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Vesicle-to-Spherical Micelle-to-Tubular Nanostructure Transition of Monomethoxy-poly(ethylene glycol)-poly(trimethylene carbonate) Diblock Copolymer So Young Kim,† Kyung Eun Lee,‡ Sung Sik Han,‡ and Byeongmoon Jeong*,† Department of Chemistry, DiVision of Nano Sciences, Ewha Womans UniVersity, Daehyun-Dong, Seodaemun-Ku, Seoul, 120-750, Korea, and DiVision of Life Science, College of Life Science and Biotechnology, Korea UniVersity, Anam-Dong, Seongbuk-Ku, Seoul, 136-701, Korea ReceiVed: NoVember 29, 2007; ReVised Manuscript ReceiVed: April 11, 2008
Recently, we reported a temperature-sensitive biodegradable diblock copolymer of monomethoxy-poly(ethylene glycol)-b-poly(trimethylene carbonate) (mPEG-PTMC; Macromolecules 2007, 40, 5519-5525). In this paper, we report the detailed morphological transition of the polymer in water as a function of polymer concentration and temperature, using cryo-transmission electron microscopy (cryo-TEM). At a low polymer concentration (0.05 wt %), the mPEG-PTMC diblock copolymers formed vesicles in water. On the other hand, vesicle-tomicelle transition was observed as the polymer concentration increased. The polymer predominantly formed micelles above 2.0 wt %. In the 2.0 wt % polymer solution, the mPEG-PTMC underwent spherical micelleto-tubular nanostructure transition as the temperature increased from 10 to 40 °C, and the transition accompanied an increase in turbidity of the polymer aqueous solution due to the increase in the apparent size of the polymer aggregates. Here, we report that the morphology of vesicles, spherical micelles, and tubular nanostructures is reversibly controlled by a thermosensitive polymer of mPEG-PTMC and the variation of the morphology can be carefully traced by using cryo-TEM. This paper will not only provide an important method for morphological control of an amphiphilic polymer but also improve our understanding of a temperature-sensitive transition mechanism of the polymer. Introduction Morphological control among the various forms of vesicles or micelles has been a hot topic during the past decade because they can be utilized to fabricate various nanostructured materials such as nanoparticles, nanofibers, and mesoporous silica.1–3 Amphiphilic block copolymers adopt various morphologies in water depending on the size and shape of each block, polymer concentration, and temperature. The predominant species of the amphiphilic block copolymer in water are micelles and vesicles. The more favorable morphology and the transition between micelles and vesicles are usually predicted by a packing parameter. which is a measure of the relative size of a hydrophobic tail to a polar headgroup of an amphiphilic surfactant.4,5 Micelle formation is favored when any factors decrease the packing parameter, whereas vesicles are formed when any factors increase the packing parameter.6–10 Thanks to the application of the cryo-transmission electron microscope (cryo-TEM) technique, the micelles or vesicles can be visualized as they are in a solvent.11,12 By the traditional solvent evaporation method, morphologies of the micelles or vesicles in the TEM images may be different from those in a solvent. Now, we are reporting control of various morphologies of an amphiphilic surfactant by use of a temperature-sensitive block copolymer of monomethoxy-poly(ethylene glycol)-b-poly(trimethylene carbonate) (mPEG-PTMC). The PEG blocks are hydrophilic and are exposed to the aqueous environment, whereas the PTMC blocks are hydrophobic and form an inner * Corresponding author: fax 82-2-3277-3411; e-mail
[email protected]. † Ewha Womans University. ‡ Korea University.
core of the self-assembled structures in water. We investigated the morphology of the block copolymer by cryo-TEM, dynamic light scattering, and UV-vis spectroscopy. Experimental Section Materials. The synthetic procedure for mPEG-PTMC (550-2750) diblock copolymers was published previously.13 The number-average molecular weights of the polymer are 3300 (as denoted 550-2750 for each block) by 1H NMR and 4700 by gel-permeation chromatography against polystyrene standards. The polydispersity index, defined by the ratio of the weight-average molecular weight to the number-average molecular weight of mPEG-PTMC was 1.4. Cryo-Transmission Electron Microscopy. Samples were equilibrated at given temperatures of 10, 20, 30, and 40 °C in the Vitrobot (FEI) for 20 min under saturated humidity conditions to avoid evaporation of water during specimen preparation. Vitrified specimens were prepared on the 200 mesh copper grid coated with a perforated form film (Ted Pella). A small drop (7 µL) was applied to the grid and blotted with filter paper to form a thin liquid film of solution, which was immediately plunged into liquid ethane at -170 °C. The procedure was performed automatically in the Vitrobot. The vitrified specimens were studied on a FEI Tecnai G2 TEM, at 200 kV with a Gatan cryoholder maintained below -170 °C, and images were recorded on an Ultrascan 2K × 2K CCD camera. In the microscopes, images were recorded with the Digital Micrograph software package under low-dose conditions to minimize damage by the electron beam radiation. Dynamic Light Scattering. The apparent size of a polymer aggregate in water was studied by a dynamic light scattering instrument (ALV 5000, 60 × 0) as a function of polymer concentration at 10 °C. A YAG DPSS-200 laser (Langen,
10.1021/jp711291x CCC: $40.75 2008 American Chemical Society Published on Web 06/04/2008
Morphological Transition of mPEG-PTMC Copolymer
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Figure 1. Cryo-TEM images of mPEG-PTMC diblock copolymer aqueous solution (0.05 wt %) as a function of temperature at (a) 10, (b) 20, (c) 30, and (d) 40 °C. The scale bar is 100 nm.
Figure 2. Cryo-TEM images of mPEG-PTMC diblock copolymer aqueous solutions (10 °C) as a function of concentration at (a) 0.05, (b) 2.0, and (c) 10 wt %. The scale bar is 100 nm.
UV-Vis Spectroscopy. Aqueous polymer solutions (1.0 mL of 0.05 and 2.0 wt %) were prepared and the absorbance at 500 nm was recorded as a function of temperature in a range of 10∼50 °C. Results and Discussion
Figure 3. Apparent aggregate size of mPEG-PTMC diblock copolymer in water as a function of concentration at 10 °C, determined by dynamic light scattering.
Germany) operating at 532 nm was used as a light source. Measurements of scattered light were made at an angle of 90° to the incident beam. The results of dynamic light scattering were analyzed by the regularized CONTIN method. The decay rate distributions were transformed to an apparent diffusion coefficient. From the diffusion coefficient, the apparent hydrodynamic size of a polymer aggregate can be obtained by the Stokes-Einstein equation.
Previously, our study based on dynamic light scattering showed a decrease in the apparent aggregate size of the mPEGPTMC diblock copolymer at 0.05 wt % as the temperature increased from 20 to 45 °C.13 Currently, we prove that such a transition is caused by the morphological change of the polymer aggregates (Figure 1). At 0.05 wt % aqueous solution, the polymers formed vesicles with diameters of 40∼100 nm (Figure 1a). As the temperature increased to 30 °C, the vesicles were shrunken to a size of about 20∼40 nm (Figure 1c). The different contrast between shell and core in the cryo-TEM images of the vesicles (Figure 1c) clearly distinguished the micelles (Figure 2b,c) that showed a uniform contrast over the entire spherical image. After a further increase in temperature to 40 °C, vesicles turned into smaller particles with a size of 5∼15 nm (Figure 1d).
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Kim et al.
Figure 4. Cryo-TEM images of mPEG-PTMC diblock copolymer aqueous solutions (2.0 wt. %) showing (a) spherical micelles at 10 °C and (b) tubular nanostructure at 40 °C. The scale bar is 100 nm.
Figure 5. Absorbance at 500 nm of the mPEG-PTMC diblock copolymer aqueous solutions (0.05 and 2.0 wt %) as a function of temperature. Inset photos show (left) the bluish translucent aqueous solution (2.0 wt %) of mPEG-PTMC at 10 °C and (right) the stable suspension at 40 °C.
PEG with a small molecular weight such as 550 is dehydrated as the temperature increases, and such a trend is more significant when it is chemically bonded to a hydrophobic molecule as reported by a decrease in intrinsic viscosity of PEG as the temperature increased.14 The shrinkage of the PEG portion of mPEG-PTMC was reflected in the 13C NMR spectra of the polymer in D2O.13 The PEG peak at 69∼71 ppm in the 13C NMR spectra collapsed as the temperature increased from 20 to 40 °C. The micelle-to-vesicle transition has been extensively reported when the relative size of the polar head to the hydrophobic tail of a surfactant system decreases.6–10,15 For example, when hydrophobic counterions were added to ionic surfactants, ion pairs formed. They were incorporated into the hydrophobic tail and the micelle-to-vesicle transition occurred due to the increase in the hydrophobic tail volume.6 As the temperature increased, the ion pairs dissociated and a vesicle-to-micelle transition occurred.7,8 In geminal surfactants containing amine groups as a polar head, the hydrodynamic size of the polar headgroup increased as the pH decreased due to the protonation of the polar
head. Thus, vesicle-to-micelle transition of the surfactant occurred as the pH decreased from 7 to 2.9 When lipids and linear surfactants were mixed, the vesicle-to-micelle transition occurred as the composition of linear surfactants increased.15 Therefore, the possibility of vesicle-to-micelle transition can be excluded for mPEG-PTMC as the temperature increases. Instead, the mPEG-PTMC underwent the vesicle-to-shrunken vesicle-to-nanoparticle transition as the temperature increased. As the polymer concentration increased, vesicle-to-micelle transition was observed at 10 °C. A concentration-dependent morphological change between micelles and vesicles was also reported for oleyldimethylamine oxide aqueous solutions.16 The oleyldimethylamine oxide underwent a micelle-to-vesicle transition as the surfactant concentration increased. Several mechanisms of vesicle-to-micelle transition, including monomer diffusion and collision of vesicles, were suggested.17,18 A twostep theoretical model for the transition involves rapid formation of disklike intermediates, growth of micelles, and closure of micelles to form vesicles.19 The current mPEG-PTMC polymer has a small hydrophilic PEG block (MW ) 550) relative to the long hydrophobic PTMC block (MW ) 2750). Therefore, crewcut vesicles are formed at low concentrations. The vesicles have a dynamic structure between unimers and vesicles. As the concentration of mPEG-PTMC increases, they are reorganized to spherical micelles. Above 2.0 wt %, micelles with a size of 20∼50 nm were the predominant species in water, and only micelles were present in the cryo-TEM images of 10 wt % polymer aqueous solution at 10 °C (Figure 2). Dynamic light scattering results supported such a trend (Figure 3). Because the TEM images at 10 °C show spherical vesicles and spherical micelles, dynamic light scattering was carried out at an angle of 90° to the incident beam. As the polymer concentration increased, the most probable size of polymer aggregates decreased from 80 nm (0.05 wt %) to 40 nm (2.0 wt %) to 25 nm (10 wt %). The apparent size is correlated to the cryo-TEM images.
Figure 6. Schematic presentation of the morphological control of mPEG-PTMC diblock copolymer by varying concentration and temperature.
Morphological Transition of mPEG-PTMC Copolymer As spherical micelles are the dominant species in water when the concentration is higher than 2.0 wt %, the morphological change of micelle was compared at 10 and 40 °C for 2.0 wt % mPEG-PTMC diblock copolymer aqueous solution. The spherical micelles at 10 °C (Figure 4a) turned into tubular nanostructures at 40 °C (Figure 4b). The tubular nanostructures with a diameter of 10∼15 nm and a length of several hundred nanometers were large enough to scatter visible light with a wavelength of 400∼700 nm, and the absorbance at 500 nm increased at 40 °C (Figure 5). This transition was reversible. As shown in the inset photos, the suspension of the polymer in water (2.0 wt %) at 40 °C is quite stable over several hours against the syneresis, which is macromolecular phase separation between water and polymer. The morphological change of mPEG-PTMC in water is shown schematically in Figure 6, along with cryo-TEM images. At low concentrations, the polymers form vesicles. As the concentration increases, they undergo vesicle-to-spherical micelle transition as confirmed by dynamic light scattering and cryo-TEM studies. The spherical micelles turn into tubular nanostructures as the temperature increases. Conclusions mPEG-PTMC is a temperature-sensitive amphiphilic polymer with a variety of morphologies depending on the concentration and temperature. By use of cryo-TEM, the morphological transition of the polymer was investigated. The mPEG-PTMC diblock copolymer formed vesicles at low concentration (0.05 wt. %) and low temperature (10 °C). The vesicle-to-micelle transition occurred as the polymer concentration increased above 2.0 wt %. The micelles underwent a spherical micelle-to- tubular nanostructure transition as the temperature increased from 10 to 40 °C. This paper not only suggested an excellent method to provide a variety of morphologies, including vesicle, spherical micelles,
J. Phys. Chem. B, Vol. 112, No. 25, 2008 7423 and tubular nanostructures, but also enhanced our understanding of the thermosensitive transition mechanism of the polymer at the same time. Acknowledgment. This work was supported by the MEST/ KOSEF (No. R11-2005-00800000-0), Seoul R & BD Program (10816), and MOEHRD (KRF-2005-041-C00300). References and Notes (1) Piao, Y.; Jang, Y.; Shokouhimehr, M.; Lee, I. S.; Hyun, T. H. Small 2007, 3, 255. (2) Hartgerink, J. D.; Beniash, E.; Stupp, S. I. Science 2001, 294, 1684. (3) Jan, J. S.; Shantz, D. F. AdV. Mater. 2007, 19, 2951. (4) Jain, S.; Bates, F. S. Science 2003, 300, 460. (5) Stuart, M. C.; Boekema, E. J. Biochim. Biophys. Acta 2007, 1768, 2681. (6) Kaler, E. W.; Herrington, K. L.; Murthy, A. K.; Zsadzinski, J. A. N. J. Phys. Chem. 1992, 96, 6698. (7) Salkar, R. A.; Hassan, P. A.; Samant, S. D.; Valaulikar, B. S.; Kumar, V. V.; Kern, F.; Candau, S. J.; Manohar, C. Chem. Commun. 1996, 1223. (8) Buwalda, R. T.; Stuart, M. C. A.; Enberts, J. B. F. N. Langmuir 2000, 16, 6780. (9) Johnsson, M.; Engberts, J. B. F. N. J. Phys. Org. Chem. 2004, 17, 934. (10) Bernheim-Groswasser, A.; Zana, R.; Talmon, Y. J. Phys. Chem. B 2000, 104, 12192. (11) Discher, B. M.; Won, Y. Y.; Ege, D. S.; Lee, J. C. M.; Bates, F. S.; Discher, D. E.; Hammer, D. A. Science 1999, 284, 1143. (12) Won, Y. Y.; Davis, H. T.; Bates, F. S. Science 1999, 283, 960. (13) Kim, S. Y.; Kim, H. J.; Lee, K. E.; Han, S. S.; Sohn, Y. S.; Jeong, B. Macromolecules 2007, 40, 5519. (14) Jeong, B., Jr.; Park, M. J.; Sohn, Y. S.; Gutowska, A.; Char, K. J. Phys. Chem. B 2003, 107, 10032. (15) Aratono, M.; Onimaru, N.; Yoshikai, Y.; Shigehisa, M.; Koga, I.; Wongwailikhit, K.; Ohta, A.; Takiue, T.; Lhoussaine, B.; Strey, R.; Takata, Y.; Villeneuve, M.; Matsubara, H. J. Phys. Chem. B 2007, 111, 107. (16) Miyahara, M.; Kawasaki, H.; Garamus, V.; Nemto, N.; Kakehasi, R.; Tanaka, S.; Annaka, M.; Maeda, H. Colloids Surf., B 2004, 38, 131. (17) Zhdanov, V. P.; Kasemo, B. Langmuir 2000, 16, 7352. (18) Johnsson, M.; Edwards, K. Biophys. J. 2003, 85, 3839. (19) Gradzielski, M. Curr. Opin. Colloid Interface Sci. 2004, 9, 256.
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