Article pubs.acs.org/Langmuir
Single-Molecule Force Spectroscopy of Selenium-Containing Amphiphilic Block Copolymer: Toward Disassembling the Polymer Micelles Xinxin Tan, Ying Yu, Kai Liu, Huaping Xu, Dongsheng Liu, Zhiqiang Wang, and Xi Zhang* Key Lab of Organic Optoelectronics and Molecular Engineering, Department of Chemistry, Tsinghua University, Beijing 100084, People’s Republic of China ABSTRACT: Selenium-containing polymers are a new type of responsive polymer material. Here, a selenium-containing amphiphilic block copolymer (PEG-PUSePEG) has been investigated using atomic force microscopy (AFM)-based singlemolecule force spectroscopy (SMFS). The deviation between force−extension curves of PEG-PUSe-PEG in water and in DMSO is found to be related to the disassembly of the micellar structures in water. SMFS experiments on PEG-PUSeox-PEG suggest that the change from selenide to oxidized selenone contributes significantly to the change in amphiphilicity, without obviously influencing the single-chain elasticity.
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INTRODUCTION Although atomic force microscopy (AFM) has been widely used to image microscale and nanoscale surface structures,1 it is also a force sensor with high sensitivity.2−6 AFM-based singlemolecule force spectroscopy (SMFS) can be employed to measure minute forces on the molecular scale and to record exceptionally small distances simultaneously. AFM-based SMFS was popularized in the study of single polymer chains7−22 because it is relatively easy to operate in comparison to other force techniques21−24 such as optical tweezers and the surfaceforces apparatus. The curves of force signals versus extension can be obtained, thus providing valuable information about intramolecular and intermolecular interactions of polymer systems on the single-chain level.23−38 Selenium-containing polymers are a new type of responsive polymer materials39 that can respond to either oxidants or reductants. Compared to other redox-responsive materials such as polysulfides, the redox responsiveness of seleniumcontaining polymers is more sensitive and faster.40−42 We have designed and synthesized a series of selenium-containing amphiphilic block copolymers that can form micellar structures by self-assembly. The micellar structures formed by seleniumcontaining amphiphilic block copolymers are promising candidates for drug-delivery vehicles.40−42 In addition, besides redox responsiveness, the micellar structures are radiationsensitive and can be destroyed under very low doses of radiation, therefore providing a kind of drug-delivery vehicle for combining chemotherapy and radiotherapy.43 In this article, we have attempted to employ AFM-based SMFS to study the single-chain mechanics of seleniumcontaining amphiphilic block copolymers. SMFS experiments of the selenium-containing amphiphilic block copolymers © 2012 American Chemical Society
before and after oxidation were carried out in different solvents. Therefore, the factors influencing the single-chain mechanics were summarized; moreover, the force that needs to disassemble the micellar structures could be obtained.
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EXPERIMENT SECTION
We have chosen a selenium-containing amphiphilic block copolymer, PEG-PUSe-PEG, as shown in Figure 1a. The molecular weight is 1.95 × 104 (Mw determined by GPC with polystyrene as a standard), and the polydispersity is 2.12.42 The selenium in PEG-PUSe-PEG could be oxidized to selenone (Figure 1b) while the polymeric structure was maintained. PEG-PUSe-PEG and its oxidation product (PEG-PUSeoxPEG in short) used in this research had been well characterized in our previous study.42 We first dissolved PEG-PUSe-PEG and PEG-PUSeox-PEG in THF and then diluted the solution in deionized water to get 1.0 × 10−4 mg/ mL PEG-PUSe-PEG and 1.0 × 10−4 mg/mL PEG-PUSeox-PEG solutions, respectively. The composition of THF/water in the solution was fixed at 1/100 v/v. Silicon wafers were treated with hot piranha solution (70:30 v/v 98% H2SO4/30% H2O2) for 1 h, sonicated in large amounts of deionized water several times, rinsed with ethanol, and dried in a steam of nitrogen. (Caution! Piranha solution is very corrosive and can react violently with organics, so security measures should be taken.) A clean silicon wafer was immersed in 1.0 × 10−4 mg/mL PEG-PUSePEG (or PEG-PUSeox-PEG) solution overnight, and then the silicon wafer was flushed thoroughly with deionized water. After being dried in a steam of nitrogen, the silicon wafer was used as a substrate in the SMFS experiment. The micelles of PEG-PUSe-PEG on a silicon substrate were observed by AFM. The observation was taken in air utilizing commercially available Si AFM cantilevers (Bruker, Santa Received: April 26, 2012 Revised: May 17, 2012 Published: June 4, 2012 9601
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Figure 1. (a) Chemical structure of PEG-PUSe-PEG. (b) The selenium in PEG-PUSe-PEG can be oxidized to selenone in 0.1% H2O2. Barbara, CA) with a sharp tip (radius of curvature 7 nm). As shown in Figure 2, PEG-PUSe-PEG does form micellar structures on the substrate, and the size of the micelles is about 50−80 nm, in agreement with the TEM result in our previous study.42
Figure 3. Typical force curves of PEG-PUSe-PEG at a stretching velocity of 2 μm/s in deionized water. (Inset) The force curves are normalized at the extension referring to the same force value of 300 pN and superimposed on each other.
rupture of a polymer bridge from the tip or substrate. Because the PEG-PUSe-PEG block copolymer is polydisperse and control of the point at which the polymer chain is picked up by the tip is not possible, the contour length of the stretched polymer chain is different. To compare the elasticity of polymer chains of different contour lengths, we have normalized the force curves at the extension referring to the same force value.22−24 As shown in the inset of Figure 3, all of the normalized force curves obtained from the block copolymer are superimposed well. This fact suggests that single-chain polymers are stretched in the experiment.7 The force curves can be simulated using a modified freely jointed chain (M-FJC) model based on the extended Langevin function.22,44 The M-FJC model treats a polymer as a chain of statistically independent rigid Kuhn segments with a length of lk (Kuhn length). The segments that can be deformed under stress are freely jointed without any long-range interactions. The extended Langevin function is shown below:
Figure 2. AFM observation of PEG-PUSe-PEG micelles formed on a silicon wafer. Commercially available V-shaped Si3N4 AFM cantilevers (Bruker, Santa Barbara, CA) with a sharp tip (radius of curvature 50 nm) at the end of a soft cantilever and a spring constant of 0.010−0.040 N/m were utilized in the experiment. The SMFS experiments were carried out at room temperature. The SMFS experiments in water were carried out by utilizing a 3D commercially available molecular force probe (Asylum Research, Santa Barbara, CA). The SMFS experiments in DMSO were carried out by utilizing a commercially available DI multimode picoforce (Bruker, Santa Barbara, CA). Details of the SMFS experiment have been described elsewhere.7−9 In brief, when an AFM tip was brought into contact with a polymer that had been physical absorbed on the substrate, the polymer could absorb onto the AFM tip as well as on the substrate, thus forming the so-called polymer bridge between the tip and the substrate. With the tip separated from the substrate, the polymer bridge was elongated, resulting in the bending of the cantilever toward the substrate. The deflection of the cantilever and the extension were recorded and converted to force−extension curves (in brief, force curves).
⎡ ⎛ Fl ⎞ k T ⎤⎛ nF ⎞ x(F ) = ⎢coth⎜ k ⎟ − B ⎥⎜Lc + ⎟ ⎢⎣ Flk ⎥⎦⎝ Ks ⎠ ⎝ kBT ⎠
Here, x represents the extension of a polymer chain (end-toend distance), F is the applied force on an individual polymer chain, Lc is the contour length of the polymer chain, lk is the length of the statistically independent segment, n is the number of segments being stretched, which equals Lc/lk, kB is the Boltzmann constant, and T is the temperature in Kelvin. The deformability of segments is characterized by the segment elasticity, Ksegment. As shown in Figure 4, in both the low-forces regime (below 40 pN) and the high-forces regime (above 600
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RESULTS AND DISCUSSION Typical force curves of PEG-PUSe-PEG block copolymer in deionized water using different cantilevers are shown in Figure 3. All of the force curves exhibit similar deformational characteristics: a sharp monotonically rising force with increasing extension and a rapid force dropping to zero upon 9602
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Figure 4. In both the low-forces regime (below 40 pN) and the highforces regime (above 600 pN), the M-FJC model can fit the normalized force curve of PEG-PUSe-PEG in water, and a deviation exists in the middle regime.
pN), the M-FJC model can fit the normalized force curve of PEG-PUSe-PEG in water. The elasticity of an M-FJC chain is dominated by either the entropic contribution at low forces or by the enthalpic contribution at high forces. Ksegment of the PEG-PUSe-PEG main chain is 50 nN/nm and lk is 0.42 nm, as fitted by the M-FJC model. It should be noted that there is a deviation between the force curve and the fitting curve in the middle regime from 40 to 600 pN. This fact suggests the disassembly of a suprastructure within the PEG-PUSe-PEG. Because water is a good solvent for PEG segments but a poor solvent for PUSe segments, PEGPUSe-PEG block copolymers can form micellar structures in water. In other words, the deviation of the force curve should be related to the disassembly of the micellar structure. To confirm the speculation above, we have done the following control experiment: single-chain stretching is carried out in DMSO, which is a good solvent for both PEG and PUSe segments. As shown in Figure 5a, it displays typical normalized force curves of PEG-PUSe-PEG in DMSO. The comparison of the force curves of PEG-PUSe-PEG in DMSO and in water is shown in Figure 5b. There are two points that need to be highlighted: (1) micellar structures cannot be formed in DMSO because it is a good solvent for both segments. Therefore, the force curves of PEG-PUSe-PEG in DMSO can be well fitted by the M-FJC model over all of the force ranges. (2) A deviation in the force curves between that in DMSO and that in water existed. These facts support our speculation that the deviation of the force curves is related to the disassembly of the micellar structures.35 We wondered if the force curves of PEG-PUSe-PEG would be different before and after oxidation. As mentioned, the dialkyl selenide in PEG-PUSe-PEG can be oxidized to selenone. The force curve of PEG-PUSeox-PEG in water is shown in Figure 6. Comparing with the force curves of PEG-PUSe-PEG in water before oxidation, we can see that there is a deviation in the middle force region. This fact indicates that PEG-PUSeoxPEG is more hydrophilic and that no micellar structures are formed in water. Notably, PEG-PUSe-PEG can be partially oxidized. In this case, the deviation in the force curve can disappear only partially as well, as seen in the green curve in between the force curves of PEG-PUSe-PEG before the oxidation and in the force curves of PEG-PUSeox-PEG in the fully oxidized state in water.
Figure 5. (a) Force curves of PEG-PUSe-PEG at a stretching velocity of 2 μm/s in DMSO superimposed on each other after normalization. (b) Comparison of force curves of PEG-PUSe-PEG in DMSO and in water. The normalized force curve of PEG-PUSe-PEG in DMSO can be well fitted by the M-FJC model.
Figure 6. Comparison of force curves of PEG-PUSe-PEG, PEGPUSeox-PEG, and partially oxidized PEG-PUSe-PEG in water. The velocity is 2 μm/s. The normalized force curve of PEG-PUSeox-PEG in water can be well fitted by the M-FJC model.
We have also compared the force curves of PEG-PUSe-PEG and PEG-PUSeox-PEG in DMSO. As shown in Figure 7a, the force curves of PEG-PUSeox-PEG in DMSO are superimposed on each other after normalization. As shown in Figure 7b, no significant difference is observed before and after oxidation. This suggests that, although it contributes significantly to the change in amphiphilicity, the change from selenide to oxidized selenone does not obviously influence the single-chain elasticity. 9603
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Dengli Qiu and Chao Zhou for help with the DI multimode picoforce measurement.
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(1) Binnig, G.; Quate, C. G.; Gerber, C. Atomic force microscope. Phys. Rev. Lett. 1986, 56, 930−933. (2) Florin, E.-L.; Moy, V. T.; Gaub, H. E. Adhesive forces between individual ligand−receptor pairs. Science 1994, 264, 415−417. (3) Lee, G. U.; Kidwell, D. A.; Colton, R. J. Sensing discrete streptavidin-biotin interactions with atomic force microscopy. Langmuir 1994, 10, 354−357. (4) Zhang, Y. H.; Zhao, D.; Tan, X. X.; Cao, T. B.; Zhang, X. AFM force mapping for characterizing patterns of electrostatic charges on SiO2 electrets. Langmuir 2010, 26, 11958−11962. (5) Liu, N. N.; Peng, B.; Yuan, L.; Su, Z. H.; Niu, Z. W.; Wang, Q.; Zhang, W. K.; Li, H. B.; Shen, J. C. Pulling genetic RNA out of tobacco mosaic virus using single-molecule force spectroscopy. J. Am. Chem. Soc. 2010, 132, 11036−11038. (6) Liu, K.; Song, Y.; Feng, W.; Liu, N. N.; Zhang, W. K.; Zhang, X. Extracting a single polyethylene oxide chain from a single crystal by a combination of atomic force microscopy imaging and single-molecule force spectroscopy: toward the investigation of molecular interactions in their condensed states. J. Am. Chem. Soc. 2011, 133, 3226−3229. (7) Rief, M.; Oesterhelt, F.; Heymann, B.; Gaub, H. E. Single molecule force spectroscopy on polysaccharides by atomic force microscopy. Science 1997, 275, 1295−1297. (8) Li, H. B.; Rief, M.; Oesterhelt, F.; Gaub, H. E. Single-molecule force spectroscopy on xanthan by AFM. Adv. Mater. 1998, 10, 316− 319. (9) Li, H. B.; Liu, B. B.; Zhang, X.; Gao, C. X.; Shen, J. C.; Zou, G. T. Single-molecule force spectroscopy on poly(acrylic acid) by AFM. Langmuir 1999, 15, 2120−2124. (10) Li, H. B.; Zhang, W. K.; Zhang, X.; Shen, J. C.; Liu, B. B.; Gao, C. X.; Zou, G. T. Single molecule force spectroscopy on poly(vinyl alcohol) by atomic force microscopy. Macromol. Rapid Commun. 1998, 19, 609−611. (11) Xu, Q. B.; Zou, S.; Zhang, W. K.; Zhang, X. Single-molecule force spectroscopy on carrageenan by means of AFM. Macromol. Rapid Commun. 2001, 22, 1163−1167. (12) Marszalek, P. E.; Li, H. B.; Oberhauser, A. F.; Fernandez, J. M. Chair-boat transitions in single polysaccharide molecules observed with force-clamp AFM. Proc. Natl. Acad. Sci. U.S.A. 2002, 99, 4278− 4283. (13) Cui, S. X.; Liu, C. J.; Zhang, W. K.; Zhang, X.; Wu, C. Desorption force per polystyrene segment in water. Macromolecules 2003, 36, 3779−3782. (14) Cui, S. X.; Liu, C. J.; Wang, Z. Q.; Zhang, X. Single molecule force spectroscopy on polyelectrolytes: effect of spacer on adhesion force and linear charge density on rigidity. Macromolecules 2004, 37, 946−953. (15) Zou, S.; Schönherr, H.; Vancso, J. Stretching and rupturing individual supramolecular polymer chains by AFM. Angew. Chem., Int. Ed. 2005, 44, 956−959. (16) Gunari, N.; Balazs, A. C.; Walker, G. C. Force-induced globulecoil transition in single polystyrene chains in water. J. Am. Chem. Soc. 2007, 129, 10046−10047. (17) Shi, W. Q.; Giannotti, M. I.; Zhang, X.; Hempenius, M. A.; Schonherr, H.; Vancso, G. J. Closed mechanoelectrochemical cycles of individual single-chain macromolecular motors by AFM. Angew. Chem., Int. Ed. 2007, 46, 8400−8404. (18) Yu, Y.; Zhang, Y. H.; Jiang, Z. H.; Zhang, X.; Zhang, H. M.; Wang, X. H. Full view of single-molecule force spectroscopy of polyaniline in oxidized, reduced and doped states. Langmuir 2009, 25, 10002−10006. (19) Li, H. B.; Cao, Y. Protein mechanics: from single molecules to functional biomaterials. Acc. Chem. Res. 2010, 43, 1331−1341. (20) Cui, S. X.; Pang, X. C.; Zhang, S.; Yu, Y.; Ma, H. W.; Zhang, X. Unexpected temperature-dependent single chain mechanics of poly(nisopropyl-acrylamide) in water. Langmuir 2012, 28, 5151−5157.
Figure 7. (a) Force curves of PEG-PUSeox-PEG at a stretching velocity of 2 μm/s in DMSO superimposed on each other after normalization. (b) Comparison of force curves of PEG-PUSe-PEG and PEG-PUSeox-PEG in DMSO.
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CONCLUSIONS We have investigated the single-chain mechanics of PEG-PUSePEG and PEG-PUSeox-PEG using AFM-based SMFS. The deviation between force−extension curves of PEG-PUSe-PEG in water and in DMSO is found to be related to the disassembly of the micellar structures in water. The change from selenide to oxidized selenone contributes significantly to the change in amphiphilicity, without obviously influencing the single-chain elasticity. Our study has given further insight into the disassembly mechanism of block-copolymer micelles. In addition, on the single-molecule level, we can obtain new information about how much small changes in the chemical structure can influence the amphiphilicity as well as the assembly behavior of selenium-containing amphiphilic block copolymers.
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REFERENCES
AUTHOR INFORMATION
Corresponding Author
*E-mail:
[email protected]. Notes
The authors declare no competing financial interest.
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ACKNOWLEDGMENTS This work was financially supported by the National Natural Foundation of China (20834003), an NSFC-DFG joint grant (TRR-61), and the National Basic Research Program (2007CB808000). We thank Dr. Ning Ma for the seleniumcontaining amphiphilic block copolymer samples and Dr. 9604
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(21) Janshoff, A.; Neitaert, M.; Oberdoerfer, Y.; Fuchs, H. Force spectroscopy of molecular systems-single molecule spectroscopy of polymers and biomolecules. Angew. Chem., Int. Ed. 2000, 39, 3212− 3237. (22) Hugel, T.; Seitz, M. The study of molecular interactions by AFM force spectroscopy. Macromol. Rapid Commun. 2001, 22, 989− 1016. (23) Zhang, W. K.; Zhang, X. Single molecule mechanochemistry of macromolecules. Prog. Polym. Sci. 2003, 28, 1271−1295. (24) Zhang, X.; Liu, C. J.; Wang, Z. Q. Force spectroscopy of polymers: studying on intramolecular and intermolecular interactions in single molecular level. Polymer 2008, 49, 3353−3361. (25) Zou, S.; Zhang, W. K.; Zhang, X.; Jiang, B. Z. Study on polymer micelles of hydrophobically modified ethyl hydroxyethyl cellulose using single-molecule force spectroscopy. Langmuir 2001, 17, 4799− 4808. (26) Fernandez, J. M.; Li, H. B. Force-clamp spectroscopy onitors the folding trajectory of a single protein. Science 2004, 303, 1674−1678. (27) Eckel, R.; Ros, R.; Decker, B.; Mattay, J.; Anselmetti, D. Supramolecular chemistry at the single-molecule Level. Angew. Chem., Int. Ed. 2005, 44, 484−488. (28) Liu, C. J.; Jiang, Z. H.; Zhang, Y. H.; Wang, Z. Q.; Zhang, X.; Feng, F.; Wang, S. Intercalation interactions between dsDNA and acridine studied by single molecule force spectroscopy. Langmuir 2007, 23, 9140−9142. (29) Zhang, Y. H.; Liu, C. J.; Shi, W. Q.; Wang, Z. Q.; Dai, L. M.; Zhang, X. Direct measurements of the interaction between pyrene and graphite in aqueous media by single molecule force spectroscopy: understanding the π−π interactions. Langmuir 2007, 23, 7911−7915. (30) Liu, C. J.; Jiang, Z. H.; Wang, Z. Q.; Zhang, X. The unwinding of surfactant-induced helical structure of carboxymethyl amylose by single molecule force spectroscopy. Polymer 2007, 48, 2030−2034. (31) Shi, W. Q.; Zhang, Y. H.; Liu, C. J.; Wang, Z. Q.; Zhang, X. Interaction between dendrons directly studied by single-molecule force spectroscopy. Langmuir 2008, 24, 1318−1323. (32) Anselmetti, D.; Bartels, F. W.; Becker, A.; Decker, B.; Eckel, R.; Mclntosh, M.; Mattay, J.; Plattner, P.; Ros, R.; Schäfer, C.; Sewald, N. Reverse engineering of an affinity-switchable molecular interaction characterized by atomic force microscopy single-molecule force spectroscopy. Langmuir 2008, 24, 1365−1370. (33) Zhang, Y. H.; Yu, Y.; Jiang, Z. H.; Xu, H. P.; Wang, Z. Q.; Zhang, X.; Oda, M.; Ishizuka, T.; Jiang, D. L.; Chi, L. F.; Fuchs, H. Single-molecule study on intermolecular interaction between C60 and porphyrin derivatives: toward understanding how strong the multivalency is. Langmuir 2009, 25, 6627−6632. (34) Zhang, Y. H.; Wang, Z. Q.; Zhang, X. Direct measurements of intermolecular interactions [J]. Acta Polym. Sin. 2009, 1, 973−979. (35) Yu, Y.; Wu, G. L.; Liu, K.; Zhang, X. Force required to disassemble block copolymer micelles in water. Langmuir 2010, 26, 9183−9186. (36) Jiang, Z. H.; Zhang, Y. H.; Yu, Y.; Wang, Z. Q.; Zhang, X.; Duan, X. R.; Wang, S. Study on intercalations between double-stranded DNA and pyrene by single-molecule force spectroscopy: toward the detection of mismatch in DNA. Langmuir 2010, 26, 13773−13777. (37) Schröder, T.; Geisler, T.; Schnatwinkel, B.; Anselmetti, D.; Mattay, J. Single-molecule force spectroscopy of supramolecular heterodimeric capsules. Phys. Chem. Chem. Phys. 2010, 12, 10981− 10987. (38) Zhang, W. K. AFM imaging and single-molecule force spectroscopy studies on macromolecular interactions at singlemolecule level [J]. Acta Polym. Sin. 2011, 0, 913−922. (39) Zhang, X.; Xu, H. P.; Dong, Z. Y.; Wang, Y. P.; Liu, J. Q.; Shen, J. C. Highly efficient dendrimer-based mimic of glutathione peroxidase. J. Am. Chem. Soc. 2004, 126, 10556−10557. (40) Ma, N.; Li, Y.; Xu, H. P.; Wang, Z. Q.; Zhang, X. Dual redox responsive assemblies formed from diselenide block copolymers. J. Am. Chem. Soc. 2010, 132, 442−443.
(41) Ren, H. F.; Wu, Y. T.; Ma, N.; Xu, H. P.; Zhang, X. Side-chain selenium-containing amphiphilic block copolymers: redox-controlled self-assembly and disassembly. Soft Matter 2012, 8, 1460−1466. (42) Ma, N.; Li, Y.; Ren, H. F.; Xu, H. P.; Li, Z. B.; Zhang, X. Selenium-containing block copolymers and their oxidation-responsive aggregates. Polym. Chem. 2010, 1, 1609−1614. (43) Ma, N.; Xu, H. P.; An, L. P.; Li, J.; Sun, Z. W.; Zhang, X. Radiation-sensitive diselenide block copolymer micellar aggregates: toward the combination of radiotherapy and chemotherapy. Langmuir 2011, 27, 5874−5878. (44) Smith, S. B.; Finzi, L.; Bustamante, C. Direct mechanical measurements of the elasticity of single DNA molecules by using magnetic bead. Science 1992, 258, 1122−1126.
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