Self-Assembly of a Thermally Responsive Double-Hydrophilic

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Self-Assembly of a Thermally Responsive DoubleHydrophilic Copolymer in Ethanol-Water Mixtures: The Effect of Preferential Adsorption and Co-Nonsolvency Victoria Iskrova Michailova, Denitsa B. Momekova, Hristiana A. Velichkova, Evgeni H. Ivanov, Rumiana Kirilova Kotsilkova, Daniela B. Karashanova, Elena D. Mileva, Ivaylo Vladimirov Dimitrov, and Stanislav Miletiev Rangelov J. Phys. Chem. B, Just Accepted Manuscript • DOI: 10.1021/acs.jpcb.8b01746 • Publication Date (Web): 14 May 2018 Downloaded from http://pubs.acs.org on May 15, 2018

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The Journal of Physical Chemistry

Self-Assembly of a Thermally Responsive DoubleHydrophilic Copolymer in Ethanol-Water Mixtures: The Effect of Preferential Adsorption and CoNonsolvency AUTHOR NAMES: Victoria I. Michailova,†* Denitsa B. Momekova,† Hristiana A. Velichkova,‡ Evgeni H. Ivanov,‡ Rumiana K. Kotsilkova,‡ Daniela B. Karashanova,# Elena D. Mileva,ǁ Ivaylo V. Dimitrov,§ Stanislav M. Rangelov§ AUTHOR ADDRESS: †Faculty of Pharmacy, Medical University of Sofia, 2, Dunav Str., Sofia 1000, Bulgaria ‡Institute of Mechanics, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. 4, Sofia 1113, Bulgaria #Institute of Optical Materials and Technologies, Bulgarian Academy of Sciences, 109 Akad. G. Bonchev Str., Sofia 1113, Bulgaria ǁInstitute of Physical Chemistry, Bulgarian Academy of Sciences, Akad. G. Bonchev Str., Bl.11, Sofia 1113, Bulgaria

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§Institute of Polymers, Bulgarian Academy of Sciences, Akad. G. Bonchev Str., Bl. 103A, Sofia 1113, Bulgaria

ABSTRACT: Lower alcohols can induce a combined collapse-swelling de-mixing transition (LCST-type co-nonsolvency) in aqueous solutions of poly(N-isopropylacrylamide) (PNIPAM) by interacting with the polymer’s amide groups. This interaction results in an increase of the total surface area of hydrophobic sites and destabilizes the chains. Here, we make use of this phenomenon to drive the counterintuitive self-assembly of a PNIPAM-containing doublehydrophilic graft copolymer in water-ethanol mixtures at T ≪ LCST. Rheological frequency sweeps are used to quantify the distinct solvation states of PNIPAM at various temperatures and ethanol concentrations. The energy stored through elastic deformation at the de-mixing transition is simply related to the solvent binding. We find that the storage modulus decreases progressively, but non-linearly with ethanol concentration, which evidences a preferential solvation pattern. Analogously, through a combination of DLS and TEM analyses we demonstrate that a low-temperature structure variation takes place by adding ethanol following a similar solvent-content morphology dependent model.


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INTRODUCTION Thermally responsive, smart materials have received significant attention in many research fields, including biomedicine, sensors and separation technology, due to their ability to undergo sharp changes in their physical properties as a result of small variations in temperature.1-3 Most of these applications rely on the self-assembled organization of lower critical solution temperature (LCST)-based macromolecules that can yield ordered structures in a broad collection of morphologies, including micelles, vesicles, and multi-scale hierarchical structures.4,5 Such nano-structures are classically formed by self-assembly of amphiphilic block copolymers in water which associates the hydrophobic block due to the hydrophobic hydration, while the responsive block sterically stabilizes the aggregates and imparts them with temperature-regulating properties.3,6 Recently, increasing attention has been given to the doublehydrophilic copolymers (DHCs) with thermally responsive blocks or segments.7,8 Their conformational transition from hydrophilic coils to hydrophobic and contracted globules above the LCST has been widely used as a driving force for construction of LCST-switchable assemblies, some of which are considered promising as catalytic nano-reactors or drug delivery nano-vectors.9-11 While the past studies on DHCs mainly focused on the solution properties in relation to the temperature response, there are only a few examples on the DHCs self-assembly at < ,12,13 where all blocks are hydrophilic and soluble and the present hydrophobic interactions are considered insufficient to drive polymer micellization. In spite of this, such copolymers are able to form unique water-in-water mesophases in concentrated aqueous solutions, including body-centered-cubic, hexagonal and lamellar lyotropic phases, provided that the blocks are incompatible and not completely hydrated.14-16 For dilute DHCs systems,


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however, generation of loose internally disordered aggregates but not defined nano-structures has only been reported below LCST until now.12,17 On the other hand, lower alcohols, such as methanol and ethanol, may provide an opportunity for DHCs self-assembly at < . Due to the dual affinity for polar and non-polar residues of amphiphilic molecules, these alcohols have a strong influence on the conformation and stability of various bio- and synthetic polymers in aqueous solutions, the most prominent example being the drop in the LCST and the re-entrant coil-to-globule-to-coil transition of poly(N-isopropylacrylamide) (PNIPAM) in mixtures of water with alcohol, a good solvent for PNIPAM, so-called the “co-nonsolvency effect”.18,19 Recent spectroscopic and scattering studies20-23 and molecular dynamics simulations24,25 have provided evidence that preferential hydrogen bonding of alcohol onto PNIPAM is a key feature that promotes the dehydration and collapse of PNIPAM in the region of co-nonsolvency. A water-alcohol solvation shell has been shown to form in solution already below LCST, in which methanol preferably resides in the vicinity of the polymer chain orientating the non-polar methyl groups toward the bulk solvent. It has been proposed that at water-rich solvent compositions and > , interfacing that hydrophobic layer with the polar bulk solvent can induce step-wise water extrusion away from the outer solvent regions, while preserving the local solvent composition around PNIPAM chains almost unchanged.20 However, in the alcohol-rich bulk solvent, with increasing methanol content in the layer structure, the misbalance weakens and the LCST ultimately disappears at a “critical” bulk solvent composition, since the mixed solvent has become now compatible with the polymer solvation shell. In the present work, we specifically took advantage of the formation of such a solvation shell to produce a range of nano-structured essentially micellar assemblies in dilute aqueous-ethanol


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solutions of a PNIPAM-based DHC and at ≪ . The DHC used here, namely PNIPAM-gPEO (PNIPAM-graft-poly(ethylene oxide)) graft copolymer, was obtained via the “grafting through” method using maleic acid monoester of PEO-based macro-monomers (Mn = 2000 Da) whose low reactivity in copolymerization with NIPAM yielded a linear copolymer with a sparse degree of PEO grafting (2.7 mol% PEO), as described previously.26 The low graft content was preferred to ensure that the copolymer could assume various morphologies if the solvent quality worsens for the backbone.27 On the other hand, the PEO chains being incompatible with PNIPAM12,28 and having higher binding affinity to hydro-ethanolic solutions than to the individual solvents29 were expected to further promote manipulation of the assemblies’ morphology by changing core-corona and corona chains repulsive interactions.30,31

EXPERIMENTAL SECTION Materials.

Poly(N-isopropylacrylamide)-graft-poly(ethylene

oxide)

(PNIPAM-g-PEO)

copolymer with a low degree of PEO grafting (2.7 mol%), weight average molecular weight of 1.1 x 105 and polydispersity index of 1.65 was synthesized and characterized according to a procedure described elsewhere (Supporting Information (SI), Figure S1, Table S1).26 Ethanol (95% v/v) was purchased from Sigma Aldrich. Pure water was obtained through Milli-Q (Millipore, Merck Germany) purification. Sample Preparation. Samples for rheological, TEM and DLS experiments were prepared from 0.5 mg/ml stock solutions of PNIPAM-g-PEO dissolved at room temperature in MilliQ water. In each case, 0.5 ml of polymer stock solution was diluted to a total volume of 2.5 ml first with water and then with ethanol (0.15 ml.min-1 addition rate; moderate stirring), at 20°C. After equilibration of 45 min, samples were subjected to further analyses.


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Instruments and Methods. Rheological Measurements. Rheological measurements were performed using AR-G2 Rheometer (TA Instruments) with a Peltier plate geometry (cone of 60 mm). The tangent of the phase angle , storage and loss " moduli were measured versus the angular frequency of 0.1 – 100 rad.s-1 at low strain amplitude of 0.01 (viscoelastic range). The gap size between plates was 29 µm. The linear viscoelastic range of the strain amplitude was determined by strain sweep test at angular frequency of 1 Hz. All the rheological measurements were carried out at a fixed temperature in the temperature range of 20-50°C unless other stated. Transmission Electron Microscopy. The morphologies of aggregates were examined by HRTEM (high resolution transmission electron microscope, JEOL JEM 2100, Japan) operated at an accelerating voltage of 200 kV. To prepare the specimens for TEM analysis a drop of the sample solution was deposited on a carbon coated copper grid. The excess solution was blotted away with a strip of filter paper resulting in the formation of thin film suspended on the mesh holes. The grid was then dried at 20°C in a chamber with controlled environment. No staining was used for the TEM observations. Dynamic Light Scattering Measurements. The average size and size distribution of the nanoparticles were estimated by dynamic light scattering (DLS) using a Malvern Zetasizer Nano 3600 (Malvern Instruments Ltd., UK) equipped with a He-Ne laser (0.4 mW; 633 nm) and a temperature-controlled cell holder, at 20oC. The intensity of the scattered light was detected at 173o to the incident beam. The average diameter (Z-ave) of the aggregates was recorded as a mean of three measurements.

RESULTS AND DISCUSSION


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Prior to probing the morphogenic effect of ethanol, we conducted rheological frequency sweep experiments on dilute solutions of PNIPAM-g-PEO (0.01 %wt) in water-ethanol mixtures aiming to assess the solvation and phase behavior of the copolymer under the co-nonsolvency constraint. In such a test, polymeric systems containing elastically active components, i.e. contracted PNIPAM segments, store more energy and generate larger dynamic moduli values through elastic deformation, which allows differentiation between the distinctive solvation states of PNIPAM.32 In the present study, the oscillatory test parameters - the storage modulus ’, the loss modulus ”, and the tangent of the phase angle δ ( = "/ ′) - were measured by means of a dynamic oscillation program, and their values obtained for various water-ethanol mixtures at 20°C and 45°C were compared at the lowest frequency of 0.1 rad.s-1 (Figure 1). The data showed that the presence of ethanol and the temperature influenced the rheological properties of the polymer solutions (see also Figures S2 and S3 in the SI for comparison with the oscillatory rheograms). At 20°C, below the polymer’s LCST, the values of the viscoelastic moduli were very low due to the solvated coil conformation of the copolymer (Figure 1a). The ’ and ” moduli decreased as the volume fraction of ethanol increased indicating that both the elastic and the viscous properties decreased when ethanol was added. The δ values were higher than 1 in mixtures with ethanol excess ( > 0.5) reflecting the propensity of ethanol to increase the viscous character of the solvent swollen coils (Figure 1c; see also Figure S2 g-i). The elastic properties ( δ < 1) dominated the solvent system at higher water concentrations (Figure 1a, c; Figure S2 a-f) consistent with the less expanded conformation of PNIPAM in water compared to that in pure ethanol.20,22


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Figure 1. Oscillatory test parameters at a frequency ω = 0.1 rad.s-1 for PNIPAM-g-PEO aqueous solutions as a function of solvent composition. Storage modulus ’ and loss modulus ” at (a) 20°C and (b) 45°C, (c) δ at 20°C and 45°C. Copolymer concentration 0.01 %wt. The lines are guides to the eye.


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The viscoelastic properties obtained upon heating at 45°C, beyond the transition temperature, were very different for the different solvent mixtures (Figure 1b, c; see also Figure S3 in the SI). At water-rich solvent compositions ( = 0 − 0.4), the samples showed the same general behavior as that at the lower temperature ( > " and ≪ 1), but the dynamic moduli values were 5-75 times higher, which showed the stiffer nature of the PNIPAM chain segments after their transition from coil conformation to globule conformation. The inclusion of ethanol significantly influenced the rheological properties of the solutions and the ’ and ” values markedly decreased with increasing . As already mentioned, cooperative dehydration of PNIPAM took place upon heating allowing the development of hydrophobic and H-bonding interactions between the chains segments.7,33 In the ethanol-water mixtures, these relations were apparently weakened due to ethanol still present along the PNIPAM segments at higher temperatures,34-36 which interfered with the chain collapse and loosened (swelled) the globules. When even more ethanol was added to the solutions ( = 0.5), the sensitivity of PNIPAM-gPEO to temperature changes became insignificant. No considerable difference between the viscoelastic behavior at 20°C and 45°C was noted (See also Figure S2 g-i and Figure S3 g in the SI); the systems exhibited nearly the same plasticity as the graft copolymer in pure ethanol, i.e., the bound solvent turned the globular PNIPAM into swollen flexible coils again. The evolution of these solvation changes was visualized by plotting the storage and the loss moduli values (ω = 0.1 rad.s-1) of the PNIPAM-g-PEO aqueous-ethanol solutions (ϕ = 0 − 0.5) as a function of temperature (Figure 2a; see also Figure S4 in the SI). Except for sample solutions with ϕ = 0.5, the ’, ” versus dependence yielded sigmoidal plots indicative of LCST-switching: an abrupt increase in the two moduli values was observed at a certain temperature, the mid-point of which was taken as the phase transition temperature, ; upon


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further heating, at about + 5°, both moduli approached almost constant values, indicating the PNIPAM main chains underwent a completely collapsed state. It is worth emphasizing that the onset temperatures of ’ and ” concurred for each ethanol concentration, meaning that the inter-related changes of the PNIPAM-solvents and PNIPAM-PNIPAM interactions started at the same temperature and did not depend on the present ethanol. However, it could be seen from Figure 2d that the elevation of the loss modulus was only moderate and quite unaffected by the change of the solvent composition in the ethanol/water mixtures. Conversely, thanks to the elastic nature of the LCST phase separation,37,38 the effect of ethanol could be clearly discerned by the shift and variation of the ’ versus temperature dependence (Figure 2c), which decreased in magnitude and moved to lower temperatures as ethanol increased from ϕ = 0 to 0.5. Specifically, the decreased from 44 (ethanol free) to 37, 34.6 and 34.4°C in the presence of

ϕ = 0.10, 0.15 and 0.20, respectively (Figure 2b), but this relation reversed for = 0.3 and 0.4, and at = 0.5 there was not a detectable . It is also worth noting that the critical transition temperature of PNIPAM-g-PEO was much higher than that of PNIPAM homopolymers (Figure 2b). Besides, UCST-type phase behavior39 was not observed in the covered temperature range (20-45°C). This depressed phase transition could be attributed to the grafted PEO chains which having good solubility in ethanol/water mixtures29 and partially mixing with PNIPAM40 may act as a compatibilizing layer between the PNIPAM backbone and the mixed solvent, thus preventing the collapse.


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Figure 2. (a) Temperature dependence of the storage modulus ’ (filled symbols) and the loss modulus ” (open symbols) at frequency ω = 0.1 rad.s-1 for PNIPAM-g-PEO in water. The critical solution temperature is marked by a vertical dashed line. The rounded rectangle indicates the position of ’ at the high temperature plateau ( () used to prepare Figure 3. (b) (filled squares) is plotted against the volume fraction of ethanol . LCST (filled triangles) and UCST (open triangles) of PNIPAM as a function of are also shown as a reference, adapted from Polymer, Vol. 43, Ricardo O.R Costa, Roberto F.S Freitas, Phase behavior of poly(Nisopropylacrylamide) in binary aqueous solutions, 2002, Pages No 5879- 5885, Copyright 2002, with permission from Elsevier. (c) Storage modulus ’ and (d) loss modulus ” (ω = 0.1 rad.s-1)


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vs. temperature for PNIPAM-g-PEO in various ethanol/water mixtures. The copolymer concentration is 0.01 %wt. The lines are guides to the eye. The re-entrant phase transition of PNIPAM in mixed solvents of methanol and water has been recently described in terms of the theoretical model of competitive hydration (preferential adsorption) in which the cooperative, but mutually exclusive and competitive H-bonding of water and methanol onto PNIPAM chains has been considered to cause co-nonsolvency.34-36,41 In the literature, there are only a few works concerning the determination of the preferential adsorption of solvents on PNIPAM.20,21 By the use of the dynamic oscillatory test, we are able here to evaluate the contribution of the bound ethanol in dilute solutions in terms of the elastic properties of PNIPAM-g-PEO after the transition, e.g., the average values of the ’ modulus at the high temperature plateau, (. The data showed (Figure 3) that as the ethanol content increased from 0 to 0.5 (the co-nonsolvency region), the plateau storage modulus ( decreased but did not obey a linear dependence, as one would expect for classical binary mixed solvents. A moderate reduction of ( was observed initially (0 < ϕ < 0.2), suggesting the occurrence of preferential adsorption of water over ethanol, followed by sharper dropping. Presumably, for entropic reasons, this result reflected the reduced possibility of ethanol to approach the Hbonding sites (amide groups) and form long bound sequences on the chains in low concentration, as pointed out by the aforesaid model. Stronger adsorption cooperativity of water and ethanol molecules increases the competition in forming H-bonds on PNIPAM and the adsorption of even small amount of alcohol could cause the cooperative dissociation of a large amount of water molecules leading to negative excess of solvents binding.34,36 In the regions of higher ϕ (> 0.2), the conformation of non H-bonded PNIPAM segments can no longer sustain the restriction of the


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sequences of ethanol-PNIPAM H-bonds36 and the chains expand abruptly due to the extensive adsorption of alcohol molecules.

Figure 3. The value of the plateau storage modulus ( is plotted against the solvent composition. The dependence displays positive deviation from linearity (dashed line). The position of ( for the analysis is shown with a rounded rectangle in Figure 2a and Figure S4 in the SI. Statistical error bars correspond to one standard deviation. The line is a guide to the eye.

Figure 4. Variations of the scattered light intensity from aqueous dispersions of PNIPAM-gPEO with ethanol volume fraction. Copolymer concentration 0.01 %wt, temperature 20°C. The line is a guide to the eye.


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Considering our observations on the solvation and phase behavior of PNIPAM-g-PEO in ethanol-water mixtures, the formation of distinct self-assembled nanostructures was anticipated at temperatures below the LCST and at lower fractions of ethanol ( < 0.5), a mixed solvent less compatible to the PNIPAM chains solvation shells. In order to avoid the pre-contracting dehydration of PNIPAM,8 the self-assembly was performed at 20°C, ≪ ), by slowly adding ethanol to aqueous solutions of the DHC to give 0.01 %wt dispersions. The sizes and morphologies of aggregates with increasing were further analyzed by dynamic light scattering (DLS) and high resolution transmission electron microscopy (HR-TEM). Figure 4 shows the variations of the scattered light intensity with ethanol volume fraction. The intensity increased step-wise in the ranges from 0.05 to 0.20 and from 0.25 to 0.40 reaching a maximum at = 0.50. Upon a further increase of the ethanol volume fraction, the scattered light intensity linearly decreased and reached a value in pure ethanol comparable to that in pure water (below 60 kcps). As the scattered light intensity is proportional to the size and mass of the particles, its variations can be related with the variations of the particle size distribution (Table 1). The distributions were typically bimodal. A third mode, corresponding to large (micron-size) particles only occasionally appeared. Although its amplitude was low (below 5.8%), it might have influenced the positions of the other two modes, particularly the second one. The first mode corresponded to relatively small particles. In dispersions with ethanol volume fractions up to 0.25, their hydrodynamic radii were in the 10 – 15 nm range, which is larger than what was expected for unimers (unassociated copolymer molecules), and can be attributed to loose aggregates of low aggregation number. At higher ethanol volume fractions, the size of these particles was doubled and their contribution to the total scattered light intensity became dominant. A second mode, corresponding to considerably larger particles (ca. 50 – 180 nm range), was invariably present.


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The size of the particles, associated with this mode, was less consistent and a clear dependence of their size variation with ethanol volume fraction was hardly observed. The reason for this could be the unsystematic appearance of an additional third mode (see above), which most probably derived from impurities such as dust or formation of short-living transient large objects. The monomodal distribution at = 0.50, at which the scattered light intensity exhibited a maximum, was noteworthy.

Table 1. Dynamic Light Scattering Data from Aqueous Dispersions of PNIPAM-g-PEO at Different Ethanol Volume Fractions*. Ethanol volume fraction

PDI

0

Rh (nm) 1st mode%

2nd mode%

0.267

11.057.0

83.642.8

0.05a

0.431

14.311.3

76.485.9

0.10

0.331

17.544.0

67.356.0

0.15 a

0.444

12.528.1

93.768.4

0.20 a

0.519

15.545.0

139.349.2

0.25 a

0.251

10.13.1

49.994.9

0.30 b

0.253

26.593.8

0.35

0.288

22.479.5

95.720.5

0.40

0.372

22.469.8

69.530.2

0.50 c

0.252

88.9100

1

0.327

32.184.5

182.115.5

*Copolymer concentration 0.01 %wt, temperature 20°C. a – presence of a third (slow) mode of low amplitude; b – the additional mode corresponds in position and amplitude to the third modes from a; c – monomodal distribution. The numbers in superscript correspond to the contribution to the overall scattered light intensity.


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Figure 5. TEM picture of the morphological transition of PNIPAM-g-PEO aggregates with ethanol volume fraction. (a) Disordered sponge-like globules at ϕ = 0; inset: magnification image of the selected globules showing micro-phase separated PNIPAM and PEO domains (the latter marked by arrow). (b, c) Hierarchical porous aggregates along with (c) worm-like intermediate structures at ethanol fraction ϕ = 0.10. (d) Bi-continuous structures at ϕ = 0.15. (e) Nano-sized and giant vesicles formed upon further increasing the ethanol content (ϕ = 0.30). The strawberry-like characteristics of the membrane are clearly visible. (f) At ϕ = 0.6, hierarchical porous structures reformed. Copolymer concentration 0.01 %wt, temperature 20°C. The TEM images revealed that in pure water the copolymer formed monodisperse small globules with a diameter of ∼ 25-30 nm and a disordered sponge-like structure indicative of micro-phase separation (Figure 5a). The PNIPAM block, which was the less hydrated copolymer component,28 appeared dark in the TEM micrographs, whereas the lighter, almost electron transparent zones (d ∼ 2 nm) represented the PEO segments of the globules. When 0.05-0.10 ethanol fractions were added to the solution, a lamellar structure (d = 2.5 - 3.5 nm) emerged near


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the surface of the globules (d = 20 - 36 nm) enclosing internally segregated porous domains (Figure 5b). We also observed a large amount of elongated worm-like objects along with complex structured connections between the hierarchical porous structures (Figure 5c), which suggested that the latter presumably formed through their coalescence.42 Upon further increase of the ethanol content up to = 0.15, the internal phase separation apparently increased and extended unilamellar aggregates with embedded branched network of rods and small globules appeared in the TEM micrographs (Figure 5d). Although these super-aggregates were inwardly highly disordered and non-axisymmetric, perforations in the encapsulating shell, connecting the internal and external aqueous phase, inferred the formation of bi-continuous phase. With further increasing between 0.3 and 0.4, the complex structures were no longer present and the major morphologies observed were unilamellar vesicles (Figure 5e), with an average diameter increasing from ca. ∼ 45 to 70 nm over that solvent composition range, as revealed by the DLS experiments. A small population of giant unilamellar vesicles (2 µm < d < 5 µm) were also observed. Independent of the size, the vesicular wall was similar in thickness (∼ 3.5 nm) to that noted before for the = 0.1 and = 0.15 ethanol systems, but exhibited strawberry-like and uneven features, as represented by the darker dots in between the brighter structure. This result was expected from previous studies on preparation of core-deposited PNIPAM capsules upon heating, where the shells formed through aggregation of nanometer-sized mesoglobules.43 Finally, at ethanol volume fractions between 0.5 and 0.7, hierarchical porous structures reformed (Figure 5f), although being a little smaller and predominantly monodisperse compared to those at 0.1 < < 0.25 (Table 1). Note that this re-entrant self-assembling behavior fell into the same solvent composition range and was quite similar to the solvent-induced LCST phase transition, but occurred at lower temperature, once again highlighting the importance of the pre-contraction


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solvation of PNIPAM to the phase transition. As already mentioned, when lower alcohols solvate PNIPAM through H-bonding interaction, the total surface area of hydrophobic sites increases and the chains are destabilized in water;25,44 therefore, the macromolecules have to micellize and adapt their morphology for a given in order to reduce the core-corona interfacial tension without stretching the two block chains excessively, in analogy with the classical force balance thermodynamic concept for micellization of amphiphilic block copolymers.30,31 From the phase transition results, vesicles were expected to emerge in solutions with = 0.15 − 0.20 (i.e., the minimum in LCST) since this would be consistent with the largest interfacial energy at the PNIPAM core-mixed solvent interface. However, the TEM results demonstrated considerable deviation to a larger indicating that something in addition to the hydrophobic effect and amphiphilic interactions was responsible for the self-assembly process. It is well-known that the interaction between the polymer chains and the organic solvent can alter the aggregate morphology by changing the dimensions of both the aggregate core and the corona: the morphology evolves from spheres to rods to vesicles as the core enlarges and stretches the chains due to solvent binding, whereas this trend translates in reverse (i.e. from vesicles to micelles) if the corona chains solvation interactions are stronger.45 From Figure 3, it could be seen that the region of low solvent binding at < 0.3 overlapped the composition range for the formation of the complex structures. Clearly, a minute amount of ethanol was enough to produce a jump of two morphological steps beyond spheres, all the way to vesicles, by preferentially adsorbing to PNIPAM and stretching the coils; the contribution of intra-corona interactions were considered less important in this case as the solvent quality for the PEO chains only marginally changed for this range of .29,46 When the ethanol content was raised further up to = 0.3 and = 0.4, the solvation of the PNIPAM chains abruptly increased (Figure 3)


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and the energetics of the system changed such that favored the formation of vesicles. On may presume that the entropy cost of having high chain stretching in the PNIPAM core has become now largest for these solvent compositions, causing the aggregates to convert into the lowest curvature of the vesicles. Generally, with further addition of solvent the stretching increases, vesicles grow in size and then may adapt “inverted” micelle morphology for free energy optimization. Contrary to that expectation, at higher ethanol contents ( = 0.5 − 0.7), the copolymer underwent a re-entrant transition from vesicles back to the complex (cage-like) structures that seemed quite related to the corona solubility morphology dependence.47-49 Note that the solvent quality for the PEO moieties markedly increases for solutions with around 0.5,29,46 leading to corona swelling. In addition, beyond a certain level, the presence of ethanol along the molecules of PNIPAM started to increase the polymer-mixed solvent attractive interactions.25 In that case, the local core-solvent interfacial tension is assumed to be low and the vesicle walls are sloppy, therefore the structures would adjust to the force balance variation by increasing their surface area, by fluctuating and wobbling around.48 This was theoretically described for small amphiphilic block copolymers with basically solvent-philic components:48,49 when the fluctuations become sufficiently strong, small micelles are eventually cut off, released and coalesced inside of the vesicle until finally internally branched micro-structure is formed.

CONCLUSIONS We demonstrated the self-assembly properties of a thermally responsive double-hydrophilic PNIPAM-g-PEO copolymer in ethanol-water co-nonsolvent mixtures and at temperature far below the hydrophobic collapse transition of the copolymer. The micellization in that system was favored due to the destabilization of PNIPAM by the alcohol molecules H-bonded onto the


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chains. Using rheological frequency sweeps to assess the solvent binding at the LCST-de-mixing transition we showed that the preferential solvation of PNIPAM caused progressive, but nonlinear swelling of the micellar core resulting in a two-step structure variation from small globules to vesicles. The present studies significantly extend our understanding of the molecular mechanisms that determine the solution properties of PNIPAM-based DHCs in complex media, which is of major importance for their use as transformable functional materials.

ASSOCIATED CONTENT Supporting Information. 1H NMR spectrum (Figure S1) and characterization data (Table S1) of PNIPAM-g-PEO copolymer; oscillatory rheograms at 20°C (Figure S2) and 45°C (Figure S3), and temperature dependence of the dynamic moduli values at a frequency of 0.1 rad.s-1 (Figure S4) for aqueous PNIPAM-g-PEO solutions with various volume fractions of ethanol. (PDF) AUTHOR INFORMATION Corresponding Author * Phone: +359 2 9236 546. Fax: +359 2 9879 874. E-mail: [email protected] (V. M.) Author Contributions The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript. Notes The authors declare no competing financial interests.


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Self-Assembly of a Thermally Responsive Double-Hydrophilic

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