Article pubs.acs.org/Langmuir
A Novel Approach for Synthesis of Zwitterionic Polyurethane Coating with Protein Resistance Chunhua Wang,† Chunfeng Ma,*,§ Changdao Mu,‡ and Wei Lin*,† †
Department of Biomass and Leather Engineering, Key Laboratory of Leather Chemistry and Engineering of Ministry of Education and ‡Department of Pharmaceutics and Bioengineering, School of Chemical Engineering, Sichuan University, Chengdu, China 610065 § Faculty of Material Science and Engineering, South China University of Technology, Guangzhou, China 510640 ABSTRACT: We have developed a novel approach to introduce zwitterions into polyurethane for the preparation of antibiofouling coating. First, the thiol−ene click reaction between 2-(dimethylamino)ethyl methacrylate (DMAEMA) and 3-mercapto-1,2-propanediol (TPG) is used to synthesize dihydroxy-terminated DMAEMA (DMA(OH)2) under UV catalysis. The product has been proved by gel permeation chromatography (GPC), Fourier transform infrared spectrum (FT-IR), proton nuclear magnetic resonance (1H NMR), and high resolution mass spectrometry (HRMS). DMA(OH)2 is then incorporated into polyurethane as side groups by polyaddition with diisocyanate and further reacts with 1,3-propane sultone to obtain the zwitterionic polyurethanes. The presence of sulfobetaine zwitterions side groups has been demonstrated by FT-IR and X-ray photoelectron spectroscopy (XPS). Thermal analysis indicates that the thermal stability is decreased with the increasing content of zwitterionions. The antibiofouling property of polyurethanes has been investigated by the measurement of adsorption of fibrinogen, bovine serum albumin (BSA), and lysozyme on the polyurethanes surface using quartz crystal microbalance with dissipation (QCM-D). The results show that the polyurethane coatings exhibit effective nonspecific protein resistance at higher content of zwitterionic side groups.
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INTRODUCTION Biofouling is of great concern in numerous applications ranging from biomedical materials1 to biological-related processes2 and from food packaging,3 to leather finishing,4 to industrial and marine equipment.5,6 For preparing antibiofouling coatings, one of the key strategies is to design the surface that can resist the nonspecific adsorption of protein and microbes. 7 Functionalization of surfaces with oligo(ethylene glycol) or poly(ethylene glycol) (PEG) is most commonly used to confer a surface protein resistance. Several PEG-based coatings showing good antifouling properties have been reported in the literature.8,9 However, PEG is susceptible to oxidation catalyzed by transition metals or chain cleavage after long-term exposure to biochemically relevant solutions.10,11 Moreover, the antiprotein property of grafted PEG brushes on membrane surface was affected by temperature.12 Zwitterionic polymers are promising alternatives to conventional PEG-functionalized surfaces because of their reasonably good biocompatibility and resistance to nonspecific protein adsorption.13 The latter is related to the hydration of polymers and strong binding with water molecules via electrostatic interactions. Therefore, increasing attention has been attracted on incorporating of zwitterion groups onto polymer surfaces to enhance the antifouling property. Polyurethane is one of the most favorable matrixes because it exhibits wide ranges of physical and chemical properties and biocompatibility and covers various high value applications as a coating material. In © 2014 American Chemical Society
leather industry, waterborne polyurethane is largely used as finishing of skins to improve surface appearance and resistance to abrasion and light.14 It has been reported that polyurethanes containing zwitterionic phosphorylcholine (PC) exhibit good antifouling property,15,16 but the phosphoester group has the tendency of hydrolysis owing to its susceptibility to moisture.17 And the modification of polyurethane with zitterionic carboxybetaine (CB) has conferred enhanced blood compatibility and the resistance to bacterial colonization.18,19 However, the responsive protein adsorption cannot be avoided due to the protonation/deprotonation properties of carboxybetaine.20 So far, grafting zwitterionic sulfobetaine (SB) onto polyurethane surface21,22 also shows good protein resistance, which is affected by the grafting density and the chains length. Nevertheless, the strategy of surface grafting is not applicable for industrial processing. Besides, interpenetrating polymer networks (IPNs) of polyurethane and SB polymer were reported, and the difficult control of protein resistance was related to the distribution and diffusion of SB in the polyurethane network.23 Recently, we have incorporated zwitterionic SB polymer into polyurethane as the side chains, which can effectively resist nonspecific protein adsorption.24 The key is to solve the Received: August 27, 2014 Revised: October 12, 2014 Published: October 13, 2014 12860
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Article
compatibility between highly polar zwitterions and the relatively hydrophobic polyurethane backbone during the synthesis process since they do not have the same solvent. Considering the length of zwitterionic side chains was relatively long, which may cause high water absorption and not suitable for leather finishing or coatings, herein we have prepared a welldefined short side chain or side group, dihydroxy-terminated 2(dimethylamino)ethyl methacrylate (DMA(OH)2), by thiol− ene click reaction.25,26 The DMA(OH)2 can be readily introduced into polyurethane as a chain extender and further react with 1,3-propane sultone to yield zwitterions. Our aim is to develop a novel and easy approach for the preparation of polyurethanes with good antibiofouling and coating properties.
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Table 2. Effect of Reaction Conditions on the Products Resulted from DMAEMA and TPG molar ratio of TPG/DMAEMA
EXPERIMENTAL SECTION
Mw (kDa)
isoelectric point (IEP)
net charge in PBS (0.14 M, pH 7.4)
340
5.5
negative
BSA
68
4.8
negative
lysozyme
14.7
11.1
positive
fibrinogen
Mn
PDI
product color
0.5 h UV, DMPA
2
576
1.02
yellow
1:1
2.0 h UV, DMPA
2
306 434
1.02 1.11
yellow
1.2:1
0.5 h UV, DMPA
2
169 414
1.02 1.07
yellow
2:1
0.5 h UV, DMPA 2 h UV, DMPA
1
211 277
1.02 1.08
colorless
1
266
1.1
colorless
TPG (10 mmol) and 1.89 g of DMAEMA (12 mmol) were introduced into a glass weighing bottle, after which 15 mL of tetrahydrofuran (THF) was added. Then 0.0297 g of DMPA (1 wt % based on the total mass of monomers) was poured into each flask, and then the solution was degassed with nitrogen for 20 min to replace the air. The samples were incubated by irradiation with a UV-lamp (emitting nominally at 350 nm, light intensity 100%, 100 mW/cm2) at room temperature for 2 h. The product was purified by precipitation into hexane and butyl acetate in succession for two times, filtered, and then dried under vacuum at 40 °C for 24 h. Synthesis of Polyurethane with DMAEMA Side Groups (NPU). NPU was synthesized via a condensation reaction in THF under a nitrogen atmosphere (Scheme 2). IPDI reacted with PTMG at 70 °C for 1 h, yielding a low-Mw prepolymer. Then DMA(OH)2 was introduced as a chain extender, and meanwhile DBTDL was added as the catalyst. The reaction was conducted at 80 °C for 1 h. Then another chain extender 1,4-BD was introduced, and the mixture was further reacted at 80 °C for 3 h. The synthesized polyurethane was precipitated in pure water and vacuum-dried for 24 h. NPU with varying contents of DMA(OH)2 and the GPC results are shown in Table 2. The polyurethane with DMAEMA side groups is designated as NPU-x, where x represents the weight percentage of DMA(OH)2 calculated according to the formula. Meanwhile, PU without DMA(OH)2 was also synthesized from IPDI, PTMG, and 1,4-BD as reference. Ionization or Betainization of NPU. 1 g of NPU and 20 mol % excess of 1,3-propane sultone (calculated based on DMA(OH)2 content in PU matrix) were dissolved in THF. The ionization reaction was conducted at 25 °C for 24 h under constant stirring. As the increasing content of DMAEMA in NPU, the resulting solutions became opaque in THF due to more sulfonate (−SO3) groups introduced. THF was removed by rotary evaporation, and the prepared ionized polyurethane (iNPU) was precipitated in the deionized water, filtered, and vacuum-dried for 24 h. Characterization. Fourier Transform Infrared Spectroscopy (FTIR). FT-IR measurements were conducted on a Bruker VECTOR-22 IR spectrometer using the KBr disk method. The spectra were obtained at 64 scans with the resolution of 4 cm−1 ranging from 400 to 4000 cm−1.
Table 1. Characteristics Data of the Used Proteins27 protein
peak no.
1:1
2:1
Materials. Poly(tetramethylene oxide) (PTMG) (Mw = 2000 g/ mol), 2-(dimethylamino)ethyl methacrylate (DMAEMA), 1,3-propane sulton (1,3-PS), 3-isocyanatomethyl-3,5,5-trimethylcyclohexyl isocyanate (IPDI), and 3-mercapto-1,2-propanediol (TPG) were purchased from Aladdin (Shanghai). 1,4-Butanediol (1,4-BD) and dibutyltindilaurate (DBTDL) were from Aldrich. DMAEMA and 1,4-BD were purified under a reduced pressure before use. PTMG was dried under a reduced pressure for 2 h before use. Tetrahydrofuran (THF) was refluxed over CaH2 and distilled. Fibrinogen (fraction I from human plasma) from Merck Chemicals and lysozyme, via chicken egg white from Sangon (Shanghai), and BSA from Hualvyuan Biotechnology Company were used as received. The characteristics data of three proteins are shown in Table 1. Certain amounts of NaCl, KCl,
condition
property fibrous, hard protein global, hard protein global, soft protein
Na2HPO4, and KH2PO4 were dissolved in Milli-Q water to obtain the physiological phosphate buffered saline (PBS, 0.14 M, pH 7.4). Each protein solution (1.0 mg/mL) in PBS buffer was prepared separately. Other reagents were used as received without any pretreatment. Synthesis of Chain Extender DMA(OH)2 via Thiol−Ene Click Reaction. Thiol−ene click reactions can be manipulated under a variety of initiating systems for varied products. Here, photoinitiating reaction has been utilized for DMA(OH)2 synthesis with UV catalysis in the presence of photoinitiator, 2,2-dimethoxy-2-phenyl acetophenone (DMPA) (Scheme 1). Thiol is accepted as a radical initiator to obtain telomere or monoadduct because it can provide a readily cleavable S−H bond by homolytic cleavage at moderate temperature. In order to make sure the reaction is nucleophilic rather than a free radical process, we adopted different conditions and different ratios of TPG and DMAEMA (shown in Table 2). In a typical experiment, 1.08 g of
Scheme 1. Thiol−Ene Click Reaction between TPG and DMAEMA
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Scheme 2. Synthesis of the Polyurethane with DMAEMA (NPU) and the Ionized NPU (iNPU)
nm was used to minimize the effects of surface roughness.29 The QCM-D technique was described in detail elsewhere.30 Briefly, it allows to simultaneous measurements of the changes in the frequency (Δf) and energy dissipation (ΔD) of an oscillating AT-cut quartz crystal in response to adsorption of material to the crystal surface. The mass of a thin layer on quartz crystal is related to the decrease in the resonant frequency of the crystal, whereas the dissipation factor is involved in the viscoelasticity of the absorbed layer. In our measurement, Δf and ΔD give the information about the protein adsorption and structural change of the protein layer. All the presented data are from the third overtone (n = 3). The polyurethane coating used for QCM measurement was prepared by spin-casting of polyurethane solution in THF (0.5 wt %) on a quartz crystal with a spin coater (CHEMAT, KW-4) at 4000 rmp in air. Then the coating was heated at 40 °C for 24 in a drying oven to remove THF before testing. All the adsorption experiments were carried out at 25 °C with PBS buffer as reference.
Proton Nuclear Magnetic Resonance Spectroscopy (1H NMR). All H NMR spectra were recorded on a Bruker AV400 NMR spectrometer. The sample was dissolved in CDCl3 or DMSO with conventional tetramethylsilane (TMS) as internal standard. Gel Permeation Chromatography (GPC). The weight- and number-average molecular weights (Mw and Mn) and corresponding polydispersity (PDI) (Mw/Mn) were determined by GPC on a Waters 1515 at 35 °C. A series of monodisperse polystyrenes were used as the standard and THF as the fluent with a flow rate of 1.0 mL/min. High-Resolution Mass Spectrometry (HRMS). HRMS was performed by using eletrospray ionization (ESI) and recorded on Bruker maxisimpact. Samples were first dissolved in THF and properly diluted by methanol before measurement. X-ray Potoelectron Spectroscopy (XPS). The analysis of C, N, O, S, and other elements on the slice surface of polyurethane coating was conducted on an X-ray photoelectron spectrometer (ESCALAB 250, Thermo-VG Scientific Corporation) with a monochromatic focused Al K X-ray source (1486.6 eV). High-resolution core-level spectra were collected by using the monochromatic Mg X-ray source (1253.6 eV). The detection was performed at 45° with respect to the sample surfaces and in the sample chamber pressure of 10−9 mbar. All spectra were charge corrected with the C1s peak at 284.7 eV. Each sample was detected at three different points on the slices to reduce the error. TGA Measurements. Thermogravimetric analysis (TGA) were performed on a SDT Q600 (TA Instruments) over the range from room temperature to 800 °C under a nitrogen atmosphere with the flow rate of 35 cm3/min. The heating rate is 10 °C/min. About 4 mg of specimen from each sample was taken for the measurement. QCM-D Measurement. The protein absorption on polyurethanes was measured by a quartz crystal microbalance with dissipation (E1, Q-sense AB, Sweden).28 The AT-cut quartz crystal sensor with a fundamental resonant frequency of 5 MHz (Q-sense AB) was mounted in a fluid cell with one side exposed to the solution. A highly polished crystals with a root-mean-square roughness less than 3 1
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RESULTS AND DISCUSSION Preparation and Characterization of Chain Extender DMA(OH)2. The thiol−ene reaction is generally initiated under radical conditions and then proceeds with free-radical propagation and chains termination. However, it is proved that aside from the radical mediated reactions, hydrothiolations can be readily accomplished by the click Michael addition of a thiol to an activated CC bond, e.g., the bond at methacrylate, with mild base or nucleophile as catalyst.25,31 Herein, we synthesize DMA(OH)2 from DMAEMA and TPG in the presence of DMPA as photoinitiator and in the combination with UV catalysis, since UV-initiated thiol−ene click reaction enables an unique spatial and temporal control in synthetic process.31 The influence of UV-irradiation time and TPG/ 12862
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DMAEMA ratio on the products was preliminarily identified by GPC determination, as shown in Table 2. It shows that low ratios (
