3010 microscope, (300 kV) and on a Titan Ultimate microscope (200 kV). For TEM, a dispersion of SiNWs in dichloromethane is drop-cast on an ultrathin carbon lacey film on a Cu grid (TedPella Pelco 1824).

Brunauer-Emmett-Teller (BET) analysis N 2 adsorption/desorption curves were acquired on a MICROMERITICS ASAP 2060 analyser. First, the samples were dried and degassed at 120 °C under high vacuum. The adsorption/desorption measurements were conducted at 77.349 K in the range of relative pressure p•( p 0 ) -1 from 0.01 to 0.995. Adsorption/desorption curves were analyzed using ASAP 2060 software.

Raman analysis

The Raman spectrometer setup consists of a confocal microscope with a 320 nm spot size for a laser wavelength of about 532 nm. Confocality of the system is ensured by using a 50 mm optical fiber for both injection and collection of light which allowed an accuracy in the mapping about 120 nm. The elastically scattered light from the sample is filtered out by using an edge filter, while the inelastically scattered light is collected and sent to a spectrometer with resolution less than 0.9 cm -1 . Spectrum acquisition was performed by using a CCD camera, and cooling down to -65 °C was performed by Peltier cooling. The Raman spectrometer (WITec alpha 500) is equipped with a piezoelectrical stage, allowing making of 2D confocal maps of the sample.

Conductivity measurements

We used the two-terminal sensing method to measure the conductivity of SiNW pellets. For this measurement, a shadow mask is used to evaporate the two 200 nm-thick and 3 mmspaced Au contacts on top of the pellet under secondary vacuum. Due to the pellet fragility, the electrical spring probes were not directly put onto the pellet: we bonded the pellet to deported electrical contacts by gluing gold wires with silver paste. Prior to any measurement, the pellets were pumped overnight under primary vacuum. The measurements were performed in a dynamic helium flow cryostat Oxford Instruments CF 1200 D from 340 K down to the lowest possible temperature, i.e. the temperature for which the sample resistance is measurable by the electronic setup (combination of a Keithley 220 current source and Keithley 6512 electrometers).

Phosphorus content determination by ICP-AES

Inductively coupled plasma atomic emission spectroscopy (ICP-AES) (Shimadzu ICP 9000 with a Mini plasma Torch in axial reading mode) was used to measure the phosphorus content. Standard solutions of P for atomic absorption spectroscopy (Sigma Aldrich) were used for quantification (calibration curve between 7.8 and 2000 µg L -1 with 10% HNO 3 (Fluka)). SiNWs (10 mg) were dissolved in NaOH aqueous solution (4 M, 0.5 mL) in a plastic vial. After 1 hour incubation in an ultrasound bath, the solution was centrifuged (2000g, 2 minutes) to remove the gold nanoparticles and diluted in 6 mL aqueous HNO 3 10%. ICP-AES was performed on this solution and a 1/100 dilution in HNO 3 10%. Results were obtained in (mg of P)/(mg of SiNW).

Thermogravimetry and mass spectrometry analysis (TG-MS analysis)

Thermogravimetry (TG) was carried out with SETARAM TAG 16 equipment, using approx. 5 mg samples within 100 μL alumina crucibles. The thermal analysis was composed of a single heating ramp of 10 K min -1 until 600 °C. It was directly coupled to a HIDEN analytical apparatus (QGA-HAL201-RC) mass spectrometry (MS) that analyses the decomposition by products. Specific masses were followed during all the thermal analyses. MS signal intensity was normalized by the sample mass for comparison.

Electronic spin resonance ESR

Spectra were recorded at 9 GHz at 77 K on an ER 200 Bruker spectrometer in a resonant cavity ER 4104 OR. For each sample, a pure powder of SiNWs (3 mg) was packed at the bottom of a 3 mm diameter quartz tube. Low temperature experiments were performed with a cryostat (ESR 900). Comparisons are made on spectra recorded under the same conditions (gain, modulation, accumulation times, accumulation number and microwave power). The operating conditions were chosen to avoid significant power saturation of each line and modulation broadening. The number of ESR lines present in the global signal, their Landé g-factors, line shapes (Gaussian or Lorentzian or both with relative g-centered contribution), peak-to-peak line width and relative intensities were determined after simulation of the experimental spectra. Separate experimental measurements of the magnetic field and of the frequency were performed in order to ensure accurate g-values (±0.0002).

Results and discussion

One key feature of our method is the use of micron-sized salt (NaCl) particles acting as a sacrificial support for the catalyst nanoparticles. The latter consist of dodecanethiol-capped 1-2 nm gold nanoparticles (AuNPs) synthesized following the Brust method. 34,35 If deposited on a large flat surface, AuNPs merge into extended gold droplets and yield Si pillars of large diameters (>200 nm, data not shown). Here, the AuNPs are de-posited from hexane onto NaCl microcrystals, which prevent them from agglomeration and allow for SiNW growth in the 10-20 µm-sized void spaces. The high porosity of the NaCl powder enables an optimal circulation of the reagents (in the gaseous phase at the reaction temperature) leading to homogeneous growth within the whole volume of the 3D support. Catalyst loading is kept at a high molar ratio of Au/Si = 1/250, together with a high organosilane loading of 0.11 mmol cm -3 of reactor volume. This results in a maximum calculated pressure of 9.2 bar in the reactor at 420 °C, far below the bursting pressure of the glass tube (14 bar). Diphenylsilane was chosen as the Si source, which has been shown by Korgel 29,33 and Ryan and coworkers 38 to be one of the organosilanes best suited for SiNW growth. Among its advantageous features, diphenylsilane is air-stable and its high boiling point (280 °C) is conveniently suited close to the Au-Si eutectic temperature (363 °C) at which SiNW growth starts. For doping, air-stable diphenylphosphine (PHPh 2 ) was applied used at molar P/Si ratios ranging from 0.025 to 1%. Standard syntheses (420 °C, 4 hours) produce 30-35 mg of SiNWs in 30 cm 3 glass tubes sealed under vacuum. Until now producing such high quantities of SiNWs in comparably small reactor volumes (1 mg cm -3 ) could only be achieved under supercritical conditions, in the so-called SuperFluid-liquid-solid (SFLS) growth. 32,39 However, SFLS requires high pressure/high temperature setups compatible with the extreme reaction conditions (typically 100 bar/500 °C). Simpler growth methods in standard glassware, on the other hand, yield very low amounts of SiNWs. 28,40 Here we attain high SiNW quantity per reactor volume due to high organosilane and catalyst loadings and catalyst stabilization on NaCl micropowder acting as a 3D sacrificial support. 41 Moreover, by using a 150 mL stainless steel reactor and simply upscaling the used precursor quantities by a factor of 15, around 500 mg of SiNWs could be produced in a single run, clearly demonstrating the facile scalability of the presented method.

The mechanistic path of the reaction as proposed by Lee et al. 33 unfolds into 3 steps: diphenylsilane disproportionation 42 to monophenylsilane (1) then silane (2), cracking of silane (3) on the catalyst yielding Si atoms dissolved in the Au droplet, and (4) crystallization of Si from supersaturated Au droplets. Note that the overall conversion of Si from diphenylsilane to SiNW along eqn (5) cannot exceed 50%.

2 SiPh 2 H 2 ! SiPh 3 H þ SiPhH 3 ð1Þ
SiPh 2 H 2 þ SiPhH 3 ! SiPh 3 H þ SiH 4 ð2Þ
SiH 4 þ Au ! 2 H 2 þ Si=Au ð3Þ
Si=Au ! Si=NW ð4Þ
Overall : 2 SiPh 2 H 2 ! SiPh 4 þ 2 H 2 þ Si=NW ð5Þ
Purification of the NWs is achieved by dissolving NaCl in water, while the tetraphenylsilane subproduct is separated by condensation at a cold-point, induced by cooling one end of the glass reactor 15 minutes before stopping the reaction. The Si atom conversion yield is 35 ± 5%, i.e. 70% of the maximum expected yield (5), independent of the presence of diphenylphosphine.

SEM and TEM analyses of the product reveal entangled SiNWs mostly assembled in cushion-like structures of 10-20 µm (Fig. 1 and S1 †). The SiNWs are long and straight (Fig. 1A andB) in a fluffy intertwined structure of ∼90% porosity. On the outer surface of the "cushions" appear a few compact agglomerates consisting of tightly associated, short SiNWs (Fig. S1 †). These agglomerates are very thin and show a flat free surface probably originating from the NaCl crystal on which the growth took place. The SiNWs contain AuNPs of identical diameter at their tips in accordance with a VLS-type growth mechanism. 10 As the AuNP catalysts are 1.5-2 nm in diameter, SiNWs as thin as 2 nm in diameter could in principle be expected. However, it has to be taken into account that at a reaction temperature of 420 °C the small Au particles are in their molten state. Therefore, although a few very short wires thinner than 5 nm are observed (Fig. 1E), the overwhelming majority of SiNWs show larger diameters. Undoped SiNWs have a highly reproducible, homogeneous diameter of 10 ± 3 nm, which is lower than reported values using other growth methods: SiNWs from CVD have usual diameters in the 20-100 nm range; 10,43 the lower diameters are difficult to obtain because the growth rate depends inversely on the radius. 22 High-resolution TEM imaging (Fig. 1C) demonstrates the highly crystalline character and the presence of a very thin (0.9 ± 0.3 nm) amorphous shell attributed to surface oxidation occurring during the purification and handling under ambient air. Importantly, when scaling up the reaction by a factor of 15 in a stainless steel reactor yielding 500 mg of SiNWs per synthesis, the morphology, diameter and diameter distribution remain the same (Fig. S2 †).

Addition of the P dopant has a strong effect on the NW morphology, increasing both the mean diameter and its distribution in 1%P-SiNWs (Fig. 1B andD). (For simplicity, SiNWs obtained with an initial molar ratio P/Si = X% are named X% P-SiNWs, with X ranging from 0.025 to 1.) The shape remains cylindrical with no change in diameter along the length, indicating the absence of lateral growth. As for undoped NWs, the core is crystalline as shown by TEM (Fig. 1D) and X-ray diffraction (Fig. S3 †). TEM analyses further indicate that growth occurs along the 〈110〉 orientation, as classically reported for SiNWs with diameters below 20 nm grown by VLS. 10 In contrast, 〈111〉 SiNWs have been reported in syntheses using diphenylsilane in supercritical toluene. 33 The diameter distribution of undoped and doped SiNWs is depicted in Fig. 2A andC, showing a gradual shift of the mean diameter to larger values with increasing dopant concentration. An increase by a factor of 4 is observed when going from undoped to 1% P-SiNWs, inducing a drop of the surface/volume ratio by a coherent factor of 5 measured using the Brunauer-Emmett-Teller (BET) technique (Table 1). The larger mean diameter and diameter distribution of doped SiNWs suggest that the initial 2 nm-sized AuNPs fuse into larger gold droplets in the presence of the P dopant. This change of the catalyst morphology is favored by slower growth kinetics, resulting in a longer lag time between Au particle melting on the NaCl surface and SiNW growth. This hypothesis is supported by two reported effects: first, phosphines are well known to bind to Au with a high affinity 44 and thus the formation of a PPh 2 H layer at the surface of the AuNPs can be expected, slowing down SiH 4 cracking. Second, P also dissolves in liquid Au so that the catalyst droplet likely consists of an Au-Si-P ternary mixture in the presence of phosphine instead of an Au-Si eutectic. In such ternary systems, the rate of Si crystallization depends on the P concentration because a high P content decreases the solubility of Si in Au. Perea et al. 45 measured this effect in germanium NW growth demonstrating that the GeNW growth rate was reduced by 16% when adding 0.6% PH 3 . Summarizing, we expect the larger diameter of the doped SiNWs to be the result of slowed down growth kinetics induced by dopant interactions with the AuNPs.

The mean diameters and diameter distributions are also evaluated by Raman spectroscopy following the shift and broadening of the LO-TO Si vibration at 520 cm -1 (Fig. 2B). A series of Raman spectra were recorded for each sample using a confocal micro-Raman setup probing populations of SiNWs dispersed on a glass slide (Fig. S4 †). The secondary peak at 498 cm -1 is attributed to strains in the Si crystal arising in the growth process. 46 The LO-TO band is sensitive to quantum confinement, a diameter below 20 nm inducing red shift and broadening. 47 As SiNW heating induces an additional shift, broadening and asymmetric deformation due to the Fano effect, 48 the laser power was tuned to a level low enough to make temperature effects negligible (Fig. S4 †). Doping can also induce shifts, which are, however, in the case of n-doped silicon very small in comparison with confinement effects. 47,48 The LO-TO band in all SiNW spectra was blue shifted as compared to bulk Si (520.8 cm -1 , FWHM of 4 cm -1 limited by the optics), nonetheless both the blue shift and broadening were less and less pronounced with increasing doping level. The average Raman shift and FWHM nicely correlate with diameter data by SEM as a function of the doping level (Fig. 2C).

A further point of fundamental interest for any type of application is the surface state of the obtained NWs. Unlike CVD-grown SiNWs showing a hydrophilic behavior, the samples obtained by the present method are hydrophobic due to an organic layer essentially consisting of phenyl groups (vide infra). TEM images of the undoped, highly crystalline SiNWs (Fig. 1C) indicate that this organic coating is very thin. 1%P-SiNWs, on the other hand, exhibit an amorphous shell of approximately 4.6 ± 2.2 nm around the crystalline core (Fig. 1D, Table 1). This shell could for example consist of a poly-phenylsilane organic layer as previously described by Chan et al., 49 or of amorphous silicon as observed by Bogart et al. 50 To gain further insight into the surface state of the obtained SiNWs, additional analyses were carried out. First, the amount of oxygen in SiNWs estimated by EDX (Table S1 †). The O content of O/Si = 0.12 in undoped SiNWs and O/Si = 0.32 in 1%P-SiNWs was fitted by a geometric model of a crystalline Si core surrounded by an amorphous shell made of an oxide SiO x and organic material. The model was adjusted to the average outer diameters as determined by SEM (Fig. 2C), the thickness of the amorphous layer as measured by TEM and the Si/C/O content by EDX. The data for undoped SiNWs suggest that the amorphous layer has global composition SiO 1.2 C 2.4 , while 1%P-SiNW data fit with a more O-rich shell of SiO 1.75 C 0.6 . Second, thermogravimetry (TG) coupled to mass spectroscopy of effluent gas was performed. The TG spectra show at 500 °C a maximum mass loss of 5% for undoped SiNWs, and of 1% for 1%P-SiNWs (Fig. 2D). This 5-times higher mass loss for the undoped vs. the 1%P-SiNWs is in agreement with their 5 times higher BET surface (Table 1). As the amount of organic species is proportional to the surface area, it follows that the organic layer thickness must be essentially the same in doped and undoped SiNWs, i.e. a very thin layer. This result together with the EDX data indicates that the thicker amorphous shell in 1%P-SiNWs is mainly composed of amorphous SiO x . The formation of amorphous Si during the growth (then oxidizing to SiO x once in air) is likely induced by the dopant addition, as (i) phosphorus introduced during Si nanocrystal 51 or nanowire 52,53 growth is known to migrate partially to the surface and (ii) 0.4% and 1% are very high doping levels compared to the limit of solubility of P in Si, reported as 0.6%. 54,55 Mass spectrometry of the TG effluent (Fig. 2D) clearly identified the thin organic layer as being constituted of phenyl radicals. In particular, the mass loss between 250 and 500 °C was accompanied by the emission of benzene (m/z = 78), as well as lower signals at m/z = 77, 52 and 39 corresponding to usual fragments from phenylsilane thermal decomposition. 56 Benzene can form during the cleavage of vicinal Si-Ph and Si-H bonds at the surface of SiNWs. 57 This process is accompanied by the oxidation of surface Si-Si bonds in the gas flow containing traces of water (<10 ppm in Ar), forming Si-O-Si and releasing H 2 as observed by MS at 300 °C (Fig. S5 †). Mass gain due to oxidation thus partially compensates mass loss from benzene emission. Oxidation proceeds during cooling at the end of the TG analysis leading to a net mass gain. By consequence the 5% and 1% mass losses observed in the TG spectra for undoped and doped SiNWs (vide supra) underestimate the total mass corresponding to the organic layer. This explains the higher carbon content in SiNWs determined by energy-dispersive X-ray spectroscopy (EDX, Table S1 †) as C/Si = 0.24 (undoped) and C/Si = 0.12 (1% P-SiNWs), which should lead to a maximal mass loss of 8% and 3%, respectively.

Quantitative EDX measurements and measurement of P content by ICP-AES provided a direct and coherent determination of the P content in the doped SiNWs (Fig. 3A). Previous studies reported the P incorporation to be proportional to the initial P/Si ratio in VLS growth 58 with a saturation at P/Si = 1.5%. 59 However, the P content was estimated indirectly, relying on SiNW conductivity and subsequent modeling. Here, a direct measurement of the atomic P/Si ratio was obtained by two methods: the EDX was recorded under quantitative conditions from SiNW pressed pellets, and P content was confirmed by P content determination by ICP-AES. The measurements are coherent and reproducible except for 1% P-SiNWs, in which P is unevenly distributed (Fig. S7 †). Phosphorus was shown to incorporate into the SiNWs with the same P/Si ratio as that used in the reagents, as previously reported in ref. 58,59.

Global P content measurements do not indicate whether the dopants are localized in the crystalline NW core or accumulated at the surface, as previously reported in ref. 51,53. Only in the former situation efficient doping takes place leading to a significant increase in electrical conductivity. In the case of germanium NW doping, it has been reported that the P distribution can be very inhomogeneous along the nanowire section, with differences in concentration as high as a factor of 100. 23,45 Electron spin resonance (ESR) measurements indicate that in our case effective doping of the crystalline SiNW core takes place. This is evidenced by the appearance of a new signal at g = 1.998, noted gP (Fig. 3B), not visible in the case of undoped NWs. The gP signal is characteristic of free conduction band electrons induced by the n-type doping of crystalline Si. 52,60 In the spectra of both the doped and undoped SiNWs, the signal located at g = 2.006, noted gSi, is attributed to defects at the interface between Si and SiO 2 . 53 The intensity of the gP band, visible in the ESR signal of all doped SiNWs, grows with P content and reaches a plateau at 1%P. The ESR measurements do not allow for a more quantitative determination of the n-type doping because both the dopant concentration and the SiNW diameter change with P content, and therefore the gP and gSi signals change simultaneously. Nonetheless, the gP/gSi signal intensity ratio quali- tatively follows the P content (Fig. 3B inset). Furthermore, the ESR cavity was increasingly difficult to tune with increasing doping level due to resonance with the SiNW samples of higher conductivity.

Electrical conductivity (σ) was measured on pressed pellets of SiNWs (compacity >80%) in a 2-probe configuration as a function of P content at room temperature (Fig. 3C). As the oxide layer was not etched, bulk conductivity is low (max. 4 × 10 -3 S cm -1 ) as compared to the conductivity of single doped SiNWs reported in the literature 43,61 (1-10 S cm -1 ). However, σ shows an impressive 3-order of magnitude increase from undoped to highly doped SiNW pellets. Two parameters contribute synergistically to this tremendous conductivity change: the increase of the doping level and of the NW diameter. As discussed earlier, undoped and low-doped SiNWs (highlighted in the shaded area) have smaller average diameters around 10 nm, corresponding to a crystalline core of around 8 nm (Table 1). Such small diameters induce a strong reduction of the charge carrier concentration by at least a factor of 10 due to depletion via proximate surface traps. 22,62 As a result, these SiNWs are essentially insulating as shown by the steep σ drop in the low-temperature conductivity measurements (Fig. 3D).

By contrast, highly doped SiNWs show reduced depletion due to a 4-fold increase in diameter (Fig. 2C). However, this factor alone cannot explain the observed strong increase in conductivity, which can only be understood by considering the additional contribution arising from the increase of the free carrier concentration induced by n-type doping. The small variation of σ at low temperature, where σ scales with T 0.7 , may suggest dopant-to-dopant hopping of electrons in the case of highly doped 0.4%P-and 1%P-SiNWs. Taken together, these results also demonstrate that the amorphous shell observed around doped SiNWs does not impede electrical contact between them.

One application benefitting strongly from both the dopinginduced enhanced conductivity and the inherent very high developed surface of 1%P-SiNWs are supercapacitors (SCs). To demonstrate the potential of the obtained materials in this application, we implemented them as electrodes in symmetrical SC devices (cf. Experimental section). Fig. 4 shows the electrochemical performance of these devices by using cyclic voltammetry (CV), galvanostatic charge-discharge (GCD) cycles and electrochemical impedance spectroscopy (EIS) techniques. 37 triangles, 63 crosses, 64 empty diamonds, 65 filled squares, 18 and empty squares. 66 a rectangular shape without any redox peak, which denotes an excellent electrochemical double layer capacitive behavior. This tendency is constant at all scan rates from 10 to 0.01 V s -1 (Fig. S9 †). The average capacitive current grows linearly with the scan rate (Fig. 4B) resulting in an areal capacity (AC) of 0.25 mF cm -2 . A slight distortion of the rectangular shape CV curve was attributed to the presence of the amorphous Si layer (SiO x ) (cf. Table 1 andS1 †), which was found in good agreement with the reported behavior of SiNW-based supercapacitors. 66 Nevertheless, the major energy storage mechanism is electrochemical double layer capacitance, i.e. adsorption/desorption of ions during charge/discharge. Accordingly, both 1%P-and undoped SiNWs exhibited symmetric and linear GCD profiles (Fig. 4C) with a similar small distortion. The areal capacitances derived from GCD profiles at current densities 0.1 to 5 mA cm -2 (Fig. S10 †) confirm the high AC obtained by CV for 1%P-SiNWs, with 0.23 mF cm -2 at 4.5 mA cm -2 (Fig. 4D). We note that 1%P-SiNWs show 2 to 4 times higher AC than undoped SiNWs, despite their 4 times lower specific area. As the capacity of an ideal conductive material is proportional to its specific area, 21,65 a 4 times lower AC could be expected for doped SiNWs. The observed high gain in capacity is therefore attributed to their strongly enhanced conductivity, reducing ohmic losses in the thick SiNW electrode.

Accordingly, doped SiNWs exhibited AC values ranging from 740 to 245 µF cm -2 , whereas undoped SiNWs displayed AC values ranging from 255 to 120 µF cm -2 (Fig. 4D). AC shows an important initial decay reaching a plateau from 2.2 mA cm -2 with an AC value of 0.25 mF cm -2 for 1% P-SiNWs, in excellent agreement with the CV data (Fig. 4B). The EIS technique is a powerful tool to evaluate the electrodeelectrolyte interface of a SC device. The Nyquist plot (Fig. 4E) shows two regions at high and low frequencies. At high frequencies, the semi-circle associated with the charge transfer resistance (R ct ) is smaller for 1%P-SiNWs, as expected from their higher conductivity. In this direction, at low frequencies, 1%P-SiNWs showed a more vertical line than undoped SiNWs, indicating better capacitive behavior. Besides, the impedance phase angle of doped SiNWs reached a value of approximately 70°at low frequencies (Fig. 4F), closer to the value of an ideal capacitor (90°) than undoped SiNWs (60°).

Long-term stability, another critical factor in SCs, was tested for 1%P-SiNWs (Fig. 4G) at a current density of 0.57 mA cm -2 . The capacitance decayed by ∼10% in the first 10 000 cycles owing to the chemical reactions of the native silicon oxide layer and the possible presence of impurities in the electrolyte as previously demonstrated for CVD-grown doped SiNW 64 and Si-nanotree 37 based SCs. During further cycling the surface reached passivation and the capacitance was stable at >80% of its initial value for (at least) one million cycles, an excellent stability performance according to the lifetime criteria for supercapacitors. 67 In addition, no structural degradation of the SiNWs after the cycling test was observed according to the SEM image depicted in the inset of Fig. 4G. The Ragone plot in Fig. 4H shows energy density values of 0.47 and 1.44 mJ cm -2 at power densities of 3.9 and 2.8 mW cm -2 , respectively. These values are higher than those reported in the literature for SC applying doped SiNWs obtained by other methods on a much smaller scale (e.g. CVD) and come close to the performance of the best reported system (CVD grown, doped Si nanotrees). 18,37,[63][64][65][66]

Conclusion

This work provides a novel chemical synthesis method for preparing SiNWs of a small diameter (<50 nm) and controlled doping level with phosphorus (up to 1%). The use of a sacrificial high-surface-area substrate (NaCl microcrystals) in combination with elevated precursor loading results in a high-yield process (1 mg SiNWs per cm 3 reactor volume), which can be easily scaled up as demonstrated by the use of a 150 mL reactor. Applying complementary characterization techniques, we show that successful n-type doping of the crystalline SiNW core is achieved. Phosphorus doping leads also to an unexpected increase of the SiNW diameter. Both effects have a synergistic impact on electrical conductivity, which sharply switches from mainly resistive at a low doping level to conductive at a doping level above 0.4%P. Symmetric supercapacitors based on electrodes fabricated with 1%P-SiNWs show a high areal capacity of 0.24 mF cm -2 combined with an excellent cycling stability (>1 million cycles), placing them at the top of the state of the art in the field of SiNW-based SCs. This performance is particularly significant considering that the present SC electrodes, made from a SiNW slurry deposited on a standard Cu current collector, lack electrical connection of the individual SiNWs. The latter is ensured in reported SCs using CVD-grown SiNWs, which have an epitaxial contact to the Si collector. To take full advantage of the good conductivity of this nanomaterial, improvements of the electrical connection between individual nanowires are required to reduce ohmic losses in macroscopic SiNW assemblies. This can be achieved by modifying the outer shell of the SiNWs. We expect that the presented approach opens the way for the use of SiNWs in a wide range of electrochemical storage devices such as SCs and lithium-ion batteries, as well as in energy conversion devices such as thermoelectric generators.

Fig. 1

 1 Fig. 1 Scanning electron microscopy (SEM) (A and B), transmission electron microscopy (TEM) (C and D) and high angle annular dark field scanning TEM (HAADF-STEM) (E) images of undoped (A, C and E) and 1% P-(B and D) SiNWs. D inset: enhanced details showing the atomic plane distance in the crystalline core.

Fig. 2 A

 2 Fig. 2 A: Histograms of SiNW diameters measured by SEM. B: Raman spectra of undoped (black), 0.2%P (green) and 0.8%P (red) SiNWs. C: Correlation between the diameter and characteristics of the Si LO-TO Raman band as a function of doping level: average diameter (dots), Raman shift (triangles) and Raman peak full width at half maximum FWHM (squares) versus initial dopant concentration. (standard deviation of diameter represented as bars). D: Thermogravimetry (TG, thick lines) and TG effluent mass spectroscopy (MS, thin lines) of undoped (crosses) and 1%P-SiNWs (circles) under Ar, as a function of temperature. Mass m/z = 78 corresponds to benzene.

Fig. 3 P

 3 Fig. 3 P content and conductivity in doped SiNWs. A: Quantitative EDX (dots) and ICP-AES (triangles) measurements of the P content in SiNWs as a function of the initial P/Si ratio in the reagents, demonstrating quantitative P incorporation (EDX spectra in Fig. S6 †). B: ESR spectra of undoped (thin black line) and 0.4%P (bold red line) SiNWs. The line at g = 2.006 (noted gSi) arises from defects in crystalline Si at the Si-SiO x interface; the line at g = 1.998 (noted gP) arises from conduction band electrons in doped Si. Inset: Variation of the intensity ratio gP/gSi as a function of P content in the reagents (Fig. S8 †). C: Electrical conductivity at room temperature of doped (filled squares) and undoped (empty squares) SiNW pellets versus P content. The shaded zone shows samples with a SiNW average diameter <12 nm. D: Electrical conductivity versus temperature for undoped (black dots), 0.05%P (blue down triangles), 0.2%P (blue up triangles), 0.4%P (brown crosses) and 1%P (red squares) SiNW pellets.

Fig. 4

 4 Fig. 4 Electrochemical performance supercapacitors applying 1%P-SiNWs (red line/filled diamonds) compared to undoped SiNWs (black line/ empty circles). (A) CV curve at a scan rate of 1 V s -1 and (B) average capacitive current density versus scan rate of 1%P-SiNWs. (C) Galvanostatic charge-discharge cycles at a current density of 1.13 mA cm -2 and (D) areal capacity versus current density for both systems. (E) Nyquist plot with a magnified region at high frequency, and (F) Bode plot. (G) Cycling stability over 10 6 GCD cycles at a current density of 0.57 mA cm -2 . The inset shows a SEM image of 1%P-SiNWs after the cycling test. (H) Ragone plot (P vs. E) of 1%P-SiNW-supercapacitors of this work (red diamonds) compared to other state-of-the-art SiNW-based SCs: stars,37 triangles,63 crosses,64 empty diamonds,65 filled squares,18 and empty squares.66

Table 1

 1 Top: BET surface area of undoped and 1%P-SiNWs; the expected surface area calculated from SEM diameter histograms Fig.2Ais given for comparison. Bottom: Average diameter of the crystalline core and thickness of the amorphous shell as measured by TEM

	SiNWs	Undoped	1%P
	Surface area (m 2 g -1 )
	BET	230	43
	Calculated from SEM	170	41
	Size by TEM (nm)
	Outer diameter	9.1 ± 2.4	26 ± 7
	Crystalline core diameter	7.4 ± 2.0	18 ± 6
	Amorphous shell thickness	0.9 ± 0.3	4.6 ± 2.2

Nanoscale

This journal is © The Royal Society of Chemistry 2019

This journal is © The Royal Society of Chemistry 2019 Nanoscale

Acknowledgements

OB, PC and PR thank the DSM Energie program of CEA for funding. This work has been performed with the use of the Hybrid-En facility at CEA-Grenoble (France). The authors thank Thibaut Gutel in BET measurements, Eric Robin in STEM-EDX analysis, Julien Pérard in ICP-AES analysis and Stéphanie Pouget in X-ray diffraction for their help.

Conflicts of interest

There is no conflict of interest.


domain: chim sdv phys

text:
Mans Broekgaarden

Anne-Laure Bulin

Estelle Porret

Benjamin Musnier

Benoit Chovelon

Corinne Ravelet

Lucie Sancey

Pierre Hainaut

Hélène Elleaume

Jean-Luc Coll

Xavier Le Guével

Surface functionalization of gold nanoclusters with arginine: a trade-off between microtumor uptake and radiotherapy enhancement

Ultra-small gold nanoclusters (AuNCs) are increasingly investigated for cancer imaging and radiotherapy enhancement. While inetuning the AuNC surface chemistry can optimize their pharmacokinetics, its effects on radiotherapy enhancement remain largely unexplored. This study demonstrates that optimizing the surface chemistry of AuNCs for increased tumor uptake can significantly affect its potential to augment radiotherapy outcomes.

Radiotherapy enhancement strategies are critically needed for the management of various cancer types. For glioblastoma, radiation regimens are challenging to design: High radiation doses are needed to control tumor growth, yet applicable radiation doses are limited to remain tolerable to surrounding healthy brain tissue 1 . Strategies that render cancer tissues more susceptible are highly desired to make lower radiation doses more effective, so that effective cancer management can be achieved with minimal damage to healthy tissues.

Ultra-small gold particles called nanoclusters (AuNCs) are being increasingly investigated for cancer imaging and have potential to augment radiotherapy 2,3 . AuNCs are typically composed of ten to hundred gold atoms corresponding to a metal core smaller than 3 nm which can be stabilized in solution by a variety of biomolecules such as peptides, proteins or DNA [3][4][5] . This flexibility in terms of core size, ligand coverage, and functionalization can be used to tune their photoluminescence in the near-infrared (NIR, 700-900 nm) 6 and shortwave infrared (SWIR, 1000-1700 nm) 7,8 for in vivo imaging and optical detection of cancer 9 , blood vasculatures 8 , and also for fluorescence-guided surgery 10 .

As Au is a high Z-atom (ZAu = 79), it is capable of absorbing orthovoltage (<250 keV) X-rays more efficiently than water and soft tissue [START_REF] Hubbell | X-Ray Mass Attenuation Coefficients[END_REF][START_REF] Butterworth | [END_REF] . By itself, radiotherapy induces DNA damage, either by direct one-electron damage to DNA bases, or by the radiolysis of water to produce highly cytotoxic hydroxyl radicals ( • OH) 13 . However, the presence of Au in tumor tissues can result in high degrees of radiation absorption, after which the energy is locally deposited to enhance radiation-induced tissue damage by increased generation of • OH 14,15 . Hainfeld et al., were the first to demonstrate the augmentation of 250 kVp radiotherapy, yielding greatly improved survival rates in syngeneic mouse models of breast cancer 16 . Similarly, 2 to 3fold improved reductions in tumor weight were achieved by AuNC-augmented radiotherapy (662 keV) in mice bearing subcutaneous ovarian cancer [17][18][19] .

Our group recently demonstrated that zwitterionic sulfobetaine stabilized AuNCs have the capacity to accumulate in brain tumors 20 . To further improve tumor uptake, we developed AuNCs functionalized with arginine (AuSG-2Arg) 21 , which demonstrated rapid accumulation in cancer cells, making them potentially interesting for radiotherapy enhancement.

This study specifically explored the radiotherapy enhancement two AuNCs with distinct surface chemistries, namely glutathione-stabilized AuNCs (AuSG), and the novel arginine-stablized AuNCs (AuSG-2Arg). The theranostic properties of these AuNCs were evaluated using 3D culture models of glioblastoma, as these more faithfully recapitulate cancer growth. We demonstrate for the first time that, despite achieving an 11-fold increase in tumor accumulation, AuSG-2Arg are less efficient at augmenting radiotherapy compared to the AuSG. Modifying the surface chemistry of AuNCs thus strongly influences their radiotherapeutic potential. These findings are critical to the design of novel AuNCs for imaging and cancer therapy.

AuSG and AuSG-2Arg nanoclusters (Fig. 1) were prepared by a bottom-up approach as described previously 21 . Suspended spheroids were established in ultra-low-adhesion plates from human U87MG and rat F98 glioblastoma cell lines. Please do not adjust margins Please do not adjust margins Radiotherapy was given using monochromatic 50 keV synchrotron radiation. Multiparametric assessment of radiotherapy outcomes was performed as described previously 22 . Assessment of necrosis was based on cell permeability using propidium iodide staining. Oxidative stress was evaluated using the dichlorodihydrofluorescein (DCFH2-DA) method 23 . A detailed Materials and Methods section is provided in the Supplementary Information. The physicochemical characteristics of the AuNCs are reported in Fig. S1. High resolution transmission electron microscopy of AuSG and AuSG-2Arg showed core sizes between 2 and 4 nm with semi-crystalline structure. The HD of both AuNCs were 1.66 ± 0.01 nm for AuSG and 2.02 ± 0.02 nm for AuSG-2Arg. The zeta potentials at pH 7 were ζ =-31.05 ± 1.12 mV for AuSG and ζ = 7.96 ± 0.44 mV for AuSG-2Arg as expected after addition of Arg. The AuNCs have similar absorption spectra with gradually declining intensities in the range of 400-900 nm. Fluorescence emission profiles are also comparable between the AuNCs, with the AuSG-2Arg exhibiting an increased emission intensity around 670 nm (Fig. S1).

Next, the acute (12 h) and prolonged (96 h) toxicity of the AuNCs was evaluated on U87MG and F98 glioblastoma spheroids. The results depict that both AuNCs exerted minor toxicity (<10% reduction in viability, 5% increase in necrosis) at concentrations exceeding 100 µg Au/mL, most notably in F98 spheroids (Fig. S2). Thus, the AuNCs were generally welltolerated and all subsequent experiments were performed using an intrinsically non-toxic dose of 100 µg Au/mL. We also evaluated the the AuNC toxicity on non-malignant endothelial cells and mesothelial cells, as these are the first tissues exposed to AuNCs during either intravenous or intraperitoneal administration. The results demonstrate that the particles were well tolerated in both cell types (Fig. S3).

The uptake of the AuNCs was evaluated using inductivelycoupled plasma mass spectrometry (ICP-MS), NIR-, and SWIR fluorescence imaging. The ICP-MS results indicate that there was a significant, 4-fold (U87MG) and 13-fold (F98) increase in uptake of AuSG-2Arg as compared to AuSG (Fig. 2A, Fig. S2 andS3). The uptake of both AuNCs typically reached a plateau after 12 h of incubation. Semi-quantitative NIR fluorescence imaging confirmed similar kinetics of AuNCs uptake for both spheroid types (Fig. 2B, Fig. S4 andS5). Fluorescence intensity plots of the NIR images revealed that AuSG-2Arg accumulated predominantly in the first few cell layers of the spheroids, with fluorescence intensity values reaching background levels at approximately 100 µm from the edge of the spheroid at 2 h and 12 h incubation times (Fig. 2D, Fig. S4). In contrast, AuSG also accumulate in the first few cell layers after 12 h of incubation, but their fluorescence emission was also detected deeper inside the spheroids (Fig. 2D). After 24 h incubation, both the AuSG-2Arg and AuSG nanoclusters demonstrated peak intensities at the spheroid periphery (Fig. S4). Similar findings were observed for the F98 spheroids (Fig. S5). As AuNCs exhibit broad fluorescence emission in the near-infrared, it was also possible to detect their presence at longer wavelengths in the SWIR window and confirm a similar trend as compared to the NIR imaging for AuSG-2Arg, and to a lesser extent for AuSG (Fig. S4 andS5). Lastly, we found that while AuSG exhibited a relatively homogeneous sub-and intercellular distribution throughout the spheroid periphery, whereas AuSG-2Arg appeared to form extracellular aggregates (Fig. S6). We next investigated the radiosensitization of the AuNCs in human (U87MG) and rat (F98) 3D cultures of glioblastoma. The X-ray absorption of Au is well-characterized to peak at an energy of 81 keV (k-edge), although the highest absorption difference with water is at 50 keV [START_REF] Hubbell | X-Ray Mass Attenuation Coefficients[END_REF] . Spheroids were incubated with AuSG-2Arg or AuSG (100 µg Au/mL) for 12 h, after which free AuNCs were washed away. Immediately thereafter, the glioblastoma spheroids were exposed to different doses of 50 keV monochromatic synchrotron radiation.

In U87MG spheroids, we demonstrate a significant dose-dependent inhibition of U87MG growth (Fig 3A -B), which was not influenced by the AuNCs. Multiparametric treatment analysis revealed that U87MG spheroid viability decreased in a radiation-dose dependent manner, and the radiotherapy Please do not adjust margins Please do not adjust margins efficacy was significantly improved by AuSG (1.4-fold), but not by AuSG-2Arg (Fig. S7). Although radiotherapy, through its induction of wide-spread DNA damage, typically associates with reduced cancer cell proliferation and apoptosis 24 , we demonstrate that both AuSG and AuSG-2Arg induced a radiation-dose-dependent increase in radiotherapy-induced necrosis compared to spheroids receiving radiotherapy alone (Fig. 3C). The dose response fits based on spheroid necrosis indicated a 1.8-and 1.5-fold increase in efficacy for AuSG and AuSG-2Arg, respectively. By comparatively investigating the and level of necrosis of each spheroid, it can be seen that radiotherapy with AuSG and AuSG-2Arg results in smaller masses with higher degrees of necrosis compared to radiotherapy alone (Fig. S5). In the F98 spheroids, quantification of the spheroid viabilities and levels of necrosis indicated that there was also a significant improvement in the radiotherapy efficacy following AuSG exposure (1.9-fold and 1.5-fold based on viability and necrosis, respectively), but not for AuSG-2Arg (Fig. S6).

These findings were surprising given the substantially elevated levels of AuSG-2Arg in the spheroids compared to AuSG. When assuming that the spheroids are spherical with a diameter of 500 µm, we estimated Au concentrations after a 12 h incubation in the U87MG spheroids in the order of 20-80 µg gold/mL for AuSG and AuSG-2Arg. Our findings are therefore unlikely to be explained by a bona-fide dose-enhancement effect, which requires far higher intratumor concentrations of gold in the order of mg/mL 25 . Our results appear best supported by the findings of Cheng et al., who reported that Au nanoparticles could act as chemical catalysts of ROS production during radiotherapy [START_REF] Butterworth | [END_REF]26 . Inspection of oxidative stress levels in the U87MG spheroids showed that radiotherapy induced sustained oxidative stress, as measured 5-days post-treatment. However, this was not elevated in the presence of the AuNCs. In contrast, the extent of oxidative stress in the AuSG-2Arg group was significantly lower compared to the control and AuSG groups (Fig. 4). Further investigations demonstrated that AuSG elevates basal levels of oxidative stress in both U87MG and F98 cells, whereas AuSG-2Arg was only able to do this in U87MG cells, and to a lesser extent compared to AuSG (Fig. S11 Recent works have demonstrated that AuNCs have high potential as theranostic agents for cancer imaging and radiosensitization. Despite exhibiting promising tumor retention 9 , their exploitation is challenged by their suboptimal pharmacokinetic profiles 10,20 . To improve this, we and others continue to develop novel functionalized AuNCs. However, the theranostic evaluation of emerging AuNCs with 2D cultures have often produced promising results that failed to be effectively translated to in vivo studies. To better br idge the gap between in vitro and in vivo research, this study focused specifically on the use of 3D culture models, as these more effectively the 3D architecture and treatment responses of cancer, as further discussed in the Supplemental Information.

Encouraged by the high uptake of AuSG-2Arg compared to AuSG in 2D cultures 28 , this study explored the therapeutic potential of both AuNCs as radiosensitizing agents for glioblastoma. We report notable enhancement in radiotherapy efficacy by AuSG, and to a lesser extent by AuSG-2Arg, which was characterized by an increased necrotic response. The findings suggest that, despite weak uptake, the AuSG are efficient radiosensitizers. In contrast, the AuSG-2Arg are weaker radiosensitizers than AuSG but more efficient diagnostic agents given their enhanced uptake. The reduced radiotherapy efficacy This journal is © The Royal Society of Chemistry 20xx

Please do not adjust margins Please do not adjust margins of AuSG-2Arg compared to AuSG may stem from its formation of extracellular aggregates, and its reduced capacity to augment ROS-induced oxidative stress. In addition, the AuSG appeared more effective to perfuse throughout the tumor compared to AuSG-2Arg (Fig. 2), potentially leading to a more homogeneous amplification of the radiotherapeutic effects throughout the microtumors by AuSG compared to AuSG-2Arg.

Our findings corresponded relatively well to previous studies, achieving a similar degree of radiotherapy augmentation with AuSG [17][18][19] . While the novel AuSG-Arg have strong potential for diagnostic imaging, their limited radiotherapeutic potential did not encourage further in vivo studies. An extended discussion on the mechanisms of radiotherapy enhancement by high-z elements is provided in the Supplemental Information. The complex interactions of AuNC on cancer tissues and their theranostic potential can thus be effectively screened in 3D culture models, benefiting from the relative high-throughput of in vitro models while more faithfully recapitulating in vivo cancer tissues. The development of novel nanoclusters may thus benefit from the use of 3D cultures and emerging high-content assays for such models 22 .

Conclusions

Functionalization with arginine moieties greatly improves the uptake of AuNCs, enabling near-and shortwave infrared imaging with high sensitivity. Functionalization with glutathione is more suitable for radiotherapy enhancement, capable of achieving a 2-fold enhancement in overall efficacy. Our findings stress the importance of careful design and optimization of the surface chemistry of AuNCs for future radiotherapy applications. 3D cultures combined with high-content imaging assays are highly suited to evaluate the theranostic properties of AuNCs, enabling accurate screening of promising particles for further in vivo studies.

Figure 1 .

 1 Figure 1. Schematic overview of the AuSG and AuSG-2Arg.

Figure 2 .

 2 Figure 2. Arginine-functionalized gold nanoclusters are efficiently taken up by U87MG glioblastoma spheroids. (A-B) Quantification of AuNCs uptake by ICP-MS (A), and NIR confocal fluorescence microscopy (B). (C) NIR fluorescence images of AuNCs in U87MG spheroids following 12h of incubation. AuNC fluorescence is depicted in yellow, scalebar = 200 µm. (D) Fluorescence intensity profiles obtained from the radius of the spheroids depicted in panel C. Data represents the mean ± SEM (N ≥ 6).

Figure 3 .

 3 Figure 3. Multiparametric evaluation of 50 keV radiation dose enhancement by AuNCs in U87MG glioblastoma spheroids. (A) Live/dead heatmaps displaying radiotherapy dose-response effects on U87MG spheroid viability. Scalebar = 200 µm. (B) Effect of 50 keV radiotherapy on U87MG spheroids growth (logarithmic growth fits). (C) Radiation dose response fits U87MG spheroids necrosis following radiotherapy alone (black), radiotherapy + AuSG (blue), and radiotherapy + AuSG-2Arg (green). Data is the mean ± SEM of N ≥ 12.

  C&F). The findings overlap well with the radiotherapy efficacy findings in both spheroid types (Fig 3 & S7). When incubated with Fe 2+ to generate • OH 27 , i.e., the same ROS produced during radiotherapy, we found significantly lower levels of oxidative stress in cells exposed to AuSG-2Arg compared to AuSG (Fig. 11 D&G).

Figure 4 .

 4 Figure 4. Detection of intracellular ROS following radiotherapy reveals reduction in radiotherapy-induced oxidative stress by AuSG-2Arg. Fluorescence microscopy images and quantification of DCF-emission of 8 day-old U87MG spheroids incubated with DCFH2-DA following radiotherapy (5 days post-treatment). Scalebar = 200 µm. Data represents the mean ± SEM (N ≥ 6).

Notes and references

This work was supported by Plan Cancer 2014-2019 (C16080CS & C18038CS), Cancéropôle Lyon Auvergne Rhône-Alpes, Fondation ARC (R17157CC), Region Rhône Alpes (Pack Ambition Recherche 2017), Fondation pour la Recherche Médicale.

Conflicts of interest

The authors have no conflicts to declare.