pubs.acs.org/Langmuir © 2010 American Chemical Society
Oxidation-Responsive Micelles Based on a Selenium-Containing Polymeric Superamphiphile Peng Han,† Ning Ma,† Huifeng Ren,† Huaping Xu,*,† Zhibo Li,‡ Zhiqiang Wang,*,† and Xi Zhang† †
Key Lab of Organic Optoelectronics & Molecular Engineering, Department of Chemistry, Tsinghua University, Beijing 100084, PR China, and ‡Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, PR China Received July 16, 2010. Revised Manuscript Received August 12, 2010 We have fabricated a polymeric superamphiphile based on the electrostatic interaction between the double hydrophilic block copolymer of poly(ethylene glycol)-b-acrylic acid (PEG-b-PAA) and a selenium-containing surfactant (SeQTA). The polymeric superamphiphiles are able to self-assemble to form micelles in solution. The micelles can be disassembled with the addition of 0.1% H2O2 because SeQTA is very sensitive to oxidation. The selenide group in SeQTA is oxidized into selenoxide (SeQTA-Ox) by H2O2, which makes the surfactant more hydrophilic, thus leading to the disassembly of the micelles. In addition, small guest molecules such as fluorescein sodium can be loaded into the micelles made from the polymeric superamphiphiles and released in a controlled way under mild oxidation conditions. This study represents a new way to fabricate stimuli-responsive superamphiphiles for controlled self-assembly and disassembly.
Introduction Amphiphilic block copolymers can self-assemble into welldefined organized aggregates, such as micelles or vesicles under different conditions, that have been widely used as containers and carriers for drug or gene delivery.1 Normally, the self-assembly of amphiphilic block copolymers involves using organic solvents and complicated preparation processes. To solve these problems, a new way to prepare block copolymer assemblies in aqueous solution based on electrostatic interactions was developed.2 In these systems, double-hydrophilic block copolymers with ionic and nonionic water-soluble segments and oppositely charged polyions or surfactants were mixed together. A polyion complex or a block ionomer complex was then formed on the basis of the electrostatic interaction between the charged blocks of copolymers and oppositely charged polyions or surfactants that make the ionic segments insoluble in water. Such complexes were found to form micelle-like or vesicle-like aggregates. The advantages of these systems include the avoidance of organic solvents, a simple preparation procedure, and the decrement of surfactants. Amphiphilicity, the basis for molecular self-assembly, can be altered by various kinds of external stimuli to tune the structure and function of the assemblies.3 The external stimuli can be light, *Corresponding authors. E-mail:
[email protected],
[email protected]. (1) (a) Zhang, L.; Eisenberg, A. Science 1995, 268, 1728. (b) Dou, H.; Jiang, M.; Peng, H.; Chen, D.; Hong, Y. Angew. Chem., Int. Ed. 2003, 42, 1516. (c) Zhao, C.; Winnik, M. A.; Riess, G.; Croucher, M. D. Langmuir 1990, 6, 514. (d) Xu, R.; Winnik, M. A.; Hallett, F. R.; Riess, G.; Croucher, M. D. Macromolecules 1991, 24, 87. (e) Zhou, Y.; Yan, D. Angew. Chem., Int. Ed. 2004, 43, 4896. (f ) Wang, Y.; Xu, H.; Ma, N.; Wang, Z.; Zhang, X.; Liu, J.; Shen, J. Langmuir 2006, 22, 5552. (2) (a) Harada, A.; Kataoka, K. Macromolecules 1995, 28, 5294. (b) Harada, A.; Kataoka, K. Science 1999, 283, 65. (c) Koide, A.; Kishimura, A.; Osada, K.; Jang, W. -D.; Yamasaki, Y.; Kataoka, K. J. Am. Chem. Soc. 2006, 128, 5988. (d) Bronich, T. K.; Kabanov, A. V.; Kabanov, V. A.; Yu, K.; Eisenberg, A. Macromolecules 1997, 30, 3519. (e) Kabanov, A. V.; Bronich, T. K.; Kabanov, V. A.; Yu, K.; Eisenberg, A. J. Am. Chem. Soc. 1998, 120, 9941. (f ) Bronich, T. K.; Popov, A. M.; Eisenberg, A.; Kabanov, V. A.; Kabanov, A. V. Langmuir 2000, 16, 481. (3) (a) Wang, Y.; Xu, H.; Zhang, X. Adv. Mater. 2009, 21, 2849. (b) Wang, Y.; Ma, N.; Wang, Z.; Zhang, X. Angew. Chem., Int. Ed. 2007, 46, 2823. (c) Wang, C.; Guo, Y.; Wang, Y.; Xu, H.; Wang, R.; Zhang, X. Angew. Chem., Int. Ed. 2009, 48, 8962. (d) Wang, C.; Guo, Y.; Wang, Y.; Xu, H.; Zhang, X. Chem. Commun. 2009, 5380. (e) Jiang, Y.; Wan, P.; Xu, H.; Wang, Z.; Zhang, X.; Smet, M. Langmuir 2009, 25, 10134.
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redox, and pH, which have been employed to adjust the amphiphilicity of the surfactants for controlled self-assembly and disassembly.4 Among them, redox-responsive aggregates have attracted wide interest because of their promising applications in controllable encapsulation and delivery in physiological environments.5 Organic selenium compounds have proven to be excellent candidates for redox-responsive materials for their good activity in the presence of oxidants or reductants.6 Recently, our group has reported a series of redox-responsive polymeric aggregates that are formed from main-chain-type selenide-containing amphiphilic triblock copolymers.7 These aggregates can respond rapidly to external redox stimuli and subsequently release the incorporated species under mild conditions. In this work, we describe how to fabricate polymeric superamphiphiles on the basis of the electrostatic interaction between the double hydrophilic block copolymer of poly(ethylene glycol)b-acrylic acid (PEG-b-PAA) and a selenium-containing surfactant (SeQTA), as shown in Scheme 1. The polymeric superamphiphiles are able to self-assemble to form micelles in solution. The micelles can be disassembled with the addition of oxidant because SeQTA is very sensitive to oxidation. The way to fabricate polymeric superamphiphiles avoids the complicated synthesis and purification of the polymers. Moreover, the simple backbone of selenium-containing surfactants allows us to control the function of the obtained aggregates precisely by simply varying the structure or amount of surfactants. (4) (a) Wang, Y.; Han, P.; Xu, H.; Wang, Z.; Zhang, X.; Kabanov, A. V. Langmuir 2010, 26, 709. (b) Wang, Y.; Zhang, M.; Moers, C.; Chen, S.; Xu, H.; Wang, Z.; Zhang, X.; Li, Z. Polymer 2009, 50, 4821. (5) (a) Napoli, A.; Valentini, M.; Tirelli, N.; M€uller, M.; Hubbell, J. A. Nat. Mater. 2004, 3, 183. (b) Rosslee, C.; Abbott, N. Anal. Chem. 2001, 73, 4808. (c) Rehor, A.; Hubbell, J. A.; Tirelli, N. Langmuir 2005, 21, 411. (d) Cerritelli, S.; Velluto, D.; Hubbell, J. A. Biomacromolecules 2007, 8, 1966. (e) Ryu, J.-H.; Roy, R.; Ventura, J.; Thayumanavan, S. Langmuir 2010, 26, 7086. (6) (a) Liotta, D. Organoselenium Chemistry, John Wiley & Sons, 1987. (b) Zhang, X.; Xu, H.; Dong, Z.; Wang, Y.; Liu, J.; Shen, J. J. Am. Chem. Soc. 2004, 126, 10556. (c) Xu, H.; Gao, J.; Wang, Y.; Smet, M.; Dehaen, W.; Zhang, X. Chem. Commun. 2006, 796. (7) (a) Ma, N.; Li, Y.; Xu, H.; Wang, Z.; Zhang, X. J. Am. Chem. Soc. 2010, 132, 442. (b) Ma, N.; Li, Y.; Ren, H.; Xu, H.; Li, Z.; Zhang, X. Polym. Chem. 2010, DOI: 10.1039/c0py00144a.
Published on Web 08/19/2010
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Scheme 1. Oxidation-Responsive Micelles Based on Polymeric Superamphiphiles Formed by SeQTA and PEG43-b-PAA153
Figure 1. Synthesis Routes of Selenium-Containing Surfactant.
Experimental Section Materials. Block copolymer PEG43-b-PAA153 was kindly provided by Dr. Yapei Wang.4 Selenium-containing surfactant was synthesized through the synthesis routes shown in Figure 1. Diphenyl diselenide (98%) was purchased from Acros. 11-Bromoundecanol (97%) was a product of Alfa Aesar. Other chemicals and solvents were analytical-grade products from Beijing Chemical Reagent Co., China, and were used without further purification unless stated. Synthesis. Synthesis of 1-Hydroxylundecylphenyl Selenide. Diphenyl diselenide (0.31 g, 1.0 mmol) was dissolved in 2 mL of anhydrous THF, and then 2 mL of an aqueous solution of 0.075 g (2.0 mmol) of sodium borohydride was added. The reaction was finished in a few minutes, and a colorless sodium phenylselenide (PhSeNa) solution was obtained. The obtained PhSeNa solution was immediately injected into 5 mL of a THF solution of 0.50 g (2.0 mmol) of 11-bromoundecanol, and the reaction was performed at 50 °C for 12 h under argon flow. Then the product was extracted with CH2Cl2 and purified by column chromatography with CH2Cl2 as the eluent, and a white powder Langmuir 2010, 26(18), 14414–14418
was obtained with a yield of 82%. 1H NMR (300 MHz, CDCl3) δ: 7.47 (2H, d, aromatic), 7.23 (3H, t, aromatic), 3.64 (2H, t, HOCH2), 2.91 (2H, t, SeCH2), 1.70 (2H, t, HOCH2CH2), 1.56 (2H, t, CH2CH2Se), 1.42-1.20 (14H, m, HOCH2CH2(CH2)7CH2CH2Se). Synthesis of 1-Bromoundecylphenyl Selenide. 1-Hydroxylundecylphenyl selenide (0.51 g, 1.6 mmol) was dissolved in 10 mL of dried CH2Cl2, and the solution was degassed with argon for 10 min. Then 0.19 mL (2.0 mmol) of phosphorus tribromide was added. The reaction was allowed to stir at room temperature (∼20 °C) for 2 h. The reaction was terminated with water, and the product was extracted with CH2Cl2 and purified by column chromatography with a mixture of 10:1 petroleum ether/CH2Cl2 as the eluent, and a white powder was obtained with a yield of 70%. 1H NMR (300 MHz, CDCl3) δ: 7.47 (2H, d, aromatic), 7.23 (3H, t, aromatic), 3.41 (2H, t, BrCH2), 2.91 (2H, t, SeCH2), 1.84 (2H, t, BrCH2CH2), 1.70 (2H, t, CH2CH2Se), 1.43-1.20 (14H, m, BrCH2CH2(CH2)7CH2CH2Se).
Synthesis of Phenylselenide-1-undecyl Triethylamonium Bromide (SeQTA). 1-Bromoundecylphenyl selenide (0.43 g, 1.1 mmol) was dissolved in 10 mL of dried CH3CN, and 0.50 mL of triethylamine was added. The solution was degassed with argon for 10 min and put into an oil bath at 70 °C to react for 8 h. The product was purified by dissolving and precipitating in diethyl ether two times, thus a white powder was obtained in a yield of 90%. 1H NMR (300 MHz, CDCl3) δ: 7.48 (2H, d, aromatic), 7.24 (3H, t, aromatic), 3.50 (6H, t, CH3CH2N), 3.27 (2H, t, Et3NCH2), 2.91 (2H, t, SeCH2), 1.70 (4H, m, Et3NCH2CH2 and CH2CH2Se), 1.40 (9H, t, CH3CH2N), 1.451.20 (14H, m, Et3NCH2CH2(CH2)7CH2CH2Se). 77Se-NMR (600 MHz, CDCl3, diphenyldiselenide as reference (δ = 464.1)) DOI: 10.1021/la102837a
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Letter δ: 290.1. ESI-MS: [M - Br]þ calculated 412.25, found 412.30. Instrumentation. 1H NMR and 77Se-NMR spectra were recorded using a JEOL JNM-ECA300 and JEOL JNMECA600 spectrometer, respectively. Electrospray ionization mass spectrometry (ESI-MS) was performed using a PE SciexAPI 3000 spectrometer. Fourier transform infrared (FT-IR) spectra were collected on a Bruker IFS 66 V instrument equipped with a DTGS detector. Cryo-TEM samples were prepared in a controlled environment vitrification system (CEVS) at 28 °C. The vitrified samples were then stored in liquid nitrogen until they were transferred to a cryogenic sample holder (Gatan 626) and examined by a JEM 2200FS TEM (200 kV) at about -174 °C. The size distribution of the aggregates in aqueous solution through dynamic light scattering (DLS) was confirmed by ALV/DLS/SLS5022F. The light wavelength was set at 632.8 nm, and the sample was centrifuged 12 000 rpm for 30 min before the measurement. Critical Micelle Concentration. The dependence of the solution conductivity on the surfactant concentration was used to determine the critical micelle concentration (cmc) of SeQTA and its oxidant. Typically, the slope of the change in conductivity versus concentration below the cmc is steeper than the slope above the cmc. Therefore, the junction of the conductivityconcentration plot represents the cmc value. Preparation of the Polymeric Superamphiphile. PEG43-bPAA153 was dissolved in a pH 10 aqueous solution with a concentration of 0.06 mg/mL in which the molar concentration of carboxylate group is 7.3 10-4 M. SeQTA was dissolved in water with a concentration of 1.2 10-4 M. To prepare the polymeric superamphiphile, 0.5 mL of PEG43-b-PAA153 was added to a tube, followed by the addition of SeQTA solution and water until the total volume reached 5 mL. The molar ratio of positive charge on SeQTA and negative charge on carboxylate groups is defined as Z (Z= [SeQTA]/[COO-]), which can represent the composition of the mixture. For example, when 1 mL of SeQTA is mixed with 0.5 mL of PEG43-b-PAA153 and further diluted to 5 mL with water, the final concentrations of SeQTA and carboxylate groups are 2.4 10-5 and 7.3 10-5 M, respectively, and the Z value is about 1:3. Loading and Releasing of Fluorescein Sodium. The loading procedure was described as follows. Four components, 1 mL of PEG43-b-PAA153 (0.06 mg/mL), 1 mL of fluorescein sodium (FluNa, 2 10-4 M), 2 mL of SeQTA (1.2 10-4 M), and 6 mL of water, were mixed. The Z value in the mixture was 1:3. Second, after the mixture was stirred for about an hour, it was dialyzed against water to remove the excess FluNa that was not incorporated into the micelles. The dialysis was performed by changing the water from time to time until it reached equilibrium (usually 2 or 3 days). For the release of FluNa, 1 mL of the mixture after dialysis was trapped in the dialysis bag. Then the dialysis bag was put into a beaker with 15 mL of 0.1% H2O2. At a certain time, the solution in the beaker was monitored by fluorescence spectroscopy.
Results and Discussion Self-Assembly of SeQTA before and after Oxidation. SeQTA is a typical surfactant and can self-aggregate in water. Because of the good redox activity of selenide moieties, it is greatly anticipated that the amphiphilicity of SeQTA can be effectively changed upon oxidation. To confirm this, the selenium-containing surfactants are first allowed to self-assemble in aqueous solutions to determine their oxidation responsiveness. The oxidants of SeQTA were depicted as SeQTA-Ox, which was obtained by complete oxidation of the SeQTA solution with 1% H2O2. From the results of cmc measurements of solution conductivity, the cmc of SeQTA was 4.1 10-5 M. However, the cmc of SeQTA-Ox increased to 2.9 10-4 M, which is around 7 times that before oxidation. These results show that the amphiphilicity 14416 DOI: 10.1021/la102837a
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Figure 2. Size distribution of SeQTA and SeQTA-Ox by DLS measurement. the concentration of the solution is 1 10-3 M.
of SeQTA has changed after oxidation (i.e., it has become more hydrophilic). The change in amphiphilicity can influence the size of the aggregates as supported by DLS. As shown in Figure 2, the average radius of the aggregates in the SeQTA 1 10-3 M solution was 61.9 nm. After oxidation with 1% H2O2, the average radius of the aggregates changed to 39.7 nm. It should be noted that DLS was performed at a concentration of 1 10-3 M because the scattered light was very weak under a lower concentration, which was beyond the detection of the machine. To confirm what SeQTA was converted to after oxidation, 77 Se-NMR, ESI-MS, and FT-IR were employed to characterize the oxidized product. From 77Se-NMR shown in Figure 3a, the peak shifted from 290.1 to 1024.9 ppm after oxidation, indicating that the products contain selenoxide groups.8 In ESI-MS, the molecular ion peak (eliminating Br-) was found at 428.29, which is about 16 m/z higher than for SeQTA, as shown in Figure 3b. These results indicate that selenide was oxidized to selenoxide (i.e., only one oxygen atom was added to the selenium). Further FT-IR results shown in Figure 3c show that an extra peak at 847 cm-1 appeared, which is a characteristic peak of selenoxide.9 From all of these results, we can conclude that the selenide group in SeQTA is oxidized to selenoxide . Formation of Polymeric Superamphiphile Micelles. The strategy employed to prepare polymeric superamphiphiles is illustrated in Scheme 1. The selenium-containing surfactant, SeQTA, and the block copolymer, PEG43-b-PAA153, used as its sodium salt are both water-soluble. PEG43-b-PAA153 cannot form any aggregates in water because of the good water solubility of the two segments. The polymeric superamphiphile is formed by mixing SeQTA and PEG43-b-PAA153 on the basis of electrostatic attraction. The polymeric superamphiphile was first prepared with Z = 1:3, and the cryo-TEM measurements were performed to observe the structure of the assemblies. As shown in Figure 4a, spherical micellar aggregates with solid cores were formed during the self-assembly process, which have an average diameter of ∼52 nm as shown in Figure 4d. To investigate further the relationship between the self-assembled structure and Z value, we also prepared the polymeric superamphiphile with Z values of 1:2 and 1:1 and observed their structures with cryo-TEM. When Z was 1:2, micellar aggregates with similar structures and sizes were obtained. Although Z became 1:1, the average size of the micelles increased to ∼61 nm, indicating that the Z value was effective for the micellar structure. It should be noted that for the Z = 1:3 and 1:2 mixtures the concentrations of SeQTA in the solution were (8) Xu, H.; Huang, K. Selenium: Its Chemistry, Biochemistry and Application in Life Science (in Chinese); Huazhong University of Science and Technology Press: Wuhan, China, 1994. (9) Socrates, G. Sulphur and Selenium Compounds. Infrared Characteristic Group Frequencies; John Wiley & Sons: New York, 1980; Chapter 16.
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Figure 3.
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Se-NMR of SeQTA and SeQTA-Ox (a), ESI-MS of SeQTA-Ox (b), and FT-IR of SeQTA and SeQTA-Ox (c).
Figure 4. Cryo-TEM results of the mixture, with Z values of (a) 1:3, (b) 1:2, and (c) 1:1. The concentration of SeQTA is (a) 2.4 10-5, (b) 3.6 10-5, and (c) 7.2 10-5 M, and the concentration of PEG43-b-PAA153 is kept at 0.006 mg/mL. (d-f ) Size distribution determined by counting the number of micelles in a-c, respectively.
under its cmc. Such concentrations ensured the formation of polymeric superamphiphiles instead of the aggregation of SeQTA itself. However, for the Z = 1:1 mixture the concentration of SeQTA was a little above its cmc but still lower than the cmc of SeQTA-Ox. Cryo-TEM results show that a similar micellar assembly was formed, which means that the formation of the polymeric superamphiphile is dominant. The above results indicate that the self-assembled structure of these polymeric superamphiphiles can be easily adjusted by simply varying the number of selenium-containing surfactants during the preparation process. We wonder what would happen to the micelles when oxidant was added. In answer to this question, we have employed cryoTEM to characterize the oxidized products once again. The micelles disappeared an hour after oxidation with 0.1% H2O2, and no aggregates were observed (data not shown). This is understandable because SeQTA became more hydrophilic after oxidation, which makes the polymeric superamphiphile not as stable as before and causes it to disassemble into free PEG43-bPAA153 and oxidized SeQTA. Therefore, the formed micelles are very sensitive to oxidants and disassemble after oxidation. Compared with the oxidation responsiveness of aggregates formed from small amphiphile SeQTA, the oxidation responsiveness of micelles formed from the polymeric superamphiphile is more sensitive. This is because after oxidation more hydrophilic molecule SeQTA-Ox has a stronger interaction with the solvent other than interacting with PEG-b-PAA, leading to the disassembly of superamphiphiles. From this point of view, the polymeric Langmuir 2010, 26(18), 14414–14418
superamphiphile is a better stimuli-responsive entity than the small surfactant amphiphile. Loading and Releasing of Fluorescein Sodium (FluNa). We wonder whether such micelles formed by the polymeric superamphiphile of PEG43-b-PAA153 and SeQTA can be used to incorporate guest molecules and release them under mild oxidation. For this purpose, fluorescein sodium was chosen as a fluorescent probe model molecule and incorporated into the polymeric superamphiphile micelles. In the experiments, FluNa was first codissolved with PEG43-b-PAA153 upon addition of SeQTA under stirring. The obtained polymeric superamphiphile aggregates were allowed to be dialyzed against water to remove the unloaded FluNa. Fluorescence spectroscopy results show that FluNa is successfully incorporated into the micelles. The maximum loading amounts of FluNa in different mixtures are 1.4 10-7 M (Z = 1:3), 2.5 10-7 M (Z = 1:2), and 2.6 10-7 M (Z = 1:1) FluNa. The release behavior of FluNa in 0.1% H2O2 was investigated by monitoring the fluorescence emission intensity of the solution outside the dialysis bag. Figure 5 shows that FluNa was released in a controlled way. It took about 10 h to reach release equilibrium. Moreover, it is interestingly shown that the release rate of FluNa is tunable upon the complex recipe during the self-assembly procedure. For example, in a micellar solution of Z = 1:3, the release of FluNa was a little faster than that of Z = 1:2. This difference can be attributed to the different contents of SeQTA in the polymeric superamphiphiles, which endow the obtained micelles with different stabilities in the presence DOI: 10.1021/la102837a
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our system is within the appropriate concentration of seleniumcontaining compounds used as drugs.8,10
Conclusions We have shown that oxidation-responsive micelles of polymeric superamphiphiles can be formed between a seleniumcontaining surfactant and double-hydrophilic block copolymers. The micelles disassembled when 0.1% H2O2 was added because selenide was oxidized into selenoxide, which is more hydrophilic. Such micelles can be used as nanocontainers to load guest molecules, and the guest molecules can be released in a controlled way under mild oxidation. It is anticipated that this system may find applications in cargo delivery and release. Figure 5. Release of FluNa in the polymeric superamphiphile micelles under the mild oxidation of 0.1% H2O2.
of an oxidation stimulus. This unique property clearly shows one advantage of the polymeric superamphiphile system by small surfactants and block copolymers, which can precisely adjust the structure and properties of the assemblies by simply changing the complex recipe in the self-assembly process. As far as the toxicity of organoselenium is concerned, it is concentration-dependent. The maximum amount of selenium (5.7 μg/mL) that we used in
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Acknowledgment. This work was financially supported by the National Basic Research Program of China (2007CB808000), the National Natural Science Foundation of China (20974059, 50973051, and 20904028), the Tsinghua University Initiative Scientific Research Program (2009THZ02-2), the Doctoral Fund of Ministry of Education of China, and the joint project between NSFC and DFG (TRR61). (10) Nogueira, C. W.; Zeni, G.; Rocha, J. B. T. Chem. Rev. 2004, 104, 6255.
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