pubs.acs.org/Langmuir © 2009 American Chemical Society
Construction of Multifunctional Coatings via Layer-by-Layer Assembly of Sulfonated Hyperbranched Polyether and Chitosan Xiaofen Hu and Jian Ji* Department of Polymer Science and Engineering, Key Laboratory of Macromolecule Synthesis and Functionalization of Minster of Education, Zhejiang University, 310027, Hangzhou, China Received July 24, 2009. Revised Manuscript Received August 29, 2009 Layer-by-layer assembly has shown a great deal of promise in biomedical coatings, as well as local drug delivery systems. The poor loading capacity of hydrophobic drugs within the multilayers is a drawback in their potential applications. Herein, sulfonated hyperbranched polyether (HBPO-SO3) with a hydrophobic core was incorporated into LBL films to provide nanoreservoirs for hydrophobic guest molecules. HBPO-SO3 was proven to form stable micelles in the sodium acetate and acetic acid buffer solution (HAc buffer) for LbL assembly. The QCM and ellipsometry experiments demonstrated that the LBL films can be fabricated via alternating deposition of HBPO-SO3 micelles and chitosan. The fluorescence emission spectra verified that the hydrophobic pyrene can be incorporated both by pre-encapsulation in HBPO-SO3 micelles and post-diffusion in preassembled multilayer films. Compared with the pre-encapsulation approach, the post-diffusion process was more efficient in incorporating hydrophobic guest molecules into the LbL films and carried out a much more controllable release of the guest molecules. A multifunctional coating with potential anticoagulation, antibacterial, and local release of hydrophobic drug Probucal, which has powerful antioxidant properties and can prevent restenosis after coronary angioplasty, was then developed via postdiffusion of the anti-restenosis agents into the multilayer films of HBPO-SO3 and chitosan.
Introduction The layer-by-layer (LbL) assembly method has attracted much attention due to its versatility and convenience.1,2 Multilayer thin films with tailored structure and composition are easily fabricated via sequential adsorption of oppositely charged species on a charged substrate with different geometry.3 Numerous charged species have been used as building blocks for LbL films, such as polyelectrolytes,4-6 biomacromolecules,7,8 nanoparticles,9 and colloids,10,11 providing functional coatings for implanted medical devices. Recently, micelles with charged coronas were involved in the development of local drug delivery systems.12-16 Sun et al. described the fabrication of LbL multilayer films based on polyelectrolyte stabilized surfactant micelles as carriers for noncharged hydrophobic dyes.12 Hammond and co-workers integrated linear-dendritic block copolymer micelles encapsulating a hydrophobic antibacterial drug into LbL films.13 Furthermore, Rubner and co-workers suggested that the interpenetration of the *To whom correspondence should be addressed. E-mail:
[email protected]. Tel./fax:þ86-571-87953729.
(1) Decher, G. Science 1997, 277, 1232. (2) Zhang, X.; Shen, J. C. Adv. Mater. 1999, 11, 1139. (3) Tang, Z.; Wang, Y.; Podsiadlo, P.; Kotov, N. A. Adv. Mater. 2006, 18, 3203. (4) Fu, J. H.; Ji, J.; Shen, L. Y.; Kueller, A.; Rosenhahn, A.; Shen, J. C.; Grunze, M. Langmuir 2009, 25, 672. (5) Ji, J.; Fu, J. H.; Shen, J. C. Adv. Mater. 2006, 18, 1441. (6) Jiao, Q.; Yi, Z.; Chen, Y. M.; Xi, F. Polymer 2008, 49, 1520. (7) Ren, K. F.; Ji, J.; Shen, J. C. Macromol. Rapid Commun. 2005, 26, 1633. (8) Ren, K. F.; Ji, J.; Shen, J. C. Biomaterials 2006, 27, 1152. (9) Kotov, N. A.; Dekany, I.; Fendler, J. H. J. Phys. Chem. 1995, 99, 13065. (10) Gao, M.; Gao, M.; Zhang, X.; Yang, Y.; Yang, B.; Shen, J. C. Chem. Commun. 1994, 2777. (11) Schmitt, J.; Decher, G. Adv. Mater. 1997, 9, 61. (12) Liu, X. K.; Zhou, L.; Geng, W.; Sun, J. Q. Langmuir 2008, 24, 12986. (13) Nguyen, P. M.; Zacharia, N. S.; Verploegen, E.; Hammond, P. T. Chem. Mater. 2007, 19, 5524. (14) Ma, N.; Zhang, H. Y.; Song, B.; Wang, Z. Q.; Zhang, X. Chem. Mater. 2005, 17, 5065. (15) Manna, U.; Patil, S. J. Phys. Chem. B 2008, 112, 13258. (16) Qi, B.; Tong, X.; Zhao, Y. Macromolecules 2006, 39, 5714.
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weak polyelectrolytes in the alternating layers is an important issue regarding the surface property of the polyelectrolyte multilayers.17 A significant number of the chain segments of the previously adsorbed layer penetrate into the outermost surface layer. The interpenetration of the biomacromolecules then provides the possibility of constructing a multilayered coating surface with synergic properties of different biomacromolecule components.18,19 We previously have proven that the multilayered thin films constructed by alternating deposition of heparin and chitosan onto aminolyzed poly(ethylene terephthalate) (PET) films exhibit strong anticoagulant and antibacterial activities.18 However, the poor loading capacity of hydrophobic drugs within these traditional polyelectrolyte multilayer films is a real drawback to their potential applications. Hyperbranched polymers have recently received much attention as one class of spherical compounds due to their easy preparation, versatility, and the capability of incorporating high loadings of different types of molecules within their imperfectly branched structures.20-22 Furthermore, the great number of external groups of hyperbranched polymers provides many possibilities to be either functionalized or multifunctionalized for potential applications in biomedicine, pharmacology, and biotechnology.23-26 Most recently, we have synthesized a heparinlike sulfonated hyperbranched polyether (HBPO-SO3) consisting (17) Yoo, D.; Shiratori, S. S.; Rubner, M. F. Macromolecules 1998, 31, 4309. (18) Fu, J. H.; Ji, J.; Yuan, W. Y.; Shen, J. C. Biomaterials 2005, 26, 6684. (19) Serizawa, T.; Yamaguchi, M.; Akashi, M. Biomacromolecules 2002, 3, 724. (20) Hong, H. Y.; Mai, Y. Y.; Zhou, Y. F.; Yan, D. Y.; Chen, Y. J. Polym. Sci., Part A: Polym. Chem. 2008, 46, 668. (21) Wan, A. J.; Kou, Y. X. J. Nanopart. Res. 2008, 10, 437. (22) Kontoyianni, C.; Sideratou, Z.; Theodossiou, T. Macromol. Biosci. 2008, 8, 871. (23) Jiang, G. H.; Chen, W. X.; Xia, W. Des. Monomers Polym. 2008, 11, 105. (24) Dong, W. Y.; Zhou, Y. F.; Yan, D. Y.; Li, H. Q.; Liu, Y. Phys. Chem. Chem. Phys. 2007, 9, 1255. (25) Zhou, Y. F.; Yan, D. Y. Angew. Chem., Int. Ed. 2005, 44, 3223. (26) Mao, J.; Ni, P. H.; Mai, Y. Y.; Yan, D. Y. Langmuir 2007, 23, 5127.
Published on Web 09/18/2009
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of a hydrophobic hyperbranched poly(3-ethyl-3-oxetanemethanol) (HBPO) core and negatively charged sulfonic acid terminal groups.27 The HBPO-SO3 could self-assemble in aqueous media to form stable micelles and displayed good hemocompatibility and low cytotoxicity. We hypothesize here that the negatively charged HBPO-SO3 micelles can be used as a building block for LbL assembly, and the hydrophobic cores of the micelles will introduce hydrophobic nanodomains into multilayer films, which can be regarded as nanoreservoirs for hydrophobic guest molecules. The layer-by-layer assembly of sulfonated hyperbranched polyether and chitosan will then provide the new possibility to develop a multifunctional coating capable of anticoagulation and antibacterial and local drug delivery.
Experimental Section Materials. Chitosan (average MW ca. 410 000, 91% deacetylation) was obtained from Qingdaos Haihui Corporation of China. Heparin (sodium salt) was purchased from Shanghais Chemical Reagent Company of China. Polyethylenimine (PEI, average Mw ca. 25 000 (LS)) and pyrene were purchased from Aldrich Chemical Co. PET membranes cut into squares 3 cm 3 cm were cleaned by sonication in acetone, methanol, and pure water for 10 min, respectively, followed by rinsing with pure water. Quartz slides (1 cm 2 cm) and silicon wafers were cleaned by treatment in hot piranha solution (H2O2/H2SO4 3:7 v/v) for 40 min (caution: piranha solution is extremely corrosive) and then thoroughly washed with pure water. Synthesis of HBPO-SO3. The synthesis and complete characterization of the sulfonated hyperbranched polyether HBPO-SO3 have been previously published.27 Briefly, the mixtures of HBPO and excess sodium hydride in tetrahydrofuran (THF) reacted under reflux overnight with stirring before adding 1,3-propane sultone, and then allowed to react for 12 h more under reflux. The resulting solution was filtered; then, the crude product was exhaustively dialyzed against pure water, and a white product HBPO-SO3 was obtained. The sulfonation degree was estimated to be around 45%. Preparation of Multilayer Films. The LbL films were assembled on PET membranes for plasma recalcification time experiments, silicon wafers for ellipsometry and atomic force microscopy (AFM) measurements, and quartz slides for UV-vis and fluorescence emission spectra, respectively. The substrate was first immersed in PEI solution for 30 min to ensure a uniform positively charged coating so that the effects of the substrate on the layer growth are minimized. After rinsing with pure water and drying under a nitrogen stream, the resulting substrate was alternately immersed into solutions of HBPO-SO3 and chitosan for 10 min each. Between each deposition step, the substrate was rinsed with pure water and blown dry with a stream of nitrogen. This cycle was repeated until the desired number of HBPO-SO3/ chitosan layers (typically 10) was reached. Multilayer films based on heparin and chitosan were fabricated as the control. All polymers, PEI, HBPO-SO3, chitosan, and heparin, were of the same concentration, 1 mg/mL in sodium acetate and acetic acid buffer (HAc buffer, pH = 4, 0.1 M). Loading and Release of Pyrene. Two different methods were used to load pyrene into HBPO-SO3/chitosan multilayer films. The first approach required the encapsulation of pyrene in HBPO-SO3 micelles before the LbL assembly, while the second involved the post-diffusion of pyrene molecules from the solution into the preassembled multilayer films. Pre-Encapsulation Approach. Ten milligrams of HBPO-SO3 was first dissolved in HAc buffer (10 mL) to form micelles; then, pyrene in THF solution (1 mg/mL, 2 mL) was added. The solution containing pyrene was vigorously stirred in the dark at room temperature. After THF was totally removed using a rotative (27) Hu, X. F.; Ji, J. Acta Polym. Sin. 2009, 8, 828.
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evaporator, the solution was filtered through a 0.45 μm membrane to eliminate the precipitated pyrene and was then ready for LbL deposition. Post-Diffusion Method. A quartz slide deposited with the LbL films (10 bilayers) was exposed to the chloroform solution of pyrene (2 mg/mL). The quartz slide was taken out after 24 h, rinsed three times with chloroform to remove the dye molecules weakly adsorbed on the surface, and dried in vacuo. The quartz slide covered with pyrene-loaded multilayer films was immersed into a vial of phosphate buffered saline (PBS, pH= 7.4, 0.1 M) at 37 °C, which was replaced by fresh solution at appropriate time point to ensure constant release conditions. The concentration of pyrene in the PBS solution was analyzed with the UV-vis spectrometer . Loading and Release of Probucal. Probucal, which has powerful antioxidant properties and can prevent restenosis after coronary angioplasty,28 was incorporated into HBPO-SO3/ chitosan and heparin/chitosan multilayer films by the post-diffusion approach. Probucal was previously dissolved in chloroform or dimethyl sulfoxide (DMSO) at 1 mg/mL. Quartz slide deposited with the multilayer films (10 bilayers) was exposed to the Probucal solution for 24 h, then rinsed three times in the pure organic solvent and dried in vacuo. The amount of Probucal loaded in multilayer films was analyzed by the measurement of the characteristic absorbance of Probucal at 240 nm. The quartz slide covered with Probucal-loaded multilayer films was immersed into a vial of phosphate buffered saline (PBS, pH= 7.4, 0.1 M) at 37 °C. At regular intervals, the quartz slide was removed from the solution, rinsed with pure water, dried under nitrogen flow followed by UV-vis absorption, and then moved to a fresh vial of PBS to maintain sink conditions. Plasma Recalcification Time (PRT). Platelet-poor plasma (PPP) was bought from Central Blood Bank in Hangzhou. PPP (50 μL) was dropped onto the surface of sample and allowed to stand for 1 min at 37 °C before the addition of 0.025 M CaCl2 solution (50 μL), at which point a stopwatch was started. The stopwatch was stopped when fibrin clotting was first visible, and the time was recorded. At least six experiments were carried out for each sample and the mean clotting time reported. Characterization Techniques. Quartz crystal microbalance (QCM) measurements were taken with a Q-Sense QCM-E4 using Au-coated resonator (Hongrong Boman Biotechnology Co. Ltd., Beijing, China). The fundamental resonant frequency of the crystal was 5 MHz. The crystal was mounted in a fluid cell with one side exposed to the solution. A measurement of LbL deposition was initiated by switching the liquid exposed to the resonator from HAc buffer to a PEI solution. PEI was allowed to adsorb onto the resonator surface for 15 min before being rinsed with buffer. Then, HBPO-SO3 (with or without pyrene encapsulation) and chitosan solutions were alternately introduced for 15 min with buffer rinsing in between. All the polymers, PEI, HBPO-SO3 (with or without pyrene encapsulation), and chitosan, were of the same concentration, 1 mg/mL in HAc buffer (pH = 4, 0.1 M) and injected into the chamber at a rate of 0.1 mL/min. Spectroscopic ellipsometry was carried out using a M-2000 (J. A. Wollam Co. Inc.) to measure the film thickness on silicon wafers. AFM images were performed in the tapping mode under ambient conditions using a commercial scanning probe microscope, Seiko SPI3800N, equipped with a silicon cantilever, Nanosensors, typical spring constant 40 N 3 m-1. Fluorescence spectra were recorded on a spectrofluorometer (FP-770, Japan Spectroscopic) at room temperature. Emission spectra were recorded over the range 350-500 nm with an excitation wavelength of 336 nm. UV-vis spectra were obtained on a Shimadzu model UV-2550 spectrometer. (28) Tardif, J. C.; C^ote, G.; Lesperance, J.; Bourassa, M.; Lambert, J.; Doucet, S.; Bilodeau, L.; Nattel, S.; deGuise, P. N. Engl. J. Med. 1997, 337, 365.
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Figure 1. Chemical structure of HBPO-SO3.
Results and Discussion Fabrication and Characterization of the LBL Films of HBPO-SO3 and Chitosan. The multilayer films were formed through an alternating deposition of negatively charged HBPOSO3 and positively charged chitosan on PEI adsorbed substrates. HBPO-SO3 used here consists of a hydrophobic hyperbranched poly(3-ethyl-3-oxetanemethanol) (HBPO) core and negatively charged sulfonic acid terminal groups (Figure 1). As shown in our previous paper, when the concentration of HBPO-SO3 increased above 0.017 mg/mL, these heparin-like hyperbranched polyethers could self-assemble in aqueous solution to form spherical micelles with negatively charged sulfonic acid groups on the exterior, and exhibited good hemocompatibility and low cytotoxicity.27 The aggregation of HBPO-SO3 in the HAc buffer solution prepared for LbL assembly was investigated by TEM and the laser particle size analyzing system (Figure 2). HBPOSO3 self-assembled to form micelles with a mean diameter of 142 nm. QCM measurement was employed to confirm the successful LbL assembly, and a linear film growth was observed (Figure 3). The LbL assembly on silicon wafer was also monitored with ellipsometry. Figure 4 indicated a similar LbL growth curve with linear growth. On average, a bilayer was approximately 3 nm. The AFM images demonstrated the stability of HBPO-SO3 micelles during the LbL process and the presence of micelles in LbL films. As indicated in Figure 5, spherical micelles occurred on the surface of PEI adsorbed silicon wafer after the deposition of HBPO-SO3, and the diameter was consistent with that measured in solution. However, the results from ellipsometry measurements revealed that the increase in thickness due to the deposition of HBPO-SO3 micelles was limited and the thickness of (HBPOSO3/chitosan)10 multilayer films was much less than the diameter of the micelles. This was probably because the imperfectly branched polyether skeleton of this designed polymer was flexible and the terminal sulfonate groups made HBPO-SO3 a strong polyanion. For these reasons, the HBPO-SO3 micelles might be crushed under the electrostatic interaction with the positively charged surface, while the shrinkage of the aggregations occurred in the deposition, and compact multilayer thin films were fabricated as shown in Figure 6. 2626 DOI: 10.1021/la902719k
Figure 2. (a) TEM image of HBPO-SO3 micelles in HAc buffer solution at 1 mg/mL. (b) Results of the size distribution by particle size distribution analyzer.
Figure 3. QCM frequency decrease (-ΔF) for the alternating deposition of HBPO-SO3 micelles (with or without pyrene) and chitosan. The LbL assembly was initiated with the adsorption of PEI on resonators. The odd and even layer numbers correspond to the deposition of HBPO-SO3 micelles and chitosan, respectively.
Drug Loading by Pre-Encapsulation and Post-Diffusion in Multilayer Films. The hydrophobic dye pyrene was used as a model drug and incorporated into the LbL films by pre-encapsulation in HBPO-SO3 micelles and post-diffusion in preassembled multilayer films, respectively. Langmuir 2010, 26(4), 2624–2629
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Figure 4. Thicknesses of assembled films at various number of bilayers. The LbL assembly was initiated with the adsorption of PEI on silicon substrates.
Figure 7. (a) Fluorescence spectra of a 10-bilayer film of pyreneloaded HBPO-SO3 micelles and chitosan. (b) Changes in the normalized fluorescence intensity of pyrene at 393 nm as a function of the number of micelle layers.
Figure 5. AFM images of PEI adsorbed silicon wafer before (a) and after (b) deposition of HBPO-SO3 layer.
Figure 6. Schematic illustration of LbL assembly of negatively charged HBPO-SO3 micelles and positively charged chitosan on a PEI adsorbed substrate.
The LbL assembly of pyrene-loaded HBPO-SO3 micelles and chitosan was carried out, and continuous film growth was monitored with QCM and ellipsometry. Unlike the LbL assembly of HBPO-SO3 micelles and chitosan mentioned above, the multilayer films based on pyrene-loaded HBPO-SO3 micelles showed a nonlinear growth pattern (Figures 3 and 4). Similar results have been reported previously in the multilayer of polyelectrolyte stabilized pyrene-loaded micelles due to the increased surface roughness.12 To ensure that pyrene remained in the HBPO-SO3 micelles during the LbL assembly process, fluorescence emissions of pyrene after the deposition of each layer of micelles were recorded (Figure 7). The fluorescence intensity of pyrene in the multilayer films increased with respect to the increase of the number of layers of pyrene-loaded micelles. On the basis of the aforementioned results, pyrene could be incorporated into the LbL films via pre-encapsulation in the HBPO-SO3 micelles, and pyrene-loaded HBPO-SO3 micelles did not induce Langmuir 2010, 26(4), 2624–2629
a significant perturbation of the LbL deposition with chitosan. The amount of pyrene loaded in the multilayer films can be controlled by simply changing the cycles of films deposited. It is well-known that the fluorescence spectrum of pyrene is sensitive to the polarity of the surrounding environment.29 The vibronic band intensities of the fluorescence emission spectra provide a new possibility for elucidating the location of the pyrene within the multilayer. The fluorescence intensity ratio of the two peaks at 383 and 373 nm (I3/I1) reflects the change of environmental polarity.29 The I3/I1 ratio of pyrene in an aqueous HBPO-SO3 micelle solution was 1.19, which indicated that the pyrene molecules were incorporated into the hydrophobic cores of HBPO-SO3 micelles. The I3/I1 ratios of the pyrene within the multilayer by pre-encapsulation in HBPO-SO3 micelles and post-diffusion in the preassembled multilayer of HBPO-SO3/ chitosan were 0.94 and 0.91, respectively, while the fluorescence intensity of pyrene within the multilayer films based on heparin and chitosan was too weak to calculate the I3/I1 ratio. Although the deformation of the hyperbranched polyether micelles driven by the electrostatic interaction with chitosan might decrease the hydrophobicity of the core, most hydrophobic pyrene molecules were still incorporated into the hydrophobic HBPO cores. The LBL films of HBPO-SO3 and chitosan offer nanoreservoirs for hydrophobic guest molecules and provide a new possibility of incorporating a hydrophobic drug. The UV-vis absorption spectra of pyrene-loaded multilayer films were presented in Figure 8a. Although the peaks were slightly hypsochromically shifted in the pyrene-loaded LbL films, the incorporation of the pyrene molecules was not questioned. For the multilayer films, both with ten deposition cycles, more pyrene loading was achieved via the post-diffusion process. (29) Kalyanasundaram, K.; Thomas, J. J. Am. Chem. Soc. 1977, 99, 2039.
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Figure 9. UV-vis spectra of (a) HBPO-SO3/chitosan multilayer films without loading; heparin/chitosan multilayer films after immersion in (b) chloroform or (c) DMSO solution of Probucal; HBPO-SO3/chitosan multilayer films after immersion in (d) DMSO or (e) chloroform solution of Probucal; and (f) Probucal in dilute solution. All the LbL films were fabricated with ten deposition cycles.
Figure 8. (a) UV-vis spectra of pyrene in CH3OH solution (dash), blank LbL films (dot), pyrene-loaded LbL films via the post-diffusion method (black solid) and the pre-encapsulation approach (gray solid), respectively. All the LbL films were fabricated with ten deposition cycles. The spectrum of pyrene in dilute solution was modified by raising the baseline, for clarity in comparison. (b) Release profile of pyrene from the multilayer films.
UV-vis spectrometry was also employed to quantify the release of pyrene from the multilayer films. The pyrene-loaded LbL films prepared via either the pre-encapsulation approach or the postdiffusion process showed continuous release over a period of several weeks. In the case of the pre-encapsulation approach, a burst release of 0.96 μg pyrene within the first 24 h can be shown, reaching a cumulative release of about 3.0 μg pyrene after 20 days and a sustainable continued release of pyrene (Figure 8b). Further inspection of Figure 8b revealed that the release of pyrene incorporated via the post-diffusion method went through three stages: with an initial slow release in the first 30 h, subsequently a more rapid release of about 0.47 μg per day, and then a sustainable slow release after 5 days. The absence of the usual burst effect indicated that the pyrene loadings in multilayer films were below the saturation concentration.30 The post-diffusion process was more efficient at incorporating hydrophobic guest molecules into the LbL films and carried out a much more controllable release of the guest molecules than the pre-encapsulation approach. Fabrication of Multifunctional Coatings. Hydrophobic drug Probucal was chosen and incorporated into HBPO-SO3/ chitosan multilayer films via post-diffusion. We investigated the (30) Guyomard, A.; Nysten, B.; Muller, G.; Glinel, K. Langmuir 2006, 22, 2281.
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Probucal uptake by HBPO-SO3/chitosan multilayer films, taking heparin/chitosan multilayer films as the control. The thickness of preassembled HBPO-SO3/chitosan multilayer films was of the same order of magnitude as the heparin/chitosan multilayer films formed under the same conditions (10 bilayers, 34.9 nm vs 31.3 nm). As indicated in Figure 9, the HBPO-SO3/chitosan multilayer films showed the capability of loading after immersion in the chloroform solution of Probucal, while there was almost no Probucal within the heparin/chitosan multilayer films (Figure 9). Successful loading of Probucal in both HBPO-SO3/chitosan and heparin/chitosan multilayer films was obtained using DMSO as the solvent. However, Probucal-loaded heparin/chitosan multilayer films obtained by the post-diffusion of DMSO solution had a burst release within the first 10 h; approximately 66% of the drug was released, and the Probucal remaining in the multilayer films was hardly detected (Figure 10). The result revealed that the drugs might be entrapped within the surface of the heparin/ chitosan multilayer. As good solvents for the hydrophobic polyether cores of HBPO-SO3 micelles, chloroform and DMSO could drive diffusion of Probucal into hydrophobic nanodomains in the HBPO-SO3/chitosan multilayer films. The Probucalloaded HBPO-SO3/chitosan multilayer films via the post-diffusion process showed a continuous release over a period of several weeks. A burst release of 20% within the first 10 h can be shown, reaching a cumulative release of about 38% after 14 days, and a sustainable release continued for more than three weeks. The high concentration of sulfate and sulfamate groups of heparin has been proven to play an important role in the anticoagulant activity of heparin.31 A number of polymers containing sulfonate groups, so-called heparin-like materials, have been designed to exhibit more or less enhanced blood compatibility.32,33 As an alternative to heparin, HBPO-SO3 should provide the multilayer films with anticoagulant capability. The PRT assay was performed to determine the blood compatibility of HBPOSO3/chitosan multilayer films. Compared with the bare PET, (31) Tamada, Y.; Murata, M.; Hayashi, T.; Goto, K. Biomaterials 2002, 23, 1375. (32) Aguilar, M. R.; Rodrı´ guez, G.; Fernandez, M.; Gallardo, A.; Roman, J. S. J. Mater. Sci.: Mater. Med. 2002, 13, 1099. (33) Kim, Y. H.; Han, D. K.; Park, K. D.; Kim, S. H. Biomaterials 2003, 24, 2213.
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surprised to find that the coagulation of HBPO-SO3-terminated multilayer films was not observed until 1974 ( 11 s. However, it is interesting that the multilayer films with chitosan as the outermost layer also presented a PRT as long as 1670 ( 121 s, although the chitosan is procoagulated. The results are consistent with our previous heparin/chitosan multilayer films, in which the multilayer films with chitosan as the outermost layer exhibited strong anticoagulant and antibacterial activities due to interpenetration of the polyelectrolytes in the alternating layers.18 The results reported here implied that the same interpenetration of layers might exist in the multilayer films of sulfonated hyperbranched polyether and chitosan. A multifunctional coating with potential anticoagulation, antibacterial, and local release of anti-restenosis drugs can be developed via post-diffusion of the hydrophobic drug Probucal into the multilayer films of HBPO-SO3 and chitosan. Figure 10. Drug release profile of Probucal incorporated in HBPO-SO3/chitosan multilayer films (solid icons) and heparin/ chitosan (hollow icons) multilayer films by the post-diffusion of a DMSO solution of drugs.
Figure 11. Comparison of PRT obtained on the surface of PET (sample 1), HBPO-SO3/chitosan multilayer films (10 bilayers) assembled on PET with chitosan as the outmost layer (sample 2), and that with HBPO-SO3 as the outermost layer (sample 3).
which has been widely used in cardiovascular implants due to its excellent mechanical properties and moderate biocompatibility, the PRT of HBPO-SO3/chitosan multilayer films deposited on PET membranes was obviously prolonged (Figure 11). While coagulation occurred on the bare PET after 383 ( 80 s, it is not
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Conclusion Sulfonated hyperbranched polyether (HBPO-SO3) was employed as a building block to fabricate multilayer films with chitosan via LbL assembly. The QCM and ellipsometry results verified the progressive growth of films. Results from the loading and release experiments demonstrated that the hydrophobic guest molecules could be incorporated into the HBPO-SO3/chitosan multilayer films by either the pre-encapsulation process or the post-diffusion approach, and released in a controlled and sustainable way. In addition, the multilayer films exhibited anticoagulant activity even with chitosan as the outermost layer. These anticoagulant HBPO-SO3/chitosan multilayer films with anticipated antibacterial activity and excellent capability for loading and controlled release of hydrophobic drugs, such as antiproliferative agents, may have good potential for surface modification of implanted medical devices. Acknowledgment. Financial support from the Natural Science Foundation of China (NSFC- 20774082, 50830106), 863 National High-Tech R&D Program (2006AA03Z329, 2006AA03Z444), Ph.D. Programs Foundation of Ministry of Education of China (No. 20070335024), Open Project of State Key Laboratory of Supramolecular Structure and Materials (SKLSSM200911) and Natural Science Foundation of China of Zhejiang Province(Y4080250) is gratefully acknowledged. The authors thank Prof. Deyue Yan and Yongfeng Zhou at Shanghai Jiao Tong University for beneficial discussions and the offer of hyperbranched poly(3-ethyl-3-oxetanemethanol) (HBPO).
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