2624 DOI: 10.1021/la902719k Langmuir 2010, 26(4), 2624–2629Published on Web 09/18/2009

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 andFunctionalization 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 deliverysystems. The poor loading capacity of hydrophobic drugs within the multilayers is a drawback in their potentialapplications. Herein, sulfonated hyperbranched polyether (HBPO-SO3) with a hydrophobic core was incorporatedinto LBL films to provide nanoreservoirs for hydrophobic guest molecules. HBPO-SO3 was proven to form stablemicelles in the sodium acetate and acetic acid buffer solution (HAc buffer) for LbL assembly. The QCM andellipsometry 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 bothby pre-encapsulation in HBPO-SO3 micelles and post-diffusion in preassembled multilayer films. Compared with thepre-encapsulation approach, the post-diffusion process was more efficient in incorporating hydrophobic guestmolecules into the LbL films and carried out a muchmore controllable release of the guest molecules. Amultifunctionalcoating with potential anticoagulation, antibacterial, and local release of hydrophobic drug Probucal, which haspowerful antioxidant properties and can prevent restenosis after coronary angioplasty, was then developed via post-diffusion of the anti-restenosis agents into the multilayer films of HBPO-SO3 and chitosan.

Introduction

The layer-by-layer (LbL) assembly method has attractedmuchattention due to its versatility and convenience.1,2 Multilayer thinfilms with tailored structure and composition are easily fabricatedvia sequential adsorption of oppositely charged species on acharged substrate with different geometry.3 Numerous chargedspecies have been used as building blocks for LbL films, such aspolyelectrolytes,4-6 biomacromolecules,7,8 nanoparticles,9 andcolloids,10,11 providing functional coatings for implanted medicaldevices. Recently, micelles with charged coronas were involved inthe development of local drug delivery systems.12-16 Sun et al.described the fabrication of LbL multilayer films based onpolyelectrolyte stabilized surfactant micelles as carriers for non-charged hydrophobic dyes.12 Hammond and co-workers inte-grated linear-dendritic block copolymermicelles encapsulating ahydrophobic antibacterial drug into LbL films.13 Furthermore,Rubner and co-workers suggested that the interpenetration of the

weak polyelectrolytes in the alternating layers is an importantissue regarding the surface property of the polyelectrolyte multi-layers.17 A significant number of the chain segments of thepreviously adsorbed layer penetrate into the outermost surfacelayer. The interpenetration of the biomacromolecules then pro-vides the possibility of constructing amultilayered coating surfacewith synergic properties of different biomacromolecule compo-nents.18,19 We previously have proven that the multilayered thinfilms constructed by alternating deposition of heparin and chito-san onto aminolyzed poly(ethylene terephthalate) (PET) filmsexhibit strong anticoagulant and antibacterial activities.18 How-ever, the poor loading capacity of hydrophobic drugs within thesetraditional polyelectrolyte multilayer films is a real drawback totheir potential applications.

Hyperbranched polymers have recently received much atten-tion as one class of spherical compounds due to their easypreparation, versatility, and the capability of incorporating highloadings of different types of molecules within their imperfectlybranched structures.20-22 Furthermore, the great number ofexternal groups of hyperbranched polymers provides manypossibilities to be either functionalized or multifunctionalizedfor potential applications in biomedicine, pharmacology, andbiotechnology.23-26 Most recently, we have synthesized a heparin-like sulfonated hyperbranched polyether (HBPO-SO3) consisting

*Towhom correspondence should be addressed. E-mail: jijian@zju.edu.cn.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.

(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.

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Hu and Ji Article

of a hydrophobic hyperbranched poly(3-ethyl-3-oxetanemethanol)(HBPO) core and negatively charged sulfonic acid terminalgroups.27 The HBPO-SO3 could self-assemble in aqueous mediato form stable micelles and displayed good hemocompatibilityand low cytotoxicity. We hypothesize here that the negativelycharged HBPO-SO3 micelles can be used as a building block forLbL assembly, and the hydrophobic cores of the micelles willintroduce hydrophobic nanodomains intomultilayer films, whichcan be regarded as nanoreservoirs for hydrophobic guest mole-cules. The layer-by-layer assembly of sulfonated hyperbranchedpolyether and chitosan will then provide the new possibility todevelop a multifunctional coating capable of anticoagulation andantibacterial and local drug delivery.

Experimental Section

Materials. Chitosan (average MW ca. 410 000, 91% deace-tylation) was obtained from Qingdaos Haihui Corporation ofChina. Heparin (sodium salt) was purchased from ShanghaisChemical Reagent Company of China. Polyethylenimine (PEI,average Mw ca. 25000 (LS)) and pyrene were purchased fromAldrich Chemical Co. PET membranes cut into squares 3 cm �3 cm were cleaned by sonication in acetone, methanol, and purewater for 10 min, respectively, followed by rinsing with purewater. Quartz slides (1 cm� 2 cm) and siliconwafers were cleanedby treatment in hot piranha solution (H2O2/H2SO4 3:7 v/v) for40 min (caution: piranha solution is extremely corrosive) and thenthoroughly washed with pure water.

Synthesis of HBPO-SO3. The synthesis and completecharacterization of the sulfonated hyperbranched polyetherHBPO-SO3 have been previously published.27 Briefly, the mix-tures of HBPO and excess sodium hydride in tetrahydrofuran(THF) reacted under reflux overnight with stirring before adding1,3-propane sultone, and then allowed to react for 12 h moreunder reflux. The resulting solution was filtered; then, the crudeproduct was exhaustively dialyzed against purewater, and awhiteproduct HBPO-SO3 was obtained. The sulfonation degree wasestimated to be around 45%.

Preparation of Multilayer Films. The LbL films wereassembled on PET membranes for plasma recalcification timeexperiments, silicon wafers for ellipsometry and atomic forcemicroscopy (AFM) measurements, and quartz slides for UV-visand fluorescence emission spectra, respectively. The substrate wasfirst immersed in PEI solution for 30 min to ensure a uniformpositively charged coating so that the effects of the substrate onthe layer growth areminimized. After rinsing with pure water anddrying under a nitrogen stream, the resulting substrate wasalternately immersed into solutions of HBPO-SO3 and chitosanfor 10 min each. Between each deposition step, the substrate wasrinsed 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 basedon heparin and chitosan were fabricated as the control. Allpolymers, PEI, HBPO-SO3, chitosan, and heparin, were of thesame concentration, 1 mg/mL in sodium acetate and acetic acidbuffer (HAc buffer, pH=4, 0.1 M).

Loading andRelease of Pyrene.Twodifferentmethodswereused to load pyrene into HBPO-SO3/chitosan multilayer films.The first approach required the encapsulation of pyrene inHBPO-SO3 micelles before the LbL assembly, while the secondinvolved the post-diffusion of pyrenemolecules from the solutioninto the preassembled multilayer films.

Pre-EncapsulationApproach.TenmilligramsofHBPO-SO3

was first dissolved in HAc buffer (10 mL) to form micelles; then,pyrene inTHF solution (1mg/mL, 2mL)was added.The solutioncontaining pyrene was vigorously stirred in the dark at roomtemperature. After THF was totally removed using a rotative

evaporator, the solution was filtered through a 0.45 μm mem-brane to eliminate the precipitated pyrene and was then ready forLbL deposition.

Post-Diffusion Method. A quartz slide deposited with theLbL films (10 bilayers) was exposed to the chloroform solution ofpyrene (2 mg/mL). The quartz slide was taken out after 24 h,rinsed three times with chloroform to remove the dye moleculesweakly adsorbed on the surface, and dried in vacuo.

The quartz slide covered with pyrene-loaded multilayer filmswas immersed into a vial of phosphate buffered saline (PBS, pH=7.4, 0.1 M) at 37 �C, which was replaced by fresh solution atappropriate time point to ensure constant release conditions. Theconcentrationof pyrene in the PBS solutionwas analyzedwith theUV-vis spectrometer .

Loading and Release of Probucal. Probucal, which haspowerful antioxidant properties and can prevent restenosis aftercoronary angioplasty,28 was incorporated into HBPO-SO3/chitosan and heparin/chitosan multilayer films by the post-diffu-sion approach. Probucal was previously dissolved in chloroformor dimethyl sulfoxide (DMSO) at 1 mg/mL. Quartz slide deposi-ted with the multilayer films (10 bilayers) was exposed to theProbucal solution for 24 h, then rinsed three times in the pureorganic solvent and dried in vacuo. The amount of Probucalloaded inmultilayer filmswas analyzedby themeasurement of thecharacteristic absorbance of Probucal at 240 nm.

The quartz slide coveredwith Probucal-loadedmultilayer filmswas immersed into a vial of phosphate buffered saline (PBS, pH=7.4, 0.1 M) at 37 �C. At regular intervals, the quartz slide wasremoved from the solution, rinsed with pure water, dried undernitrogen flow followedbyUV-vis absorption, and thenmoved toa 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 allo-wed to stand for 1 min at 37 �C before the addition of 0.025 MCaCl2 solution (50 μL), at which point a stopwatch was star-ted. The stopwatch was stopped when fibrin clotting wasfirst visible, and the time was recorded. At least six experimentswere carried out for each sample and the mean clotting timereported.

Characterization Techniques. Quartz crystal microbalance(QCM)measurements were taken with a Q-Sense QCM-E4 usingAu-coated resonator (HongrongBomanBiotechnologyCo. Ltd.,Beijing, China). The fundamental resonant frequency of thecrystal was 5 MHz. The crystal was mounted in a fluid cell withone side exposed to the solution. A measurement of LbL deposi-tionwas initiated by switching the liquid exposed to the resonatorfrom HAc buffer to a PEI solution. PEI was allowed to adsorbonto the resonator surface for 15 min before being rinsedwith buffer. Then, HBPO-SO3 (with or without pyrene en-capsulation) and chitosan solutions were alternately introducedfor 15 min with buffer rinsing in between. All the polymers, PEI,HBPO-SO3 (with or without pyrene encapsulation), and chit-osan, were of the same concentration, 1 mg/mL in HAc buffer(pH = 4, 0.1 M) and injected into the chamber at a rate of0.1 mL/min.

Spectroscopic ellipsometry was carried out using a M-2000(J. A. Wollam Co. Inc.) to measure the film thickness on siliconwafers. AFM images were performed in the tapping mode underambient conditions using a commercial scanning probe micro-scope, Seiko SPI3800N, equipped with a silicon cantilever, Na-nosensors, typical spring constant 40 N 3m

-1. Fluorescencespectra were recorded on a spectrofluorometer (FP-770, JapanSpectroscopic) at room temperature. Emission spectra wererecorded over the range 350-500 nm with an excitation wave-length of 336 nm. UV-vis spectra were obtained on a Shimadzumodel UV-2550 spectrometer.

(27) Hu, X. F.; Ji, J. Acta Polym. Sin. 2009, 8, 828.(28) Tardif, J. C.; Cot�e, G.; Lesp�erance, J.; Bourassa, M.; Lambert, J.; Doucet,

S.; Bilodeau, L.; Nattel, S.; deGuise, P. N. Engl. J. Med. 1997, 337, 365.

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Results and Discussion

Fabrication and Characterization of the LBL Films of

HBPO-SO3 and Chitosan. The multilayer films were formedthrough an alternating deposition of negatively charged HBPO-SO3 and positively charged chitosan on PEI adsorbed substrates.HBPO-SO3 used here consists of a hydrophobic hyperbranchedpoly(3-ethyl-3-oxetanemethanol) (HBPO) core and negativelycharged sulfonic acid terminal groups (Figure 1). As shown inour previous paper, when the concentration of HBPO-SO3

increased above 0.017 mg/mL, these heparin-like hyperbranchedpolyethers could self-assemble in aqueous solution to form sphe-rical micelles with negatively charged sulfonic acid groups on theexterior, and exhibited good hemocompatibility and low cyto-toxicity.27 The aggregation of HBPO-SO3 in the HAc buffersolution prepared for LbL assembly was investigated by TEMand the laser particle size analyzing system (Figure 2). HBPO-SO3 self-assembled to formmicelleswith amean diameter of 142 nm.

QCM measurement was employed to confirm the successfulLbL assembly, and a linear film growth was observed (Figure 3).The LbL assembly on silicon wafer was also monitored withellipsometry. Figure 4 indicated a similar LbL growth curve withlinear growth.On average, a bilayer was approximately 3 nm. TheAFM images demonstrated the stability of HBPO-SO3 micellesduring the LbL process and the presence of micelles in LbL films.As indicated in Figure 5, spherical micelles occurred on thesurface of PEI adsorbed silicon wafer after the deposition ofHBPO-SO3, and the diameter was consistent with that measuredin solution.However, the results from ellipsometrymeasurementsrevealed that the increase in thickness due to the deposition ofHBPO-SO3 micelles was limited and the thickness of (HBPO-SO3/chitosan)10 multilayer films was much less than the diameterof the micelles. This was probably because the imperfectlybranched polyether skeleton of this designed polymer was flexibleand the terminal sulfonate groups made HBPO-SO3 a strongpolyanion. For these reasons, the HBPO-SO3 micelles might becrushed under the electrostatic interaction with the positivelycharged surface, while the shrinkage of the aggregations occurredin the deposition, and compact multilayer thin films were fabri-cated as shown in Figure 6.

Drug Loading by Pre-Encapsulation and Post-Diffusion

inMultilayer Films. The hydrophobic dye pyrene was used as amodel drug and incorporated into the LbL films by pre-encapsu-lation inHBPO-SO3micelles and post-diffusion in preassembledmultilayer films, respectively.

Figure 1. Chemical structure of HBPO-SO3.

Figure 2. (a) TEM image of HBPO-SO3 micelles in HAc buffersolution at 1mg/mL. (b) Results of the size distribution by particlesize distribution analyzer.

Figure 3. QCM frequency decrease (-ΔF) for the alternatingdeposition of HBPO-SO3 micelles (with or without pyrene) andchitosan. The LbL assembly was initiated with the adsorption ofPEI on resonators. The odd and even layer numbers correspond tothe deposition of HBPO-SO3 micelles and chitosan, respectively.

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The LbL assembly of pyrene-loaded HBPO-SO3 micelles andchitosan was carried out, and continuous film growth wasmonitoredwithQCMand ellipsometry.Unlike the LbL assemblyof HBPO-SO3 micelles and chitosan mentioned above, themultilayer films based on pyrene-loaded HBPO-SO3 micellesshowed a nonlinear growth pattern (Figures 3 and 4). Similarresults have been reported previously in the multilayer of poly-electrolyte stabilized pyrene-loaded micelles due to the increasedsurface roughness.12 To ensure that pyrene remained in theHBPO-SO3 micelles during the LbL assembly process, fluores-cence emissions of pyrene after the deposition of each layer ofmicelles were recorded (Figure 7). The fluorescence intensity ofpyrene in the multilayer films increased with respect to theincrease of the number of layers of pyrene-loaded micelles. Onthe basis of the aforementioned results, pyrene could be incorpo-rated into the LbL films via pre-encapsulation in the HBPO-SO3

micelles, and pyrene-loaded HBPO-SO3 micelles did not induce

a significant perturbation of the LbL deposition with chitosan.The amount of pyrene loaded in the multilayer films can becontrolled by simply changing the cycles of films deposited.

It is well-known that the fluorescence spectrum of pyrene issensitive to the polarity of the surrounding environment.29 Thevibronic band intensities of the fluorescence emission spectraprovide a newpossibility for elucidating the location of the pyrenewithin the multilayer. The fluorescence intensity ratio of the twopeaks at 383 and 373 nm (I3/I1) reflects the change of environ-mental polarity.29 The I3/I1 ratio of pyrene in an aqueousHBPO-SO3 micelle solution was 1.19, which indicated that thepyrene molecules were incorporated into the hydrophobic coresof HBPO-SO3 micelles. The I3/I1 ratios of the pyrene within themultilayer by pre-encapsulation in HBPO-SO3 micelles andpost-diffusion in the preassembled multilayer of HBPO-SO3/chitosan were 0.94 and 0.91, respectively, while the fluorescenceintensity of pyrene within the multilayer films based on heparinand chitosan was too weak to calculate the I3/I1 ratio. Althoughthe deformation of the hyperbranched polyether micelles drivenby the electrostatic interaction with chitosan might decrease thehydrophobicity of the core, most hydrophobic pyrene moleculeswere still incorporated into the hydrophobic HBPO cores. TheLBL films of HBPO-SO3 and chitosan offer nanoreservoirs forhydrophobic guest molecules and provide a new possibility ofincorporating a hydrophobic drug.

The UV-vis absorption spectra of pyrene-loaded multilayerfilms were presented in Figure 8a. Although the peaks wereslightly 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, morepyrene loading was achieved via the post-diffusion process.

Figure 4. Thicknesses of assembled films at various number of bi-layers. The LbL assembly was initiated with the adsorption of PEIon silicon substrates.

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 negativelychargedHBPO-SO3micelles and positively charged chitosan on aPEI adsorbed substrate.

Figure 7. (a) Fluorescence spectra of a 10-bilayer film of pyrene-loaded HBPO-SO3 micelles and chitosan. (b) Changes in thenormalized fluorescence intensity of pyrene at 393 nmas a functionof the number of micelle layers.

(29) Kalyanasundaram, K.; Thomas, J. J. Am. Chem. Soc. 1977, 99, 2039.

2628 DOI: 10.1021/la902719k Langmuir 2010, 26(4), 2624–2629

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UV-vis spectrometry was also employed to quantify the releaseof pyrene from themultilayer films. The pyrene-loaded LbL filmsprepared via either the pre-encapsulation approach or the post-diffusion process showed continuous release over a period ofseveral weeks. In the case of the pre-encapsulation approach, aburst 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 daysand a sustainable continued release of pyrene (Figure 8b). Furtherinspection of Figure 8b revealed that the release of pyreneincorporated via the post-diffusion method went through threestages: with an initial slow release in the first 30 h, subsequently amore rapid release of about 0.47 μg per day, and then a sustain-able slow release after 5 days. The absence of the usual burst effectindicated that the pyrene loadings in multilayer films were belowthe saturation concentration.30 The post-diffusion process wasmore efficient at incorporating hydrophobic guest molecules intothe LbL films and carried out amuchmore controllable release ofthe 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

Probucal uptake by HBPO-SO3/chitosan multilayer films, tak-ing heparin/chitosanmultilayer films as the control. The thicknessof preassembledHBPO-SO3/chitosanmultilayer filmswas of thesame order of magnitude as the heparin/chitosan multilayer filmsformedunder the same conditions (10 bilayers, 34.9 nmvs 31.3 nm).As indicated in Figure 9, the HBPO-SO3/chitosan multilayerfilms showed the capability of loading after immersion in thechloroform solution of Probucal, while there was almost no Pro-bucal within the heparin/chitosan multilayer films (Figure 9).Successful loading of Probucal in bothHBPO-SO3/chitosan andheparin/chitosan multilayer films was obtained using DMSO asthe solvent. However, Probucal-loaded heparin/chitosan multi-layer films obtained by the post-diffusion of DMSO solution hada burst release within the first 10 h; approximately 66% of thedrug was released, and the Probucal remaining in the multilayerfilms was hardly detected (Figure 10). The result revealed that thedrugs might be entrapped within the surface of the heparin/chitosan multilayer. As good solvents for the hydrophobic poly-ether cores of HBPO-SO3 micelles, chloroform and DMSOcould drive diffusion of Probucal into hydrophobic nanodomainsin the HBPO-SO3/chitosan multilayer films. The Probucal-loaded HBPO-SO3/chitosan multilayer films via the post-diffu-sion process showed a continuous release over a period of severalweeks. A burst release of 20% within the first 10 h can be shown,reaching a cumulative release of about 38% after 14 days, and asustainable release continued for more than three weeks.

The high concentration of sulfate and sulfamate groups ofheparin has been proven to play an important role in the anti-coagulant activity of heparin.31 A number of polymers containingsulfonate groups, so-called heparin-like materials, have been de-signed to exhibit more or less enhanced blood compatibility.32,33

As an alternative to heparin, HBPO-SO3 should provide themultilayer films with anticoagulant capability. The PRT assaywas performed to determine the blood compatibility of HBPO-SO3/chitosan multilayer films. Compared with the bare PET,

Figure 8. (a) UV-vis spectra of pyrene in CH3OH solution(dash), blank LbL films (dot), pyrene-loaded LbL films via thepost-diffusion method (black solid) and the pre-encapsulationapproach (gray solid), respectively. All the LbL films were fabri-cated with ten deposition cycles. The spectrum of pyrene in dilutesolution was modified by raising the baseline, for clarity in com-parison. (b) Release profile of pyrene from the multilayer films.

Figure 9. UV-vis spectra of (a) HBPO-SO3/chitosan multilayerfilms without loading; heparin/chitosan multilayer films afterimmersion in (b) chloroform or (c) DMSO solution of Probucal;HBPO-SO3/chitosanmultilayer filmsafter immersion in (d)DMSOor (e) chloroform solution of Probucal; and (f) Probucal in dilutesolution. All the LbL films were fabricated with ten depositioncycles.

(30) Guyomard, A.; Nysten, B.; Muller, G.; Glinel, K. Langmuir 2006, 22, 2281.

(31) Tamada, Y.; Murata, M.; Hayashi, T.; Goto, K. Biomaterials 2002, 23,1375.

(32) Aguilar, M. R.; Rodrıguez, G.; Fern�andez, M.; Gallardo, A.; Rom�an, 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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Hu and Ji Article

which has been widely used in cardiovascular implants due to itsexcellent mechanical properties and moderate biocompatibility,the PRT of HBPO-SO3/chitosan multilayer films deposited onPET membranes was obviously prolonged (Figure 11). Whilecoagulation occurred on the bare PET after 383 ( 80 s, it is not

surprised to find that the coagulation of HBPO-SO3-terminatedmultilayer films was not observed until 1974( 11 s. However, it isinteresting that the multilayer films with chitosan as the outer-most layer also presented a PRT as long as 1670( 121 s, althoughthe chitosan is procoagulated. The results are consistent with ourprevious heparin/chitosan multilayer films, in which the multi-layer films with chitosan as the outermost layer exhibited stronganticoagulant and antibacterial activities due to interpenetrationof the polyelectrolytes in the alternating layers.18 The results repor-ted here implied that the same interpenetration of layers mightexist in the multilayer films of sulfonated hyperbranched poly-ether and chitosan. Amultifunctional coating with potential anti-coagulation, antibacterial, and local release of anti-restenosis drugscan be developed via post-diffusion of the hydrophobic drugProbucal into the multilayer films of HBPO-SO3 and chitosan.

Conclusion

Sulfonated hyperbranched polyether (HBPO-SO3) was emp-loyed as a building block to fabricate multilayer films withchitosan via LbL assembly. The QCM and ellipsometry resultsverified the progressive growth of films. Results from the loadingand release experiments demonstrated that the hydrophobic guestmolecules could be incorporated into the HBPO-SO3/chitosanmultilayer films by either the pre-encapsulation process or thepost-diffusion approach, and released in a controlled and sustain-ableway. In addition, themultilayer films exhibited anticoagulantactivity even with chitosan as the outermost layer. These antic-oagulant HBPO-SO3/chitosan multilayer films with anticipatedantibacterial activity and excellent capability for loading andcontrolled release of hydrophobic drugs, such as antiproliferativeagents, may have good potential for surface modification ofimplanted medical devices.

Acknowledgment.Financial support from theNatural ScienceFoundationofChina (NSFC- 20774082, 50830106), 863NationalHigh-Tech R&D Program (2006AA03Z329, 2006AA03Z444),Ph.D. Programs Foundation of Ministry of Education of China(No. 20070335024), Open Project of State Key Laboratory ofSupramolecular Structure and Materials (SKLSSM200911) andNatural Science Foundation of China of Zhejiang Province-(Y4080250) is gratefully acknowledged. The authors thank Prof.Deyue Yan and Yongfeng Zhou at Shanghai Jiao Tong Uni-versity for beneficial discussions and the offer of hyperbranchedpoly(3-ethyl-3-oxetanemethanol) (HBPO).

Figure 10. Drug release profile of Probucal incorporated inHBPO-SO3/chitosan multilayer films (solid icons) and heparin/chitosan (hollow icons) multilayer films by the post-diffusion of aDMSO 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).