Abstract
The present invention relates to a process for preparing poly(3,4-ethylenedioxythiophene):polystyrenesulfonate (PEDOT:PSS) particles at least comprising the steps: a) providing a mixture comprising poly(3,4-ethylenedioxythiophene) and polystyrene sulfonate in a solvent at least comprising water; b) forming one or more PEDOT:PSS droplets by introducing the mixture from process step a) into an organic solvent A, wherein the aqueous PEDOT:PSS mixture forms the droplet interior and the organic solvent A forms the droplet exterior; c) contacting the PEDOT:PSS droplets obtained from process step b) with a coagulating solution comprising a curing agent and at least one further solvent B, the density of the coagulating solution being greater than the density of the organic solvent A and less than the density of the aqueous poly(3,4-ethylenedioxythiophene) and polystyrene sulfonate mixture; with curing of the PEDOT:PSS droplets to PEDOT:PSS particles. Furthermore, the present invention discloses spherical PEDOT:PSS particles without further mechanically solidifying substances and the use of the particles, for example, as cell culture microcarriers or suspension electrodes.
Claims
1. Process for preparing poly(3,4ethylenedioxythiophene):polystyrenesulfonate (PEDOT:PSS) particles at least comprising the steps: a) providing a mixture comprising poly(3,4-ethylenedioxythiophene) and polystyrene sulfonate in a solvent at least comprising water; b) forming one or more PEDOT:PSS droplets by introducing the mixture from process step a) into an organic solvent A, wherein the aqueous PEDOT:PSS mixture forms the droplet interior and the organic solvent A forms the droplet exterior; c) contacting the PEDOT:PSS droplets obtained from process step b) with a coagulating solution comprising a curing agent and at least one further solvent B, the density of the coagulating solution being greater than the density of the organic solvent A and less than the density of the aqueous poly(3,4-ethylenedioxythiophene) and polystyrene sulfonate mixture; with curing of the PEDOT:PSS droplets to PEDOT:PSS particles.
2. Process according to claim 1, wherein the organic solvent A is selected from the group consisting of branched or unbranched C5-C10 alkanes, branched or unbranched C5-C10 alcohols or mixtures of at least two solvents thereof.
3. Process according to claim 1, wherein the further solvent B is selected from the group consisting of branched or unbranched C1-C5 alcohols or mixtures of at least two solvents thereof.
4. Process according to claim 1, wherein the solvent A comprises octanol and the coagulating solution in process step c) comprises isopropanol as solvent B and sulfuric acid as curing agent.
5. Process according to claim 1, wherein the weight ratio of curing agent and solvent B in the coagulating solution, expressed as weight of curing agent divided by weight of solvent B, is greater than or equal to 0.005 and less than or equal to 0.2.
6. Process according to claim 1, wherein the PEDOT:PSS mixture in process step a) does not comprise any further mechanically solidifying substances.
7. Process according to claim 1, wherein the PEDOT:PSS mixture in process step a) comprises, in addition to water an organic solvent A as a further solvent component.
8. Poly(3,4-ethylenedioxythiophene)-polystyrenesulfonate particles, characterized in that the particles are spherical and do not contain any other mechanically solidifying substances in addition to PEDOT:PSS.
9. The particles according to claim 8, wherein the particles are porous and have a porosity greater than 0 volume % and less than or equal to 95 volume %.
10. The particles according to claim 8, wherein the particle has a modulus of elasticity greater than or equal to 0.05 MPa and less than or equal to 15 MPa.
11. The particles according to claim 8, wherein the particle is at least partially crystalline with Bragg reflections in an XRD spectrum at 4.3 (+−0.2) nm.sup.−1 and 18.4 (+−0.2) nm.sup.−1.
12. The particles according to claim 8, wherein the surface of the particle has a zeta potential of less than or equal to 0 mV.
13. Use of the particles according to claim 8, selected from the group consisting of cell culture microcarriers, suspension electrodes, switchable redox absorber material, catalyst supports, or combinations thereof.
14. Use according to claim 13, wherein the particles are used as cell culture microcarriers, wherein the surface of the particles is coated prior to cultivation with one or more molecules selected from the group consisting of poly-L-lysine, laminin, collagen, fibronectin, vitronectin or mixtures thereof.
15. Use according to claim 14, wherein the surface of the particles is first coated with poly-L-lysine and then with laminin.
Description
[0050] The figures show:
[0051] FIG. 1 an FeSEM image of a porous PEDOT:PSS particle prepared according to the invention;
[0052] FIG. 2 an FeSEM image of a porous PEDOT:PSS particle with fibroblast colonization prepared according to the invention;
[0053] FIG. 3 the gravimetric capacity of PEDOT:PSS particles according to the invention as a function of porosity;
[0054] FIG. 4 the dependence of the redox kinetics of PEDOT:PSS particles according to the invention as a function of porosity;
[0055] FIG. 5 the dependence of the particle diameters of PEDOT:PSS particles according to the invention as a function of porosity;
[0056] FIG. 6 the size distribution of PEDOT:PSS particles according to the invention prepared with a volume fraction of 30% of 1-octanol in the aqueous PEDOT:PSS mixture;
[0057] FIG. 7 the pore size distribution of PEDOT:PSS particles prepared according to the invention with a volume fraction of 30% 1-octanol in the aqueous PEDOT:PSS mixture;
[0058] FIG. 8 the proliferation of L929 cells on particles according to the invention as a function of time and as a function of the crystallinity of the support material;
[0059] FIG. 9 the influence of crystallinity on the aspect ratio on particles of proliferating L929 cells according to the invention;
[0060] FIG. 10 the influence of the crystallinity of particles according to the invention on the propagation area of L929 cells.
[0061] FIG. 1 shows an FeSEM image of a porous PEDOT:PSS particle prepared according to the invention. The PEDOT:PSS particle was prepared with a 1-octanol volume fraction of 30% in the aqueous PEDOT:PSS mixture. Since PEDOT:PSS particles are hydrogels and thus collapse in anhydrous environments, the particle was freeze-dried prior to optical analysis.
[0062] FIG. 2 shows an FeSEM image of a porous PEDOT:PSS particle colonized with L929 mouse fibroblasts. Colonization of the microcarrier is shown after 4 days of cultivation at 37° C., 95% humidity and 5% CO.sub.2. The culture medium was RPMI supplemented with 10% fetal calf serum and 1% penicillin-streptomycin. The inoculation concentration was 10,000 cells/cm.sup.2. Since PEDOT:PSS particles are hydrogels and thus collapse in anhydrous environments, the particle was dried with an ethanol series (35, 50, 70, 100%) followed by treatment in hexamethyldisilazane (HMDS) prior to optical analysis.
[0063] FIG. 3 shows the gravimetric electrical capacitance of the PEDOT:PSS particles as a function of the 1-octanol volume fraction in the 1-octanol PEDOT:PSS emulsion and as a function of the sampling rate. Higher 1-octanol fractions denote a larger proportion of pore volume to particle volume (porosity) and thus a higher specific surface area. Electrical capacitances were determined from cyclic voltametry measurements in a 3-electrode setup as a function of scan rate. Since the specific surface area of the particles is directly proportional to the particle capacitance, more porous particles show a higher gravimetric capacitance. Higher scan rates result in smaller capacitances because the faster cycling of voltages means that the complete surface area of the particle contributing to the capacitance is not used.
[0064] FIG. 4 shows the current curve as a function of time. The redox kinetics of the PEDOT:PSS particles were recorded by chronoamperometry measurements as a function of the volume fraction of 1-octanol and thus of the porosity in a 3-electrode setup. The reaction time of the particles shortens with increasing porosity, although the charge density increases with increasing porosity. The shortened reaction time is attributed to the high specific surface area as well as the good accessibility of the pore system, which allows for fast redox kinetics. The measurements were performed over 9 cycles, with only one cycle shown in the diagram.
[0065] FIG. 5 shows the average particle diameter as a function of the 1-octanol volume fraction in the 1-octanol PEDOT:PSS emulsion as a measure of particle porosity. All particles shown in the diagram were prepared with a 1-octanol flow rate (continuous phase) of 0.5 mL/min and a PEDOT:PSS dispersion/emulsion flow rate of 0.05 mL/min The particles have a particle diameter of approximately 540 pm regardless of particle porosity. The small standard deviation in particle diameter likely stems from the fact that the droplets in the co-flow device are produced monodisperse Smaller variations in particle diameter come from a very small difference in separation kinetics of the protective 1-octanol shell in the coagulation bath.
[0066] FIG. 6 shows the size distribution of high (left) and low (right) crystalline PEDOT:PSS particles prepared with a volume fraction of 1-octanol of 30% in the aqueous PEDOT:PSS mixture. A rather narrow particle size distribution is obtained for both cases.
[0067] FIG. 7 shows the porogen and pore size distributions for high and low crystallinity porous PEDOT:PSS particles prepared with a volume fraction of 1-octanol of 30%. Most of the pores show a size between 15 and 20 μm. More than 90% of the pores show a pore size between 10 and 30 μm.
[0068] FIGS. 8-10 show the results in cell colonization of particles according to the invention. For the culture experiments, spherical PEDOT:PSS particles were prepared from a 30 vol % 1-octanol in PEDOT:PSS (1.1-1.3 wt %) emulsion, which was brought to droplet breakup in a continuous 1-octanol phase. The emulsion was obtained via an ultrasonic homogenizer (Hierschler UP100H). Both phases were added together via a syringe pump (Chemyx, Nexus Fusion 4000) at flow rates of 0.05 and 0.5 ml/min, respectively. The coagulation bath consisted of 5 vol % sulfuric acid in isopropanol unless otherwise specified.
[0069] FIG. 8 shows the results of the proliferation of L929 cells on particles according to the invention as a function of time and as a function of the crystallinity of the support material. Using different coagulation with different amounts of acid, porous particles with different degree of crystallinity were produced. The low-crystalline particles were coagulated with 5 vol % and the high-crystalline particles were coagulated with 95 vol % sulfuric acid. Particles with different mechanical properties are obtained. The properties result as follows:
TABLE-US-00001 E-modulus Breaking load/ Elongation at Crystallinity in MPa kPa break % High 0.07 28 (+/−13) 36 (+/−6) Low 9.85 626 (+/−32) 13 (+/−6)
[0070] The different mechanical properties are a strong indication that the structure of the two samples, despite having the same composition, is different. These different properties of the spherical particles also lead to changes in the biological properties. FIG. 8 shows the results of cell proliferation of L929 mousefibroblasts with a seeding density of 2,600 cells/cm.sup.2, N=5 on pure PEDOT:PSS microcarriers, where viability was quantified using an XTT proliferation assay. It can be clearly seen that the crystallinity of the support material has an effect on cell proliferation. Proliferation from day 5 is significantly higher on high crystallinity samples (triangles) than on low crystallinity samples (circles).
[0071] FIG. 9 shows the influence of crystallinity on the aspect ratio on particles of proliferating L929 cells according to the invention. The degree of crystallinity of the particles also seems to have an influence on the achievable morphology of the cell lines used. By means of confocal microscopy, DAPI/phalloidin-stained L929 cells can be assessed morphologically. One way to visualize cell symmetry is to determine the aspect ratio of the L929 cells. Different cell morphologies are found on low and high crystallinity particles, with more rounded cell morphologies developing on low crystallinity particles and more elongated cell morphologies developing on high crystallinity particles. The seeding density was 2,600 cells/cm.sup.2, the measurement was performed on 250 cells on the second day.
[0072] FIG. 10 shows the influence of the crystallinity of particles according to the invention on the propagation area of L929 cells. The different degrees of crystallinity of the particles were obtained via different coagulation treatment of spherical particles. It can be seen that a single L929 cell colonizes a significantly larger area on crystalline particles. In contrast, the spread of cells on particles with low crystallinity is much more limited. Furthermore, it results that the cells on particles with low crystallinity probably proliferate deeper into the particle interior. The colonization density inside the particles, on the other hand, appears to be reduced in the case of highly crystalline particles.
