pubs.acs.org/Langmuir © 2010 American Chemical Society
Ultrasound-Triggered Smart Drug Release from Multifunctional Core-Shell Capsules One-Step Fabricated by Coaxial Electrospray Method Yujia Jing, Yihua Zhu,* Xiaoling Yang, Jianhua Shen, and Chunzhong Li* Key Laboratory for Ultrafine Materials of Ministry of Education, School of Materials Science and Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai 200237, China Received September 9, 2010. Revised Manuscript Received December 6, 2010 Multifunctional core-shell capsules that triggered release by ultrasound stimulus were one-step fabricated by the coaxial electrospray method. The TiO2 shell suppressed the initial burst release of the paclitaxel. Fe3O4 and graphene quantum dots inside the oil core functioned successfully for magnetic targeting and fluorescence imaging, respectively. Paclitaxel was trigger released when the dual layer of titania shell cracked under the ultrasound stimulation, and the releasing profile could be controlled by the length of applied ultrasound time.
1. Introduction Smart multifunctional microcapsules are receiving increasing attention owing to their capability of targeting/imaging a specific organ/tissue and triggering drug release through the application of an external stimulus. The capsules can sense changes in physical or chemical characteristics, thereby offering the merits of a repeatable and accurate dosing or a controllable constant release rate.1 Smart capsules with multifunctional components can interact with an external magnetic field positioned at a specified area targeted for drug delivery and also fluorescence imaging.2 Magnetic microcapsules with monodisperse size distribution and uniform structures have been fabricated by microfluidic method, but the relative larger sizes limited its practical application.3 Electrohydrodynamic cojetting also has been used to make highly ordered spheres and tube structures with multicompartmental materials, but the fluorescent materials were simply compounded without a proper protection provided from a shell layer.4 Some polymeric or inorganic delivery systems developed are triggered by changes in surrounding environment such as ultrasound,5 pH,6 temperature,7 and magnetic fields.8 These types of smart delivery systems allow real-time control of drug dosage according to alterations in chemical and physical status.2 Ultrasonic irradiation is one of the most promising external triggers for smart drug release because it is noninvasive and can penetrate deep into the interior of the body and be accurately controlled by altering a number of parameters including frequency, power density, duty cycles, and time of application. *Corresponding author. Tel: þ86-21-64252022. Fax: þ86-21-64250624. E-mail:
[email protected] (Y.Z.);
[email protected] (C.L.). (1) Gong, X. Q.; Peng, S. L.; Wen, W. J.; Sheng, P.; Li, W. H. Adv. Funct. Mater. 2009, 19, 292–297. (2) Kim, H. J.; Matsuda, H.; Zhou, H. S.; Honma, I. Adv. Mater. 2006, 18, 3083– 2088. (3) Okushima, S.; Nisisako, T.; Torii, T.; Higuchi, T. Langmuir 2004, 20, 9905– 9908. (4) George, M. C.; Braun, P. V. Angew. Chem., Int. Ed. 2009, 48, 8606–8609. (5) Kooiman, K.; B€ohmer, M. R.; Emmer, M.; Vos, H. J.; Chlon, C.; Shi, W. T.; :: Hall, C.; Winter, S.; Schroen, S. K.; Versluis, M.; de Jong, N.; Wamel, A. J. Controlled Release 2009, 133, 109–118. (6) Soppimath, K. S.; Tan, D. C. W.; Yang, Y. Y. Adv. Mater. 2005, 17, 318–323. (7) Pradhan, P.; Giri, J.; Rieken, F.; Koch, C.; Mykhaylyk, O.; D€oblinger, M.; Banerjee, R.; Bahadur, D.; Plank, C. J. Controlled Release 2010, 142, 108–121. (8) Hu, S. H.; Chen, S. Y.; Liu, D. M.; Hsiao, C. S. Adv. Mater. 2008, 20, 2690– 2695.
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From a clinical point of view, it is more desirable that a “zero or near-zero release profile” can be achieved as practically needed before the drug-containing carriers reach the targeted sites because the nature of drug molecules diffusing from the capsules to the environment is an unavoidable thermodynamic procedure under the condition of a drug concentration gradient.9 Once such a mechanism is triggered right after administration, the risk of undesired clinical complications may result, including a reduced therapeutic efficacy. Unfortunately lots of drug carriers that are made of polymers reveal a naturally burst release behavior at an uncontrollable time point. Numerous attempts have been made to find a better candidate, among which metal oxide with mesoporous structure has been great attention.10 Core-shell structure capsules are always chosen as the perfect candidate to encapsulate the drug; both natural7 and artificial materials3 have been used to fabricate the capsule matrix. The shell should have proper mechanical properties for physically supporting and remarkable chemical characteristics for desirable therapeutic release. The shell layer interfacial solidifying methods include photopolymerization,11 hydrolysis condensation,12 dewetting coacervation,13 or other chemical reactions. Some of the core-shell capsules’ preparation methods have been widely reported, such as emulsion method,14 microfluidic method,3 selfassembly method,15 and coaxial electrospray.16 Recently, electrospray technique is applied to produce monodisperse droplets with diameters ranging from nanometers to micrometers, but the single-capillary electrosprayed capsules show initial burst release at the surface/near-surface loading of drug; this drawback is caused by the surfactant introduced in the (9) Hu, S. H.; Liu, D. M.; Tung, W. L.; Liao, C. F.; Chen, S. Y. Adv. Funct. Mater. 2008, 18, 2946–2955. (10) Zhao, W. R.; Chen, H. R.; Li, Y. S.; Li, L.; Lang, M. D.; Shi, J. L. Adv. Funct. Mater. 2008, 18, 2780–2788. (11) Kim, E.; Kim, D.; Jung, H.; Lee, J.; Paul, S.; Selvapalam, N.; Yang, Y.; Lim, N.; Park, C. G.; Kim, K. Angew. Chem., Int. Ed. 2010, 49, 4405–4408. (12) Chen, H. Y.; Zhao, Y.; Song, Y. L.; Jiang, L. J. Am. Chem. Soc. 2008, 130, 7800–7801. (13) Zhao, H.; Chen, J. F.; Zhao, Y.; Jiang, L.; Sun, J. W.; Yun, J. Adv. Mater. 2008, 20, 3682–2686. (14) Buyukozturk, F.; Benneyan, J. C.; Carrier, R. L. J. Controlled Release 2010, 142, 22–30. (15) Gai, S. L.; Yang, P. P.; Li, C. X.; Wang, W. X.; Dai, Y. L.; Niu, N.; Lin, J. Adv. Funct. Mater. 2010, 20, 1166–1172. (16) Wu, Y.; Fei, Z. Z.; Lee, L. J.; Wyslouzil, B. E. Biotechnol. Bioeng. 2010, 105, 834–842.
Published on Web 12/23/2010
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solution.17 The above-mentioned issue can be overcome by using a coaxial electrospray system.18 Loscertales et al. first reported that two media flowing simultaneously under electrohydrodynamic conditions can result in encapsulation, and the two materials have to be immiscible.19 The mechanisms of the immiscible liquids’ coaxial electrospraying have been discussed,20,21 and applications about drug releasing from polymer shells also have been reported.22 Unlike the other hollow capsules’ synthesizing methods that require a template or a preorganized structure,23,24 coaxial electrospray is a one-step method without using extra surfactants or lots of solvents. Herein, we report a facile, surfactant-free, multifunctional capsule platform for simultaneous fluorescence imaging, magnetically guided delivery, and ultrasound-triggered releasing that is fabricated by two coaxial electrospray immiscible liquids. This system is composed of olive oil as the reservoir of oil-soluble drug, magnetite (Fe3O4), carbon (graphene) quantum dots, and duallayer porous titania (TiO2) shell as the sensitive vehicle. Olive oil is used throughout the world and considered to be a safe and healthy material in cooking, cosmetics, and pharmaceutics. The graphene quantum dots encapsulated in the multifunctional carriers provide optical information for in situ monitoring of the drug release.25
2. Experimental Section 2.1. Materials. Poly(vinylpyrrolidone) (PVPk-30), tetrabutyl titanate (TBT), ethanol, acetate acid, dimethyl formamide (DMF), and olive oil were purchased from Sinopharm Chemical Reagent. Paclitaxel T7191 (purity 97%) was purchased from CPG Biotech (Shanghai, China). Fe3O4 was synthesized by the solvent-thermal method, as described elsewhere; then, 0.5 g sample was suspended in ethanol (50 mL). Under pH 5.0 and 80 °C conditions, the suspension was mixed with sodium oleate (0.15 g) and was agitated for 1 h to fabricate modified oil-soluble magnetic particles. Poly(ethylene glycol) (PEG)-coated graphene quantum dots were synthesized as reported.26 2.2. Instruments. Scanning electron microscope (SEM) images were taken using a JEOL JSM-6360LV instrument. The surfaces of the samples were sputtered with gold before testing. Transmission electron microscope (TEM) images were taken on a JEOL JEM 2011 microscope (JEOL, Tokyo, Japan) at an acceleration voltage of 200 kV. The specimen was prepared by dropcasting the sample dispersion onto a carbon-coated copper grid, followed by drying at room temperature. Thermal gravity analysis (TGA) was carried out by TGA/SDTA/DSC851e (Mettler-Toledo, Swiss) in N2 circumstance. We have scanned the 13.7537 mg sample from 25 to 1000 °C. UV-vis absorption spectra were recorded by a Unico UV-2102PC spectrophotometer. All spectra were recorded with quartz cells of 10 mm path length. Ultrasound was applied in a SK3200H ultrasound cleaner (KUDOS) with a frequency of 59 kHz. Vibrating sample magnetometer (model (17) Wu, Y. Q.; MacKay, J. A.; McDaniel, J. R.; Chilkoti, A.; Clark, R. L. Biomacromolecules 2009, 10, 19–24. (18) Lee, Y. H.; Mei, F.; Bai, M. Y.; Zhao, S. L.; Chen, D. R. J. Controlled Release 2010, 145, 58–65. (19) Loscertales, I. G.; Barrero, A.; Guerrero, I.; Cortijo, R.; Marquez, M.; Ga~nan-Calvo, A. M. Science 2002, 129, 1695–1698. (20) Ahmad, Z.; Nangrejo, M.; Edirisinghe, M.; Stride, E.; Colombo, P.; Zhang, H. B. Appl. Phys. A: Mater. Sci. Process. 2009, 97, 31–37. G.; Loscertales, I. G.; Marquez, M.; Barrero, A. Phys. Rev. Lett. (21) Marı´ n, A. 2007, 98, 014502–1. (22) Hwang, Y. K.; Jeong, U.; Cho, E. C. Langmuir 2008, 24, 2446–2451. (23) Sivakumar, S.; Bansal, V.; Cortez, C.; Chong, S. F.; Zelikin, A. N.; Caruso, F. Adv. Mater. 2009, 21, 1820–1824. (24) Wang, Y. J.; Yan, Y.; Cui, J. W.; Rigau, L. H.; Heath, J. K.; Nice, E. C.; Caruso, F. Adv. Mater. 2010, 22, 4293–4297. (25) Hu, S. H.; Kuo, K. T.; Tung, W. L.; Liu, D. M.; Chen, S. Y. Adv. Funct. Mater. 2009, 19, 3396–3403. (26) Pan, D. Y.; Zhang, J. C.; Li, Z.; Wu, M. H. Adv. Mater. 2010, 22, 734–738.
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7407, LakeShore, Westerville, OH, USA) was tested at room temperature. 2.3. Capsule Preparation. Tetrabutyl titanium (3 g) and PVPk-30 (0.5 g) were blended in premixed solution composed of ethanol (1 g), DMF (1 g), and acetic acid (1 g). Ethanol acted as the main solvent, DMF was added to increase the electric conductivity, acetic acid played an important role as a controller for hydrolysis speed, and PVP was used to adjust the viscosity and create a porous structure. The as-prepared solution was the water phase flowing in outer capillary. Paclitaxel (10 mg), modified Fe3O4 (0.3 g), and carbon quantum dots (0.5 g) were added to olive oil (2 mL) to make the oil phase as the inner fluid. After these two immiscible solutions were loaded in a 5 mL syringe, the feeding rate of the syringe pump connected to the outer capillary was set to 0.8 mL/h. The liquid flowing speed in the inner needle was 0.4 mL/h. The high voltage applied on the homemade coaxial spraying nozzle orifice was 22 kV. The distance from the nozzle to the round-shaped grounded collector was 30 cm. 2.4. Drug Content and Encapsulation Efficiency. The amount of paclitaxel loaded in 50 mg capsules was measured by UV-vis spectrophotometry by recording the absorbance at 227 nm. The water-insoluble paclitaxel was extracted by dimethyl sulfoxide (DMSO) prior to the assay. The concentration of the drugs was determined by correlation with a calibration curve. 2.5. Drug Releasing Profile. Capsules (50 mg) were suspended in PBSD medium (5 mL) and removed to the dialysis bag. The as-prepared dialysis bag was submerged in PBSD (10 mL) in a flask. The release system was placed in a 37.5 °C, 100 rpm shaking table. PBSD in the flask was replaced with 10 mL of fresh medium every hour. At the end of the 5th and 11th hours, 15 min of ultrasound were applied to the capsules suspension. All drugcontaining PBSD solution was collected and labeled in a centrifuge tube ready for the UV test.
3. Results and Discussion Figure 1 presents the morphology and structure of the capsule before and after ultrasound treatment and the schematic ultrasound triggering mechanism. The shell of the capsule was chosen to be TiO2 for the ultrafast hydrolytic condensation reaction of TBT to make sure the capsules’ sphere shape can be formed and kept after spraying out. The sizes of the capsules ranged from 600 nm to 6 μm, most of which were 1.5 to 2 μm. The thickness of the shell layer was influenced by viscosity, electrical conductivity, liquid flow rate,27,28 and applied voltage. The thickness of the dual layer capsules’ shell was ∼300 nm, 100 nm for each layer of the shell, as shown in Figure 1d. The formation mechanism of the dual-layer shell structure properly is that after the two immiscible liquids have been electrosprayed. The spherical core-shell structure formed immediately when TBT contacted the moisture in the air, and as a result of hydrolysis and condensation, the TiO2 shell appeared. The other reaction product, butanol, kept effusing and evaporating with the unreacted TBT. The secondary reaction happened, and the outside layer of titania shell formed. A similar process has been mentioned by Zhao’s report.13 In the TEM image, the lighter dark part on the capsule sphere was the cavity between the two shell layers, and an uneven number of Fe3O4 particles was encapsulated inside because the complicated electrospray system was easily disturbed by changes in jet interface tension. With the longer ultrasound time, increasing cracked ratio appeared on a capsule surface, shown in Figure 1c,d. In the picture of the 15 min ultrasound sample, several Fe3O4 particles can be seen attaching to the inner shell layer because the particles (27) Berkland, C.; Pollauf, E.; Varde, N.; Pack, D. W.; Kim, K. Pharm. Res. 2007, 24, 1007–1013. (28) Snider, C.; Lee, S. Y.; Yeo, Y.; Gregori, G. J.; Robinson, J. P.; Park, K. Pharm. Res. 2008, 25, 5–15.
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Figure 1. (a) SEM image capsules and (b) TEM image. (The insert is the magnification of a single capsule.) (c) SEM image of the capsules ultrasound for 5 min and (d) SEM image of ultrasound 15 min sample.
Figure 2. Distribution of shell thickness (inner and outer).
were accelerated by the magnetic field created by the high-power electric field29 and contacting the solidifying inner shell. Figure 2 displays the thickness distribution of the two shell layers. The thickness of each shell layer is relatively uniform, and it is ca. 100 nm by statistically measuring 45 pieces of incomplete capsule spheres or segments from the SEM images. This indicates that the synthetic method of multifunctional core-shell capsules is viable and stable by coaxial electrospray. Because of its normal distribution, the releasing profile could be controlled by the ultrasound time. The thinnest porous titania shell was first fractured and crumbled by ultrasound for a short time. With (29) Chang, M. W.; Stride, E.; Edirsinghe, M. Langmuir 2010, 26, 5115–5121.
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Figure 3. Magnetization curve of the magnetic titania capsule. (The insert shows the photographs of the capsules suspension’s magnetic property.)
increasing ultrasound time, the thicker shell would be destroyed. The organic/inorganic contents weight percentage of multicomponent capsules is exhibited by the TGA curve in Figure S1 of the Supporting Information. The weight of the complex capsules decreased slowly initially because of the evaporation of solvent residue and absorbed moisture. The sharp descent occurred at 300 °C and ends at 450 °C, when the organics start chain-breaking and burning; finally, 12% inorganic materials like TiO2 and Fe3O4 was left. The final organic/inorganic weight percentage DOI: 10.1021/la1042734
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Figure 4. Photographs of (a) cuvette with graphene quantum-dots-loaded capsule suspension, (b) cuvette with a magnet-targeted adsorption, and (c) cuvette after ultrasound treatment under 365 nm UV light irradiation.
of capsules from TGA is 6.8125, corresponding to a theoretical ratio of 7.4708. Figure 3 illuminates the magnetic property of the multicomponent capsules. We estimated dried sample (10 mg) by vibrating the sample magnetometer at room temperature with the magnetic field sweeping from -18 000 to þ18 000 G. Superparamagnetic behavior of the microspheres was evidently proved by the measured results because the Fe3O4 particle encapsulated inside was an aggregation of 10 nm Fe3O4 nanospheres,30 where the saturation magnetization was ∼3 emu g-1. The presence of titania shell, oil, and other components dilutes the concentration of Fe3O4 particles, resulting in a lower saturation magnetization of the capsules than that of the pure Fe3O4 particles.31 The magnetic properties of the complex capsules were also tested by applying a magnet beside the cuvette, where the capsules were completely attracted to the side of the cuvette nearest to the magnet in ∼2 min, as displayed by the insert of Figure 3. Similar results can be found in references 9 and 15. Figure 4 vividly demonstrates the release behavior. A cuvette was charged with capsules (20 mg) in PBSD solution (5 mL), which was composed of 70 wt % PBS (pH 7.4) and 30 wt % DMSO. The blue fluorescence was generated by graphene quantum dots. The graphene QDs were chosen as the fluorescent particles for their excellent biocompatibility. As attractive and promising carbon QDs, they are much safer than traditional quantum dots such as inorganic QDs composed of heavy metals or toxic organic fluorescent dyes. Besides biocompatibility, the amphiphilic PEG layer outside of the graphene QDs provides better solubility in olive oil. The modified graphene QDs acts as tracer for capsule targeting and a marker for drug releasing when capsules are broken. Fluorescence penetrated through the shell of capsules can be detected by UV light excitation; after the capsules were adsorbed to the side of the cuvette near the magnet, the suspension became clear, and fluorescence disappeared; then, 1 h ultrasound was applied to the solution, resulting in graphene quantum dots released from the capsule, and stronger fluorescence appeared with a magnet placed beside the cuvette. With a drug loading content of 3.2 wt %, the encapsulation efficiency achieved 95%. The applied ultrasonic treatment condition and the releasing profile of paclitaxel from titania capsule were given in Figure 5. Under the control condition (without (30) Liu, J.; Sun, Z. K.; Deng, Y. H.; Zou, Y.; Li, C. Y.; Guo, X. H.; Xiong, L. Q.; Gao, Y.; Li, F. Y.; Zhao, D. Y. Angew. Chem., Int. Ed. 2009, 48, 5875–5879. (31) Liu, T. Y.; Hu, S. H.; Liu, K. H.; Liu, D. M.; Chen, S. Y. J. Controlled Release 2008, 126, 228–236.
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Figure 5. Drug release profile of paclitaxel-loaded capsules with and without ultrasound treatment and the applied ultrasound profile.
ultrasound treatment), paclitaxel was detected slightly released within the first 2 h, the subsequent release rate was rare. The initial leaking may be due to shell damage that happened in the processes of gathering or shaking during dissipation. Chen et al. have mentioned that the electrosprayed titania multichamber capsules can be broken easily by grinding for taking the SEM image.12 Under ultrasound condition, the initial period release behavior was almost the same as that in the control sample. We applied 15 min of ultrasound to sample suspension 5 h later; burst release appeared, then quiczkly stopped when all paclitaxel released from the cracked capsules. Another 5 h later, the left unbroken capsules was treated with 15 min of ultrasound again; the same burst release showed, but the amount of released paclitaxel decreased after the second irradiation. The reason that the drug release amount differed with the same irradiation period was that bigger capsules with thinner shell thickness collapsed in the first wave of ultrasound attack. The left smaller ones were loading with a smaller amount of drugs, leading to less release.32 After the repeated ultrasound application, no paclitaxel could be detected. Figure 6a-c shows the morphology of capsules changing with increasing ultrasound time. The insert in part b shows a magnified (32) Li, Z. Z.; Xu, S. A.; Wen, L. X.; Liu, F.; Liu, A. Q.; Wang, Q.; Sun, H. Y.; Yu, W.; Chen, J. F. J. Controlled Release 2006, 111, 81–88.
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Figure 6. (a) SEM image of the capsules ultrasound for 5 min. (b) SEM image of the capsules ultrasound for 15 min; the insert is a magnification of the capsules. (c) SEM image of the capsules ultrasound for 30 min. (d) Size distribution and capsules’ status of sample ultrasound for 5 min. (e) Size distribution and capsules’ status of sample ultrasound for 15 min. (f) Schematic illustration for procedures of shell cracking until drug release from the capsules with increasing ultrasound time.
SEM picture of capsule ultrasound for 15 min. Spheres that were >2 μm cracked both the inner and outer shell layers; the smaller one marked with the red circle just came off the outer layer shell. With the increasing ultrasound time, capsules kept cracking and finally all collapsed, as shown in Figure 6c. Figure 6d,e statistically analyze the capsules’ status corresponding to different sizes of group ultrasound for 5 and 15 min, respectively. Figure 6f presents the schematic ultrasound triggering mechanism. The porous titania shell was fractured and finally crumbled by the cavitation generated by ultrasound. Cavitation is a well-known effect of ultrasound: the formation, growth, and collapse of gas/vapor-filled microbubbles in liquids.33 In cavitation, the (33) Flint, E. B.; Suslick, K. S. Science 1991, 253, 1397–1399.
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bubble collapse is adiabatic, so the bubble serves to concentrate the acoustic energy leading to the local generation of extreme conditions of temperature and pressure within a short time. PVPk-30 in the outer layer of the titania shell dissolved in phosphatebuffered solution (PBS) buffer (pH 7.4), and at the same time, liquids penetrated through the porous shell and infused into the space between two shells. Cavitations created at both sides of the outer shell, leading to local “explosion” and cracking the surface of the capsule. With the increasing ultrasound time, the inner shell broke stepwise, and drugs in the oil phase got chance to interact with the releasing medium shown in Figure 6f. There is a tendency in Figure 6d,e that a higher percentage of bigger capsules’ shell comes off at the end of 5 min of ultrasound irradiation; then, more middle-sized ones start cracking at ultrasound of 15 min. At the DOI: 10.1021/la1042734
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same time, lesser intact capsules
