Nanospheres Prepared by Self-Assembly of Random Copolymers in Supercritical Carbon Dioxide

The synthesis of spherical particles was attained by the direct self-assembly of poly[2-(perfluorooctyl)ethyl acrylate-random-acrylic acid], P(POA-r-AA), and by the indirect self-assembly poly[POA-random-2-(dimethylamino)ethyl acrylate], P(POA-r-DAA), with dicarboxylic acids in supercritical carbon dioxide (scCO2). The copolymers formed spherical particles with hundreds of nanometer diameters in a heterogeneous state at pressures lower than the cloud point pressure. The formation of spherical particles was also dependent on the temperature. The formation of spherical particles could be optimized through varying the solvent quality by the manipulation of the CO2 pressure and temperature for the different copolymer compositions. The dynamic light scattering and 1H NMR studies demonstrated that the nanospheres had the micellar structures consisting of the CO2-philic POA shells and the CO2-phobic AA or DAA cores including the main chain cores. The nanospheres produced the superhydrophobic surfaces based on the water-proof shells of the POA units.

We found a novel and convenient method to prepare nanospheres through the direct and indirect self-assembly of random copolymers in scCO 2 [24][25][26] [18][19][20] Scheme 1: The self-assembly of the block copolymers and surfactants in scCO 2 .describes the preparation of nanospheres by the selfassembly of random copolymers containing the CO 2 -philic 2-(perfluorooctyl)ethyl acrylate (POA) units.

Instrumentation.
The 1 H NMR measurement was conducted using a Varian 300 FT NMR spectrometer.The gel permeation chromatography (GPC) was performed using a Tosoh GPC-8020 instrument equipped with a DP-8020 dual pump, a CO-8020 column oven, and an RI-8020 refractometer.Two polystyrene gel columns, Tosoh TSKgel GMH HR -M, were used with hexafluoroisopropanol (HFIP) as the eluent at 40      of methanol to precipitate a polymer.The polymer was collected and was dried in vacuo for several hours to obtain P(POA-r-TBA) (4.44 g).The P(POA-r-TBA) (2.00 g) was dissolved in HFIP (60 mL).Concentrated hydrochloric acid (2 mL) was added to the copolymer solution at room temperature, and the solution was refluxed for 3 h.After the solution was cooled to room temperature, it was concentrated to one-third of its original volume with an evaporator.The residual solution was poured into hexane (1 L) to precipitate a polymer.The polymer was purified by reprecipitation from hexafluorobenzene (5 mL) into hexane (500 mL).The resulting polymer was dried in vacuo for 5 h to obtain P(POA-r-AA) (1.76 g). were placed in ampule.After the contents were degassed, the ampule was sealed in vacuo.The polymerization was carried out at 60 • C for 10 min and was terminated by cooling with liquid nitrogen.The reaction mixture was dissolved in hexafluorobenzene and poured into hexane to precipitate a polymer.The precipitate was dried in vacuo for several hours to obtain P(POA-r-DAA) (0.994 g).

Cloud Point Measurement.
The cloud-point measurement was performed with a variable volume view cell (Scheme 2).P(POA-r-AA) (30 mg) was placed in the cell, then CO 2 liquefied with a cooler was added to it.The cloud point was defined as the point at which the contents of the cell turned opaque, indicating precipitation of the polymer from solution.
2.6.Preparation of Particles in ScCO 2 .P(POA-r-AA) (POA/ AA = 7/3, 30 mg) was placed in the variable volume view cell, then liquid CO 2 was added to it.The solution was stirred at 3620 psi and 45.0 • C for 10 min.The pressure of the homogeneous solution was reduced until the cloud point (3059 psi) was reached by volume expansion.The pressure was further reduced to 2759 psi, and the heterogeneous solution was sprayed into a plastic bag made of polyethylene to collect the polymer particles (29 mg).

SEM Measurement.
The polymer particles were put on a carbon adhesive tape and were subjected to SEM measurement after coated with Pt.

Contact Angle Measurement.
The surfaces completely covered with the nanospheres were prepared on a carbon adhesive tape attached to a slide glass.The contact angles were measured by dropping water (10 μL) on the nanosphere-coated surface at ambient temperature.The surface on the cast film of the copolymer was prepared directly on a slide glass from a copolymer solution in hexafluorobenzene.

Direct Self-Assembly through Hydrogen Bond
The P(POA-r-AA) random copolymers were obtained by the hydrolysis of P(POA-r-TBA) prepared by the radical polymerization (Scheme 3).The hydrolysis of P(POA-r-TBA) was carried out in HFIP using concentrated hydrochloric acid under reflux for 3 h.The characterizations of the P(POA-r-TBA) and P(POA-r-AA) copolymers are listed in Tables 1 and 2. The molecular weight distributions of P(POA-r-AA) could not be estimated due to the very broad distribution.
The P(POA-r-AA) copolymers completely dissolved in scCO 2 .Figure 1 shows the CO 2 density at the cloud points plotted versus temperature.The CO 2 density at the cloud points decreased as the temperature increased, indicating that the solubility of the copolymers increased with the increasing temperature.And further, the copolymer with a higher AA ratio had a lower solubility, suggesting that the hydrogen bonding based on the AA units prevented the copolymer from dissolving in scCO 2 .SEM observations revealed that the P(POA-r-AA) copolymers self-assembled into spherical particles in scCO 2 .Figure 2 shows the SEM images of the particles obtained from the copolymer solution at 45 • C in a heterogeneous state at a pressure lower than the cloud point pressure.The images included spherical particles independent of the POA/AA ratio.However, the proportion of the spherical particles was different among the various ratios.The copolymers with the 9/1 and 8/2 ratios contained some spherical particles of several micrometers in diameter and a lot of small particles with nonspecific forms.These copolymers had too low AA content to aggregate into spherical forms.The copolymer with the 6/4 ratio also contained both the spherical particles and the small particles.However, the small particles also had a spherical shape with several hundreds of nanometer diameters.It is likely that the intramolecular aggregation is preferable for the 6/4 sample due to large contribution of the hydrogen bonding based on the higher AA content.The copolymer with the 7/3 ratio produced the largest amount of spherical particles among the copolymers.Based on the SEM image of the 7/3 sample, the size of the spherical particles was estimated to be 820 nm on average, and the size distribution was 1.20 [27].These SEM images are for the polymer particles obtained by spraying the heterogeneous solutions into the atmosphere, based on the assumption that the shapes of the particles formed in scCO 2 are maintained in this atmosphere at ambient temperature by rapidly releasing the CO 2 pressure.
In order to confirm the formation of the particles in solution, the light scattering measurements of the copolymer were performed in hexafluorobenzene.Hexafluorobenzene is similar to scCO 2 in having low polarity and the ability to dissolve fluoropolymers.Figure 3 shows the scattering intensity distribution of the hydrodynamic diameter of the particles formed by the copolymer with the 7/3 ratio in hexafluorobenzene.The intensity distribution was obtained by a Marquadt analysis of the dynamic light scattering results.The hydrodynamic diameter of the particles was estimated to be 600 nm based on the Marquadt analysis.Compared with the particle size based on the SEM image, the copolymer formed smaller particles in hexafluorobenzene.This can be accounted for by the fact that the AA units more tightly formed the particle cores in hexafluorobenzene than in scCO 2 because of the lower affinity of hexafluorobenzene to the AA units and also that the particles swelled when rapidly sprayed into the atmosphere.The 1 H NMR analysis confirmed that the particles consisted of the POA unit shells and the AA unit cores.The 1 H NMR spectra of the copolymer in the presence and absence of trifluoroacetic acid are shown in Figures 4(a) and 4(b).The copolymer showed no aggregation in the presence of trifluoroacetic acid.The signals based on the methyne and methylene of the main chain were broadened in the absence of trifluoroacetic acid, as compared to those in its presence.This broadening of the signals indicates that the insoluble AA units and main chains were shielded from the magnetic field.Consequently, it is expected that the copolymers formed random copolymer micelles consisting of the shells of the CO 2 -philic perfluorooctyl groups and the cores of the CO 2 -phobic AA units and main chains when the copolymers were placed in scCO 2 as well as in hexafluorobenzene (Scheme 4).

International Journal of Polymer Science
The shapes of the polymer particles were dependent on temperature based on the formation by hydrogen bonding.Figure 5 shows the SEM images of the polymer particles obtained at three different temperatures in the heterogeneous state.The particles were not completely spherical at 35 • C. The copolymer could not completely take a spherical shape because of the low mobility of the copolymer at this low temperature.On the other hand, the copolymer produced large International Journal of Polymer Science   spherical particles of several micrometers at 55 • C, along with small particles with hundreds of nanometer diameters.It was considered that an increase in the mobility of the copolymer at the high temperature promoted both the intermolecular aggregation and the intramolecular association.
The self-assembly of the copolymers into spherical particles was controlled by the CO 2 pressure.Figure 6 shows the SEM images of the polymer particles produced under the different pressures.The particles produced at a pressure higher than the cloud point pressure had nonspecific forms.Spherical particles were hardly observed in the image.The particles prepared at the cloud point pressure included a small amount of spherical parts, however, most of the particles were still nonspecific.When the pressure was reduced to 100 psi lower than the cloud point pressure, the copolymer somewhat aggregated into spherical particles.At a pressure 300 psi lower than the cloud point, most of the particles had spherical shapes.However, it was observed that the spherical particles partly combined at the cloud point pressure minus 900 psi.Completely spherical and larger-sized particles were produced at the cloud point minus 1500 psi.It was found that more stable spherical particles were produced at the lower pressure and that the self-assembly of the copolymer was controlled through varying the solvent quality manipulated by the CO 2 pressure.
The copolymers with the 9/1 and 8/2 ratios were found to produce nearly spherical particles at the pressure 1500 psi lower than the cloud point pressure, although a few spherical particles were obtained at the cloud point pressure minus 300 psi (Figure 7).The hydrogen bonding interaction among the AA units increased with the decreasing pressure, thus the self-assembly of the copolymers into spherical particles was promoted.At the pressure 1500 psi lower than the cloud point, the copolymer with the 6/4 ratio formed not only spherical particles (Figure 8(a)), but also a small amount of nonspecific aggregates of dozens or hundreds micrometers

Indirect Self-Assembly through Hydrogen Bond
It is possible to induce self-assembly for a copolymer that shows no self-assembly by itself through hydrogen bond interaction with additives.P(POA-r-DAA) random copolymers dissolve in scCO 2 , however, these copolymers show no self-assembly in it.The characterizations of the P(POAr-DAA) copolymers prepared by radical copolymerization are listed in Table 3 (Scheme 5). Figure 9 shows the plots of the cloud points versus the CO 2 density.There was a slight difference in the cloud point among the POA/DAA ratios of the copolymers.It was found that the solubility of the copolymers in scCO 2 decreased in the presence of perfluoroazelaic acid (PA).Figure 10 shows the cloud points of the copolymer with 7/3 of POA/DAA in the absence and presence of PA at 0.3 and 0.5 as a molar ratio of PA to the DAA unit.As a result of increasing the amount of PA, the cloud points were shifted to a higher side of the CO 2 density.The copolymer became less soluble in the presence of PA.The decrease in the solubility should be caused by hydrogen bond cross-linking between the amino groups in the copolymer via the carboxylic groups of PA.SEM observation demonstrated that the copolymer formed different shapes in the absence and presence of PA in scCO 2 .Figure 11 shows the SEM images of the copolymer with and without PA.The images were obtained for the polymer particles produced in the heterogeneous state below the cloud points.The copolymer took unspecific forms in the absence of PA.There was a slight difference in the shape at 0.3 as the PA/DAA ratio.On the other hand, the copolymer had spherical forms at the 0.5 ratio.The size of the particles was ca.700 nm.At 0.5 of PA/DAA, all the amino groups interact with the carboxylic group of PA, so that no free amino groups are expected to exist in the copolymer.Therefore, the nanospheres should be formed by cross-linking between the amino groups in the copolymer through the electrostatic interaction with PA (Scheme 6).
We explored the stability of the nanospheres versus pressure.Figure 12 shows the SEM images of polymer particles obtained from three different states: the heterogeneous state below the cloud point, the state at the exact cloud point, and the homogeneous state over it.The pressure at the heterogeneous state was 300 psi lower than the cloud point pressure, while that at the homogeneous state was 500 psi higher.The particles included PA at the 0.5 ratio.The spherical particles formed in the heterogeneous state partly changed to unspecific forms as a result of increasing the pressure to the cloud point pressure.The nanospheres completely changed to unspecific forms over the cloud point.These changes were based on the cross-linking by the weak electrostatic interaction and on an increase in the solubility of the nanospheres into scCO 2 as a result of increasing the density of CO 2 .Consequently, the shape of the polymer particles was dependent not only on the acid concentration but also on the pressure of CO 2 .
The spherical shapes of the polymer particles were independent of the temperature.Figures 13 shows the SEM images of the polymer particles obtained at 35, 45, and 60 • C at 0.5 of PA/DAA.The particles were prepared in the heterogeneous states at which the pressures were 300 psi lower than the cloud point pressure at each temperature.
The polymer particles maintained their spherical shapes at 45 and 60 • C. In addition, the copolymer with PA took more completely spherical shape with an increase in the temperature.It may be accounted for by the fact that the perfluoroalkyl chains extend much more at higher temperature, resulting in that the copolymer more easily form spherical particles at the higher temperature.
The POA/DAA ratio in the copolymer affected the cloud point and size of the nanospheres.Figure 14 shows the cloud points of three different copolymers with POA/DAA of 7/3, 8/2, and 9/1 in the presence of PA at 0.3 and 0.5 of PA/DAA.Whereas there was a negligible difference in the cloud point among the copolymers in the absence of PA, the copolymers made a marked difference in the presence of it.This difference was much greater as the amount of PA increased.The copolymer having a lower DAA content showed lower cloud point in the presence of PA.The SEM observation revealed that the copolymer with a lower DAA content formed smaller nanospheres.Figure 15 shows the SEM images of the nanospheres obtained from the copolymers with PA at the 0.5 ratio.The size of the nanospheres decreased as the DAA content decreased.The copolymer with the 8/2 ratio formed the nanospheres with ca.600 nm as the particle size, while the 9/1 copolymer produced those with 400 nm.The decrease in the size of the nanospheres with the decrease in the DAA content may be account for by the fact that the copolymer with lower DAA content forms a lower degree of cross-linking due to the nonspecific forms made some contribution to enhancing the hydrophobicity of the surface.On the other hand, the smooth surface of the cast films showed the small contact angle.

Conclusions
The synthesis of spherical particles was attained by the direct self-assembly of P(POA-r-AA) and by the indirect self-assembly of P(POA-r-DAA) with the dicarboxylic acids in scCO 2 .The copolymers formed spherical particles with hundreds of nanometer-diameters in a heterogeneous state at pressures lower than the cloud point pressure.The formation of spherical particles was also dependent on the temperature.The formation of spherical particles could be optimized through varying the solvent quality by the manipulation of the CO 2 pressure and temperature for the different copolymer compositions.The dynamic light scattering and 1 H NMR studies demonstrated that the nanospheres had the micellar structures consisting of the CO 2 -philic POA shells and the CO 2 -phobic AA or DAA cores including the main chain cores.The nanospheres had the superhydrophobicity based on the water-proof shells of the POA units.This study demonstrated that the nanospheres prepared by the self-assembly in scCO 2 produced the superhydrophobic surfaces.

Figure 2 :Figure 3 :
Figure 2: SEM images of the polymer particles obtained from the copolymers in scCO 2 at 45 • C and the cloud point pressure −300 psi.

Figure 4 : 1 H
Figure 4: 1 H NMR spectra of P(POA-r-AA) with a 7/3 POA/AA ratio in the presence (a) and absence (b) of CF 3 COOH.Solvent: C 6 F 6 including C 6 D 6 .

Figure 5 :
Figure 5: SEM images of the polymer particles obtained at 35, 45, and 55 • C at the cloud point pressure −300 psi.

Figure 7 :
Figure 7: SEM images of the polymer particles obtained from the copolymers with 9/1 and 8/2 POA/AA ratios at 45 • C and the cloud point pressure −1500 psi.

Figure 8 :
Figure 8: SEM images of the polymer particles obtained from the copolymer with a 6/4 POA/AA ratio at 45 • C and the cloud point pressure −1500 psi.

Figure 9 :Figure 10 :Figure 11 :
Figure 9: Plots of the experimental cloud points of the copolymers at each temperature versus the CO 2 density.

Figure 12 :
Figure 12: SEM images of polymer particles obtained from the heterogeneous state below the cloud point, the state at the exact cloud point, and the homogeneous state over it.POA/DAA = 7/3, PA/DAA = 0.5.

Figure 14 :Figure 15 :
Figure 14: Plots of the cloud points of the copolymers with POA/DAA of 7/3, 8/2, and 9/1 in the absence and presence of PA at 0.3 and 0.5 of PA/DAA.

Figure 16 :
Figure 16: Plots of the experimental cloud points of the P(POA-r-DAA) copolymer in the presence of the dicarboxylic acids at each temperature.The acid/DAA = 0.5.

Figure 17 :
Figure 17: SEM images of the particles produced by the copolymer in the presence of Glu and Psuc, and photographs of water droplets placed on the surface coated with the particles.

Figure 19 :
Figure 19: SEM images of the random forms produced by the copolymer and of the surface on the cast films.Also shown are photographs of water droplets placed on the surface of the random forms produced by the copolymer and of the surface on the cast films.
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a Estimated by GPC based on poly(methyl methacrylate) standard.b Estimated by 1 H NMR.

Table 4 :
Contact angles (CAs) on the surface coated with the nanospheres.Acid/DAA = 0.5.b Acid/DAA = 1.0.c Treated in scCO 2 .d The cast film prepared from a solution in hexafluorobenzene under atmosphere.
a N Scheme 6: A schematic of the formation of the nanospheres.