Supporting Information

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1 Supporting Information One-Dimensional Fluorescent Silicon Nanorods Featuring Ultrahigh Photostability, Favorable Biocompatibility and Excitation Wavelength-Dependent Emission Spectra Bin Song, Yiling Zhong, Sicong Wu, Binbin Chu, Yuanyuan Su, and Yao He * Jiangsu Key Laboratory for Carbon-Based Functional Materials and Devices, Institute of Functional Nano & Soft Materials (FUNSOM) and Collaborative Innovation Center of Suzhou Nano Science and Technology (NANO-CIC), Soochow University, Suzhou, Jiangsu , China yaohe@suda.edu.cn S1

2 1.1 Experimental methods Materials and devices (3-Aminopropyl) trimethoxysilane (97%) was purchased from Sigma-Aldrich. Trisodium citrate ( 99.0%) was purchased from Sinopharm Chemical Reagent Co., Ltd (China). All chemicals were used without additional purification. Fresh milk was supplied by local markets. All solutions were prepared using Milli-Q water (Millipore) as the solvent. The microwave system NOVA used for synthesizing materials was made by Preekem of Shanghai, China. UV-vis absorption spectra were recorded with a Perkin Elmer Lambda 750 UV-vis-near-infrared spectrophotometer. Photoluminescence (PL) measurements were performed using a HORIBA JOBIN YVON FLUOROMAX-4 spectrofluorometer. High-resolution X-ray photoelectron spectroscopy (XPS) analyses were performed using a Kratos AXIS Ultra DLD ultrahigh vacuum (UHV) surface analysis system, which consists of a fast entry air lock (base pressure < Torr), a multiport carousel chamber (< Torr), a deposition chamber (< Torr), and an analysis chamber (< Torr). A monochromatic Al Kα source ( ev) with a resolution of 0.1 ev was used to irradiate the samples. XPS samples were prepared by drop-casting SiNRs ( 2 mg) onto aluminum substrates and degassing at 10-7 Torr for 15 hours prior to analysis. FTIR measurement, KBr was pressed into a slice, onto which the SiNRs sample was dropped. FTIR spectra were recorded on a Bruker HYPERION FTIR spectrometer and cumulated 32 scans at a resolution of 4 cm -1. A Raman microscope (HR800) equipped with a 633 nm He-Ne 20 mw laser (polarized 500:1) was employed for obtaining the Raman spectra. The S2

3 acquisition and analysis of Raman data were performed by using the LabSpec5 software. The powder X-ray diffraction (XRD) spectra were recorded on a PANalytical, Empyrean, X-ray diffractometer, operated at 40 ma and 40 kv. The materials were placed dropped on zero-background sample holder made of monocrystal silicon plate. Energy-dispersive X-ray (EDX) spectroscopy was utilized to determine the fraction of the resultant materials. TEM and HRTEM samples were prepared by dispersing the sample onto carbon-coated copper grids with the excess solvent evaporated. The TEM/HRTEM overview images were recorded using Philips CM 200 electron microscope operated at 200 kv. Time-resolved fluorescence decay curves were attained on HORIB-FM-2015 spectrofluorometer using 370 nm lasers as the excitation source. White-LED characteristics was measured by a constant current source (Keithley 2400s Source Meter) combined with a photometer (Photo Research PR 655 spectrophotometer) and an integrating sphere (LED300 + HAAS2000_V1_USB). TGA analysis was performed on a METTLER TOLEDO TGA/STDA 851 instrument. The samples, which ranged in weight from 5 to 12 mg, were placed in a porcelain crucible and heated under air atmosphere from 30 to 800 C at a rate of 10 C/min. Multi-color bioimaging was examined under a confocal laser micro-scope (Leica, TCS-SP5) equipped with diode laser 405 nm, 458 nm and 514 nm. Images were captured and processed with image analysis software. S3

4 1.2 Microwave synthesis of SiNPs Fluorescent SiNPs were readily achieved through microwave synthesis based on previous reports. 1 In details, The SiNPs precursor solution was prepared by adding 1mL (3-aminopropyl) trimethoxysilane to 8 ml N 2 -saturated aqueous solution dispersed with g trisodium citrate dihydrate. The mixture was stirred for 10 min. The resultant precursor solution was transferred into the exclusive vitreous vessel with a volume of 30 ml. The SiNPs with the maximum emission at ~460 nm were prepared under 150 C/60 min. After microwave irradiation, the SiNPs sample was removed when the temperature cooled to <30 C naturally. To exclude impurities influence, such as (3-aminopropyl)-trimethoxysilane molecules and trisodium citrate dihydrate in solution, the residual reagents were removed by dialysis (1 kda). The purified SiNPs aqueous solution was used for characterizations. 1.3 Microwave synthesis of carbon NPs Fluorescent carbon NPs were facilely prepared via microwave-assisted synthetic strategy using our previously reported protocol. 2 In details, 2 mg milk was firstly put into the exclusive vitreous vessel and heated to 150 C for 60 min. When cooled down to room temperature, the solution was centrifuged at rpm for three times to fully remove impurities and larger particles, and then the supernatant was collected. The obtained red-brown and foamy solid was dialyzed using pure water through a dialysis membrane (1 kda), then further filtered with ultra-filtration membrane (220 S4

5 nm). The final products were collected for characterizations, which were noted as carbon NPs. 1.4 Photoluminescent quantum yields (PLQY) measurements PLQY, well considered as an important factor for quantitatively evaluating fluorescent intensity of materials, equals to the ratio of the number of emitted photons and the number of absorbed photons. In our experiment, a well-established reference method was employed for determining the PLQY value of SiNRs as follow (Quinine sulfate in 0.1 M H 2 SO 4 (literature quantum yield: 58%)). 1 Φx=Φst (Ix/Ist ) (ηx /ηst ) 2 (Ast /Ax) Where Φ is the QY, "I" is the integrated emission intensity, "η" is the refractive index of the solvent, and A is the optical density. The subscript "st" and "x" stand for standard with known QY and the SiNRs sample, respectively. To minimize reabsorption effects, absorption value at the excitation wavelength is required to be smaller than The MTT assay of cell viability HeLa (Henrietta Lacks) cells (in H-DMEM medium) were dispersed in 96-well plates (100 μl in each well containing cells per well). Serial concentrations (0.125, 0.25, 0.5, 1, 2 mg/ml) of ~140 nm-length SiNRs solutions as that used in the following cellular imaging were added to each well (10 μl). Incubation was carried out for 0.5, 3, 6, 12, 24 and 48 h in a humidified atmosphere at 37 C with 5% CO 2. S5

6 The cytotoxicity of the SiNRs was evaluated by the MTT (3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide) assay (Thiazolyl blue tetrazolium bromide (M5655); Sigma). The assay was based on the accumulation of dark blue formazan crystals inside living cells after exposure to MTT, which is well-established for assessment of cellular viability. The destruction of cell membranes by the addition of sodium dodecylsulfate (SDS) resulted in the liberation and solubilization of the crystals. The number of viable cells was thus directly proportional to the level of the initial formazan product created. The formazan concentration was finally quantified using a spectrophotometer by measuring the absorbance at 570 nm (ELISA reader). A linear relationship between cell number and optical density was established, thus allowing an accurate quantification of changes in the rate of cell proliferation. For the SiNRs with different length of ~100, ~140, ~180 and ~250 nm, all the manipulations were identical to those as mentioned above. S6

7 2. Additional data Table S1 shows comparisons between silicon nanorods (SiNRs) and silicon nanoparticles (SiNPs). Figure S1 shows TEM images and fluorescence spectra of the prepared carbon nanoparticles and SiNPs. Figure S2 shows TEM images of the SiNRs. Figure S3 shows Raman spectra of the SiNRs. Figure S4 shows EDX pattern and EDX elemental maps of the SiNRs. Figure S5 shows XPS spectra of the SiNRs. Figure S6 shows TGA profiles of the SiNRs. Figure S7 shows PLQYs measurements of the SiNRs. Figure S8 shows PL decay curves of the prepared SiNRs. Figure S9 shows fluorescence spectra of the SiNRs under different excitation wavelength. Figure S10 shows cell viability of HeLa cells treated by the SiNRs with different lengths for different incubation time. Figure S11 shows EL spectra of SiNRs-based white LED under various currents. S7

8 Table S1. Comparisons between silicon nanorods (SiNRs) and silicon nanoparticles (SiNPs). SiNPs (Ref. 1., i.e., J. Am. Chem. Features SiNRs (this work) Soc. 2013, 135, ) Structure Zero dimension One dimension Tunable lengths from ~100 to ~250 Size ~2.5-4 nm nm with the diameter of ~20 nm Emission property Single color (Blue emission) Selective emission peaks in a broad range over the visible region from ~450 nm to ~600 nm. Photoluminescent quantum yield (PLQY) ~20-25% ~15% Application Single-color cell imaging Multi-color bioimaging in vitro and in vivo; light-emitting diodes (LED) In summary, compared with published work introducing zero-dimensional fluorescent SiNPs for single-color cell imaging application, 1 this current work, for the first time, presents a kind of biocompatible fluorescent one-dimensional SiNRs showing robust photo-stability and excitation wavelength-dependent emission. Taking advantage of unique optical properties and feeble toxicity, we further explore the prepared SiNRs as biocompatible fluorescent labels for multi-color bioimaging in vitro and in vivo, and present the first proof-of-concept LED device fabricated using the one-dimensional fluorescent silicon nanostructures as new color converters. These results would open exciting avenues for the design of one-dimensional silicon nanostructures featuring novel optical functionality, potentially promoting the long-awaited silicon-based optical applications. S8

9 Figure S1. (a, b) TEM images and (c, d) fluorescence spectra of the prepared carbon nanoparticles (a, c) and silicon nanoparticles (b, d). Insets in (a) and (b) represent corresponding HRTEM images. As a control, C 6 H 17 NO 3 Si molecules or pure milk is respectively used as reaction precursor alone, which can only produce SiNPs or carbon NPs instead (Please see experiment details in Experimental methods section). TEM and HRTEM images show that the prepared SiNPs assume a spherical structure with good monodispersity, with lattice planes (220) of 0.19 nm spacing in the HRTEM image, revealing high crystallinity of the resultant SiNPs. 1 As shown in Figure S1 (c), while the carbon NPs exhibit excitation wavelength-dependent emission spectra, the red-shift range only reaches 70 nm (maximum emission wavelength ranges from ~490 nm to ~560 nm under continuous excitation from 400 to 510 nm, respectively, consistent with previously published results 2 ), much narrower than that (~120 nm) of the SiNRs. Moreover, for the fluorescent SiNPs featuring excitation-independent emission behavior, the maximal emission wavelength maintains ~495 nm in spite of serial excitation wavelengths ranging nm (Figure S1 (d)). S9

10 Figure S2. (a-d) TEM images of SiNRs with different length. As presented in Figure S2 (a-d), the resultant SiNRs show distinct rod-shaped structure with good monodispersibility. The TEM images also display that the length of the SiNRs is ~250 nm (Figure S2 (a), ~180 nm (Figure S2 (b)), ~140 nm (Figure S2 (c)) and ~100 nm (Figure S2 (d)), respectively. The aspect ratio of these SiNRs are systematically tuned from 12.5 to 4.9, simply by adjusting the milk concentration from 1 to 4 mg/ml (12.5 (1 mg/ml), 9.2 (2 mg/ml), 6.9 (3 mg/ml), 4.9 (4 mg/ml)). S10

11 Figure S3. Raman spectra of the prepared SiNRs with different length. Notably, a peak at 520 cm -1 confirms the presence of crystalline silicon. Moreover, the peaks at 1315 cm -1 and 1595 cm -1 are resolved to the D band (sp 3 -hybridized) and G band (sp 2 -hybridized) of carbon, respectively. And the peaks at 607 cm -1 and 963 cm -1 are ascribed to CaP. 3 S11

12 Figure S4. (a) EDX spectra and (b-e) EDX elemental maps of the prepared SiNRs with different length. To further analyze the chemical compositions of SiNRs with different length, the EDX analyses were performed, confirming the presence of Si, Ca, P and O. It is worthwhile to point out that, for EDX measurement, a quantitative analysis of the elemental ratios is not possible since the supporting substrate (e.g., carbon-coated copper girds) was carbon containing a measurable amount of residual oxygen. The SiNRs with aspect ratio of 6.9 are employed as a model; the EDX elemental maps of SiNRs confirm that the SiNRs contain Si, P and Ca element, as indicated by yellow (c), green (d) and blue (e) color, respectively. 4 S12

13 Figure S5. (a) XPS spectra and (b) C 1s, (c) Ca 2p and (d) P 2p spectra of the prepared SiNRs with different length. As shown in Figure S5 (a), five peaks at 103 ev, 133 ev, 286 ev, 350 ev and 533 ev (ascribed to Si 2p, P 2p, C 1s, Ca 2p and O 1s) are observed in the XPS spectrum, demonstrating the existence of Si, P, C, Ca and O elements in the SiNRs. In particular, Figure S5 (b) presents the typical C 1s spectrum, exhibiting four distinct peaks located at 285.7, 286.8, 287.5, and ev, which are ascribed to C-H, C-O, C-N, and C=O, respectively. The spectrum of Ca 2p is displayed in Figure S5 (c), in which 2p 1/2 and 2p 3/2 produce two peaks at and ev, respectively. For the P 2p spectrum, two peaks at and ev are observed, which are ascribed to the 2p 1/2 and 2p 3/2 (Figure S5 (d)), respectively. 5 S13

14 Figure S6. Thermal gravity analysis (TGA) of SiNRs with different lengths. Figure S6 gives the TGA thermograms of SiNRs under air from room temperature to 800 C. Rapid mass loss between 150 and 480 C is observed for as-prepared SiNRs, which is ascribed to the burning of carbonous residues. Comparatively, the as-prepared SiNRs exhibit a noticeable weight increase above 600 C resulted from Si oxidaetion (Si + O 2 SiO 2 ). 6 It is evident that the SiNRs obtained from higher concentration of milk yield greater weight losses. At 800 C, the residual weight percentage is 87.4% for 250 nm, 79.5% for 180 nm, 76.5% for 140 nm, and 71.4% for 100 nm. S14

15 Figure S7. PLQYs measurements of the SiNRs with different length. The solid lines represent the fitting results for each set of data. PLQYs of nanorods were calculated by using the equation: Φx=Φst (Ix/Ist ) (ηx /ηst ) 2 (Ast /Ax), 1 where φ is the QY, K is the slope determined by the curves and η is the refractive index. The subscript st refers to the standards and x refers to the unknown samples. For these aqueous solutions, η x /η st =1. The highest quantum yield of 15% is observed for the sample with the largest aspect ratio (with length of 250 nm), while the sample with length of 180 nm had a quantum yield of 13%. When the length decreases to 100 nm, the quantum yield went down to 9%. Because the milk content carbonaceous resource may bring about the generation of C-related impurity levels localized within the optical band gap of nanorods. The C-related impurity levels act as nonradiative recombination sites under the recombination processes of electron-hole pairs which lead to the decrease in the fluorescence QY. 7 S15

16 Figure S8. Time-resolved fluorescence-decay curves of SiNRs with different length (λ exitation =370 nm). The corresponding emission decay curves can be fit to a third-order exponential decay. The average lifetime was calculated according to ԏ av = (α 1 ԏ 2 1 +α 2 ԏ 2 2 +α 3 ԏ 2 3 )/ (α 1 ԏ 1 +α 2 ԏ 2 +α 3 ԏ 3 ). Under these conditions, the lifetimes of SiNRs with different lengths are ~6.1 ns (100 nm), ~6.9 ns (140 nm), ~7.2 ns (180 nm) and ~7.9 ns (250 nm), which is consistent with the time-scale (ns) of defect-related emission from silicon nanomaterials. Furthermore, such accelerated PL decay can be due to increased rates of either radiative recombination (induced by interactions between the semiconductor and the metal components) or nonradiative decay. 8 S16

17 Figure S9. Fluorescence spectra of the prepared SiNRs with different length under different excitation wavelengths. The resultant SiNRs samples exhibit various maximum emission wavelengths from ~450 to ~600 nm corresponding to different excitation wavelengths from 390 to 560 nm. 9 S17

18 Figure S10. Cell viability of HeLa cells treated with the SiNRs of different lengths (100 nm (a), 180 nm (b) and 250 nm (c)) for serial incubation time. The concentration of SiNRs is from to 2 mg/ml. The viability of the control groups is considered as 100%. Specifically, for the HeLa cells treated by the three-lengths SiNRs (i.e., 100 nm, 180 nm and 250 nm), the cell viabilities all preserve >90% under different SiNRs concentrations ( mg/ml) and incubation time ( h). These results indicate that the as-prepared SiNRs with different lengths exhibit negligible cytotoxicity. S18

19 Figure S11. EL spectra of SiNRs-based white LED under various currents. EL spectra of the constructed SiNRs-based LED device under various currents are also investigated, as shown in Figure S13. Based on previous reports, 10,11 a blue LED chip packaged with SrGa 2 S 4 :Eu 2+ and (Ca 1-x Sr x ) S: Eu 2+ exhibit a large variation in its CIE color coordinates upon an increase in forward current in a narrow range (from 20 ma to 60 ma). In sharp contrast, in our devices, the PL bands of SiNRs increase accompanied with the increase of applied forward current. Therefore, SiNRs-based white LED shows an even better color coordinate stability than commercial white LEDs. Based on these results, it is clear that these as-prepared SiNRs can be efficiently excited in the UV and blue range. S19

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