Langmuir 2008, 24, 8501-8506
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Self-Assembly of Poly(ethylene oxide)-block-poly(acrylic acid) Induced by CaCl2: Mechanistic Study H. Ronny Sondjaja,† T. Alan Hatton,‡ and K. C. Tam*,§ Singapore-MIT Alliance, 4 Engineering DriVe 3, National UniVersity of Singapore, Singapore 117576, Singapore, Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA, Department of Chemical Engineering, UniVersity of Waterloo, Waterloo, Ontario, N2L3G1, Canada ReceiVed March 7, 2008. ReVised Manuscript ReceiVed May 6, 2008 The interaction between CaCl2 and double hydrophilic block copolymer, poly(ethylene oxide)45-block-poly(acrylic acid)70, PEO45-b-PAA70, was investigated. At a stoichiometric ratio of Ca2+:COO- ) 0.5, Ca2+ ions were bound to COO- groups on PAA segments via electrostatic interaction. Small particles of 4-8 nm in diameter were observed, suggesting the formation of coil-like polymeric globule induced by charge neutralization. At Ca2+:COO- g 2.5, monodispersed aggregates of average hydrodynamic diameter of 52.0 ( 7.4 nm were produced. The ISE, ITC, surface tension and fluorescence spectroscopic data confirmed that the formation of these aggregates is not the result of interaction between excess Ca2+ ions and the polymer, but rather it is due to changes in the water activity that triggers the structural rearrangement of Ca2+/PEO45-b-PAA70 complex.
Introduction Current progress in the controlled synthesis of nanomaterials using self-assembled double hydrophilic block copolymers (DHBCs) has attracted significant attention. These block copolymers are composed of two hydrophilic segments that can form self-associating structures when the system is triggered by changes in the physicochemical environment.1,2 Usually one block is composed of a polyelectrolyte while the second block promotes the stability of the nanostructure.3 The polyelectrolyte block provides interaction with various species, such as metal cations,4–6 drugs,7–10 proteins, and genes,11 thus facilitating the use of DHBCs as nanoscale carriers.9 Recently, investigations on the interaction between Ca2+ and DHBC containing acrylic-based polyanions were carried out to produce stable nanosized self-assembled structures.5,8,12,13 One common example of such a polymer is poly(ethylene oxide)block-poly(methacrylic acid) (PEO-b-PMAA), where the interaction between Ca2+ and COO- on the PMAA segment was * Corresponding author. E-mail:
[email protected]. † National University of Singapore. ‡ Massachusetts Institute of Technology. § University of Waterloo. (1) Cohen Stuart, M. A.; Hofs, B.; Voets, I. K.; De Keizer, A. Curr. Opin. Colloid Interface Sci. 2005, 10, 30. (2) Nakashima, K.; Bahadur, P. AdV. Colloid Interface Sci. 2006, 123-126, 75. (3) Co¨lfen, H. Macromol. Rapid Commun. 2001, 22, 219. (4) Bronstein, L. M.; Sidorov, S. N.; Gourkova, A. Y.; Valetsky, P. M.; Hartmann, J.; Breulmann, M.; Co¨lfen, H.; Antoinietti, M. Inorg. Chim. Acta 1998, 280, 348. (5) Li, Y.; Gong, Y.-K.; Nakashima, K.; Murata, Y. Langmuir 2002, 18, 6727. (6) Sanson, N.; Bouyer, F.; Ge´rardin, C.; In, M. Phys. Chem. Chem. Phys. 2004, 6, 1463. (7) Kataoka, K.; Kwon, G. S.; Yokoyama, M.; Okano, T.; Sakurai, Y. J. Controlled Release 1993, 24, 119. (8) Bronich, T. K.; Keifer, P. A.; Shlyakhtenko, L. S.; Kabanov, A. V. J. Am. Chem. Soc. 2005, 127, 8236. (9) Jiang, X.; Zhang, J.; Zhou, Y.; Xu, J.; Liu, S. J. Polym. Sci., A 2008, 46, 860. (10) Cheng, C.; Wei, H.; Shi, B.-X.; Cheng, H.; Li, C.; Gu, Z.-W.; Cheng, S.-X.; Zhang, X.-Z.; Zhuo, R.-X. Biomaterials 2008, 29, 497. (11) Kakizawa, Y.; Kataoka, K. AdV. Drug DeliVery ReV. 2002, 54, 203. (12) Bronich, T. K.; Bontha, S.; Shlyakhtenko, L. S.; Bromberg, L.; Hatton, T. A.; Kabanov, A. V. J. Drug Target. 2006, 14, 357. (13) Tjandra, W.; Yao, J.; Ravi, P.; Tam, K. C.; Alamsjah, A. Chem. Mater. 2005, 17, 4865.
reported to induce the formation of nanoaggregates containing Ca2+-PMAA cores stabilized by PEO shells. These nanoaggregates can be used directly as nanotemplates to produce nanosized biomaterials, such as calcium phosphate nanoparticles.13 The PMAA cores can also be cross-linked and dialyzed to form nanogels that can be further used as carriers in delivery systems.8 The argument that electrostatic interactions between Ca2+ ions and COO- produce a more hydrophobic environment, resulting in the onset of self-assembly is debatable. Although, in theory, the electrostatic interaction occurs between 1 mol of Ca2+ ions and 2 mol of COO- (Ca2+:COO- ) 0.5), the reported selfassembled nanoaggregates had to be prepared in the presence of excess CaCl2.5,8,12 Li et al.5 found that nanoaggregates of Ca2+/ PEO-b-PMAA with an average diameter of about 130 nm were formed when the ratio Ca2+:COO- was equal to or greater than 1, where it was believed that the methyl group on PMAA segments contributed to the enhanced hydrophobicity of the complex. Similarly, the Ca2+/PEO-b-PMAA and Ca2+/Pluronic-b-PAA nanogels reported by Bronich and co-workers8,12 were prepared at Ca2+:COO- ) 1.3. In this paper, we examined the self-assembly of DHBC nanoaggregates induced by Ca2+ ions using a well-defined PEOb-PAA block copolymer. Unlike PMAA, PAA does not possess additional methyl groups that contributes to the hydrophobicity of the system, and thus the formation of the aggregates should only be due to the presence of Ca2+ ions. Although the use of PEO-b-PAA as a calcite controlling agent has been studied,14 the mechanism of the self-assembly of nanoaggregates caused by the interaction between Ca2+ and PEO-b-PAA has not been reported. In the present study, we seek to elucidate the formation of such hybrid organic-inorganic systems and to propose a mechanism for the self-assembly behavior supported by new experimental data.
Experimental Section Materials. CaCl2 and other chemicals needed for polymer synthesis, such as triethyleneamine, 2-bromoisobutyryl bromide, (14) Guillemet, B.; Faatz, M.; Gro¨hn, F.; Wegner, G.; Gnanou, Y. Langmuir 2006, 22, 1875.
10.1021/la800727e CCC: $40.75 2008 American Chemical Society Published on Web 07/23/2008
8502 Langmuir, Vol. 24, No. 16, 2008 monomer tert-butylacrylate (t-BA), and Cu(I)Br, were purchased from Sigma-Aldrich. Methylated polyethylene oxide (mPEO) with hydroxyl end groups (Carbowax 2000) was donated by Dow Chemicals. Atom transfer radical polymerization (ATRP) was used to synthesize PEO-b-PtBA from mPEO followed by hydrolysis to transform it into PEO-b-PAA, as described previously.15 The synthesized polymer contained 45 PEO units and 70 PAA units, as analyzed by 1H NMR and potentiometric titration, and it is designated as PEO45-b-PAA70. Preparation of Ca2+/PEO-b-PAA System. In a typical experiment, 2 mL of 0.05% w/v PEO45-b-PAA70 was prepared in a test tube. The pH was adjusted according to the desired degree of deprotonation of the polymer R, and the polymer solution was filtered using a PTFE syringe filter of 0.45 µm pore size. The required amount of 0.5 M CaCl2 of similar pH was added to the polymer solution, where the volume of CaCl2 added was based on the desired Ca2+:COO- ratio, where the moles of COO- available was calculated from potentiometric titration data. The sample was vigorously mixed using a vortex mixer before the analysis was carried out. Ion Selective Electrode (ISE) Titration. The interactions between PEO-b-PAA and Ca2+ were investigated using the electromotive force (emf) technique coupled with a Ca2+ ion selective electrode (ISE) and Ag/AgCl electrode as the reference. In this experiment, a 50 mL solution containing 0.05% w/v PEO45-b-PAA70 prepared at the desired pH and R value was titrated with 0.5 M CaCl2 solution of identical pH. The titrations were conducted using a Radiometer Copenhagen ABU93 triburette system. Dynamic Light Scattering (DLS). The laser light scattering experiments were carried out using the Brookhaven laser light scattering system, consisting of a BI200SM goniometer, a vertically polarized 488 nm argon ion laser light source, a BI-9000AT digital correlator, and other supporting data acquisition and analysis software and accessories. Surface Tension Titration. A Data Physics DCAT 21 system tensiometer was employed to perform the surface tension measurements. The tensiometer is equipped with a 10 × 19.9 × 0.2 mm platinum-iridium Wilhelmy plate. In a typical experiment, a 50 mL solution of 0.05% w/v PEO45-b-PAA70 was prepared at the desired R value. A solution of 0.5 M CaCl2 of identical pH was used as a titrant with an injection schedule performed by SCAT interactive software. The surface tension measurement was conducted 30 s after each injection. Fluorescence Spectroscopy. The sample was prepared by mixing 6 µL of 0.1 mmol/L pyrene directly into test tubes containing 1 mL of polymer solution (pyrene concentration in the sample ) 6 × 10-7 M) at different concentrations of CaCl2. Steady state fluorescence spectra were recorded using the AMINCO Bowman Series 2 luminescence spectrometer with the detector set at 650 V. Excitation and emission wavelengths were preset at 335 and 373 nm, respectively. The bandwidth was set to 4 nm for excitation and 2 nm for emission. The emission profiles were scanned from 360 to 430 nm with a scan rate of 1.0 nm/s. The ratio of fluorescent intensity I1 at the 0-0 band (λ ) 372 nm) and I3 at the third vibronic peak (λ ) 382 nm), I1/I3, in the emission spectra of pyrene was calculated. All measurements were performed at room temperature. Isothermal Titration Calorimetry (ITC). The calorimetric data were acquired using a Microcal ITC equipped with 1.35 mL of reference and sample cells. Both cells are insulated by an adiabatic shield. The titration was performed at 25 °C, by injecting 0.1 M CaCl2 solution from a 250 µL injection syringe into the sample cell filled with 0.05% w/v polymer solution. The syringe is tailor-made such that the tip acts as a blade-type stirrer to ensure an optimum mixing efficiency at 400 rpm. An injection schedule was carried out automatically using the injection software to set the number of injections (55 injections) and volume of each injection (1.3 µL for injections 1-20, 3 µL for injections 21-35, and 7.5 µL for injections 36-55). The observed heat flow data contained the heat of dilution for CaCl2 in water, which can be determined by performing a blank (15) Liu, J.; Sondjaja, H. R.; Tam, K. C. Langmuir 2007, 23, 5106.
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Figure 1. (a) The ISE titration data obtained from (4) titrating CaCl2 0.5 M into 0.05% w/v PEO45 homopolymer and (2) titrating CaCl2 0.5 M into 0.05% w/v PEO45-b-PAA70 at pH 9.7. (b) The titration data obtained from titrating CaCl2 0.5 M into 0.05% w/v PEO45-b-PAA70 illustrated as mmol of free Ca2+ vs Ca2+/COO- ratio.
titration in aqueous solution without polymer. The differential enthalpy curves were generated after subtracting the heat of dilution from the data using Microcal ORIGIN software. Transmission Electron Microscopy (TEM). A JEOL JEM-2010 transmission electron microscope operating at 160 kV was used to image the Ca2+/PEO45-b-PAA70 aggregates. The sample was prepared by placing one drop of the solution on a copper grid precoated with Formvar. The sample was immediately frozen and then freeze-dried. Approximately 30 min prior to placing the sample in the TEM chamber, a drop of phosphotungstic acid was added onto the surface of the copper grid to enhance the contrast.
Results and Discussion Ion Selective Electrode. Under alkaline pH conditions, (pH 9.7 and higher) the polymer PEO45-b-PAA70 was fully deprotonated (R ) 1), such that all the carboxylic groups on the PAA segments existed as COO-. Figure 1a shows the Ca2+ ISE data for the titration of 0.05% w/v PEO45-b-PAA70 and 0.05% w/v PEO45 solutions with 0.5 M CaCl2 under alkaline pH conditions. Initially no free Ca2+ ions were detected, but when [CaCl2] approached 2 mM, free Ca2+ ions were observed, suggesting that the first 2 mM of Ca2+ ions added was bound to the polymeric chains. For PEO45 homopolymer solution, the amount of Ca2+ ions detected by the ISE was equal to the added Ca2+ ions, confirming that no interaction occurred between Ca2+ ions and PEO chains. This suggests that the Ca2+ ions added were bound to PAA segments of PEO45-b-PAA70 to form Ca2+/PEO45-bPAA70 complexes. Figure 1b shows that the interaction between Ca2+ ions and polymer occurred up to Ca2+:COO- ) 0.5, which confirmed that electrostatic interactions between Ca2+ ions and COO- are the major driving force for aggregation. Beyond the ratio of 0.5, all the added Ca2+ ions were present in the bulk solution since all the COO- groups on PAA segments had already been saturated with Ca2+ ions. Size Measurements. Figure 2 displays the size distribution obtained from DLS measurements for samples containing different Ca2+/COO- ratios prepared by introducing 0.1 M CaCl2 to 0.05%w/v PEO45-b-PAA70 at R ) 1. In the absence of Ca2+ ions, PEO45-b-PAA70 existed as charged unimeric chains; hence, the decay time distribution function cannot be experimentally determined due to its small size. Addition of Ca2+ ions induced the formation of aggregates, as evident from the size distribution in Figure 2. When Ca2+ ions were added at ratios of 0.4 and 0.5, small aggregates with average hydrodynamic diameter (Dh) of 4.9 and 7.3 nm, respectively, were observed, corresponding to the formation of unimeric Ca2+/PEO45-b-PAA70 complexes. Since the theoretical length of a single PEO45-b-PAA70 molecular chain
Self-Assembly of PEO-b-PMAA Induced by CaCl2
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Figure 4. The TEM images of the Ca2+/PEO45-b-PAA70 sample (A) obtained at Ca2+:COO- ) 0.5 and (b) obtained at Ca2+:COO- ) 3:1, both at magnification 80 000×, and (inset) enlargement of panel B at magnification 120 000×. All samples were freeze-dried and stained with phosphotungstic acid 1% w/v in ethanol.
Figure 2. The size distribution of Ca2+/PEO45-b-PAA70 system at fully deprotonated state of the polymer. The size is represented as the hydrodynamic diameter (Dh) of the particles.
Figure 3. Plot of the average scattering intensity (in kilocounts per seconds, kcps) vs the mmol ratio of Ca2+ added/COO- available in the Ca2+/PEO45-b-PAA70 system in the fully deprotonated state.
is approximately 30 nm when fully stretched,16 the measured diameters of these single unimeric complexes indicate a coillike structure resulting from the condensation of Ca2+ ions on PAA chains.17 Remarkably, when the amounts of Ca2+ ions introduced exceeded the stoichiometric ratio of 0.5, the particle size increased significantly to a stable size of 52 ( 7.4 nm at Ca2+:COO- ) 3.0. On the basis of the ISE titration data, which confirmed the stoichiometric binding ratio, these larger particles are not due to the interaction between excess Ca2+ and the polymeric chains. The appearance of these larger particles could be due to aggregation of unimeric Ca2+/PEO45-b-PAA70 complexes, and this will be discussed in more detail later. Figure 3 shows the plot of average scattering intensity vs the Ca2+:COO- ratio. Initially, the intensity of the scattered light was low (22.3 kcps), but it began to increase with an increase in the Ca2+/COO- ratio, consistent with the concurrent size (16) Lide, D. R. CRC Handbook of Chemistry & Physics, 88th ed. (Internet Version 2008); CRC Press/Taylor & Francis: Boca Raton, FL, 2007. (17) Schweins, R.; Huber, K. Eur. Phys. J., E 2001, 5, 117.
increment of the nanoaggregates shown in Figure 2. Since the intensity is a function of the number of particles and the diameter raised to the sixth power, the intensity increased substantially when the larger self-assembled aggregates comprising Ca2+ and PEO-b-PAA chains began to form. The addition of Ca2+ up to the stoichiometric ratio Ca2+/COO- ) 0.5 caused the average count rate (ACR) at Ca2+/COO- ) 0.5 to increase to 52.7 kcps, slightly above the ACR for the pure polymer, indicating the formation of small aggregates. For the stoichiometric ratio greater than 0.5, the ACR increased significantly, suggesting the progressive formation of larger aggregates. The ACR of the sample increased 2-fold with the addition of Ca2+ to give a Ca2+: COO- ratio of 0.75 as the size of the nanoaggregates increased from 7.3 to 18.0 nm. The ACR continued to increase until it approached a constant value of (800 kcps at a Ca2+:COO- ratio of 2.5, significantly higher than the initial value. This result is consistent with the finding in Figure 2 where the size of Ca2+/PEO45-b-PAA70 complexes increased and approached a constant value at Ca2+/COO- greater than 2.5, as further confirmed by the TEM analyses shown in Figure 4. As indicated by light scattering, unimeric Ca2+/PEO45-b-PAA70 complex was formed at the stoichiometric ratio (Figure 4A), whereas at higher Ca2+/COO- (e.g., 3.0), larger aggregates were observed (Figure 4B). Isothermal Titration Calorimetry. When an aqueous solution of a polymer is titrated with a second solution containing a metal ion that can form a chelated complex, heat energy is either absorbed or released, both because of the dilution effect and the complexation itself. The total heat absorbed or released (Qb) during the titration can be determined by using the following model:18,19
(
)
M - √M 2 + 4K[Ca2+]T Qb ) ∆H [Ca2+]T + Vcell 2K
(1)
M ) 1 + nK[COO-]T - K[Ca2+]T
(2)
where [Ca2+]T ) the total Ca2+ the total COO- concentration, n
ion concentration, [COO-]T ) ) the number of Ca2+ attached
to 1 mol of COO-, K ) the binding equilibrium constant, ∆H ) the total enthalpy of binding, and Vcell ) the volume of the solution in the ITC cell. The relevant parameters characterizing the binding, n, K, and ∆H, can then be determined as those values giving the best fit of this expression to the cumulative heat release curve. Figure 5 displays the ITC data acquired by titrating 0.05%w/v PEO-b-PAA at R ) 1 with 0.1 M CaCl2, after the heat of dilution (18) Garcia-Valls, R.; Hatton, T. A. Chem. Eng. J. 2003, 94, 99. (19) Wiseman, T.; Willison, S.; Brandts, J.; Lin, L. W. Anal. Biochem. 1989, 179, 131.
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Figure 5. Heat released per injection (∆Q) and cumulative heat released for the titration of a 0.05%w/v PEO45-b-PAA70 with 0.1 M solution of CaCl2 at R ) 1. The solid line marks the curve fitting of the model equation to the experimental data. Regression paramaters are given in the figure.
Figure 6. Heat released per injection (∆Q) data acquired from the titration of a 0.05% w/v PEO45-b-PAA70 with 0.1 M solution of CaCl2 at R ) 1 in the presence of (b) 0.004 M, (4) 0.05 M, and (×) 0.5 M NaCl.
Table 1. Binding Characteristics (n, K) and Thermodynamic Parameters of the Complexation (∆G, ∆Η, ∆S)a
without NaCl +0.004 M NaCl +0.05 M NaCl +0.5 M NaCl a
n
K (mM-1)
∆H (kJ/mol)
∆G (kJ/mol)
∆S (J/mol)
0.69 0.62 0.89 1.65
230 147 6.9 17.3
+124.3 +109.3 +73.3 +24.2
-30.6 -29.5 -21.9 -1.4
+519.8 +465.7 +319.5 +153.6
The calculation detail was described previously.18
was subtracted. A positive heat flow was observed when the polymer solution was titrated with Ca2+, due to the electrostatic interactions between Ca2+ ions and COO- groups. The slight deviation of the curve fit of the model equation to the experimental data was due to the steric effect of the polymer and the heat gained from the release of the condensed Na+ counterions.20,21 Table 1 gives the binding parameters obtained from the fitting of the curve using the above model equation and the thermodynamic parameters (∆G, ∆S) estimated using the well-known relation:
-RT lnK ) ∆G ) ∆H - T∆S
Figure 7. Surface tension versus CaCl2 concentration at alkaline pH condition.
(3)
The estimated thermodynamic parameters yielded a positive entropy, indicating an entropically driven process. It has been reported that the electrostatic binding between cations and carboxylate sites on PAA segments is an endothermic process driven by a positive entropy gain resulting from the release of the condensed Na+ counterions from the NaOH used to adjust the pH.20,21 The ITC results obtained from titration experiments conducted in the presence of various concentrations of NaCl summarized in Figure 6 show that the magnitude of the initial endothermic peak varied inversely with salt concentration. Table 1 shows that the total binding enthalpy decreased while the Gibbs energy increased with the increase of salt concentration. Because of mass action effects, the increased concentration of Na+ ions in the solute suppressed the electrostatic interactions between Ca2+ and carboxyl groups, which weakened the binding of Ca2+ to the polymer.22 Surface Tension. Figure 7 shows that the surface tension data of a dilute alkaline CaCl2 solution (pH 9.7) exhibits the Jones-Ray effect23 that is commonly observed with salt solutions at high (20) Wang, C.; Tam, K. C. Langmuir 2002, 18, 6484. (21) Wang, C.; Tam, K. C.; Jenkins, R. D.; Tan, C. B. J. Phys. Chem. B 2003, 107, 4667. (22) Wang, C.; Tam, K. C. J. Phys. Chem. B 2005, 109, 5156. (23) Jones, G.; Ray, W. A. J. Am. Chem. Soc. 1937, 59, 187. (24) Petersen, P. B.; Johnson, J. C.; Knutsen, K. P.; Saykally, R. J. Chem. Phys. Lett. 2004, 397, 46.
Figure 8. The surface tension data obtained from titrating 0.5 M CaCl2 into 0.05% w/v PEO45-b-PAA70 solution at R ) 1.
pH.24,25 Dole26 conjectured that the water interface contains “active spots” that attract anions at low electrolyte concentration, and at high pH, the positive adsorption of OH- generates a surface charge, which is responsible for the accumulation of cations near the surface.27 The surface tension attained a minimum at approximately 0.01 M CaCl2 before it gradually increased as prescribed by the Langmuir model. Figure 8 illustrates the surface tension data of a 0.05% w/v PEO45-b-PAA70 solutions as a function of CaCl2 concentration at pH 9.7 (R ) 1). The initial surface tension of the polymer solution was 64.13 ( 0.009 mN/m and it remained constant until a ratio Ca2+:COO- of 0.4 was reached (point A), beyond which it started to decrease. The ISE result confirmed that the Ca2+ ions were bound to the polymeric chains at Ca2+:COO- e 0.5, such that no free Ca2+ ions were present to affect the surface tension. If the Ca2+-bound PAA segments were hydrophobic, we would (25) Petersen, P. B.; Saykally, R. J. J. Am. Chem. Soc. 2005, 127, 15446. (26) Dole, M. J. Am. Chem. Soc. 1938, 60, 904. (27) Manciu, M.; Ruckenstein, E. AdV. Colloid Interface Sci. 2003, 105, 63.
Self-Assembly of PEO-b-PMAA Induced by CaCl2
Figure 9. The change of I1/I3 value of 6 × 10-7 M pyrene with the addition of 0.5 M CaCl2 in (O) DI water and (b) PEO45-b-PAA70 0.05% w/v. All samples were prepared at pH 9.7.
expect to observe a significant reduction in the surface tension. The absence of this surface tension reduction suggests that the bound polymer chains did not become more hydrophobic, as previously postulated.8 Above point A, the reduction in the surface tension was observed up to a CaCl2 concentration of 0.01 M (point B). The formation of the Ca2+/PEO-b-PAA complex leads to an excess of Cl- ions, which are attracted to the active spots at the interface. Thus, the surface tension reduction in the presence of the polymer ((6.4 mN/m, Figure 8) was greater than that observed in water ((2.4 mN/m, Figure 7). The concentration difference over the region where the surface tension decreased (i.e., from A to B) was 0.0085 M, which is close to the concentration at which the Jones-Ray effect dominates (0.01 M, Figure 7), which suggested that this reduction in the surface tension is the result of free Ca2+ cations. Fluorescence Spectroscopy. The fluorescence spectrum of pyrene exhibits four emission bands between 370 and 430 nm. The ratio of the 0-0 band at λ ) 372 nm (I1) to the third vibronic band (I3), the I1/I3 ratio, is known to be dependent on the polarity of the environment.28–30 In aqueous self-assembled systems, the fluorescence spectrum of pyrene is the sum of the spectra from the three states of pyrene: (1) pyrene in water, (2) pyrene incorporated in the hydrophobic cores of the micelles, and (3) pyrene interacting with the preaggregated unimers.28 Additional information on the Ca2+/PEO-b-PAA system can be derived by monitoring the changes in I1/I3 with the addition of CaCl2, as shown in Figure 9. It has been reported that the addition of simple electrolytes decreases the pyrene monomer fluorescence at the 0-0 band (λ ) 372 nm) due to a change of solute-solvent interaction in the sample,31,32 as observed in Figure 9 in both water and polymer solution with changes in CaCl2 concentration. In the presence of the polymer, the I1/I3 values were slightly lower, but show a similar trend, indicating that the fluorescence spectrum of pyrene was affected only by the pyrene activity in water. This result further confirmed that the electrostatic interaction between Ca2+ and the polymer did not enhance the hydrophobicity within the aggregates. The values of I1/I3 obtained in the Ca2+/PEO45-b-PAA70 samples were not as low as those reported for micelles with hydrophobic cores28,30,33,34 but were similar to those reported (28) Itoh, H.; Ishido, S.; Nomura, M.; Hayakawa, T.; Mitaku, S. J. Phys. Chem. 1997, 100, 9047. (29) Kwon, G. S.; Naito, M.; Kataoka, K.; Yokoyama, M.; Sakurai, Y.; Okano, T. Colloids Surf., B 1994, 2, 429. (30) Xu, G.-Y.; Chen, A.-M.; Yang, Y.-L.; Yuan, S.-L.; Zheng, L. Colloids Surf., A 2005, 256, 69. (31) Lee, J. H.; Carraway, E. R.; Hur, J.; Yim, S.; Schlautman, M. A. J. Photochem. Photobiol., A 2007, 185, 57. (32) Kalyanasundaram, K.; Thomas, J. K. J. Am. Chem. Soc. 1977, 99, 2039. (33) Chiu, H.-C.; Hu, C.-H.; Chern, C.-S. Polym. J. 1999, 31, 535. (34) Vasilescu, M.; Anghel, D. F.; Almgren, M.; Hansson, P.; Saito, S. Langmuir 1997, 13, 6951.
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for loosely aggregated pyrene-labeled PAA chains.35 These I1/I3 values imply that water is still able to permeate to the core, suggesting that the pyrene molecules were incorporated within the “chelated” Ca2+-PAA core, which is common in a PAA-metallic system.36–38 Induced Self-Assembly Mechanism. The DLS and TEM results suggested that self-assembly of Ca2+/PEO45-b-PAA70 aggregates is induced when CaCl2 exceeds the stoichiometric ratio Ca2+/COO- of 0.5. On the basis of the results from the various physicochemical analyses, we propose a mechanism describing the self-assembly of PEO45-b-PAA70 block copolymers in the presence of Ca2+ cations as shown in Figure 10. When CaCl2 is introduced to the PEO45-b-PAA70 solution, electrostatic interactions between Ca2+ ions and COO- groups on PAA segments lead to an entropically driven binding of Ca2+ resulting in the release of bound Na+ counterions that regain their translational entropy as confirmed by ITC. The DLS and TEM results indicate that the binding of Ca2+ ions to the polymer chains induces the formation of unimeric Ca2+/PEO45-b-PAA70 complexes with a coil-like structure (Figure 10b). The formation of this coil structure is possible because the chelation of Ca2+ with nonadjacent carboxyl groups on the PAA segments39,40 leads to a gain in entropy as the water molecules that previously enclosed the polymer chains are expelled. The surface tension and fluorescence spectroscopy measurements show that these electrostatic interactions, however, do not trigger the formation of nanoaggregates nor do they increase the hydrophobicity of the polymer. Above the stoichiometric ratio, additional CaCl2 exists as Ca2+ and Cl- ions in the bulk solution. These well-hydrated ions prefer to exclude themselves from the polymer, thus making the water structure more ordered (Figure 10c). As more and more CaCl2 is added, these ions compete for water molecules to surround the polymeric chain, which induces the dehydration of the polymer chains.41 The Ca2+-PAA segments, which are less hydrophilic than PEO segments, become dehydrated, and the desolvated Ca2+-PAA segments cluster together to compensate for the entropy change (Figure 10d,e). The suggested mechanism implies that although the electrostatic binding between Ca2+ ions and PAA segments does not enhance the hydrophobicity of the polymer complex, this interaction is needed to form the Ca2+/PEO45-b-PAA70 complex system. Nevertheless, the presence of excess ions provides the entropic driving force responsible for the formation of the selfassembled nanoaggregates. These Ca2+/PEO45-b-PAA70 selforganized nanoaggregates can also be cross-linked and, as previously reported,8,12 the Ca2+ ions inside this structure can be removed, enabling the structure to be used as a template for drug delivery systems.
Conclusions We have studied the self-assembly of a double hydrophilic block copolymer, PEO45-b-PAA70, induced by CaCl2 in alkaline condition. As suggested by ITC data, two sequential phenomena (35) Anghel, D. F.; Alderson, V.; Winnik, F. M.; Mizusaki, M.; Morishima, Y. Polymer 1998, 39, 3035. (36) Kuila, D.; Blay, G. A.; Borjas, R. E.; Hughes, S.; Maddox, P.; Rice, K.; Stansbury, W.; Laurel, N. J. Appl. Polym. Sci. 1999, 73, 1097. (37) Roma-Luciow, R.; Sarraf, L.; Morcellet, M. Eur. Polym. J. 2001, 37, 1741. (38) Sebastian, N.; George, B.; Mathew, B. Polym. Degrad. Stab. 1998, 60, 37. (39) Blay, J. A.; Ryland, J. H. Anal. Lett. 1971, 4, 653. (40) Chang, D. M. J. Am. Oil Chem. Soc. 1983, 60, 618. (41) Collins, K. D.; Neilson, G. W.; Enderby, J. E. Biophys. Chem. 2007, 128, 95.
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Figure 10. Schematic representation of the self-assembly mechanism of PEO-b-PAA induced by Ca2+: (a) In aqueous solution, at R ) 1, PEO-b-PAA forms unimer with Na+ ions condensed on the PAA segment, whereas CaCl2 dissociates into Ca2+ and Cl- ions. (b) When CaCl2 is added to PEO-b-PAA solution at stoichiometric ratio, the electrostatic interaction between the polymer chains and Ca2+ ions induces the formation of Ca2+/PEO-b-PAA coil structure. (c) The addition of CaCl2 in excess causes the water structure to be less disordered. (d) To compensate for the entropy change, the Ca2+/PEO-b-PAA polymer complex begins to rearrange and (e) form the larger self-assembly nanoaggregates.
occur, electrostatic interactions between the Ca2+ ions and COOon the PAA segments, resulting in the formation of coil-like structures, after which nanoclusters of around 50-100 nm begin to form. However, in contrast to the previous reported selfassembly of aggregates between Ca2+ and PEO-b-PMAA, the formation of Ca2+/PEO-b-PAA is not triggered by a sudden
hydrophobicity caused by charge neutralization. Rather, the solvation of excess CaCl2 ions disrupts the water structure and, to compensate for this entropic change, the polymeric chains cluster to form micellar-like self-assembled nanostructures with Ca2+ cations. LA800727E
