Article pubs.acs.org/IC
Self-Assembled Upconversion Nanoparticle Clusters for NIRcontrolled Drug Release and Synergistic Therapy after Conjugation with Gold Nanoparticles Huijuan Cai,† Tingting Shen,† Alexander M. Kirillov,‡ Yu Zhang,† Changfu Shan,† Xiang Li,† Weisheng Liu,† and Yu Tang*,† †
State Key Laboratory of Applied Organic Chemistry, Key Laboratory of Nonferrous Metal Chemistry and Resources Utilization of Gansu Province, College of Chemistry and Chemical Engineering, Lanzhou University, Lanzhou 730000, P. R. China ‡ Centro de Química Estrutural, Complexo I, Instituto Superior Técnico, Universidade de Lisboa, Av. Rovisco Pais, Lisbon 1049-001, Portugal S Supporting Information *
ABSTRACT: Fabricated three-dimensional (3D) upconversion nanoclusters (abbreviated as EBSUCNPs) are obtained via an emulsion-based bottom-up selfassembly of NaGdF4:Yb/Er@NaGdF4 nanoparticles (abbreviated as UCNPs), which comprise a NaGdF4:Yb/Er core and a NaGdF4 shell. The EBSUCNPs were then coated with a thin mesoporous amino-functionalized SiO2 shell (resulting in EBSUCNPs@SiO2 precursor) and further conjugated with gold nanoparticles to give the novel EBSUCNPs@SiO2@Au material. Finally, EBSUCNPs@SiO2@Au was applied as a biocompatible and efficient drug carrier for doxorubicin (DOX), thus giving rise to a multifunctional EBSUCNPs@SiO2−DOX@Au nanocomposite. This final material, EBSUCNPs@SiO2−DOX@Au, and the precursor nanoparticles, EBSUCNPs@SiO2@Au, were both fully characterized and their luminescence was investigated in detail. In addition, the drug release properties and photothermal effects of EBSUCNPs@SiO2−DOX@Au were also discussed. Interestingly, when under NIR irradiation, an increasing DOX release was achieved owing to the thermal effect of the Au NPs after absorbing the green light from the upconversion nanoclusters based on the fluorescence resonance energy transfer (FRET) effect. Thus, a near-infrared (NIR)-controlled “on−off” pattern of drug release behavior can be achieved. Moreover, compared with a single therapy method, the assembled nanocomposites exhibit a good synergistic therapy against cancer cells that combines chemotherapy with photothermal therapy. In addition, the in vitro fluorescence microscopy images of EBSUCNPs@SiO2−DOX@Au show a higher enhancement in the red region due to the loading of DOX molecules with respect to EBSUCNPs@SiO2@Au. Therefore, this novel multifunctional 3D cluster architecture can be used in the biomedical field after modification and may pave a new way in other application areas of UCNPs clusters.
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INTRODUCTION In the past decades, one-dimensional (1D), two-dimensional (2D), or three-dimensional (3D) colloidal nanostructures have been successfully fabricated by using a variety of “top-down”, “bottom-up”, or drying-driven dynamic assembly methods.1−3 Among such materials, 3D nanoclusters driven by the bottomup directed self-assembly have attracted special attention due to their several advantages, namely easy synthesis, simple removal of the organic solvent, possibility of postsynthetic functionalization, and notable applicability for various nanoparticles (NPs) and potential applications.4 Bottom-up self-assembly of NPs may endow materials with well-controlled sizes, various shapes, and new properties,5 thus making them interesting candidates for applications as electronic and optoelectronic materials,6 lithium ion batteries,7,8 and in water treatment9 or photocatalysis.10 However, it is still a challenge to make use of this kind of clusters in cancer therapy. Recently, many different kinds of 3D nanoparticle clusters have been successfully © 2017 American Chemical Society
synthesized from various building blocks (Fe3O4, BaCrO4, Ag2Se, CdS, PbS, NaYF4).11 Although Li and co-workers12 have developed an easy method to form binary superstructures with dual-mode luminescence properties, the self-assembly of such materials and their application in biomedical field remains poorly explored. Given that upconversion NPs (UCNPs) can convert the near-infrared irradiation (NIR) to short-wavelength emissions, they possess potential applications in imaging, sensing, detection, and theranostics,13−16 also on account of their intriguing characteristics such as large anti-Stokes shifts, nonautofluorescence, no photobleaching, and deep tissue penetration. Interestingly, the emission wavelength can cover from the ultraviolet to visible light region according to the different host/activator combinations, the amount of doped Received: February 10, 2017 Published: April 12, 2017 5295
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Inorganic Chemistry
controlled “on−off” release pattern when the NIR light is on or off, which can allow designing a stimuli-controlled drug release system that combines the photothermal therapy with the chemotherapy under a single wavelength NIR light. Although Liu and co-workers34 have presented a novel Au nanorod/block copolymer/DOX drug delivery system, which can also achieve a NIR-controlled drug release, a smart single nanocomposite for simultaneous drug release and bioimaging still remains a challenge. In our work, the final nanocomposite can not only achieve a NIR-controlled “on−off” pattern of drug release, but also an in vitro fluorescence imaging from EBSUCNPs. Stepwise assembly process, full characterization, luminescence, drug release, and photothermal properties, as well as in vitro anticancer activity of EBSUCNPs@SiO2−DOX@Au and precursor materials are reported in the present study. This novel multifunctional 3D cluster architecture may pave a new way in other application areas of UCNPs clusters.
lanthanide ions, the controlled crystallite size, phase, and associated defect state,17−21 thus benefiting other materials (e.g., Au NPs, carbon dots, fluorescence dyes, photosensitizers, graphene, and graphene oxide) to absorb these emissions and make use of the fluorescence resonance energy transfer (FRET) process. Up to now, a variety of applications have been developed that utilize the UCNPs-based FRET, namely biosensing,22,23 cell adhesion,24 drug release,25 cancer therapy,26−28 nucleic acid hybridization assay,29 and detection of metal ions30,31 or hydroxyl radicals.32 Within the development of nanomedicine, the photothermal therapy that employs photoabsorbing agents to generate heat after absorbing light (based on the FRET process) has been a topic area in recent years in cancer therapy owing to its lower toxic and side effects.33 However, chemotherapy as a traditional therapy method still has its importance in synergistic therapy, especially in the stimuli-controlled drug release. Taking into consideration the above-mentioned points, we present herein a bottom-up fabrication of the 3D upconversion nanoparticle clusters using the emulsion-based self-assembly method, and demonstrate that this model nanostructure can be used in cancer therapy after modification (Scheme 1). Our
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EXPERIMENTAL SECTION
Materials and Instruments. Materials. Gadolinium chloride (GdCl3·6H2O), ytterbium chloride (YbCl3·6H2O), and erbium chloride (ErCl3·6H2O) were prepared by reacting Gd2O3, Yb2O3, or Er2O3 (99.99%, Shanghai Yuelong) with hydrochloric acid, and then superfluous hydrochloric acid was removed to obtain LnCl3·6H2O. Oleic acid (OA, > 90%) and 1-octadecene (ODE, > 90%) were purchased from Sigma Adrich. NaOH and NH4F were obtained from Sinopharm. Cetyltrimethylammonium bromide (CTAB) and tetraethoxysilane (TEOS) were purchased from J&K Scientific Ltd. HAuCl4· 4H2O was purchased from Beijing HWRK Chem Co., Ltd. Doxorubicin hydrochloride (DOX·HCl) was obtained from Beijing Huafeng United Technology Co., Ltd. (Beijing, China). Instruments. Powder X-ray diffraction (PXRD) patterns were recorded over the 2θ range from 10 to 70° using a Rigaku-Dmax 2400 diffractometer with Cu Kα radiation. Fourier transform infrared (FTIR) spectra were conducted within the 4000−400 cm−1 wavenumber range by using a Nicolet 360 FTIR spectrometer with the KBr pellet technique. Transmission electron microscopy (TEM) images were taken on a Tecnai-G2-F30 (300 kV) instrument. Inductively coupled plasma-atomic emission spectroscopy (ICP-AES) measurements were conducted using an IRIS Advantage ER/S spectrophotometer. The morphological, structural, and chemical characterization of all samples was performed at the nano/atomic scale using field emission HRTEM (Tecnai G2 F30; FEI Company, USA) working at 120 kV, which was equipped with EDX (AMETEK Inc., USA) and high-angle annular dark-field scanning transmission electron microscopy (HAADF-STEM). Dynamic light scattering (DLS) measurements were conducted on a Zetasizer Nanoseries (Nano ZS90) instrument. The UC emission spectra were acquired by using a 980nm laser as the irradiation source and detected by a PMT detector on the fluorescence instrument (FLSP920, Edinburgh instruments, the UK) from 400 to 700 nm (slit width 1.5 nm). The instrument of UCLM was rebuilt on an inverted fluorescence microscopy and an external 980-nm diode laser was illuminated. All the above-mentioned measurements were performed at room temperature. Synthesis of Nanocomposites. Synthesis of NaGdF4:Yb/Er NPs. The β-phase NaGdF4:Yb/Er nanoparticles were synthesized via a coprecipitation method.35 A slightly modified procedure was used. GdCl3·6H2O (0.78 mmol), YbCl3·6H2O (0.20 mmol), and ErCl3· 6H2O (0.02 mmol) were dispersed in oleic acid (OA, 10 mL) and 1octadecene (ODE, 15 mL), and then the mixture was heated to 160 °C for 1 h with a gentle flow of argon gas to obtain lanthanide-oleate precursors. After the formation of a homogeneous solution, the mixture was cooled to room temperature. Then, a mixture of 5 mL of methanol solution containing NaOH (2.5 mmol) and 1 mL of methanol solution containing NH4F (2.75 mmol) was quickly added. The reaction temperature was increased to 50 °C and the mixture was kept at this temperature for 30 min. To evaporate methanol, the reaction mixture was heated for 2 h at 120 °C. The obtained mixture
Scheme 1. Schematic Illustration of the Stepwise Assembly of the EBSUCNPs@SiO2−DOX@Au Nanocomposite and its NIR-controlled Drug Release
strategy comprises the synthesis of the NaGdF4:Yb/Er NPs with an average size of about 23 nm using a coprecipitation method. To enhance the upconversion luminescence intensity, NaGdF4 shell of 10 nm was then coated on the NaGdF4:Yb/Er core through the epitaxial growth process to give NaGdF4:Yb/ Er@NaGdF4 (called UCNPs). In the next step, the emulsionbased self-assembled 3D upconversion nanoparticle clusters of NaGdF4:Yb/Er@NaGdF4 (denoted as EBSUCNPs) were fabricated and coated with a thin mesoporous SiO2 shell. Then, the Au NPs with an absorption peak at 524 nm were conjugated to the silica shell by electrostatic interaction to give a final EBSUCNPs@SiO2@Au nanocomposite. It exhibits good biocompatibility and can be applied as a potential drug carrier. Doxorubicin (DOX) was chosen as the model drug and was encapsulated into the pores of the silica shell. The obtained multifunctional EBSUCNPs@SiO2−DOX@Au nanocomposite has several advantages. First, compared to the conventional UCNPs, the assembled EBSUCNPs clusters show a new structure. Second, based on the FRET process, the acceptor Au NPs can absorb the green emission from the upconverting luminescence and generate heat under 980-nm irradiation. Using this thermal effect, a rapid DOX release was achieved. On the other hand, the nanocomposite exhibits a NIR5296
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Inorganic Chemistry was then further heated for 2.5 h at 320 °C in an argon gas atmosphere. After that, it was cooled to room temperature. The resulting nanoparticles were precipitated by adding ethanol, collected by centrifugation, and then washed three times with cyclohexane/ ethanol (1:1 v/v) solution. The obtained product was finally dispersed in 4 mL of cyclohexane, which was used directly in the next step. Synthesis of NaGdF4:Yb/Er@NaGdF4 NPs (denoted as UCNPs). The coating of the NaGdF4 shell on the NaGdF4:Yb/Er core was achieved through epitaxial growth via a coprecipitation method as mentioned above. First, a mixture of GdCl3·6H2O (1.00 mmol), OA (10 mL), and ODE (15 mL) was heated for 1 h at 160 °C with a gentle flow of argon gas, and then cooled to room temperature. Second, 5 mL of methanol solution containing NaOH (2.5 mmol), 1 mL of methanol solution containing NH4F (2.75 mmol), and 4 mL of cyclohexane solution containing the as-prepared NaGdF4:Yb/Er nanoparticles were mixed. The temperature was raised to 50 °C and kept at this temperature for 30 min. The subsequent procedures were the same as that in the synthesis of NaGdF4:Yb/Er NPs. Finally, the obtained product was centrifuged, washed, and then dried overnight in a vacuum oven at 50 °C. Emulsion-Based Bottom-up Self-Assembled 3D Upconversion Nanoparticle Clusters NaGdF4:Yb/Er@NaGdF4 (denoted as EBSUCNPs). The 3D colloidal UCNPs were synthesized according to a procedure described by Li’s group.11 Briefly, 10 mg of the oleatecapped NaGdF4:Yb/Er@NaGdF4 nanoparticles were dispersed in 1 mL of chloroform, and then 15 mL of aqueous solution containing 0.06 g of CTAB was added dropwise. To evaporate chloroform, the mixture was subjected to 2.5 h of ultrasonic treatment and then the 3D CTAB-UCNPs dispersion was obtained. This dispersion was used directly in the next step without any purification. Synthesis of EBSUCNPs@SiO2 with a Silica Shell Thickness of 9 nm. After adding 30 mL of distilled water into the CTAB-UCNPs dispersion, the mixture was stirred for 2 h at room temperature. Afterward, 4 mL of ethyl acetate, 200 μL of aqueous ammonia solution (25 wt %), and 30 μL of TEOS were successively added to the above mixture under vigorous stirring, and then kept at room temperature for 4 h. In the next step, 0.08 mL of APTES were added to modify the silica shell with amino groups; the solution was kept stirring for 2 h. Finally, the product was centrifuged, washed with ethanol and distilled water for several times, then dried overnight in the vacuum oven at 50 °C. The obtained white product was the NH2-modified EBSUCNPs@ SiO2. To clean the pores of NPs and allow loading of the drug, further purification of EBSUCNPs@SiO2 was conducted to remove the CTAB surfactant. The as-synthesized material was dispersed in solution that contained 20 mL of ethanol and 1 mL of acetic acid. The mixture was refluxed for 12 h at 60 °C, then collected by centrifugation and washed with ethanol three times. Afterward, the product was redispersed in newly prepared solution. This process was repeated two times in order to extract all the remaining CTAB as fully as possible. Finally, the product was put in the vacuum oven for overnight to give the white powder. Synthesis of EBSUCNPs@SiO2 with a Silica Shell Thickness of 65 nm. EBSUCNPs@SiO2 with a silica shell thickness of 65 nm was prepared by using the same method as for EBSUCNPs@SiO2 with a silica shell thickness of 9 nm, but changing the TEOS amount from 30 to 200 μL. Synthesis of Gold NPs. The gold NPs were synthesized by the citrate reduction method according to a described procedure.36 Typically, 250 μL of 0.05 mol/L HAuCl4 was mixed with 50 mL of deionized water under stirring, followed by heating the mixture. After boiling, 1 mL of 38.8 mmol/L sodium citrate was quickly added into the solution and kept for 15 min. During this process, the color changed from light yellow to wine red. After that, the heating device was removed and the solution was kept stirring for an additional 15 min. The as-synthesized dispersion of gold nanoparticles was used directly in the next step. Preparation of EBSUCNPs@SiO2@Au. Ten mg of EBSUCNPs@ SiO2 was dispersed in 8 mL of deionized water under ultrasonic treatment. Six mL of the gold suspension was added into the mixture
and then stirred for 24 h at room temperature. The obtained EBSUCNPs@SiO2@Au material was collected by centrifugation, washed with deionized water, and freeze-dried. DOX Loading Experiment (Preparation of EBSUCNPs@SiO2− DOX@Au). The purchased DOX·HCl was treated before use to obtain DOX. First, DOX·HCl (25 mg) was dispersed in 10 mL of dichloromethane, and then 600 μL of triethylamine was slowly injected into the solution. This mixture was stirred overnight at room temperature in dark. After that, the solvent dichloromethane was evaporated to give DOX which was then dried in the vacuum oven overnight. To be used directly and conveniently, DOX was then dispersed into ultrapure water to give a DOX solution (0.25 mg/mL). To load the DOX, 30 mg of EBSUCNPs@SiO2@Au was dispersed in 10 mL of DOX solution (0.25 mg/mL) under ultrasonic treatment, followed by stirring for 24 h at room temperature in dark. The product was centrifuged, washed with water three times, and freeze-dried. Meanwhile, the supernatant was collected to measure the loading content and the entrapment efficiency. The drug loading content and entrapment efficiency were calculated by the following equations: Loading content = (weight of drug in EBSUCNPs@SiO 2−DOX@Au)/(weight of EBSUCNPs@SiO2− DOX@Au); Entrapment efficiency = (weight of drug in EBSUCNPs@SiO2−DOX @Au) /(initial feed weight of drug). The DOX loading content and the entrapment (encapsulation efficiency) were 4.73% and 59.6%, respectively. Experimental Methods. In-vitro Drug Release Study. EBSUCNPs@SiO2−DOX@Au nanocomposites (2 mg) were dispersed in 3 mL of phosphate-buffered saline (PBS) solution with pH value of 5.0 under NIR irradiation. The other control experiments (pH = 5.0, without NIR irradiation; pH = 7.4, without NIR irradiation; pH = 7.4, with NIR irradiation, respectively) were conducted by using the same amount of nanocomposite and the same volume of PBS solution. The mixture was centrifuged at predetermined time intervals. Two mL of the supernatant was then collected and replaced with an equal volume of fresh PBS buffer solution. The amount of released DOX was determined by measuring the absorption peak at 482 nm of the supernatant at different release times. Cell Culture Preparation. HeLa cell lines were provided by the Institute of Biochemistry and Cell Biology, SIBS, CAS (China). Cells were cultured in a regular growth medium consisting of RPMI 1640 supplemented with 10% FBS (fetal bovine serum) at 37 °C in a humidified and 5% CO2 incubator. The cells were routinely harvested by treatment with a trypsin−ethylenediaminetetraacetic acid (EDTA) solution (0.25%). Cell Viability Assays. To obtain the cell viability under different conditions, the standard MTT assay was conducted on HeLa cells. Briefly, the HeLa cells were first seeded in 96-well plates at a density of 1 × 104 cells per well and cultured in 5% CO2 at 37 °C for 24 h. Then, EBSUCNPs@SiO2@Au, EBSUCNPs@SiO2−DOX@Au, or free DOX were added into the cultured cells at various concentrations that were predetermined. After that, the cells were cultured in 5% CO2 at 37 °C for additional 24 h. To assess the synergistic effect of chemotherapy and photothermal therapy, 980-nm irradiation was introduced when the incubation time was 4 h and irradiation was continued for 5 min under the pump power of 1 W/cm2. Subsequently, the medium was removed and washed with a fresh culture medium, followed by treating with 100 μL of MTT (0.5 mg/mL) and incubating for another 4 h at the same conditions. Finally, the supernatant was removed and 100 μL of dimethyl sulfoxide (DMSO) was added into the cells per well and the wells were shaken for 10 min. The optical density was measured by the microtiter plate reader at 490 nm. The percentages of cell viabilities were calculated compared to the untreated cells as the control experiment. In-vitro Fluorescence Microscopy Images. The HeLa cells were incubated for 24 h and then treated with the as-prepared EBSUCNPs@SiO2@Au or EBSUCNPs@SiO2−DOX@Au at 37 °C for 0.5, 1, and 3 h, respectively. After that, the cells were washed with PBS buffer solution for three times to remove the attached nanocomposites, and then fixed with 2.5% formaldehyde at 37 °C for 10 min, then washed with PBS buffer solution three times again. 5297
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Inorganic Chemistry The fixed cells were finally immersed in 0.5 mL of PBS for analysis. All images were collected under 980-nm irradiation (1 W/cm2).
Figure S3) patterns also demonstrate the successful synthesis of β-NaGdF4. The emulsion-based bottom-up self-assembled 3D upconversion nanoparticle clusters (abbreviated as EBSUCNPs) composed of NaGdF4:Yb/Er@NaGdF4 have a wide size distribution (Figure S1c) and were successfully fabricated according to a reported procedure.11 The corresponding TEM image is shown in Figure 1c. The formation of these upconversion nanoparticle clusters is based on an oil-in-water process. First, UCNPs dispersed in chloroform were emulsified into an aqueous solution containing cetyltrimethylammonium bromide (CTAB). Subsequently, the solvent was evaporated in the emulsion droplet under ultrasonic treatment to give the EBSUCNPs (for comparison, mechanical stirring was used as an alternative method). For potential biomedical applications, a biocompatible mesoporous SiO2 shell was coated on the EBSUCNPs clusters. In fact, in this work, we synthesized the EBSUCNPs@SiO2 samples of different thicknesses, ∼9 and ∼65 nm (Figures 1d and 1e), by adding different amounts of the tetraethoxysilane (TEOS). As expected, the shell becomes thicker along with the increase of TEOS amount, while the morphology of the EBSUCNPs core remains almost unchanged, thus demonstrating that the coating of SiO2 has no effect on the shape of EBSUCNPs. Meanwhile, the effect of ultrasonic treatment on the self-assembly process compared with the mechanical stirring should be discussed.11 Figures 1f and 1g refer to the EBSUCNPs@SiO2 samples obtained using a mechanical stirring or under an ultrasonic treatment, respectively. Despite using the same amount of TEOS to achieve coating by the SiO2 shell, the difference between the obtained samples is obvious. Using an ultrasonic treatment, the shape of the obtained clusters is better ordered and the size is more uniform to some extent. On the basis of this observation, in all further experiments we used the EBSUCNPs@SiO2 nanoparticle clusters prepared by self-assembly under ultrasonic treatment. To achieve the synergistic therapy that combines a photothermal therapy based on the FRET process with a chemotherapy, we conjugated the citrate-capped gold NPs on the SiO2 shell of EBSUCNPs@SiO2 by electrostatic interaction to obtain the EBSUCNPs@SiO2@Au material (Figure 2a; this and all the subsequent experiments were performed using EBSUCNPs@SiO2 with a silica shell thickness of 9 nm). It can be seen from the magnified TEM image (Figure 2b) that Au NPs with an average size of 14 nm are successfully attached to the surface of EBSUCNPs@SiO2 without any free Au NPs left nearby; this indicates a good attachment of the Au NPs. The HRTEM image of Au NPs in EBSUCNPs@SiO2@Au (Figure 2c) confirms a lattice spacing value of 0.235 nm, which well agrees with the d111 value of the parent gold NPs.36 Meanwhile, the element mapping of EBSUCNPs@SiO2@Au indicates that the lanthanide elements (e.g., Gd; Figure 2d) are mainly located in the core of the nanocomposite, whereas the Au NPs are conjugated onto the surface of SiO2. On the other hand, all the Gd, Yb, Er, Na, F, Si, and Au elements can be found in the EDS spectrum (Figure S4), which also demonstrates a successful preparation of EBSUCNPs@SiO2@Au nanoparticle. In addition, the zeta potential changes from +75.4 mV to +41.6 mV (Table S2), which further proves the successful conjugation of Au NPs by electrostatic interaction. It has the Au weight percentage of 2.73% on the SiO2 shell, as determined by inductively coupled plasma-atomic emission spectrometry (ICP-AES).
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RESULTS AND DISCUSSION Synthesis, Morphology, and Characterization of the Nanocomposites. The stepwise assembly process of the final EBSUCNPs@SiO2−DOX@Au nanocomposites and precursor materials is shown in Scheme 1. The transmission electron microscopy (TEM) image of the NaGdF4:Yb/Er (20% Yb, 2% Er) NPs indicates that the nanoparticles are uniform (Figure 1a) and have an average size of about 23 nm, according to the
Figure 1. TEM images: (a) NaGdF4:Yb/Er (inset: high-resolution image), (b) NaGdF4:Yb/Er@NaGdF4 (inset: high-resolution image), (c) EBSUCNPs, (d) EBSUCNPs@SiO2 with a shell thickness of 9 nm, (e) EBSUCNPs@SiO2 with a shell thickness of 65 nm (c, d, and e were all obtained under ultrasonic treatment). (f, g): TEM images of EBSUCNPs@SiO2 prepared under mechanical stirring (f) or under ultrasonic treatment (g), respectively.
dynamic light scattering (DLS) analysis (Figure S1a). After coating the NaGdF4:Yb/Er NPs with a thin NaGdF4 shell through the epitaxial growth to improve the upconverting luminescence efficiency, it can be seen that the obtained NaGdF4:Yb/Er@NaGdF4 nanoparticles (UCNPs) remain monodisperse except the particle size increase from 23 to ∼34 nm (Figures 1b, S1b). The high-resolution TEM (HRTEM) images of the both NaGdF4:Yb/Er and NaGdF4:Yb/Er@NaGdF4 samples confirm a high crystallinity of the particles with a lattice spacing value of 0.52 nm, which is in agreement with the d100 value of the NaGdF4 hexagonal phase.37 Meanwhile, the powder X-ray diffraction (PXRD, Figure S2) patterns are in conformity with β-NaGdF4 (JCPDS: 27-0699), and the selected area electron diffraction (SAED, 5298
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Figure 2. (a) TEM image of EBSUCNPs@SiO2@Au with a mesoporous SiO2 shell thickness of 9 nm. (b) Enlarged image of (a). (c) High resolved TEM image of Au nanoparticle in EBSUCNPs@SiO2@Au. (d) Selected elemental mapping images of EBSUCNPs@SiO2@Au (the dotted circles stand for the main area of different elements).
Figure 3. (a) Upconversion emission of NaGdF4:Yb/Er, NaGdF4:Yb/Er@NaGdF4, and EBSUCNPs@SiO2 with an equal concentration of NaGdF4:Yb/Er. (b) Upconversion emission of EBSUCNPs@SiO2 (black line), and absorption spectra of Au NPs (blue line) and DOX (red line). (c) UV−vis spectra of various materials. (d) Upconversion emission of EBSUCNPs@SiO2, EBSUCNPs@SiO2@Au, and EBSUCNPs@SiO2− DOX@Au with an equal concentration of EBSUCNPs@SiO2. Emission spectra were obtained with the 980-nm irradiation.
In the final step of the assembly process (Scheme 1), the EBSUCNPs@SiO2@Au nanocomposite was treated with DOX as a model drug, which was loaded into the silica pores to furnish a final EBSUCNPs@SiO2−DOX@Au material. The DOX loading content of 4.73% was obtained, while the DOX entrapment (encapsulation) efficiency attained 59.6%. Photoluminescence and UV−vis Spectra. The upconversion luminescence properties of various nanocomposites were investigated under the 980-nm irradiation. As shown in Figure 3a, the upconversion emissions at 521, 540, and 654 nm (the 2H11/2→ 4I15/2, 4S3/2→ 4I15/2, and 4F9/2→ 4I15/2 transitions of Er3+, respectively)20 were observed for the NaGdF4:Yb/Er core, the NaGdF4:Yb/Er@NaGdF4 core−shell, and the
upconversion EBSUCNPs@SiO2 NPs. Obviously, compared to the upconversion emission of the undecorated NaGdF4:Yb/ Er core, the emission of the core−shell NaGdF4:Yb/Er@ NaGdF4 NPs was significantly enhanced (about 16-fold). This enhancement is of great significance for applications in biological systems. Compared with NaGdF4:Yb/Er@NaGdF4, the upconversion emission of the EBSUCNPs@SiO2 significantly decreases due to the hydrophilic pores and the energy transfer (ET) process from Er3+ to mesoporous channels which is due to the OH oscillators;38 however, the emission is still higher than that of the NaGdF4 core alone. Figure 3b shows that the absorption of Au NPs (blue line) has a good overlap with the upconversion emission. Meanwhile, 5299
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Figure 4. (a) DOX release profiles of EBSUCNPs@SiO2−DOX@Au with and without NIR irradiation in PBS buffer solution at different pH values. (b) DOX release efficiency of EBSUCNPs@SiO2−DOX@Au with the NIR irradiation on or off every 1 h. (c) The slope of (b). (d) Upconversion emission spectra at different release times.
Drug Release, Photothermal Effect, and Cell Cytotoxicity. After loading DOX into the pores, the drug release efficiencies of EBSUCNPs@SiO2−DOX@Au with and without the NIR (980 nm) irradiation were tested under different pH values. As shown in Figure 4a, the DOX release efficiency was 16.1% at the physiological pH (7.4) and 38.6% at a slightly acidic pH (5.0) without the NIR irradiation, which can be due to a weak physical adsorption in an acidic environment. In contrast, in the presence of the 980-nm irradiation, the system shows a better releasing behavior at both pH 5.0 and 7.4. Especially, at the pH value of 5.0, the DOX release efficiency reached 78.9%, which is due to the synergistic thermal effect induced by the Au NPs and the weakness of the physical adsorption of DOX. This result demonstrates that the thermal effect of the gold NPs may promote the drug release from the nanocomposite. In addition, all the release curves (Figure 4a) show a relatively rapid release of DOX within the first 5−10 h, which then gradually slows down and reaches a plateau. Thus, the drug was released within the first 2−3 h, and this can efficiently inhibit the growth of cancer cells. Then the release rate will become slow. On the other hand, we also provide a control experiment of EBSUCNPs@SiO2−DOX without Au NPs for drug release. As shown in Figure S8, it can be seen that the release efficiency shows a slower trend compared with EBSUCNPs@SiO2−DOX@Au no matter whether dispersed in PBS solution with a pH value of 5.0 or under NIR irradiation, which demonstrates the function of Au NPs in the fabricated system. Moreover, to verify that this thermal effect produced by the FRET process under the 980-nm irradiation can be used to realize the NIR-controlled “on−off” pattern, we monitored the drug release by switching on or off the NIR irradiation every 1 h during 10 h, and at different pH values (Figure 4b). As expected, when the irradiation is on for 5 min, the drug release
the distance between the EBSUCNPs core and the Au NPs is about 9 nm (
