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Cite this: DOI: 10.1039/d0tb01873b
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Highly efficient photothermal heating via distorted edge-defects in boron quantum dots† Li Wang,‡ab Si-Min Xu,‡c Shanyue Guan,*a Xiaozhong Qu, Geoffrey I. N. Waterhouse,d Shan Hee and Shuyun Zhoua
*b
Quantum dots (QDs) are increasingly being utilized as near infrared (NIR) active photothermal agents for cancer diagnosis and therapy, with the main emphasis of current research being the enhancement of photothermal conversion efficiencies. Herein, we report the facile synthesis of 2–3 nm boron quantum dots (B QDs), which demonstrated a remarkable photothermal conversion efficiency of 57% under NIR excitation. This outstanding performance can be attributed to the alteration of the electronic structure, which was a result from the distorted edge-effect induced by the unique empty orbit of B atoms in the B QDs. These results can be verified by B K-edge near edge X-ray absorption fine structure (NEXAFS), Received 2nd August 2020, Accepted 14th September 2020 DOI: 10.1039/d0tb01873b
high-resolution transmission electron microscopy (HR-TEM) and density functional theory (DFT) calculations. The results demonstrate that B QDs represent a promising new and non-toxic agent for both multimodal NIR-driven cancer imaging and photothermal therapy. This work thus identifies B QDs as an exciting new and theranostic agent for cancer therapy. Furthermore, the synthetic strategy used
rsc.li/materials-b
here to synthesize the B QDs was simple and easily scalable.
Introduction Photothermal therapy (PTT) is a non-invasive method for cancer therapy, and has been the subject of investigation for decades.1–5 Initial PTT research focused on organic and inorganic compounds which caused localized heating under appropriate light excitation,6 with recent research targeting nanoparticle-based systems which offer higher photothermal conversion efficiencies. Both the composition of nanoparticle-based PTT agents and their form influence their performance. Compared with 2D (nanosheets) materials, 0D materials (i.e. quantum dots) have attracted much recent attention due to their very high surface-to-volume ratio, large surface area, and improved stability.7 Quantum dots derived from 2D materials are typically favoured as efficient materials over their bulk counterparts for PTT due to the abundance of a
Key Laboratory of Photochemical Conversion and Optoelectronic Materials, Technical Institute of Physics and Chemistry, Chinese Academy of Sciences, Beijing, 100190, China. E-mail: [email protected]; Fax: +86 10 62554670; Tel: +86 10 82543428 b Center of Materials Science and Optoelectronics Engineering, College of Materials Science and Opto-Electronic Technology, University of Chinese Academy of Sciences, Beijing 100049, China. E-mail: [email protected] c State Key Laboratory of Chemical Resource Engineering, Beijing University of Chemical Technology, Beijing 100029, China d School of Chemical Sciences, The University of Auckland, Auckland 1142, New Zealand e Beijing Technology and Business University, Beijing, 100148, China † Electronic supplementary information (ESI) available. See DOI: 10.1039/d0tb01873b ‡ These authors contributed to this work equally.
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defects at exposed edges, a high proportion of coordinatively unsaturated atoms, and quantum confinement effects. Typically, this exposed edge effect reveals a tuneable molecular structure, which can have an influence on the band gap modulation, which can further affect the photothermal properties.8 Seeking to exploit these phenomena, researchers have explored the PTT properties of a number of quantum dot (QD) systems, including carbon QDs,9,10 Ge QDs11 and antimonene QDs,12 amongst others. Interestingly, boron is one kind of non-toxic, earth-abundant, lightweight, metal-free element,13–15 with electron-deficient p orbitals,16 and thus exhibits strong electron-withdrawing properties, which have been widely utilized towards the boronbased catalyst area. These strong electron-withdrawing/affinity properties of boron can induce the distortion of the edge effect, which is conducive to the formation of defect states and then able to tune the electron structure, and thus may have an influence on the properties. In a previous study, we established that the band gap energy and band edge positions of certain semiconductors could facilitate the electron transfer from excited photosensitizers, thereby lowering the activation energy for non-radiative electron– hole recombination in the semiconductor and thus improving the PTT performance.17,18 Inspired by our previous work, we fabricated boron quantum dots (denoted as B QDs) from 2D boron nanosheets. We hypothesized that the electronic structure and band gap energy of B QDs would differ greatly from those of 2D forms of boron due to the distorted band edge-effects, further tuning their electronic structure, and thus significantly improving
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Paper the photothermal conversion performance. Boron nanosheets (denoted herein as 2D B) have been recently investigated for multimodal imaging-guided cancer therapy.13 In this study, we utilized a topotactic transformation method (ultrasonication) to fragment 2D B into uniformly sized B QDs (diameter 2–3 nm). The obtained B QDs showed high photothermal conversion efficiency (57% under 808 nm excitation). The outstanding performance of B QDs can be attributed to band distorted edge-effects, which was confirmed by both B K-edge near edge X-ray absorption fine structure (NEXAFS) and X-ray photoelectron spectroscopy (XPS). HRTEM revealed that a significant disorder existed in B QDs. Furthermore, density functional theory (DFT) calculations showed that the B QDs possess a narrower band gap than the 2D B nanosheets from which they were synthesized, which is explained by interband gap states in the B QDs. As a result of this band gap narrowing, the B QDs displayed superior photoacoustic imaging performance and photothermal tumour performance in vivo under NIR (808 nm) compared to other B-based reference materials tested (i.e. 2D forms of boron) (Scheme 1).
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Fig. 1 (a) Schematic illustration of the synthesis of the B QDs. Characterization data for the B QDs. (b) TEM image of B QDs at low magnification. (c) and (d) HRTEM of B QDs. (e) Histograms showing the size distribution of B QDs (determined from the TEM image shown in Fig. 1c). (f) The zeta potential of B QDs in buffered saline solutions of weak acid or weak base. (g) UV-vis absorbance spectrum of B QDs.
Results and discussion B QDs were synthesized via a facile topotactic transformation method starting from 2D B, as depicted in Fig. 1a. An aqueous dispersion of boron nanosheets was prepared, and then treated by ultrasonication in an ice bath for 10 h, followed by isolation of the B QDs by centrifugation (see the Experimental part for full synthesis details). Powder XRD (Fig. S1, ESI†) patterns revealed that B QDs possessed the same space group as elemental boron (JCPDS card No. 80-0323). High-resolution transmission electron microscopy (HRTEM) showed that the B QDs were uniform in shape and size with a mean particle size of 2–3 nm (Fig. 1b, c and e), which was similar to that determined by dynamic light scattering (around 20 nm) (Fig. S2, ESI†). Lattice fringes of 0.197 nm in Fig. 1d can be assigned to the (318) plane of boron. Interestingly, boron vacancies and the rough edges on the boundaries at the outermost surface of the QDs can be seen (as indicated by the yellow
Scheme 1 Schematic illustration of boron quantum dots with good photoacoustic imaging and photothermal performance, acting as a powerful theranostic agent for cancer detection and photothermal therapy.
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arrows), which can verify the structural distortions as a consequence of defect sites. These structural distortions are universal in ultrathin nanomaterial systems due to the abundant surface defects via coordinative unsaturation of atoms, which allow minimization of the total surface energy. Structural distortions allow a more thermodynamically stable structure to be adopted. It can therefore be concluded that B QDs have enhanced coordinative unsaturation of B atoms and more disorder relative to the 2D B nanosheets from which the QDs were prepared. The zeta potential of the B QDs in phosphate buffer saline (PBS) at pH 7 was determined to be 30.5 mV (Fig. S3, ESI†), sufficient for charge stabilize of the QDs against aggregation. A similar zeta potential (approximately 30 mV) was observed for the B QDs in both weak acids and weak bases (Fig. 1f). As shown in Fig. 1g, aqueous dispersions of the B QDs absorbed strongly in the NIR region. Indeed, the absorption maximum of the B QDs falls perfectly within the biological NIR window (750–850 nm), which has been considered as the ideal source to trigger a photothermal effect and is highly beneficial for biomedical applications19,20 including PTT and photoacoustic imaging. Fourier transform infrared spectroscopy (FT-IR) was used to probe the functional groups on the surface of 2D B and B QDs (Fig. S4, ESI†). The FT-IR spectrum of the B QDs showed two peaks at 664 and 485 cm 1 which were not observed in the spectrum of the 2D B precursor. These features can be assigned to B–O vibrations,21 implying that partial oxidation of B occurred on the edges of the B QDs. The observation of a well-defined peak at 3420 cm 1, readily assigned to O–H stretching vibrations, points to the presence of surface B–OH groups.13 All the other peaks in the FT-IR spectrum of the B QDs were due to B–B networks, with the same peaks also being present in the spectrum of the 2D B. The Raman spectrum of the B QDs (Fig. S5, ESI†) showed a peak at 1100 cm 1, typical
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Fig. 2 (a) B 1s XPS spectra of B QDs and 2D B. (b) and (c) B K-edge NEXAFS spectra of B QDs and 2D B. (d) Photothermal heating curves for dispersions of B QDs at various concentrations (50, 100, 200 and 400 mg mL 1) under 808 nm laser irradiation (1.0 W). (e) Heating and cooling profiles for a dispersion of B QDs (400 mg mL 1) over 3 laser on/off cycles. (f) Time constant for heat transfer from the system determined by applying a linear fit to data from the cooling period.
for two-center B–B bonds in amorphous boron with a mediumrange order. The shoulders at 650 and 800 cm 1 are due to intra-icosahedral vibrations.22,23 The thickness of B QDs determined using atomic force microscopy (AFM) ranged from 0.6–1.6 nm (Fig. S6, ESI†). The ultrasmall diameter, surface hydroxyl groups and the negative surface charge gave the B QDs excellent dispersibility and stability in aqueous solution. Negligible aggregation or sedimentation of B QDs occurred in PBS on standing unstirred for 7 days (Fig. S7, ESI†). The digital photograph (inset of Fig. S7, ESI†) showed that the B QDs remained well dispersed in water, PBS and cell culture medium (DMEM) on standing for 24 h. XPS was used to examine the chemical speciation of boron in the 2D B and the B QDs. The B 1s XPS spectra for each sample are shown in Fig. 2a (other XPS data for the B QDs are presented in Fig. S8 (ESI†), where the elements identified in the sample were B, O and C, the latter being due to adventitious hydrocarbons). The B 1s XPS spectrum of the B QDs was deconvoluted into three peaks at 187.4 eV (Peak I), 188.9 eV (Peak II) and 192.7 eV (Peak III), while the 2D B sample shows similar features, albeit in slightly different relative intensity ratios. The peaks at 187.4 eV and 188.9 eV are assigned to the elemental crystalline boron24 (i.e. B0 with B–B bonds) and a defective B0 state (i.e. associated with coordinatively unsaturated B atoms at the surface of the QDs), respectively. This may be caused by the distorted edge-effect of the B QDs. The peak at
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Paper 192.7 eV is typical for B3+ in B2O3 or B(OH)3 (B 1s = 192.5–193.0 eV),25 which was more intense in spectrum of the B QDs than in the spectrum of 2D B, further inferring that the edges of B QDs were partially oxidized. The three B-related peaks for the B QDs were at slightly lower binding energies than the corresponding peaks for the 2D B, suggesting that the two samples possessed distinct electronic structures,26 which is explored and discussed below. B K-edge near edge X-ray absorption fine structure (NEXAFS) was applied to further investigate the electronic structure of the B QDs and 2D B. B K-edge NEXAFS involves the photoexcitation of a K-shell (i.e. B 1s) electron into the unoccupied electronic states of a material, and is routinely applied to examine the sp2/sp3 hybridization in boron-containing compounds. The B K-edge spectra for both B QDs and 2D B contained an intense and sharp feature at 191.6 eV (Fig. 2b), which represents the 1s - p* transition of elemental boron with sp2 hybridization. According to the dipole selection rules, the final states for B K-edge NEXAFS need to have s + pz character (for 1s - p* transitions) or px + py character (for 1s - s* transitions). The weak feature at 194.5 eV is assigned to the electronic transitions into unoccupied B 2pz orbitals of B–O groups,27,28 whereas the broad and relatively intense peaks centered at 197.0 and 201.4 eV are assigned to the 1s - s* transitions of tetrahedral sp3 B.29 Comparing the spectra of the B QDs and 2D B, it is evident that the ratio of B sp3/sp2 states (i.e. the intensity ratio (I197.0 eV + I201.4 eV)/ I191.6 eV) was higher for B QDs (see Fig. 2c where the spectra are overlaid), whilst the B–O related feature at 194.5 eV was also more prominent for B QDs. These results infer that significant disorder exists in the B QDs, creating more sp3 B atoms and B atoms in higher oxidation states, which was caused by the introduction of defective B and leading to the distortion of the edge B QDs (both of which were likely located at the edges of B QDs). Both XPS and NEXAFS results are in good qualitative agreement with the HRTEM observation where disorder was seen at the distorted edges of the B QDs. Clearly, the B QDs possess significant disorder due to their small size and thus abundance of coordinatively unsaturated B atoms at their surface. Such disorder was expected to modify the electronic structure of the B QDs by introducing interband gap states.28,30 Such intergap states were expected to endow the B QDs with enhanced light absorption in the NIR region relative to 2D B, which was confirmed quantitatively below. Density functional theory (DFT) calculations were performed to explore the photothermal mechanism and properties of the B QDs. 2D B and graphene quantum dots (graphene QDs) were selected as references. The optimized geometries of the ground state for B QDs, 2D B, and graphene QDs are displayed in Fig. S9 and Tables S1 S3, ESI†. The relative energy levels of the ground state S0 (set as the zero point) and the first singlet excited state (S1), together with their molecular orbitals for each material, are shown in Fig. 3. The efficiency of photothermal conversion mainly depends on two aspects, the efficiency of photo absorption and the efficiency of unwanted radiative transitions that result in undesirable energy dissipation in the form of light instead of heat. In Fig. 3, it is seen that graphene QDs had the highest oscillator strength for light absorption, and also a high oscillator strength for radiative
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Fig. 3 Relative energy levels of the ground state (S0) and the first singlet excited state (S1) for B QDs, 2D B, and graphene QDs. The molecular energy levels associated with the S0 and S1 states are shown. The oscillator strengths for vertical photo absorption and radiative relaxation transitions are labeled in red and blue, respectively.
relaxation, the latter preventing graphene QDs from having a high photothermal conversion efficiency. Conversely, the oscillator strength for photo absorption in B QDs (0.0036) is significantly larger than in 2D B (0.0016), whilst B QDs and 2D B had weak and comparable oscillator strengths for radiative transitions. Accordingly, of the three materials studies, the B QDs were expected to offer the highest photothermal conversion efficiency. To verify that the B QDs could be excited under NIR (808 nm) excitation, the absorbance spectrum of B QDs in water was collected (Fig. S10, ESI†). Under 808 nm excitation, the absorbance of an aqueous B QD dispersion was 2.002 L g 1 cm 1. This implied that most of the incident laser light could be absorbed by the B QD dispersion with the B QD concentration being B500 mg L 1. The solvation energy of B QDs was calculated to be 16.55 eV per B QDs, once again suggesting excellent water dispersibility. Effective PTT agents need to absorb strongly in the NIR region and then convert the energy of the absorbed NIR photons into heat, thus generating local hyperthermia. Inspired by the strong absorbance of the B QDs across the UV, visible and NIR regions, we photothermally heated dispersions of B QDs under 808 nm irradiation. As indicated in Fig. 2d, pure H2O showed a negligible temperature increase under 808 nm irradiation for 10 min. In contrast, a dispersion of B QDs (400 mg mL 1) showed a significant temperature increase under 808 nm laser (1.0 W) treatment, reaching 72.5 1C after 10 min of continuous irradiation. At a lower B QD concentration (50 mg mL 1), the temperature reached 52.3 1C after 10 min. These results revealed that the B QDs could efficiently convert NIR photons into hyperthermia. Subsequently, the effect of 808 nm laser power on the heating of a dispersion of B QDs (400 mg mL 1) was examined (Fig. S11 and S12, ESI†). At laser powers of 0.5, 0.8 and 1.0 W, the dispersion temperature reached 43.0, 57.0 and 72.5 1C, respectively. The photothermal temperature of the B QD dispersion thus exhibited a linear relationship with the 808 nm laser power (Fig. S13, ESI†). To examine the thermal stability of the B QDs, cycles of NIR heating (808 nm, 1.0 W, 10 min)
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Journal of Materials Chemistry B followed by natural cooling (no NIR excitation) were performed on a B QD dispersion (400 mg mL 1). As shown in Fig. 2e, no change in the photothermal performance of the B QDs occurred over three test cycles, indicating that the B QDs were a very stable photothermal agent. The photothermal efficiency (Z) of the B QDs was calculated using the data in Fig. 2f, following the method described in the ESI.† The Z of the B QDs was calculated to be 57%, which was appreciably higher than the values reported for many other QD-based PTT agents, including antimonene QDs (45.5%),12 carbon QDs (38.5%),31 black phosphorus QDs (28.4%)32 and other photothermal materials such as Au NRs33 (Table S4, ESI†). The efficient internalization of B QDs in cancer cells was probed using TEM. The results suggest that a cell uptake of B QDs occurs via an endocytosis pathway since the B QDs were mainly located in the cytoplasm and partially in the nucleus (Fig. 4a–f and Fig. S14, ESI†). As indicated in Fig. S14 (ESI†), the morphology of the B QDs can be changed into clusters of B QDs under the tumor environment. Furthermore, the in vitro toxicity of the B QDs was investigated via a typical Cell Counting Kit-8 (CCK8) assay. HeLa cells were incubated with B QDs at various concentrations. As displayed in Fig. S15 (ESI†), negligible toxicity was observed when the B QDs were incubated with HeLa cells (no NIR treatment), even at high B QD concentrations (400 mg mL 1). We further incubated HeLa cells with B QDs at various concentrations for 24 h, and then exposed the HeLa cells to an 808 nm laser at different laser powers (0.5, 0.8, and 1.0 W) for 10 min. As shown in Fig. S15, S16 (ESI†) and Fig. 4g, the
Fig. 4 (a–f) TEM images of B QDs inside HeLa cells. The viability of (g) HeLa, (h) MCF-7 and (i) HepG-2 cells incubated with B QDs (concentrations 50, 100, 200 and 400 mg mL 1). For the NIR treatments, a 808 nm laser operating at a power of 1.0 W was used. (j) Flow cytometry data for HeLa cells incubated with PBS or B QDs, with or without NIR (808 nm) irradiation. The NIR treatment was at 808 nm (1.0 W) for 10 min.
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Journal of Materials Chemistry B viability of HeLa cells decreased as the NIR laser power was increased. The HeLa cell viability was only 16% after incubation with B QDs (200 mg mL 1) followed by 1.0 W NIR irradiation treatment (Fig. 4g). These results demonstrate that cancer cell viability can be altered simply by changing the power of the NIR laser (808 nm), consistent with the results of the photothermal conversion experiments (Fig. S11, S12 and Fig. 2d, ESI†). Similar results were obtained for the MCF-7 and HepG-2 cell lines, demonstrating that the B QDs have excellent phototherapy effects under NIR irradiation (Fig. 4h and i). B QDs + NIR triggered cell death was clearly observed in laser scanning confocal microscopy (LSCM) studies using living/dead cell staining (Fig. S17 and S18, ESI†).34–36 As shown in Fig. S17 (ESI†), cells incubated with B QDs showed green fluorescence (without NIR), confirming the excellent biocompatibility of B QDs. However, cells incubated with B QDs then NIR irradiated (808 nm, 1.0 W) showed strong red fluorescence. At B QD concentrations of 200 mg mL 1 or higher, all cells were dead following NIR treatment. The control group (no B QDs) showed strong green fluorescence following NIR treatment, indicating that cell death seen in the B QDS + NIR group was not caused simply by NIR exposure. Furthermore, the viability of cells incubated with B QDs and then irradiated at different 808 nm laser powers was also investigated. As shown in Fig. S18 (ESI†), as the laser power was increased, the cell viability decreased. These results demonstrate the remarkable in vitro photothermal effect of B QDs on promoting the cancer cell ablation. Furthermore, flow cytometry was utilized to quantify the cell apoptosis, with the data being analysed using Annexin V-FITC/PI staining. As shown in Fig. 4j, the PBS, PBS + NIR, and B QDs groups had negligible effect on cell apoptosis. In contrast, B QD treated cells subjected to 808 nm irradiation exhibited strong inhibition of cell growth. As a result, intracellular hyperthermia generated by B QDs under NIR light irradiation was found to clearly inhibit cell proliferation. Due to their strong absorbance in the NIR region, the B QDs were expected to be useful imaging agents37,38 including photoacoustic imaging (PAI) agents and thermal imaging contrast agents. Photoacoustic images were recorded in a time dependent method following i.v. injection of B QDs (Fig. 5a). The results revealed that B QDs can permeate into the deep tumor region and reached the highest accumulation at 6 h postinjection. The in vivo imaging data are consistent with the PAI results (Fig. S19, ESI†). Subsequently, the thermal imaging demonstrated that under 808 nm laser, the tumor temperature increased rapidly to 56 1C within 10 min whilst the temperature change was negligible for the control mice (Fig. S20, ESI†). These results confirm the effective distribution of B QDs in tumors and also the efficient penetration of NIR light to cause strong photothermal conversion. In vivo PTT experiments were performed by intravenous injection of B QD dispersions into tumors. HeLa tumor-bearing mice were divided into three groups: control, B QDs only, and B QDs + NIR. The tumors in the B QDs + NIR laser group disappeared after PTT, leaving only black scars in the initial tumor sites (Fig. S21, ESI†). Tumor volumes and mice weights for the three groups were monitored
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Fig. 5 In vivo photoacoustic images of tumor tissue (arrows) containing B QDs at (a) different times and 3D-images 6 h after the intravenous injection of B QDs. (b) Tumor growth rates of groups following different treatments (n = 5, ***P o 0.001). (The laser: 808 nm, 1 W.) (c) Change of body weights following different treatments (n = 5). (d) TUNEL- and (e) H&E-stained images of tumors from the different groups.
every 2 days for 20 days (Fig. 5b and c, respectively). The body weights of the three groups increased steadily over the 20 days, showing little fluctuation. Thus, the treatments (i.v. injection of B QDs and NIR irradiation for 10 min) had no adverse effects on the mice. However, the B QDs + NIR treatment had a remarkable effect on the tumor site, with no tumor being found in that group after 20 days (Fig. 5b). In comparison, the control and B QD (no NIR) groups showed a progressive increase in tumor volumes over the 20 days. H&E and TUNEL staining results showed severe necrosis of tumor cells in the B QDs + NIR group (Fig. 5d and e). Fig. S22 (ESI†) showed no apparent impairment to the main organs in the B QDs + NIR group. Based on the findings presented here, it can be concluded that B QDs show outstanding biocompatibility and PTT properties for in vivo applications.
Conclusions In summary, B QDs of size 2–3 nm were successfully synthesized via a facile method from 2D B. The B QDs possess substantial disorder due to the high fraction of coordinatively unsaturated edge B atoms, thus favourably modifying the electronic structure of B to allow very strong NIR photoexcitation and enhanced non-radiative transitions. Accordingly, the B QDs delivered very effective hyperthermia under 808 nm irradiation.
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Paper In addition, the B QDs were an effective, low cost PAI contrast agent. This work thus identifies B QDs as an exciting new and theranostic agent for cancer therapy. Furthermore, the synthetic strategy used here to synthesize the B QDs was simple and easily scalable.
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Experimental Model construction for DFT calculations A structural model for bulk boron was constructed according to X-ray crystallographic data reported for crystalline boron,39 based on the space group R3% mH and lattice parameters: a = b = 4.908 Å, c = 12.567 Å, a = b = 901 and g = 1201. Models for the boron quantum dots (B QDs) and 2D B were obtained by selective cleavage of bonds in the bulk Boron model. The chemical formulae of the B QD model and 2D B model were B13 and B72, respectively. The B QD cluster model was carefully constructed to ensure that the center B atom had saturation coordination. A model of graphene quantum dots (Graphene QDs)9 was also constructed by cleaving graphene, with edge C atoms saturated by the addition of H atoms. The chemical formula of the graphene QDs was C24H12. Fig. S10 (ESI†) shows the optimized geometries for the B QDs, 2D B, and graphene QD models. Computational method The electronic structures of the ground state (S0) and the first singlet excited state (S1) for the B QDs, 2D B, and graphene QD models were calculated using the Gaussian 09 software package. The exchange–correlation functional of B3LYP40 and the 6-311+G(d) basis set were applied.41 The S1 state of each model was optimized using time-dependent density functional theory (TD-DFT). The optimized geometries were confirmed to be minimum-energy points by harmonic frequency calculations, where all the frequencies were positive. The oscillator strengths of photo-absorption and radiative transitions were calculated using the Gaussian 09 software package. The solvation energy and absorbance of the B QD model was calculated with the DMol3 code in the Accelrys materials studio version 5.5 (Accelrys Software Inc.: San Diego, CA). Density functional theory calculations were performed with a plane wave implementation.42 The generalized gradient approximation (GGA) Perdew–Burke–Ernzerhof (PBE) functional43 was applied as the exchange–correlation functional. The ionic cores were described with ultrasoft pseudopotentials to improve transferability and decrease the number of plane waves needed for the expansion of the Kohn–Sham orbitals.44 The Broyden–Fletcher–Goldfarb–Shanno (BFGS) algorithm was used to search the potential energy surface during geometry optimization.45 The geometry optimization was based on the following three points: (i) an energy tolerance of 1 10 5 eV per atom, (ii) a maximum force tolerance of 0.03 eV Å 1, and (iii) a maximum displacement tolerance of 1 10 3 Å. For the calculation of the optical properties of the B QDs, the G-point-centered k-point meshes used for the Brillouin zone
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Journal of Materials Chemistry B integrations were 3 3 3 k-points in the x-, y-, and z- directions. The solvation energy of B QDs was calculated as follows. First, the model of B QDs was immersed in a solvent box filled with H2O. The solvation energy Esol was then calculated according to eqn (1) Esol = EBQD
in water
Ewater
EBQD
(1)
where EBQD in water is the energy of the solvent box with B QDs inside, Ewater is the energy of the solvent box filled only with water, and EBQD is the energy of the B QDs.
Conflicts of interest There are no conflicts to declare.
Acknowledgements The authors are grateful for the financial support from the National Natural Science Foundation of China (21805293, 51873214), and the Director Foundation of the Technical Institute of Physics and Chemistry, Chinese Academy of Sciences (Grant No. 2018-GSY) and the Youth Innovation Promotion Association of Chinese Academy of Sciences (No. 2019027). GINW acknowledges funding support from the Dodd Walls Centre for Photonic and Quantum Technologies, the MacDiarmid Institute for Advanced Materials and Nanotechnology, and the Energy Education Trust of New Zealand.
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Journal of Materials Chemistry B 11 J. Ouyang, C. Feng, X. Ji, L. Li, H. K. Gutti, N. Y. Kim, D. Artzi, A. Xie, N. Kong, Y. N. Liu, G. J. Tearney, X. Sui, W. Tao and O. C. Farokhzad, Angew. Chem., Int. Ed., 2019, 58, 1–7. 12 W. Tao, X. Ji, X. Xu, M. A. Islam, Z. Li, S. Chen, P. E. Saw, H. Zhang, Z. Bharwani, Z. Guo, J. Shi and O. C. Farokhzad, Angew. Chem., Int. Ed., 2017, 129, 12058–12062. 13 X. Ji, N. Kong, J. Wang, W. Li, Y. Xiao, S. T. Gan, Y. Zhang, Y. Li, X. Song, Q. Xiong, S. Shi, Z. Li, W. Tao, H. Zhang, L. Mei and J. Shi, Adv. Mater., 2018, 1803031. 14 Y. Fang and X. Wang, Angew. Chem., Int. Ed., 2017, 56, 15506–15518. 15 C. Huang, C. Chen, M. Zhang, L. Lin, X. Ye, S. Lin, M. Antonietti and X. Wang, Nat. Commun., 2015, 6, 7698. 16 M. L. Drummond, V. Meunier and B. G. Sumpter, J. Phys. Chem. A, 2007, 111, 6539–6551. 17 L. Wang, S. Guan, Y. Weng, S. M. Xu, H. Lu, X. Meng and S. Zhou, ACS Appl. Mater. Interfaces, 2019, 11, 6267–6275. 18 Y. Weng, S. Guan, L. Wang, H. Lu, X. Meng, G. I. N. Waterhouse and S. Zhou, Small, 2019, 16, 1905184. 19 C. Liu, B. Liu, J. Zhao, Z. Di, D. Chen, Z. Gu, L. Li and Y. Zhao, Angew. Chem., Int. Ed., 2020, 59, 2634–2638. 20 H. Lin, X. Wang, L. Yu, Y. Chen and J. Shi, Nano Lett., 2017, 17, 384–391. 21 G. Liu, X. Meng, H. Zhang, G. Zhao, H. Pang, T. Wang, P. Li, T. Kako and J. Ye, Angew. Chem., Int. Ed., 2017, 56, 5570–5574. ¨m and W. Roberg, 22 U. Kudlmann, H. Werheit, T. Lundsto J. Phys. Chem. Solids, 1994, 55, 579–587. 23 S. Sarkar, Y. S. Ha, N. Soni, G. I. An, W. Lee, M. H. Kim, P. T. Huynh, H. Ahn, N. Bhatt, Y. J. Lee, J. Y. Kim, K. M. Park, I. Ishii, S. G. Kang and J. Yoo, Angew. Chem., Int. Ed., 2016, 55, 9365–9370. 24 X. Liu, Z. Zhang, L. Wang, B. I. Yakobson and M. C. Hersam, Nat. Mater., 2018, 17, 783–788. 25 B. Feng, J. Zhang, Q. Zhong, W. Li, S. Li, H. Li, P. Cheng, S. Meng, L. Chen and K. Wu, Nat. Chem., 2016, 8, 563–568. 26 A. M. Smith and S. Nie, Acc. Chem. Res., 2010, 43, 190–200.
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PAPER
Cite this: DOI: 10.1039/d0tb01873b
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Highly efficient photothermal heating via distorted edge-defects in boron quantum dots† Li Wang,‡ab Si-Min Xu,‡c Shanyue Guan,*a Xiaozhong Qu, Geoffrey I. N. Waterhouse,d Shan Hee and Shuyun Zhoua
*b
Quantum dots (QDs) are increasingly being utilized as near infrared (NIR) active photothermal agents for cancer diagnosis and therapy, with the main emphasis of current research being the enhancement of photothermal conversion efficiencies. Herein, we report the facile synthesis of 2–3 nm boron quantum dots (B QDs), which demonstrated a remarkable photothermal conversion efficiency of 57% under NIR excitation. This outstanding performance can be attributed to the alteration of the electronic structure, which was a result from the distorted edge-effect induced by the unique empty orbit of B atoms in the B QDs. These results can be verified by B K-edge near edge X-ray absorption fine structure (NEXAFS), Received 2nd August 2020, Accepted 14th September 2020 DOI: 10.1039/d0tb01873b
high-resolution transmission electron microscopy (HR-TEM) and density functional theory (DFT) calculations. The results demonstrate that B QDs represent a promising new and non-toxic agent for both multimodal NIR-driven cancer imaging and photothermal therapy. This work thus identifies B QDs as an exciting new and theranostic agent for cancer therapy. Furthermore, the synthetic strategy used
rsc.li/materials-b
here to synthesize the B QDs was simple and easily scalable.
Introduction Photothermal therapy (PTT) is a non-invasive method for cancer therapy, and has been the subject of investigation for decades.1–5 Initial PTT research focused on organic and inorganic compounds which caused localized heating under appropriate light excitation,6 with recent research targeting nanoparticle-based systems which offer higher photothermal conversion efficiencies. Both the composition of nanoparticle-based PTT agents and their form influence their performance. Compared with 2D (nanosheets) materials, 0D materials (i.e. quantum dots) have attracted much recent attention due to their very high surface-to-volume ratio, large surface area, and improved stability.7 Quantum dots derived from 2D materials are typically favoured as efficient materials over their bulk counterparts for PTT due to the abundance of a
Key Laboratory of Photochemical Conversion and Optoelectronic Materials, Technical Institute of Physics and Chemistry, Chinese Academy of Sciences, Beijing, 100190, China. E-mail: [email protected]; Fax: +86 10 62554670; Tel: +86 10 82543428 b Center of Materials Science and Optoelectronics Engineering, College of Materials Science and Opto-Electronic Technology, University of Chinese Academy of Sciences, Beijing 100049, China. E-mail: [email protected] c State Key Laboratory of Chemical Resource Engineering, Beijing University of Chemical Technology, Beijing 100029, China d School of Chemical Sciences, The University of Auckland, Auckland 1142, New Zealand e Beijing Technology and Business University, Beijing, 100148, China † Electronic supplementary information (ESI) available. See DOI: 10.1039/d0tb01873b ‡ These authors contributed to this work equally.
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defects at exposed edges, a high proportion of coordinatively unsaturated atoms, and quantum confinement effects. Typically, this exposed edge effect reveals a tuneable molecular structure, which can have an influence on the band gap modulation, which can further affect the photothermal properties.8 Seeking to exploit these phenomena, researchers have explored the PTT properties of a number of quantum dot (QD) systems, including carbon QDs,9,10 Ge QDs11 and antimonene QDs,12 amongst others. Interestingly, boron is one kind of non-toxic, earth-abundant, lightweight, metal-free element,13–15 with electron-deficient p orbitals,16 and thus exhibits strong electron-withdrawing properties, which have been widely utilized towards the boronbased catalyst area. These strong electron-withdrawing/affinity properties of boron can induce the distortion of the edge effect, which is conducive to the formation of defect states and then able to tune the electron structure, and thus may have an influence on the properties. In a previous study, we established that the band gap energy and band edge positions of certain semiconductors could facilitate the electron transfer from excited photosensitizers, thereby lowering the activation energy for non-radiative electron– hole recombination in the semiconductor and thus improving the PTT performance.17,18 Inspired by our previous work, we fabricated boron quantum dots (denoted as B QDs) from 2D boron nanosheets. We hypothesized that the electronic structure and band gap energy of B QDs would differ greatly from those of 2D forms of boron due to the distorted band edge-effects, further tuning their electronic structure, and thus significantly improving
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Paper the photothermal conversion performance. Boron nanosheets (denoted herein as 2D B) have been recently investigated for multimodal imaging-guided cancer therapy.13 In this study, we utilized a topotactic transformation method (ultrasonication) to fragment 2D B into uniformly sized B QDs (diameter 2–3 nm). The obtained B QDs showed high photothermal conversion efficiency (57% under 808 nm excitation). The outstanding performance of B QDs can be attributed to band distorted edge-effects, which was confirmed by both B K-edge near edge X-ray absorption fine structure (NEXAFS) and X-ray photoelectron spectroscopy (XPS). HRTEM revealed that a significant disorder existed in B QDs. Furthermore, density functional theory (DFT) calculations showed that the B QDs possess a narrower band gap than the 2D B nanosheets from which they were synthesized, which is explained by interband gap states in the B QDs. As a result of this band gap narrowing, the B QDs displayed superior photoacoustic imaging performance and photothermal tumour performance in vivo under NIR (808 nm) compared to other B-based reference materials tested (i.e. 2D forms of boron) (Scheme 1).
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Fig. 1 (a) Schematic illustration of the synthesis of the B QDs. Characterization data for the B QDs. (b) TEM image of B QDs at low magnification. (c) and (d) HRTEM of B QDs. (e) Histograms showing the size distribution of B QDs (determined from the TEM image shown in Fig. 1c). (f) The zeta potential of B QDs in buffered saline solutions of weak acid or weak base. (g) UV-vis absorbance spectrum of B QDs.
Results and discussion B QDs were synthesized via a facile topotactic transformation method starting from 2D B, as depicted in Fig. 1a. An aqueous dispersion of boron nanosheets was prepared, and then treated by ultrasonication in an ice bath for 10 h, followed by isolation of the B QDs by centrifugation (see the Experimental part for full synthesis details). Powder XRD (Fig. S1, ESI†) patterns revealed that B QDs possessed the same space group as elemental boron (JCPDS card No. 80-0323). High-resolution transmission electron microscopy (HRTEM) showed that the B QDs were uniform in shape and size with a mean particle size of 2–3 nm (Fig. 1b, c and e), which was similar to that determined by dynamic light scattering (around 20 nm) (Fig. S2, ESI†). Lattice fringes of 0.197 nm in Fig. 1d can be assigned to the (318) plane of boron. Interestingly, boron vacancies and the rough edges on the boundaries at the outermost surface of the QDs can be seen (as indicated by the yellow
Scheme 1 Schematic illustration of boron quantum dots with good photoacoustic imaging and photothermal performance, acting as a powerful theranostic agent for cancer detection and photothermal therapy.
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arrows), which can verify the structural distortions as a consequence of defect sites. These structural distortions are universal in ultrathin nanomaterial systems due to the abundant surface defects via coordinative unsaturation of atoms, which allow minimization of the total surface energy. Structural distortions allow a more thermodynamically stable structure to be adopted. It can therefore be concluded that B QDs have enhanced coordinative unsaturation of B atoms and more disorder relative to the 2D B nanosheets from which the QDs were prepared. The zeta potential of the B QDs in phosphate buffer saline (PBS) at pH 7 was determined to be 30.5 mV (Fig. S3, ESI†), sufficient for charge stabilize of the QDs against aggregation. A similar zeta potential (approximately 30 mV) was observed for the B QDs in both weak acids and weak bases (Fig. 1f). As shown in Fig. 1g, aqueous dispersions of the B QDs absorbed strongly in the NIR region. Indeed, the absorption maximum of the B QDs falls perfectly within the biological NIR window (750–850 nm), which has been considered as the ideal source to trigger a photothermal effect and is highly beneficial for biomedical applications19,20 including PTT and photoacoustic imaging. Fourier transform infrared spectroscopy (FT-IR) was used to probe the functional groups on the surface of 2D B and B QDs (Fig. S4, ESI†). The FT-IR spectrum of the B QDs showed two peaks at 664 and 485 cm 1 which were not observed in the spectrum of the 2D B precursor. These features can be assigned to B–O vibrations,21 implying that partial oxidation of B occurred on the edges of the B QDs. The observation of a well-defined peak at 3420 cm 1, readily assigned to O–H stretching vibrations, points to the presence of surface B–OH groups.13 All the other peaks in the FT-IR spectrum of the B QDs were due to B–B networks, with the same peaks also being present in the spectrum of the 2D B. The Raman spectrum of the B QDs (Fig. S5, ESI†) showed a peak at 1100 cm 1, typical
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Fig. 2 (a) B 1s XPS spectra of B QDs and 2D B. (b) and (c) B K-edge NEXAFS spectra of B QDs and 2D B. (d) Photothermal heating curves for dispersions of B QDs at various concentrations (50, 100, 200 and 400 mg mL 1) under 808 nm laser irradiation (1.0 W). (e) Heating and cooling profiles for a dispersion of B QDs (400 mg mL 1) over 3 laser on/off cycles. (f) Time constant for heat transfer from the system determined by applying a linear fit to data from the cooling period.
for two-center B–B bonds in amorphous boron with a mediumrange order. The shoulders at 650 and 800 cm 1 are due to intra-icosahedral vibrations.22,23 The thickness of B QDs determined using atomic force microscopy (AFM) ranged from 0.6–1.6 nm (Fig. S6, ESI†). The ultrasmall diameter, surface hydroxyl groups and the negative surface charge gave the B QDs excellent dispersibility and stability in aqueous solution. Negligible aggregation or sedimentation of B QDs occurred in PBS on standing unstirred for 7 days (Fig. S7, ESI†). The digital photograph (inset of Fig. S7, ESI†) showed that the B QDs remained well dispersed in water, PBS and cell culture medium (DMEM) on standing for 24 h. XPS was used to examine the chemical speciation of boron in the 2D B and the B QDs. The B 1s XPS spectra for each sample are shown in Fig. 2a (other XPS data for the B QDs are presented in Fig. S8 (ESI†), where the elements identified in the sample were B, O and C, the latter being due to adventitious hydrocarbons). The B 1s XPS spectrum of the B QDs was deconvoluted into three peaks at 187.4 eV (Peak I), 188.9 eV (Peak II) and 192.7 eV (Peak III), while the 2D B sample shows similar features, albeit in slightly different relative intensity ratios. The peaks at 187.4 eV and 188.9 eV are assigned to the elemental crystalline boron24 (i.e. B0 with B–B bonds) and a defective B0 state (i.e. associated with coordinatively unsaturated B atoms at the surface of the QDs), respectively. This may be caused by the distorted edge-effect of the B QDs. The peak at
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Paper 192.7 eV is typical for B3+ in B2O3 or B(OH)3 (B 1s = 192.5–193.0 eV),25 which was more intense in spectrum of the B QDs than in the spectrum of 2D B, further inferring that the edges of B QDs were partially oxidized. The three B-related peaks for the B QDs were at slightly lower binding energies than the corresponding peaks for the 2D B, suggesting that the two samples possessed distinct electronic structures,26 which is explored and discussed below. B K-edge near edge X-ray absorption fine structure (NEXAFS) was applied to further investigate the electronic structure of the B QDs and 2D B. B K-edge NEXAFS involves the photoexcitation of a K-shell (i.e. B 1s) electron into the unoccupied electronic states of a material, and is routinely applied to examine the sp2/sp3 hybridization in boron-containing compounds. The B K-edge spectra for both B QDs and 2D B contained an intense and sharp feature at 191.6 eV (Fig. 2b), which represents the 1s - p* transition of elemental boron with sp2 hybridization. According to the dipole selection rules, the final states for B K-edge NEXAFS need to have s + pz character (for 1s - p* transitions) or px + py character (for 1s - s* transitions). The weak feature at 194.5 eV is assigned to the electronic transitions into unoccupied B 2pz orbitals of B–O groups,27,28 whereas the broad and relatively intense peaks centered at 197.0 and 201.4 eV are assigned to the 1s - s* transitions of tetrahedral sp3 B.29 Comparing the spectra of the B QDs and 2D B, it is evident that the ratio of B sp3/sp2 states (i.e. the intensity ratio (I197.0 eV + I201.4 eV)/ I191.6 eV) was higher for B QDs (see Fig. 2c where the spectra are overlaid), whilst the B–O related feature at 194.5 eV was also more prominent for B QDs. These results infer that significant disorder exists in the B QDs, creating more sp3 B atoms and B atoms in higher oxidation states, which was caused by the introduction of defective B and leading to the distortion of the edge B QDs (both of which were likely located at the edges of B QDs). Both XPS and NEXAFS results are in good qualitative agreement with the HRTEM observation where disorder was seen at the distorted edges of the B QDs. Clearly, the B QDs possess significant disorder due to their small size and thus abundance of coordinatively unsaturated B atoms at their surface. Such disorder was expected to modify the electronic structure of the B QDs by introducing interband gap states.28,30 Such intergap states were expected to endow the B QDs with enhanced light absorption in the NIR region relative to 2D B, which was confirmed quantitatively below. Density functional theory (DFT) calculations were performed to explore the photothermal mechanism and properties of the B QDs. 2D B and graphene quantum dots (graphene QDs) were selected as references. The optimized geometries of the ground state for B QDs, 2D B, and graphene QDs are displayed in Fig. S9 and Tables S1 S3, ESI†. The relative energy levels of the ground state S0 (set as the zero point) and the first singlet excited state (S1), together with their molecular orbitals for each material, are shown in Fig. 3. The efficiency of photothermal conversion mainly depends on two aspects, the efficiency of photo absorption and the efficiency of unwanted radiative transitions that result in undesirable energy dissipation in the form of light instead of heat. In Fig. 3, it is seen that graphene QDs had the highest oscillator strength for light absorption, and also a high oscillator strength for radiative
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Fig. 3 Relative energy levels of the ground state (S0) and the first singlet excited state (S1) for B QDs, 2D B, and graphene QDs. The molecular energy levels associated with the S0 and S1 states are shown. The oscillator strengths for vertical photo absorption and radiative relaxation transitions are labeled in red and blue, respectively.
relaxation, the latter preventing graphene QDs from having a high photothermal conversion efficiency. Conversely, the oscillator strength for photo absorption in B QDs (0.0036) is significantly larger than in 2D B (0.0016), whilst B QDs and 2D B had weak and comparable oscillator strengths for radiative transitions. Accordingly, of the three materials studies, the B QDs were expected to offer the highest photothermal conversion efficiency. To verify that the B QDs could be excited under NIR (808 nm) excitation, the absorbance spectrum of B QDs in water was collected (Fig. S10, ESI†). Under 808 nm excitation, the absorbance of an aqueous B QD dispersion was 2.002 L g 1 cm 1. This implied that most of the incident laser light could be absorbed by the B QD dispersion with the B QD concentration being B500 mg L 1. The solvation energy of B QDs was calculated to be 16.55 eV per B QDs, once again suggesting excellent water dispersibility. Effective PTT agents need to absorb strongly in the NIR region and then convert the energy of the absorbed NIR photons into heat, thus generating local hyperthermia. Inspired by the strong absorbance of the B QDs across the UV, visible and NIR regions, we photothermally heated dispersions of B QDs under 808 nm irradiation. As indicated in Fig. 2d, pure H2O showed a negligible temperature increase under 808 nm irradiation for 10 min. In contrast, a dispersion of B QDs (400 mg mL 1) showed a significant temperature increase under 808 nm laser (1.0 W) treatment, reaching 72.5 1C after 10 min of continuous irradiation. At a lower B QD concentration (50 mg mL 1), the temperature reached 52.3 1C after 10 min. These results revealed that the B QDs could efficiently convert NIR photons into hyperthermia. Subsequently, the effect of 808 nm laser power on the heating of a dispersion of B QDs (400 mg mL 1) was examined (Fig. S11 and S12, ESI†). At laser powers of 0.5, 0.8 and 1.0 W, the dispersion temperature reached 43.0, 57.0 and 72.5 1C, respectively. The photothermal temperature of the B QD dispersion thus exhibited a linear relationship with the 808 nm laser power (Fig. S13, ESI†). To examine the thermal stability of the B QDs, cycles of NIR heating (808 nm, 1.0 W, 10 min)
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Journal of Materials Chemistry B followed by natural cooling (no NIR excitation) were performed on a B QD dispersion (400 mg mL 1). As shown in Fig. 2e, no change in the photothermal performance of the B QDs occurred over three test cycles, indicating that the B QDs were a very stable photothermal agent. The photothermal efficiency (Z) of the B QDs was calculated using the data in Fig. 2f, following the method described in the ESI.† The Z of the B QDs was calculated to be 57%, which was appreciably higher than the values reported for many other QD-based PTT agents, including antimonene QDs (45.5%),12 carbon QDs (38.5%),31 black phosphorus QDs (28.4%)32 and other photothermal materials such as Au NRs33 (Table S4, ESI†). The efficient internalization of B QDs in cancer cells was probed using TEM. The results suggest that a cell uptake of B QDs occurs via an endocytosis pathway since the B QDs were mainly located in the cytoplasm and partially in the nucleus (Fig. 4a–f and Fig. S14, ESI†). As indicated in Fig. S14 (ESI†), the morphology of the B QDs can be changed into clusters of B QDs under the tumor environment. Furthermore, the in vitro toxicity of the B QDs was investigated via a typical Cell Counting Kit-8 (CCK8) assay. HeLa cells were incubated with B QDs at various concentrations. As displayed in Fig. S15 (ESI†), negligible toxicity was observed when the B QDs were incubated with HeLa cells (no NIR treatment), even at high B QD concentrations (400 mg mL 1). We further incubated HeLa cells with B QDs at various concentrations for 24 h, and then exposed the HeLa cells to an 808 nm laser at different laser powers (0.5, 0.8, and 1.0 W) for 10 min. As shown in Fig. S15, S16 (ESI†) and Fig. 4g, the
Fig. 4 (a–f) TEM images of B QDs inside HeLa cells. The viability of (g) HeLa, (h) MCF-7 and (i) HepG-2 cells incubated with B QDs (concentrations 50, 100, 200 and 400 mg mL 1). For the NIR treatments, a 808 nm laser operating at a power of 1.0 W was used. (j) Flow cytometry data for HeLa cells incubated with PBS or B QDs, with or without NIR (808 nm) irradiation. The NIR treatment was at 808 nm (1.0 W) for 10 min.
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Journal of Materials Chemistry B viability of HeLa cells decreased as the NIR laser power was increased. The HeLa cell viability was only 16% after incubation with B QDs (200 mg mL 1) followed by 1.0 W NIR irradiation treatment (Fig. 4g). These results demonstrate that cancer cell viability can be altered simply by changing the power of the NIR laser (808 nm), consistent with the results of the photothermal conversion experiments (Fig. S11, S12 and Fig. 2d, ESI†). Similar results were obtained for the MCF-7 and HepG-2 cell lines, demonstrating that the B QDs have excellent phototherapy effects under NIR irradiation (Fig. 4h and i). B QDs + NIR triggered cell death was clearly observed in laser scanning confocal microscopy (LSCM) studies using living/dead cell staining (Fig. S17 and S18, ESI†).34–36 As shown in Fig. S17 (ESI†), cells incubated with B QDs showed green fluorescence (without NIR), confirming the excellent biocompatibility of B QDs. However, cells incubated with B QDs then NIR irradiated (808 nm, 1.0 W) showed strong red fluorescence. At B QD concentrations of 200 mg mL 1 or higher, all cells were dead following NIR treatment. The control group (no B QDs) showed strong green fluorescence following NIR treatment, indicating that cell death seen in the B QDS + NIR group was not caused simply by NIR exposure. Furthermore, the viability of cells incubated with B QDs and then irradiated at different 808 nm laser powers was also investigated. As shown in Fig. S18 (ESI†), as the laser power was increased, the cell viability decreased. These results demonstrate the remarkable in vitro photothermal effect of B QDs on promoting the cancer cell ablation. Furthermore, flow cytometry was utilized to quantify the cell apoptosis, with the data being analysed using Annexin V-FITC/PI staining. As shown in Fig. 4j, the PBS, PBS + NIR, and B QDs groups had negligible effect on cell apoptosis. In contrast, B QD treated cells subjected to 808 nm irradiation exhibited strong inhibition of cell growth. As a result, intracellular hyperthermia generated by B QDs under NIR light irradiation was found to clearly inhibit cell proliferation. Due to their strong absorbance in the NIR region, the B QDs were expected to be useful imaging agents37,38 including photoacoustic imaging (PAI) agents and thermal imaging contrast agents. Photoacoustic images were recorded in a time dependent method following i.v. injection of B QDs (Fig. 5a). The results revealed that B QDs can permeate into the deep tumor region and reached the highest accumulation at 6 h postinjection. The in vivo imaging data are consistent with the PAI results (Fig. S19, ESI†). Subsequently, the thermal imaging demonstrated that under 808 nm laser, the tumor temperature increased rapidly to 56 1C within 10 min whilst the temperature change was negligible for the control mice (Fig. S20, ESI†). These results confirm the effective distribution of B QDs in tumors and also the efficient penetration of NIR light to cause strong photothermal conversion. In vivo PTT experiments were performed by intravenous injection of B QD dispersions into tumors. HeLa tumor-bearing mice were divided into three groups: control, B QDs only, and B QDs + NIR. The tumors in the B QDs + NIR laser group disappeared after PTT, leaving only black scars in the initial tumor sites (Fig. S21, ESI†). Tumor volumes and mice weights for the three groups were monitored
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Fig. 5 In vivo photoacoustic images of tumor tissue (arrows) containing B QDs at (a) different times and 3D-images 6 h after the intravenous injection of B QDs. (b) Tumor growth rates of groups following different treatments (n = 5, ***P o 0.001). (The laser: 808 nm, 1 W.) (c) Change of body weights following different treatments (n = 5). (d) TUNEL- and (e) H&E-stained images of tumors from the different groups.
every 2 days for 20 days (Fig. 5b and c, respectively). The body weights of the three groups increased steadily over the 20 days, showing little fluctuation. Thus, the treatments (i.v. injection of B QDs and NIR irradiation for 10 min) had no adverse effects on the mice. However, the B QDs + NIR treatment had a remarkable effect on the tumor site, with no tumor being found in that group after 20 days (Fig. 5b). In comparison, the control and B QD (no NIR) groups showed a progressive increase in tumor volumes over the 20 days. H&E and TUNEL staining results showed severe necrosis of tumor cells in the B QDs + NIR group (Fig. 5d and e). Fig. S22 (ESI†) showed no apparent impairment to the main organs in the B QDs + NIR group. Based on the findings presented here, it can be concluded that B QDs show outstanding biocompatibility and PTT properties for in vivo applications.
Conclusions In summary, B QDs of size 2–3 nm were successfully synthesized via a facile method from 2D B. The B QDs possess substantial disorder due to the high fraction of coordinatively unsaturated edge B atoms, thus favourably modifying the electronic structure of B to allow very strong NIR photoexcitation and enhanced non-radiative transitions. Accordingly, the B QDs delivered very effective hyperthermia under 808 nm irradiation.
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Paper In addition, the B QDs were an effective, low cost PAI contrast agent. This work thus identifies B QDs as an exciting new and theranostic agent for cancer therapy. Furthermore, the synthetic strategy used here to synthesize the B QDs was simple and easily scalable.
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Experimental Model construction for DFT calculations A structural model for bulk boron was constructed according to X-ray crystallographic data reported for crystalline boron,39 based on the space group R3% mH and lattice parameters: a = b = 4.908 Å, c = 12.567 Å, a = b = 901 and g = 1201. Models for the boron quantum dots (B QDs) and 2D B were obtained by selective cleavage of bonds in the bulk Boron model. The chemical formulae of the B QD model and 2D B model were B13 and B72, respectively. The B QD cluster model was carefully constructed to ensure that the center B atom had saturation coordination. A model of graphene quantum dots (Graphene QDs)9 was also constructed by cleaving graphene, with edge C atoms saturated by the addition of H atoms. The chemical formula of the graphene QDs was C24H12. Fig. S10 (ESI†) shows the optimized geometries for the B QDs, 2D B, and graphene QD models. Computational method The electronic structures of the ground state (S0) and the first singlet excited state (S1) for the B QDs, 2D B, and graphene QD models were calculated using the Gaussian 09 software package. The exchange–correlation functional of B3LYP40 and the 6-311+G(d) basis set were applied.41 The S1 state of each model was optimized using time-dependent density functional theory (TD-DFT). The optimized geometries were confirmed to be minimum-energy points by harmonic frequency calculations, where all the frequencies were positive. The oscillator strengths of photo-absorption and radiative transitions were calculated using the Gaussian 09 software package. The solvation energy and absorbance of the B QD model was calculated with the DMol3 code in the Accelrys materials studio version 5.5 (Accelrys Software Inc.: San Diego, CA). Density functional theory calculations were performed with a plane wave implementation.42 The generalized gradient approximation (GGA) Perdew–Burke–Ernzerhof (PBE) functional43 was applied as the exchange–correlation functional. The ionic cores were described with ultrasoft pseudopotentials to improve transferability and decrease the number of plane waves needed for the expansion of the Kohn–Sham orbitals.44 The Broyden–Fletcher–Goldfarb–Shanno (BFGS) algorithm was used to search the potential energy surface during geometry optimization.45 The geometry optimization was based on the following three points: (i) an energy tolerance of 1 10 5 eV per atom, (ii) a maximum force tolerance of 0.03 eV Å 1, and (iii) a maximum displacement tolerance of 1 10 3 Å. For the calculation of the optical properties of the B QDs, the G-point-centered k-point meshes used for the Brillouin zone
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Journal of Materials Chemistry B integrations were 3 3 3 k-points in the x-, y-, and z- directions. The solvation energy of B QDs was calculated as follows. First, the model of B QDs was immersed in a solvent box filled with H2O. The solvation energy Esol was then calculated according to eqn (1) Esol = EBQD
in water
Ewater
EBQD
(1)
where EBQD in water is the energy of the solvent box with B QDs inside, Ewater is the energy of the solvent box filled only with water, and EBQD is the energy of the B QDs.
Conflicts of interest There are no conflicts to declare.
Acknowledgements The authors are grateful for the financial support from the National Natural Science Foundation of China (21805293, 51873214), and the Director Foundation of the Technical Institute of Physics and Chemistry, Chinese Academy of Sciences (Grant No. 2018-GSY) and the Youth Innovation Promotion Association of Chinese Academy of Sciences (No. 2019027). GINW acknowledges funding support from the Dodd Walls Centre for Photonic and Quantum Technologies, the MacDiarmid Institute for Advanced Materials and Nanotechnology, and the Energy Education Trust of New Zealand.
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