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Importance of bound water in hydration–dehydration behavior of hydroxylated poly(N-isopropylacrylamide)
Journal of Colloid and Interface Science 302 (2006) 467–474 www.elsevier.com/locate/jcis
Importance of bound water in hydration–dehydration behavior of hydroxylated poly(N-isopropylacrylamide) Tomohiro Maeda a , Kazuya Yamamoto a , Takao Aoyagi a,b,∗ a Department of Nanostructure and Advanced Materials, Graduate School of Science and Engineering, Kagoshima University,
1-21-40, Korimoto, Kagoshima 890-0065, Japan b PRESTO, Japan Science and Technology Agency, 4-1-8 Honcho Kawaguchi, Saitama, Japan
Received 21 April 2006; accepted 27 June 2006 Available online 30 June 2006
Abstract In this study, a differential scanning calorimetric analysis was performed to investigate the role of water existing around the polymer chains on their thermoresponsive behaviors using the novel functional temperature-sensitive copolymer, poly(N-isopropylacrylamide-co-2-hydroxyisopropylacrylamide) (poly(NIPAAm-co-HIPAAm)). The HIPAAm comonomers were incorporated into the polymeric chains as hydrophilic parameters, and then the hydration states of poly(NIPAAm-co-HIPAAm) with various HIPAAm compositions were examined. Bound water, which is affected by the polymeric chains to some extent, was produced by adding the copolymers to the water, and the temperature due to the melting of the bound water decreased as the HIPAAm content increased. On the basis of this result, we considered that the interaction between the bound water and the polymeric chains is reinforced by the increasing HIPAAm composition. In addition, the cloud points of the copolymers shifted to a higher temperature, and the endothermic enthalpy for the phase transition decreased with increasing the HIPAAm content, suggesting that the number of water molecules disassociated from the polymeric chains decreased. Based on these results, we postulate that the changes in the interaction between the thermosensitive polymer and bound water exert a strong influence on its phase transition and/or separation, such as the cloud point or dehydration behavior. © 2006 Published by Elsevier Inc. Keywords: Thermoresponsive polymers; N -isopropylacrylamide; Cloud point; Differential scanning calorimetry; Bound water
1. Introduction Stimuli-responsive polymers abruptly transform their physicochemical properties in response to external environmental changes such as temperature, light, electric field, pH, or concentration of the chemical species. A thermoresponsive polymer, which is a kind of stimuli-responsive polymer and responds to temperature, has been studied by many researchers. In particular, poly(N -isopropylacrylamide) (PNIPAAm) has been used in various fields, such as nanotechnology or biological systems [1–5]. When the temperature is below the lower critical solution temperature (LCST, ∼31 ◦ C), PNIPAAm exists in a random coil and dissolves in water; however, above the LCST, its conformation is transformed into a globule and becomes in* Corresponding author. Fax: +81 99 285 7794.
E-mail address: [email protected] (T. Aoyagi). 0021-9797/$ – see front matter © 2006 Published by Elsevier Inc. doi:10.1016/j.jcis.2006.06.047
soluble [6]. This hydration–dehydration behavior of PNIPAAm in water is reversible. Schild reviewed some physicochemical studies concerning the PNIPAAm in order to understand its phase transition mechanism [7]. Moreover, many researchers have investigated the unique solution properties of PNIPAAm and other thermoresponsive polymers [8–12]. It has been reported that introducing hydrophilic comonomers into a thermosensitive polymer results in increasing the LCST, while hydrophobic comonomers decrease the LCST [13]. Maeda et al. systematically investigated the hydration states of PNIPAAm and its copolymers as well as other poly(N alkylacrylamide)s above and below their LCSTs by Fourier transform infrared (FTIR) spectroscopy [14–17]. In these papers, they examined the influences of ions on the behaviors of the IR spectra of these polymers, and showed that the IR spectra profiles of these polymer solutions containing salts resemble those of polymer solutions in the absence of salts,
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Fig. 1. Chemical structures of (a) CIPAAm, AIPAAm, HIPAAm, and (b) poly(NIPAAm-co-HIPAAm).
even though their LCSTs were changed by adding the salts. In other words, the amide I band or the amide II band did not shift, which suggested that the added salts do not directly interact with the amide group, but change the structure of water existing around the polymeric chains. Furthermore, it was demonstrated that the conformational change of PNIPAAm in water is governed by the dynamics of water molecules according to a computer simulation [18]. Based on these results, we considered that the thermosensitive behaviors of the polymers would be closely related to the water molecules around the polymer chains. To establish the hypothesis, we then tried to analyze the hydration state of a thermosensitive polymer using the novel temperature-responsive polymer, poly(NIPAAm-co2-hydroxyisopropylacrylamide) (poly(NIPAAm-co-HIPAAm)) (Fig. 1b). In our previous research, we hypothesized that the following three factors: (1) preserving the continuous and repeated structure of the isopropylamide group after copolymerization, (2) random monomer alignment of the polymer chains, and (3) uniformity of the comonomer content in each copolymer chain, are very important for constructing a functional polymer with a sensitive temperature response. Therefore, we have newly designed three types of monomers, namely, an anionic monomer, 2-carboxyisopropylacrylamide (CIPAAm) [19–21], a cationic monomer, 2-aminoisopropylacrylamide (AIPAAm) [22], and a nonionic monomer, HIPAAm (Fig. 1a) [23]. Since the structures of these monomers are very similar to that of NIPAAm, the obtained copolymers by free radical copolymerization with NIPAAm could retain the continuous and repeated structure of the isopropylamide group after copolymerization, and the monomer reactivity ratios were very similar to the values (in poly(NIPAAm-co-HIPAAm), r1 = 1.08 and r2 = 0.60, determined by the Kelen–Tüdös method, where NIPAAm is (1) and HIPAAm is (2)) [23]. As expected, the obtained copolymers showed very sensitive thermoresponses in aqueous media even though they contained many hydrophilic side chains.
In this study, we focused on water molecules around the poly(NIPAAm-co-HIPAAm) chains and analyzed the hydration behaviors of the copolymers by differential scanning calorimetry (DSC). The surrounding water molecules were classified into three distinct modes, i.e., free water, bound water and non-freezing water [24], and the correlation between these three types of water and temperature-responsive behaviors of the acrylamide-type thermosensitive polymers was investigated. We employed poly(NIPAAm-co-HIPAAm) due to the following reasons: (1) it is possible to introduce the HIPAAm comonomers into the NIPAAm-based copolymers as hydrophilic parameters without losing their sharp thermoresponses, (2) because the chemical structures of NIPAAm and HIPAAm very closely resemble each other, the original hydration state of PNIPAAm would not be totally destroyed, and (3) since poly(NIPAAm-co-HIPAAm) is nonionic, we could not take into account the changes in the water structures by dissociation of an ionic moiety, therefore simplifying the system. Based on these important points, the HIPAAm could be very suitable monomer for investigation of the hydration state of an acrylamide-type thermoresponsive polymer by introducing the hydrophilic comonomer into the NIPAAm-based copolymers as a hydrophilic parameter. 2. Experimental 2.1. Materials NIPAAm was kindly supplied by Kohjin (Tokyo, Japan) and purified by recrystallization from benzene–hexane. HIPAAm was synthesized from D , L-2-amino-1-propanol and acryloyl chloride which were purchased from Tokyo Kasei Kogyo (Tokyo, Japan) and used without further purification. This monomer synthesis was described in detail in our previous paper [23]. Poly(NIPAAm-co-HIPAAm) with various HIPAAm
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compositions (NH-0, NH-10, NH-30, NH-50, NH-80, NH-100, where NH-X denotes the copolymers containing X mol% of HIPAAm unit) were prepared by free radical copolymerization of NIPAAm with HIPAAm, and obtained in high yields. This preparation method was also previously mentioned [23]. The chemical structures and the HIPAAm compositions of the copolymers were determined using 1 H NMR spectroscopy (JEOL JNM-GSX400, 400 MHz spectrometer). The numberaverage molecular weights (Mn ) and molecular weight distributions (Mw /Mn ) of the copolymers were determined by gel permeation chromatography (GPC, Jasco LC-2000 Plus, Tokyo, Japan). DMF containing LiBr (10 mM) was used as an eluent, and poly(ethylene glycol) was employed as the standard for calibration. The number-average molecular weights and molecular weight distributions of the obtained copolymers were about 2.0–4.0 × 104 and 2.0–3.0, respectively. 2.2. Differential scanning calorimetric measurement of poly(NIPAAm-co-HIPAAm) aqueous solution A differential scanning calorimeter (DSC, DSC 6100, Seiko Instruments, Tokyo, Japan) was used to analyze the hydration state of poly(NIPAAm-co-HIPAAm) in water. Copolymers with various HIPAAm compositions were dissolved in ultrapure water and the sample solutions were prepared at concentrations of 10, 20, and 30 w/v%. These polymer aqueous solutions were kept at 4 ◦ C for 2 days, and about 30 µL of the copolymer solutions were then transferred to silver pans. To prevent water from evaporating during the measurements, these pans were completely sealed. Subsequently, the temperatures of the copolymer aqueous solutions were lowered to −60 ◦ C, and these solutions were frozen. We then carried out the DSC measurements by gradually raising the temperatures of the solutions. The measurements were performed between −60 and 120 ◦ C at a scanning rate of 2.0 ◦ C/min while heating. The reproducibility was checked by running 3 experiments. Hm (J/g), which is the apparent enthalpy of melting of water, was estimated by dividing the total enthalpy of melting for the copolymer solution (J) by the mass of water (g). 2.3. Analysis of dehydration behavior of poly(NIPAAm-co-HIPAAm) solution with 1 H NMR spectroscopy 1H
NMR measurements of the copolymers in deuterium oxide (D2 O) were carried out above and below the cloud points. The copolymers of various compositions were dissolved in D2 O (Wako Co., Tokyo, Japan, 99%) at a concentration of 1.0 w/v%. 3-(Trimethylsilyl)propionic acid-d4 sodium salt (Aldrich Chem. Co.) was added to each copolymer solution as the internal standard. Subsequently, we performed 1 H NMR measurements of the copolymer solutions at 25 ◦ C (below their cloud points) and at 10 ◦ C higher than each cloud point of the copolymers (above their cloud points).
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3. Results and discussion 3.1. Thermoresponsive behavior of poly(NIPAAm-co-HIPAAm) in water In our previous research, we demonstrated that poly(NIPAAm-co-HIPAAm) aqueous solutions showed very sharp thermosensitive behaviors, and even the copolymers with a high content of hydroxyl groups (50 or 80 mol% for HIPAAm unit) were capable of exhibiting clear and discontinuous transmittance changes in aqueous media [23]. NH-100, i.e., the HIPAAm homopolymer, did not show a transmittance change below 100 ◦ C at ambient pressure, but could cause a turbidity change by adding salts. On the basis of these results, poly(NIPAAm-co-HIPAAm) would have a strong intra- or intermolecular association force. In general, introducing the hydrophilic comonomers, such as acrylic acid, into temperatureresponsive polymers or hydrogels, makes their thermosensitive behaviors insensitive, and in some cases, cannot cause them [19,25–27]. However, since poly(NIPAAm-co-HIPAAm) is capable of maintaining the sharp temperature-response behavior even in the copolymer containing a high content of hydrophilic comonomers, this characteristic is very useful for the purpose of analyzing the effects of introducing the hydrophilic comonomers into the NIPAAm-based copolymers on the hydration states and the temperature-responsive behaviors of the copolymers. Furthermore, the cloud points of poly(NIPAAmco-HIPAAm) closely depended on the HIPAAm content, that is, the cloud points increased with increasing the HIPAAm content. These results are summarized in Table 1. 3.2. The correlation between the temperature-responsive behavior of poly(NIPAAm-co-HIPAAm) and water molecules around the copolymer chains In some polymer aqueous solutions, it has been reported that the condition of the water molecules around the polymeric chains is different from normal one [28,29]. Uedaira and other researchers have described that three kinds of waters, i.e., free water, bound water, and non-freezing water, would participate in the hydration of the water-soluble polymers [24, 28–30]. Therefore, we also divided water molecules existing in the copolymer solutions into three distinct states (free water, bound water, and non-freezing water) in imitation of them. Free water is hardly affected by the polymer chains and its behavior is identical with ordinary water molecules. This water Table 1 The cloud points for 1.0 w/v% aqueous solutions of poly(NIPAAm-coHIPAAm) Code
Cloud point (◦ C)
NH-0 NH-10 NH-30 NH-50 NH-80 NH-100
31.6 36.7 41.8 55.0 80.0 –
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Fig. 2. DSC curves for NH-10 aqueous solutions of poly(NIPAAm-coHIPAAm) with various polymer concentrations. Hm denotes the apparent enthalpy of melting of water.
freezes at 0 ◦ C under the atmospheric pressure. Bound water is somewhat influenced by the polymeric chains and is known to freeze at a lower temperature than the usual freezing point [24,31]. Non-freezing water is strongly associated with the polymer chains and cannot freeze at even −196 ◦ C [24]. Therefore, the stronger the water molecules interact with the polymeric chains, the more difficult they tend to crystallize. In other words, the stronger interaction between water molecules and the polymer chains results in lowering the melting point (or freezing point). We examined the relationship between these three distinct waters and the thermo-responsive behavior of the nonionic poly(NIPAAm-co-HIPAAm) by differential scanning calorimetry. DSC measurements were carried out between −60 and 120 ◦ C during a heating process, and, as described below, two endothermic peaks were observed. The one is an endothermic peak arising from the melting of water, and the other is due to dehydration of the polymer chains that occurred during the phase transition and/or separation. The former consists of two components: the one is the endothermic peak at ca. 0 ◦ C, which originates in the melting of free water, and the other is the shoulder peak observed at the lower temperature (∼ −10 ◦ C), being due to melting of the bound water. Fig. 2 shows DSC thermograms due to the melting of water in the copolymer aqueous solutions and only water. In only water (Fig. 2 (a)), a very sharp endothermic peak at ca. 0 ◦ C was observed, however, in the poly(NIPAAm-co-HIPAAm) aqueous solutions (Fig. 2 (b–d)), the broader endothermic peaks having the shoulder peaks at lower temperature were found. Accordingly, it was revealed that the bound water was produced in the polymer aqueous milieu by adding the copolymers to water. Moreover, the apparent enthalpy of melting of water, Hm , decreased by adding the copolymers to water (Fig. 2 (a, b)). We
Fig. 3. DSC curves for 30 w/v% aqueous solutions of poly(NIPAAm-coHIPAAm) with various compositions. Tm denotes the temperature due to the melting of the bound water.
considered that this result is due to not only the appearance of the bound water whose Hm might be smaller than that of free water, but also the generation of non-freezing water. It has been reported that the Hm of bound water is different from that of free water [28], and because bound water melts at a lower temperature than the melting point of free water, the Hm of the bound water seems to be lower than that of free water. Besides, since non-freezing water cannot freeze, its Hm could be equal to zero. Therefore, it is assumed that the Hm of free water (Hm,f ) is the largest among the three types of waters, and the Hm of bound water (Hm,b ) is greater than that of the non-freezing water (Hm,n ) (that is, Hm,f > Hm,b > Hm,n = 0). Moreover, Hm of the copolymer aqueous solutions decreased with increasing the polymer concentration because the fractions of bound water and non-freezing water relative to the free water could increase by adding the polymers to water (Fig. 2 (b–d)). This is also suggested by the fact that the endothermic peaks due to free water decreased, whereas the shoulder peaks resulting from the bound water increased with increasing the polymer concentration. Also, Hm measured in only water (Hm = 333 J/g, Fig. 2 (a) was consistent with the value in the literature (6.008 kJ/mol, which corresponds to ca. 333.5 J/g). Fig. 3 shows DSC thermograms and the Hm of the copolymer aqueous solutions of various HIPAAm compositions. It should be noted that the Hm values were almost same although the HIPAAm contents were different. As described above, it has been reported that the Hm,b is different from Hm,f [28], and estimating the value of Hm,b is exceedingly difficult. In this study, hence, we could not calculate the amount of each water existing in the copolymer solutions. How-
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Fig. 4. Relationship between Tm and the HIPAAm content for poly(NIPAAm-co-HIPAAm) aqueous solutions. Polymer concentration: 10 w/v% (!), 20 w/v% (P), 30 w/v% (1).
ever, on the basis of these results that the Hm values were almost constant, it seems that the quantitative balance of free water, bound water, and non-freezing water was unchanged in spite of the variation in the HIPAAm composition. Since the HIPAAm monomer is hydrophilic, we expected that the Hm might decrease with increasing the HIPAAm composition due to the change in the bound water and non-freezing water. Consequently, these data were unexpected results. Interestingly however, as seen in Fig. 3, the shoulder peaks assigned to the bound water became broader and shifted to a lower temperature as the HIPAAm content increased. We then defined the Tm , which is the temperature resulting from the melting of the bound water as shown in Fig. 3, and plotted it in Fig. 4 as a function of the HIPAAm composition. It is realized that the Tm closely depends on the HIPAAm content and decreases with an increase in the HIPAAm composition. Moreover, it was found that the Tm is independent of the polymer concentration, suggesting that the shift in Tm is not due to the freezing point depression. As described above, water molecules, which stronger associate with the polymeric chains, become difficult to crystallize, that is, leading to a lower melting point. Therefore, we consider that the interaction between the bound water and the copolymer chains is reinforced as the HIPAAm composition increases. 3.3. Dehydration behavior induced in the phase transition and/or separation for poly(NIPAAm-co-HIPAAm) aqueous solution In our previous research, we performed DSC measurements for the purpose of investigating the phase transition and/or separation behavior of poly(NIPAAm-co-HIPAAm) in aqueous media [23]. From these measurements, it became clear that although the copolymers showed very sensitive transmittance changes in water, the endothermic peaks due to dehydration of the polymeric chains that occurred during the phase transition and/or separation gradually became small with increasing the HIPAAm content, and finally could not be observed above NH-50. In this study, the polymer concentration dependency for the phase transition and/or separation be-
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Fig. 5. Ht for aqueous solutions of poly(NIPAAm-co-HIPAAm) as a function of polymer concentration: NH-0 ("), NH-10 (Q), NH-30 (2). Ht denotes the endothermic enthalpy per mol of monomer unit, which is due to the dehydration induced during the phase transition and/or separation.
havior of poly(NIPAAm-co-HIPAAm) was examined. Fig. 5 shows the endothermic enthalpy, Ht , resulting from dehydration induced during the phase transition and/or separation, as a function of the polymer concentration. In NH-0–NH-30, no noticeable changes in Ht were observed even though the polymer concentration changed, which implied that the dehydration profiles of the copolymers were independent of the polymer concentration. However, Ht decreased with increasing the HIPAAm content, and NH-50 and NH-80 did not show any endothermic peak for all the polymer concentrations. We have already demonstrated that poly(NIPAAm-co-HIPAAm) with a high HIPAAm content, such as NH-50, showed a liquid– liquid phase separation accompanied by coacervation [23]. It has been reported that the Ht of temperature-responsive polymers causing coacervation in the phase separation are quite small [32,33]. Thus, the Ht of NH-50 and NH-80 were too small to be estimated in these measurements. These results indicate that dehydration of the polymeric chains becomes insufficient with an increase in the HIPAAm composition. In other words, it is considered that the number of water molecules, which are dissociated from the copolymer chains in the phase transition and/or separation, decreases as the HIPAAm content increases. To further investigate this consideration, we performed 1 H NMR measurements of the copolymers in deuterium oxide above and below their cloud points. By using 1 H NMR spectroscopy, we can obtain information concerning the hydration states of the copolymers. Below their cloud points, since the copolymers dissolve in the solvent (D2 O), the peaks assigned to each proton of the copolymer are detected. However above their cloud points, the copolymer chains would be more or less dehydrated, therefore, the peak intensities of the copolymers might decrease or the peaks might be undetectable. Fig. 6 shows the 1 H NMR spectra of NH-0 and NH-50 in D2 O above and below each of their cloud points. Below their cloud points, the peaks assignable to each proton of the copolymers were clearly detectable. When the polymer solutions were heated to 10 ◦ C higher than each cloud point of the copolymers, the peak intensities were drastically reduced in NH-0
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Fig. 6. 1 H NMR spectra of poly(NIPAAm-co-HIPAAm) in D2 O: NH-0 (a) at room temperature (r.t.) and (b) above the cloud point; NH-50 (c) at r.t. and (d) above the cloud point. Polymer concentration: 1.0 w/v%.
(Fig. 6 (c)), however, in NH-50, although a certain degree of decrease in the peak intensity was observed, it did not dramatically decrease (Fig. 6 (d)). Because PNIPAAm gives rise to an abrupt hydration–dehydration change accompanied by a coilto-globule transition at its LCST, the polymeric chains were dehydrated above the LCST and a large decrease in the peak intensity should occur. On the other hand, in view of the result that the peaks of the copolymer remained up to a point, we postulated that NH-50 does not cause a drastic dehydration at its cloud point. In fact, NH-50 shows coacervation and its dehydration could partially occur. Accordingly, we demonstrated that there is a significant difference in the 1 H NMR spectrum between the coil–globule transition and coacervation. Also, these results were in good agreement with those of the DSC measurements which suggested that NH-0 showed a large enthalpy of transition (Ht ), whereas the NH-50 copolymer did not exhibit an endothermic peak at its cloud point for every polymer concentration. Fig. 7 shows the 1 H NMR spectra of poly(NIPAAm-co-HIPAAm) with various HIPAAm compositions in D2 O above their cloud points (10 ◦ C higher than each cloud point of the copolymers). It was revealed that the remaining peaks gradually increased with an increase in the HIPAAm content. Consequently, we suggest that dehydration of the copolymer chains in the phase transition and/or separation would become more difficult to induce with increasing the HIPAAm content, that is, the amount of water molecules disassociated from the polymeric chains decreases. These results were also consistent with the results obtained by the DSC measurements.
3.4. The relationship between the cloud point, Tm , Ht , and the HIPAAm composition for poly(NIPAAm-co-HIPAAm) aqueous solutions Based on the results obtained in this study, we derived the correlation between the Tm and the thermoresponsive behavior of poly(NIPAAm-co-HIPAAm) in water. Fig. 8 shows the relationship between Tm , Ht , and the HIPAAm composition, and Fig. 9 shows the relationship between Tm , the cloud points (determined by UV–vis measurements), and the HIPAAm content. It was revealed that the Tm and Ht decreased, whereas the cloud points of the copolymers increased as the HIPAAm composition increased. Based on these results, we postulated the following: the hydrophilicity of the copolymers increases with increasing the HIPAAm content, thus reinforcing the interaction between the bound water and copolymer chains (that is, Tm decreases). Moreover, the stronger interaction between the bound water and the copolymers results in becoming more difficult for the polymeric chains to cause dehydration in the phase transition and/or separation. Consequently, the number of water molecules, which are dissociated from the polymeric chains, decreases (that is, Ht decreases), and the cloud points of the copolymer solutions should shift to a higher temperature. It has been demonstrated that PNIPAAm includes water molecules to some extent even in the globule state, and is not completely dehydrated [14,34]. Thus, water molecules, such as non-freezing water, which strongly associate with the polymeric chains, may not be dissociated and partly remain even during the phase transition and/or separation. In addition, because free water is hardly influenced by the polymer chains, it could not affect the
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Fig. 7. 1 H NMR spectra of poly(NIPAAm-co-HIPAAm) with a variety of compositions in D2 O: (a) NH-0, (b) NH-10, (c) NH-30, (d) NH-50, (e) NH-80 above the cloud points. Polymer concentration: 1.0 w/v%.
Fig. 8. Tm and Ht for aqueous solutions of poly(NIPAAm-co-HIPAAm) as a function of the HIPAAm content. Polymer concentration: 10 w/v%; Tm (!), Ht (").
phase transition and/or separation behaviors of the temperatureresponsive polymers. Hence, it is presumed that the behavior of bound water plays an important role in the thermoresponsive profiles of the polymers. So far, it has been reported that the LCSTs of thermosensitive copolymers depend on their Ht , i.e., the LCST increases with decreasing the Ht [13,15,16]. In this study, however, it was first demonstrated that Tm , which is the temperature resulting from the melting of bound water, affected the Ht and the cloud points of the thermoresponsive copolymers, therefore, the temperature-responsive behaviors of the polymers significantly depend on the bound water. 4. Conclusion In this study, we analyzed the hydration state of a thermoresponsive polymer with a novel nonionic temperature-
Fig. 9. Tm and the cloud points for aqueous solutions of poly(NIPAAmco-HIPAAm) as a function of the HIPAAm content. Polymer concentration: 10 w/v% for Tm and 1.0 w/v% for the cloud points; Tm (!), cloud points (2); Tm and the cloud points were determined from the DSC and UV–vis measurements, respectively.
sensitive polymer, poly(NIPAAm-co-HIPAAm). Particularly, water molecules existing around the copolymer chains were determined and divided into three distinct states, namely, free water, bound water, and non-freezing water. By adding the copolymers to water, the shoulder peaks resulting from the melting of the bound water appeared in the DSC thermograms. Also, the Tm , which is the temperature due to the melting of the bound water, shifted to lower temperatures with increasing the HIPAAm composition. Based on these results, it was believed that the bound water more strongly interacts with the polymeric chains with an increase in the HIPAAm composition. Furthermore, the endothermic enthalpy for the phase transition and/or separation, Ht , decreased with increasing the HIPAAm composition. These results indicate that the amount
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of water molecules dissociated from the polymeric chains decreases with increasing the HIPAAm content. In 1 H NMR spectra, the intensities of the peaks arising from each proton of the copolymers with a low HIPAAm content, such as NH0 and NH-10, were dramatically reduced above their cloud points. These results suggest that the polymeric chains brought about an abrupt dehydration during the phase transition and the copolymers were insoluble. On the other hand, the peak intensity was not drastically reduced in the copolymer containing a high content of HIPAAm above each cloud point of the copolymer solutions, and the remaining peaks progressively increased as the HIPAAm composition increased. Consequently, we suggest that the dehydration of the copolymer chains that occurred in the phase transition and/or separation becomes incomplete with increasing the HIPAAm content, and these results were correlated with the ones obtained from the DSC measurements. We determined the correlation between the Tm , the cloud points of the copolymers, Ht and the HIPAAm content for poly(NIPAAm-co-HIPAAm) aqueous solutions. With an increase in the HIPAAm composition, the Tm and Ht decreased, and the cloud points shifted to a higher temperature. An increase in the hydrophilic comonomer (HIPAAm) content raises the hydrophilicity of the copolymer chains, thus, the interaction between the bound water and the polymeric chains could become stronger. This occurrence accounts for the fact that the dehydration of the polymeric chains induced during the phase transition and/or separation becomes more difficult to occur. Therefore, it was demonstrated that the amount of dissociated water molecules is reduced and the cloud points of the copolymer solutions increase. That is, the relationship between the Tm , the cloud point, Ht and the HIPAAm composition is determined for poly(NIPAAm-co-HIPAAm), and it becomes clear that the thermosensitive behavior of a temperature-responsive polymer is dependent on the bound water. Acknowledgment This research was partially supported by the Ministry of Education, Science, Sports and Culture, through a Grant-in-Aid for Scientific Research (B) 15300167, 2003.
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Importance of bound water in hydration–dehydration behavior of hydroxylated poly(N-isopropylacrylamide) Tomohiro Maeda a , Kazuya Yamamoto a , Takao Aoyagi a,b,∗ a Department of Nanostructure and Advanced Materials, Graduate School of Science and Engineering, Kagoshima University,
1-21-40, Korimoto, Kagoshima 890-0065, Japan b PRESTO, Japan Science and Technology Agency, 4-1-8 Honcho Kawaguchi, Saitama, Japan
Received 21 April 2006; accepted 27 June 2006 Available online 30 June 2006
Abstract In this study, a differential scanning calorimetric analysis was performed to investigate the role of water existing around the polymer chains on their thermoresponsive behaviors using the novel functional temperature-sensitive copolymer, poly(N-isopropylacrylamide-co-2-hydroxyisopropylacrylamide) (poly(NIPAAm-co-HIPAAm)). The HIPAAm comonomers were incorporated into the polymeric chains as hydrophilic parameters, and then the hydration states of poly(NIPAAm-co-HIPAAm) with various HIPAAm compositions were examined. Bound water, which is affected by the polymeric chains to some extent, was produced by adding the copolymers to the water, and the temperature due to the melting of the bound water decreased as the HIPAAm content increased. On the basis of this result, we considered that the interaction between the bound water and the polymeric chains is reinforced by the increasing HIPAAm composition. In addition, the cloud points of the copolymers shifted to a higher temperature, and the endothermic enthalpy for the phase transition decreased with increasing the HIPAAm content, suggesting that the number of water molecules disassociated from the polymeric chains decreased. Based on these results, we postulate that the changes in the interaction between the thermosensitive polymer and bound water exert a strong influence on its phase transition and/or separation, such as the cloud point or dehydration behavior. © 2006 Published by Elsevier Inc. Keywords: Thermoresponsive polymers; N -isopropylacrylamide; Cloud point; Differential scanning calorimetry; Bound water
1. Introduction Stimuli-responsive polymers abruptly transform their physicochemical properties in response to external environmental changes such as temperature, light, electric field, pH, or concentration of the chemical species. A thermoresponsive polymer, which is a kind of stimuli-responsive polymer and responds to temperature, has been studied by many researchers. In particular, poly(N -isopropylacrylamide) (PNIPAAm) has been used in various fields, such as nanotechnology or biological systems [1–5]. When the temperature is below the lower critical solution temperature (LCST, ∼31 ◦ C), PNIPAAm exists in a random coil and dissolves in water; however, above the LCST, its conformation is transformed into a globule and becomes in* Corresponding author. Fax: +81 99 285 7794.
E-mail address: [email protected] (T. Aoyagi). 0021-9797/$ – see front matter © 2006 Published by Elsevier Inc. doi:10.1016/j.jcis.2006.06.047
soluble [6]. This hydration–dehydration behavior of PNIPAAm in water is reversible. Schild reviewed some physicochemical studies concerning the PNIPAAm in order to understand its phase transition mechanism [7]. Moreover, many researchers have investigated the unique solution properties of PNIPAAm and other thermoresponsive polymers [8–12]. It has been reported that introducing hydrophilic comonomers into a thermosensitive polymer results in increasing the LCST, while hydrophobic comonomers decrease the LCST [13]. Maeda et al. systematically investigated the hydration states of PNIPAAm and its copolymers as well as other poly(N alkylacrylamide)s above and below their LCSTs by Fourier transform infrared (FTIR) spectroscopy [14–17]. In these papers, they examined the influences of ions on the behaviors of the IR spectra of these polymers, and showed that the IR spectra profiles of these polymer solutions containing salts resemble those of polymer solutions in the absence of salts,
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Fig. 1. Chemical structures of (a) CIPAAm, AIPAAm, HIPAAm, and (b) poly(NIPAAm-co-HIPAAm).
even though their LCSTs were changed by adding the salts. In other words, the amide I band or the amide II band did not shift, which suggested that the added salts do not directly interact with the amide group, but change the structure of water existing around the polymeric chains. Furthermore, it was demonstrated that the conformational change of PNIPAAm in water is governed by the dynamics of water molecules according to a computer simulation [18]. Based on these results, we considered that the thermosensitive behaviors of the polymers would be closely related to the water molecules around the polymer chains. To establish the hypothesis, we then tried to analyze the hydration state of a thermosensitive polymer using the novel temperature-responsive polymer, poly(NIPAAm-co2-hydroxyisopropylacrylamide) (poly(NIPAAm-co-HIPAAm)) (Fig. 1b). In our previous research, we hypothesized that the following three factors: (1) preserving the continuous and repeated structure of the isopropylamide group after copolymerization, (2) random monomer alignment of the polymer chains, and (3) uniformity of the comonomer content in each copolymer chain, are very important for constructing a functional polymer with a sensitive temperature response. Therefore, we have newly designed three types of monomers, namely, an anionic monomer, 2-carboxyisopropylacrylamide (CIPAAm) [19–21], a cationic monomer, 2-aminoisopropylacrylamide (AIPAAm) [22], and a nonionic monomer, HIPAAm (Fig. 1a) [23]. Since the structures of these monomers are very similar to that of NIPAAm, the obtained copolymers by free radical copolymerization with NIPAAm could retain the continuous and repeated structure of the isopropylamide group after copolymerization, and the monomer reactivity ratios were very similar to the values (in poly(NIPAAm-co-HIPAAm), r1 = 1.08 and r2 = 0.60, determined by the Kelen–Tüdös method, where NIPAAm is (1) and HIPAAm is (2)) [23]. As expected, the obtained copolymers showed very sensitive thermoresponses in aqueous media even though they contained many hydrophilic side chains.
In this study, we focused on water molecules around the poly(NIPAAm-co-HIPAAm) chains and analyzed the hydration behaviors of the copolymers by differential scanning calorimetry (DSC). The surrounding water molecules were classified into three distinct modes, i.e., free water, bound water and non-freezing water [24], and the correlation between these three types of water and temperature-responsive behaviors of the acrylamide-type thermosensitive polymers was investigated. We employed poly(NIPAAm-co-HIPAAm) due to the following reasons: (1) it is possible to introduce the HIPAAm comonomers into the NIPAAm-based copolymers as hydrophilic parameters without losing their sharp thermoresponses, (2) because the chemical structures of NIPAAm and HIPAAm very closely resemble each other, the original hydration state of PNIPAAm would not be totally destroyed, and (3) since poly(NIPAAm-co-HIPAAm) is nonionic, we could not take into account the changes in the water structures by dissociation of an ionic moiety, therefore simplifying the system. Based on these important points, the HIPAAm could be very suitable monomer for investigation of the hydration state of an acrylamide-type thermoresponsive polymer by introducing the hydrophilic comonomer into the NIPAAm-based copolymers as a hydrophilic parameter. 2. Experimental 2.1. Materials NIPAAm was kindly supplied by Kohjin (Tokyo, Japan) and purified by recrystallization from benzene–hexane. HIPAAm was synthesized from D , L-2-amino-1-propanol and acryloyl chloride which were purchased from Tokyo Kasei Kogyo (Tokyo, Japan) and used without further purification. This monomer synthesis was described in detail in our previous paper [23]. Poly(NIPAAm-co-HIPAAm) with various HIPAAm
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compositions (NH-0, NH-10, NH-30, NH-50, NH-80, NH-100, where NH-X denotes the copolymers containing X mol% of HIPAAm unit) were prepared by free radical copolymerization of NIPAAm with HIPAAm, and obtained in high yields. This preparation method was also previously mentioned [23]. The chemical structures and the HIPAAm compositions of the copolymers were determined using 1 H NMR spectroscopy (JEOL JNM-GSX400, 400 MHz spectrometer). The numberaverage molecular weights (Mn ) and molecular weight distributions (Mw /Mn ) of the copolymers were determined by gel permeation chromatography (GPC, Jasco LC-2000 Plus, Tokyo, Japan). DMF containing LiBr (10 mM) was used as an eluent, and poly(ethylene glycol) was employed as the standard for calibration. The number-average molecular weights and molecular weight distributions of the obtained copolymers were about 2.0–4.0 × 104 and 2.0–3.0, respectively. 2.2. Differential scanning calorimetric measurement of poly(NIPAAm-co-HIPAAm) aqueous solution A differential scanning calorimeter (DSC, DSC 6100, Seiko Instruments, Tokyo, Japan) was used to analyze the hydration state of poly(NIPAAm-co-HIPAAm) in water. Copolymers with various HIPAAm compositions were dissolved in ultrapure water and the sample solutions were prepared at concentrations of 10, 20, and 30 w/v%. These polymer aqueous solutions were kept at 4 ◦ C for 2 days, and about 30 µL of the copolymer solutions were then transferred to silver pans. To prevent water from evaporating during the measurements, these pans were completely sealed. Subsequently, the temperatures of the copolymer aqueous solutions were lowered to −60 ◦ C, and these solutions were frozen. We then carried out the DSC measurements by gradually raising the temperatures of the solutions. The measurements were performed between −60 and 120 ◦ C at a scanning rate of 2.0 ◦ C/min while heating. The reproducibility was checked by running 3 experiments. Hm (J/g), which is the apparent enthalpy of melting of water, was estimated by dividing the total enthalpy of melting for the copolymer solution (J) by the mass of water (g). 2.3. Analysis of dehydration behavior of poly(NIPAAm-co-HIPAAm) solution with 1 H NMR spectroscopy 1H
NMR measurements of the copolymers in deuterium oxide (D2 O) were carried out above and below the cloud points. The copolymers of various compositions were dissolved in D2 O (Wako Co., Tokyo, Japan, 99%) at a concentration of 1.0 w/v%. 3-(Trimethylsilyl)propionic acid-d4 sodium salt (Aldrich Chem. Co.) was added to each copolymer solution as the internal standard. Subsequently, we performed 1 H NMR measurements of the copolymer solutions at 25 ◦ C (below their cloud points) and at 10 ◦ C higher than each cloud point of the copolymers (above their cloud points).
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3. Results and discussion 3.1. Thermoresponsive behavior of poly(NIPAAm-co-HIPAAm) in water In our previous research, we demonstrated that poly(NIPAAm-co-HIPAAm) aqueous solutions showed very sharp thermosensitive behaviors, and even the copolymers with a high content of hydroxyl groups (50 or 80 mol% for HIPAAm unit) were capable of exhibiting clear and discontinuous transmittance changes in aqueous media [23]. NH-100, i.e., the HIPAAm homopolymer, did not show a transmittance change below 100 ◦ C at ambient pressure, but could cause a turbidity change by adding salts. On the basis of these results, poly(NIPAAm-co-HIPAAm) would have a strong intra- or intermolecular association force. In general, introducing the hydrophilic comonomers, such as acrylic acid, into temperatureresponsive polymers or hydrogels, makes their thermosensitive behaviors insensitive, and in some cases, cannot cause them [19,25–27]. However, since poly(NIPAAm-co-HIPAAm) is capable of maintaining the sharp temperature-response behavior even in the copolymer containing a high content of hydrophilic comonomers, this characteristic is very useful for the purpose of analyzing the effects of introducing the hydrophilic comonomers into the NIPAAm-based copolymers on the hydration states and the temperature-responsive behaviors of the copolymers. Furthermore, the cloud points of poly(NIPAAmco-HIPAAm) closely depended on the HIPAAm content, that is, the cloud points increased with increasing the HIPAAm content. These results are summarized in Table 1. 3.2. The correlation between the temperature-responsive behavior of poly(NIPAAm-co-HIPAAm) and water molecules around the copolymer chains In some polymer aqueous solutions, it has been reported that the condition of the water molecules around the polymeric chains is different from normal one [28,29]. Uedaira and other researchers have described that three kinds of waters, i.e., free water, bound water, and non-freezing water, would participate in the hydration of the water-soluble polymers [24, 28–30]. Therefore, we also divided water molecules existing in the copolymer solutions into three distinct states (free water, bound water, and non-freezing water) in imitation of them. Free water is hardly affected by the polymer chains and its behavior is identical with ordinary water molecules. This water Table 1 The cloud points for 1.0 w/v% aqueous solutions of poly(NIPAAm-coHIPAAm) Code
Cloud point (◦ C)
NH-0 NH-10 NH-30 NH-50 NH-80 NH-100
31.6 36.7 41.8 55.0 80.0 –
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Fig. 2. DSC curves for NH-10 aqueous solutions of poly(NIPAAm-coHIPAAm) with various polymer concentrations. Hm denotes the apparent enthalpy of melting of water.
freezes at 0 ◦ C under the atmospheric pressure. Bound water is somewhat influenced by the polymeric chains and is known to freeze at a lower temperature than the usual freezing point [24,31]. Non-freezing water is strongly associated with the polymer chains and cannot freeze at even −196 ◦ C [24]. Therefore, the stronger the water molecules interact with the polymeric chains, the more difficult they tend to crystallize. In other words, the stronger interaction between water molecules and the polymer chains results in lowering the melting point (or freezing point). We examined the relationship between these three distinct waters and the thermo-responsive behavior of the nonionic poly(NIPAAm-co-HIPAAm) by differential scanning calorimetry. DSC measurements were carried out between −60 and 120 ◦ C during a heating process, and, as described below, two endothermic peaks were observed. The one is an endothermic peak arising from the melting of water, and the other is due to dehydration of the polymer chains that occurred during the phase transition and/or separation. The former consists of two components: the one is the endothermic peak at ca. 0 ◦ C, which originates in the melting of free water, and the other is the shoulder peak observed at the lower temperature (∼ −10 ◦ C), being due to melting of the bound water. Fig. 2 shows DSC thermograms due to the melting of water in the copolymer aqueous solutions and only water. In only water (Fig. 2 (a)), a very sharp endothermic peak at ca. 0 ◦ C was observed, however, in the poly(NIPAAm-co-HIPAAm) aqueous solutions (Fig. 2 (b–d)), the broader endothermic peaks having the shoulder peaks at lower temperature were found. Accordingly, it was revealed that the bound water was produced in the polymer aqueous milieu by adding the copolymers to water. Moreover, the apparent enthalpy of melting of water, Hm , decreased by adding the copolymers to water (Fig. 2 (a, b)). We
Fig. 3. DSC curves for 30 w/v% aqueous solutions of poly(NIPAAm-coHIPAAm) with various compositions. Tm denotes the temperature due to the melting of the bound water.
considered that this result is due to not only the appearance of the bound water whose Hm might be smaller than that of free water, but also the generation of non-freezing water. It has been reported that the Hm of bound water is different from that of free water [28], and because bound water melts at a lower temperature than the melting point of free water, the Hm of the bound water seems to be lower than that of free water. Besides, since non-freezing water cannot freeze, its Hm could be equal to zero. Therefore, it is assumed that the Hm of free water (Hm,f ) is the largest among the three types of waters, and the Hm of bound water (Hm,b ) is greater than that of the non-freezing water (Hm,n ) (that is, Hm,f > Hm,b > Hm,n = 0). Moreover, Hm of the copolymer aqueous solutions decreased with increasing the polymer concentration because the fractions of bound water and non-freezing water relative to the free water could increase by adding the polymers to water (Fig. 2 (b–d)). This is also suggested by the fact that the endothermic peaks due to free water decreased, whereas the shoulder peaks resulting from the bound water increased with increasing the polymer concentration. Also, Hm measured in only water (Hm = 333 J/g, Fig. 2 (a) was consistent with the value in the literature (6.008 kJ/mol, which corresponds to ca. 333.5 J/g). Fig. 3 shows DSC thermograms and the Hm of the copolymer aqueous solutions of various HIPAAm compositions. It should be noted that the Hm values were almost same although the HIPAAm contents were different. As described above, it has been reported that the Hm,b is different from Hm,f [28], and estimating the value of Hm,b is exceedingly difficult. In this study, hence, we could not calculate the amount of each water existing in the copolymer solutions. How-
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Fig. 4. Relationship between Tm and the HIPAAm content for poly(NIPAAm-co-HIPAAm) aqueous solutions. Polymer concentration: 10 w/v% (!), 20 w/v% (P), 30 w/v% (1).
ever, on the basis of these results that the Hm values were almost constant, it seems that the quantitative balance of free water, bound water, and non-freezing water was unchanged in spite of the variation in the HIPAAm composition. Since the HIPAAm monomer is hydrophilic, we expected that the Hm might decrease with increasing the HIPAAm composition due to the change in the bound water and non-freezing water. Consequently, these data were unexpected results. Interestingly however, as seen in Fig. 3, the shoulder peaks assigned to the bound water became broader and shifted to a lower temperature as the HIPAAm content increased. We then defined the Tm , which is the temperature resulting from the melting of the bound water as shown in Fig. 3, and plotted it in Fig. 4 as a function of the HIPAAm composition. It is realized that the Tm closely depends on the HIPAAm content and decreases with an increase in the HIPAAm composition. Moreover, it was found that the Tm is independent of the polymer concentration, suggesting that the shift in Tm is not due to the freezing point depression. As described above, water molecules, which stronger associate with the polymeric chains, become difficult to crystallize, that is, leading to a lower melting point. Therefore, we consider that the interaction between the bound water and the copolymer chains is reinforced as the HIPAAm composition increases. 3.3. Dehydration behavior induced in the phase transition and/or separation for poly(NIPAAm-co-HIPAAm) aqueous solution In our previous research, we performed DSC measurements for the purpose of investigating the phase transition and/or separation behavior of poly(NIPAAm-co-HIPAAm) in aqueous media [23]. From these measurements, it became clear that although the copolymers showed very sensitive transmittance changes in water, the endothermic peaks due to dehydration of the polymeric chains that occurred during the phase transition and/or separation gradually became small with increasing the HIPAAm content, and finally could not be observed above NH-50. In this study, the polymer concentration dependency for the phase transition and/or separation be-
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Fig. 5. Ht for aqueous solutions of poly(NIPAAm-co-HIPAAm) as a function of polymer concentration: NH-0 ("), NH-10 (Q), NH-30 (2). Ht denotes the endothermic enthalpy per mol of monomer unit, which is due to the dehydration induced during the phase transition and/or separation.
havior of poly(NIPAAm-co-HIPAAm) was examined. Fig. 5 shows the endothermic enthalpy, Ht , resulting from dehydration induced during the phase transition and/or separation, as a function of the polymer concentration. In NH-0–NH-30, no noticeable changes in Ht were observed even though the polymer concentration changed, which implied that the dehydration profiles of the copolymers were independent of the polymer concentration. However, Ht decreased with increasing the HIPAAm content, and NH-50 and NH-80 did not show any endothermic peak for all the polymer concentrations. We have already demonstrated that poly(NIPAAm-co-HIPAAm) with a high HIPAAm content, such as NH-50, showed a liquid– liquid phase separation accompanied by coacervation [23]. It has been reported that the Ht of temperature-responsive polymers causing coacervation in the phase separation are quite small [32,33]. Thus, the Ht of NH-50 and NH-80 were too small to be estimated in these measurements. These results indicate that dehydration of the polymeric chains becomes insufficient with an increase in the HIPAAm composition. In other words, it is considered that the number of water molecules, which are dissociated from the copolymer chains in the phase transition and/or separation, decreases as the HIPAAm content increases. To further investigate this consideration, we performed 1 H NMR measurements of the copolymers in deuterium oxide above and below their cloud points. By using 1 H NMR spectroscopy, we can obtain information concerning the hydration states of the copolymers. Below their cloud points, since the copolymers dissolve in the solvent (D2 O), the peaks assigned to each proton of the copolymer are detected. However above their cloud points, the copolymer chains would be more or less dehydrated, therefore, the peak intensities of the copolymers might decrease or the peaks might be undetectable. Fig. 6 shows the 1 H NMR spectra of NH-0 and NH-50 in D2 O above and below each of their cloud points. Below their cloud points, the peaks assignable to each proton of the copolymers were clearly detectable. When the polymer solutions were heated to 10 ◦ C higher than each cloud point of the copolymers, the peak intensities were drastically reduced in NH-0
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Fig. 6. 1 H NMR spectra of poly(NIPAAm-co-HIPAAm) in D2 O: NH-0 (a) at room temperature (r.t.) and (b) above the cloud point; NH-50 (c) at r.t. and (d) above the cloud point. Polymer concentration: 1.0 w/v%.
(Fig. 6 (c)), however, in NH-50, although a certain degree of decrease in the peak intensity was observed, it did not dramatically decrease (Fig. 6 (d)). Because PNIPAAm gives rise to an abrupt hydration–dehydration change accompanied by a coilto-globule transition at its LCST, the polymeric chains were dehydrated above the LCST and a large decrease in the peak intensity should occur. On the other hand, in view of the result that the peaks of the copolymer remained up to a point, we postulated that NH-50 does not cause a drastic dehydration at its cloud point. In fact, NH-50 shows coacervation and its dehydration could partially occur. Accordingly, we demonstrated that there is a significant difference in the 1 H NMR spectrum between the coil–globule transition and coacervation. Also, these results were in good agreement with those of the DSC measurements which suggested that NH-0 showed a large enthalpy of transition (Ht ), whereas the NH-50 copolymer did not exhibit an endothermic peak at its cloud point for every polymer concentration. Fig. 7 shows the 1 H NMR spectra of poly(NIPAAm-co-HIPAAm) with various HIPAAm compositions in D2 O above their cloud points (10 ◦ C higher than each cloud point of the copolymers). It was revealed that the remaining peaks gradually increased with an increase in the HIPAAm content. Consequently, we suggest that dehydration of the copolymer chains in the phase transition and/or separation would become more difficult to induce with increasing the HIPAAm content, that is, the amount of water molecules disassociated from the polymeric chains decreases. These results were also consistent with the results obtained by the DSC measurements.
3.4. The relationship between the cloud point, Tm , Ht , and the HIPAAm composition for poly(NIPAAm-co-HIPAAm) aqueous solutions Based on the results obtained in this study, we derived the correlation between the Tm and the thermoresponsive behavior of poly(NIPAAm-co-HIPAAm) in water. Fig. 8 shows the relationship between Tm , Ht , and the HIPAAm composition, and Fig. 9 shows the relationship between Tm , the cloud points (determined by UV–vis measurements), and the HIPAAm content. It was revealed that the Tm and Ht decreased, whereas the cloud points of the copolymers increased as the HIPAAm composition increased. Based on these results, we postulated the following: the hydrophilicity of the copolymers increases with increasing the HIPAAm content, thus reinforcing the interaction between the bound water and copolymer chains (that is, Tm decreases). Moreover, the stronger interaction between the bound water and the copolymers results in becoming more difficult for the polymeric chains to cause dehydration in the phase transition and/or separation. Consequently, the number of water molecules, which are dissociated from the polymeric chains, decreases (that is, Ht decreases), and the cloud points of the copolymer solutions should shift to a higher temperature. It has been demonstrated that PNIPAAm includes water molecules to some extent even in the globule state, and is not completely dehydrated [14,34]. Thus, water molecules, such as non-freezing water, which strongly associate with the polymeric chains, may not be dissociated and partly remain even during the phase transition and/or separation. In addition, because free water is hardly influenced by the polymer chains, it could not affect the
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Fig. 7. 1 H NMR spectra of poly(NIPAAm-co-HIPAAm) with a variety of compositions in D2 O: (a) NH-0, (b) NH-10, (c) NH-30, (d) NH-50, (e) NH-80 above the cloud points. Polymer concentration: 1.0 w/v%.
Fig. 8. Tm and Ht for aqueous solutions of poly(NIPAAm-co-HIPAAm) as a function of the HIPAAm content. Polymer concentration: 10 w/v%; Tm (!), Ht (").
phase transition and/or separation behaviors of the temperatureresponsive polymers. Hence, it is presumed that the behavior of bound water plays an important role in the thermoresponsive profiles of the polymers. So far, it has been reported that the LCSTs of thermosensitive copolymers depend on their Ht , i.e., the LCST increases with decreasing the Ht [13,15,16]. In this study, however, it was first demonstrated that Tm , which is the temperature resulting from the melting of bound water, affected the Ht and the cloud points of the thermoresponsive copolymers, therefore, the temperature-responsive behaviors of the polymers significantly depend on the bound water. 4. Conclusion In this study, we analyzed the hydration state of a thermoresponsive polymer with a novel nonionic temperature-
Fig. 9. Tm and the cloud points for aqueous solutions of poly(NIPAAmco-HIPAAm) as a function of the HIPAAm content. Polymer concentration: 10 w/v% for Tm and 1.0 w/v% for the cloud points; Tm (!), cloud points (2); Tm and the cloud points were determined from the DSC and UV–vis measurements, respectively.
sensitive polymer, poly(NIPAAm-co-HIPAAm). Particularly, water molecules existing around the copolymer chains were determined and divided into three distinct states, namely, free water, bound water, and non-freezing water. By adding the copolymers to water, the shoulder peaks resulting from the melting of the bound water appeared in the DSC thermograms. Also, the Tm , which is the temperature due to the melting of the bound water, shifted to lower temperatures with increasing the HIPAAm composition. Based on these results, it was believed that the bound water more strongly interacts with the polymeric chains with an increase in the HIPAAm composition. Furthermore, the endothermic enthalpy for the phase transition and/or separation, Ht , decreased with increasing the HIPAAm composition. These results indicate that the amount
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of water molecules dissociated from the polymeric chains decreases with increasing the HIPAAm content. In 1 H NMR spectra, the intensities of the peaks arising from each proton of the copolymers with a low HIPAAm content, such as NH0 and NH-10, were dramatically reduced above their cloud points. These results suggest that the polymeric chains brought about an abrupt dehydration during the phase transition and the copolymers were insoluble. On the other hand, the peak intensity was not drastically reduced in the copolymer containing a high content of HIPAAm above each cloud point of the copolymer solutions, and the remaining peaks progressively increased as the HIPAAm composition increased. Consequently, we suggest that the dehydration of the copolymer chains that occurred in the phase transition and/or separation becomes incomplete with increasing the HIPAAm content, and these results were correlated with the ones obtained from the DSC measurements. We determined the correlation between the Tm , the cloud points of the copolymers, Ht and the HIPAAm content for poly(NIPAAm-co-HIPAAm) aqueous solutions. With an increase in the HIPAAm composition, the Tm and Ht decreased, and the cloud points shifted to a higher temperature. An increase in the hydrophilic comonomer (HIPAAm) content raises the hydrophilicity of the copolymer chains, thus, the interaction between the bound water and the polymeric chains could become stronger. This occurrence accounts for the fact that the dehydration of the polymeric chains induced during the phase transition and/or separation becomes more difficult to occur. Therefore, it was demonstrated that the amount of dissociated water molecules is reduced and the cloud points of the copolymer solutions increase. That is, the relationship between the Tm , the cloud point, Ht and the HIPAAm composition is determined for poly(NIPAAm-co-HIPAAm), and it becomes clear that the thermosensitive behavior of a temperature-responsive polymer is dependent on the bound water. Acknowledgment This research was partially supported by the Ministry of Education, Science, Sports and Culture, through a Grant-in-Aid for Scientific Research (B) 15300167, 2003.
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