Antifouling Polycarbonate Coatings: Role of Block

Feb 9, 2015 - IBM Almaden Research Center, 650 Harry Road, San Jose, ... efficacy as antimicrobial and antifouling agents investigated on coated surfa...
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Antimicrobial/Antifouling Polycarbonate Coatings: Role of Block Copolymer Architecture Zhi Xiang Voo,† Majad Khan,† Karthikeyan Narayanan,† Desmond Seah,† James L. Hedrick,*,‡ and Yi Yan Yang*,† †

Institute of Bioengineering and Nanotechnology, 31 Biopolis Way, The Nanos, #04-01, Singapore 138669, Singapore IBM Almaden Research Center, 650 Harry Road, San Jose, California 95120, United States

‡

S Supporting Information *

ABSTRACT: The high prevalence of catheter-associated infections accounts for more than 3 billion dollars annually in hospitals, and antimicrobial polymer coatings on catheter surface may serve as an attractive weapon to mitigate infections. Triblock polycarbonate polymers consisting of three critical components including antifouling poly(ethylene glycol) (PEG), antimicrobial cationic polycarbonate, and a tethering or adhesive functional block were synthesized. In this study, the block topology or placement of the distinctive blocks was varied and their efficacy as antimicrobial and antifouling agents investigated on coated surfaces. The individual blocks were designed to have comparable lengths that were subsequently grafted onto a prefunctionalized catheter surface through covalent bonding under mild conditions. The anchoring/adhesive functional moiety based on a maleimide functional carbonate was positioned at either the center or end of the polymer block and subsequently tethered to the surface via Michael addition chemistry. The placement of the adhesive block was investigated in terms of its effect on antimicrobial and antifouling properties. The surface coated with the polymer containing the center-positioned tethering block (2.4k-V) was unable to prevent bacteria fouling, even though demonstrated higher bacteria killing efficacy in solution as compared to the surface coated with the polymer containing the end-positioned tethering block (2.4k-S). In contrast, the 2.4k-S coating resisted fouling of both Gram-positive S. aureus and Gram-negative E. coli effectively under conditions that simulate the device lifetime (1 week). Moreover, the coating prevented protein fouling and platelet adhesion without inducing significant hemolysis. Consequently, this antibacterial and antifouling polymer coating is an interesting candidate to prevent catheter-associated bloodstream infections. results in the formation of biofilm on the surface.11 The biofilm increases bacteria survivability and tolerance to antibiotics by manyfold.18,19 Moreover, removing the biofilm-infected devices may not solve the problem completely due to residual microbes that cause recurring infections.19,20 Several strategies have been devised to prevent biofilm formation. Some of these strategies employ antibiotics,16,21 silver ions,22 or quaternized ammonium ions23,24 in the medical device. However, these strategies also suffer from burst release,25 drug resistance,26,27 and increase in biofilm formation.28 Another technique utilizes antifouling agents, such as zwitterions9 or hydrophilic poly(ethylene glycol).29 These methods may prevent the microbes from attaching to the surface for a certain period of time without killing the microbes, eventually leading to fouling. Therefore, there is an urgent need to develop novel methods and materials that possess robust antibacterial and antifouling properties in a sustained manner that simulate the typical lifetime of the targeted device.

1. INTRODUCTION Silicone-based elastomers are ubiquitous materials for many different devices, such as stents,1 catheters,2 prostheses,3 contact lenses,4 and microfluidics.5 They possess a low transition temperature, and their hydrophobicity makes the materials inert to intravenous applications and body fluids.6,7 Silicone rubber is also nontoxic and has both thermal and chemical stability; hence, it is an attractive material for biomedical applications.8 However, it is prone to protein adsorption due to its hydrophobic nature, and protein fouling can occur in a matter of seconds after implantation and exposure to body fluids,9 resulting in blood clots and subsequent thrombosis.10 Once proteins form the topmost layer on the silicone surface, microbes such as bacteria and fungi can easily anchor onto the surface, generating a biofilm.11 As such, catheter-associated nosocomial infections, which account for most hospital-related infections, are expensive to treat and amount for more than 3 billion dollars annually in the U.S.A. alone.12 Among various types of bacteria, Staphylococcus aureus13−15 and Escherichia coli1,16,17 are common bacteria found to foul the silicone surface via nonspecific and specific adhesion.18 Eventually, bacterial cell proliferation and adhesion © XXXX American Chemical Society

Received: November 6, 2014 Revised: January 19, 2015

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DOI: 10.1021/ma5022488 Macromolecules XXXX, XXX, XXX−XXX

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Figure 1. Polymer synthesis and coating: (a) synthesis of triblock copolymers of PEG, cationic, and maleimide-functionalized polycarbonates with different structures (2.4k-V, 2.4k-Vshort, and 2.4k-S); (b) polymer coating process.

physical dip coating technique.35 To improve the durability and efficacy of antibacterial and antifouling coatings, we rationally designed triblock copolymers having three critical components including (1) antifouling mPEG, (2) antibacterial polycarbonate containing quaternary ammonium, and (3) polycarbonate containing maleimide groups that were used as anchorage points and chemically coated these polymers onto thiolfunctionalized silicone surface via Michael addition reaction. The triblock copolymers were synthesized using monomethyl ether PEG (mPEG) with 2.4 kDa as a macroinitiator to ringopen the cyclic carbonate monomers MTC-furan protected maleimide (MTC-FPM) and MTC-benzyl chloride (MTC-

Recently, we demonstrated the synthesis of well-defined antimicrobial polycarbonates via organocatalytic metal-free ring-opening polymerization (ROP).30−32 The antimicrobial polycarbonates effectively eradicated multidrug-resistant microbes via a membrane-lytic mechanism with minimal toxicity. Poly(carbonate-b-ethylene oxide) copolymers having a thiol functional group on the ethylene oxide component were coated and reacted onto the silicone rubber through the reaction of the thiol moiety with a polydopamine adhesive layer via Michael addition, which was physically deposited onto the silicone rubber prior to polymer coating.33,34 Subsequent study simplified the coating process from a two-step to a single B

DOI: 10.1021/ma5022488 Macromolecules XXXX, XXX, XXX−XXX

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Macromolecules Table 1. Compositions of the Triblock Copolymers 2.4k-V and 2.4k-Sa polymer

feed molar ratio (PEG:MTC-FPM:MTC-OCH2BnCl) with TU/DBU 5 mol %

2.4k-V

1:10:140

2.4k-Vshort

1:10:35

2.4k-S

1:10:140

compositionb PEG-(MTC-FPM)7-(MTCOCH2BnCl)100 PEG-(MTC-FPM)7-(MTCOCH2BnCl)23 PEG-(MTC-OCH2BnCl)80(MTC-FPM)3

a c

composition after quaternizationb

Mwc (PDI: before and after deprotection)

PEG-(MTC-M)5-(MTCOCH2BnCl)90 PEG-(MTC-M)6-(MTCOCH2BnCl)22 PEG-(MTC-OCH2BnCl)80(MTC-M)3

35 kDa (1.23/1.22)

MTC-FPM: furan-protected maleimide monomers; MTC-M: deprotected maleimide monomers. Determined using by GPC using polystyrene standards.

b

12 kDa (1.26/1.24) 27 kDa (1.28/1.26)

Determined from 1H NMR spectrum.

added dropwise to the flask over a duration of 30 min and allowed to stir at room temperature (∼22 °C) for an additional 2.5 h immediately after complete addition. The reacted mixture was quenched by addition of 50 mL of brine, and the organic solvent was collected after separation. After removal of solvent, the crude product was purified by silica gel flash column chromatography via a hexane−ethyl acetate solvent system (gradient elution up to 80 vol % ethyl acetate) to yield MTC-OCH2BnCl as a white solid. The crude product was further purified by recrystallization. The solid was dissolved in 1 mL of CH2Cl2 and ethyl acetate, followed by addition of 50 mL of diethyl ether. The crystals are allowed to form at 0 °C for 2 days and are subsequently obtained by washing the crystals with cold diethyl ether. 1 H NMR (400 MHz, CDCl3, 22 °C): δ 7.37 (dd, J = 20.2, 8 Hz, 4H, Ph−H), 5.21 (s, 2H, −OCH2), 4.69 (d, J = 13.6 Hz, 2H, −OCH2C−), 4.59 (s, 2H, −CH2Cl), 4.22 (d, J = 14.8 Hz, 2H, −OCH2C−), 1.32 (s, 3H, −C2CH3). 2.3. Synthesis of MTC-Furan-Protected Maleimide Cyclic Carbonate Monomer.39,40 MTC-Cl was synthesized as described above and redissolved in dry CH2Cl2 (50 mL), followed by immersing the flask in an ice bath at 0 °C. A mixture of exo-3a,4,7,7a-tetrahydro2-(3-hydroxypropyl)-4,7-epoxy-1H-isoindole-1,3(2H)-dione41 (3.97 g, 17.8 mmol) and triethylamine (1.77 mL, 19.3 mmol) was dissolved in dry CH2Cl2 (50 mL), which was added dropwise to the flask over a duration of 30 min and allowed to stir at room temperature for an additional 24 h immediately after complete addition. The reacted mixture was quenched by addition of 50 mL of water, and the organic solvent was collected after separation. After removal of solvent, the crude product was dissolved in 4 mL of CH2Cl2, followed by addition of 50 mL of diethyl ether for recrystallization. The crystals were allowed to form at room temperature and are subsequently obtained by washing with cold diethyl ether. 1H NMR (400 MHz, CDCl3, 22 °C): δ 6.51 (s, 2H, −CHCH), 5.25 (s, 2H, −OCHC2−), 4.74 (d, 2H, J = 14.4 Hz, −OCH2CC2−), 4.22 (d, 2H, J = 14.8 Hz, −OCH2CC2−), 4.11 (t, 2H, J = 6.0 Hz, −OCH2CH2−), 3.58 (t, 2H, J = 6.6 Hz, CH2CH2NC−), 2.85 (s, 2H, −COCHC−), 1.96 (quin, 6.4 Hz, 2H, −CONCCOCHC−), 1.38 (s, 3H, −C2CH3). 2.4. Polymer Synthesis. Details of the metal-free organocatalytic ring-opening polymerization for the triblock copolymer with the tethering block as the middle block (polymer 2.4k-V) are given as an example. In a glovebox, 24.1 mg (0.010 mmol) of 2.4 kDa MPEG-OH initiator and 36.7 mg (0.10 mmol) of MTC-FPM were charged in a 20 mL glass vial equipped with a stir bar. Dichloromethane was added, and the monomer concentration was adjusted to 2 M. Once the initiator and monomer were completely dissolved, 1.5 μL (0.01 mmol) of DBU was added to initiate the polymerization. After 45 min, 0.3 g (1.0 mmol) of MTC-OCH2BnCl was added to the reaction mixture and followed by addition of 6 μL (0.040 mmol) of DBU and 18.6 mg (0.050 mmol) of TU. The reaction proceeded at room temperature under stirring for another 40 min before it was quenched with 30 μL of trifluoroacetic acid. Subsequently, the polymer intermediate was purified via precipitation twice in cold diethyl ether and was dried on a vacuum line until a constant weight was achieved. 1H NMR (400 MHz, CDCl3, 22 °C) δ: 7.38−7.27 (m, 400H, −C6H4CH2Cl), 6.51− 6.42 (m, 14H, −CHOC 2 H 4 CHO−), 5.27−5.21 (m, 14H, −R2CHOCHR2−), 5.15−5.12 (m, 200H, −COOCH2−), 4.64−4.49 (m, 200H, −C6H4CH2Cl), 4.46−4.39 (m, 14H, −COOCH2CH2−), 4.37−3.96 (m, 426H, −CH2OCOO−), 3.87−3.60 (m, 217H,

OCH2BnCl) in sequential order, followed by deprotection to expose maleimide anchoring groups. The copolymers were subsequently quaternized with dimethylbutylamine. Two different triblock copolymers were synthesized where the placement of the anchoring block was varied, i.e., PEG-poly(carbonatemaleimide)-cationic polycarbonate (2.4k-V) and PEG-cationic polycarbonate-poly(carbonate-maleimide) (2.4k-S) (Figure 1 and Table 1), to elucidate the role of maleimide anchoring positions. Each of the polymers was designed to have a PEG block of the same molecular weight as well as cationic and maleimide-functionalized polycarbonate blocks of comparable length to facilitate comparison. In addition, 2.4k-Vshort with a shorter cationic block was synthesized for comparison. The polymers were analyzed by 1H NMR and GPC, and the polymer-coated surfaces were characterized by XPS and static water contact angle. In addition, the coated surfaces were tested against different strains of bacteria, proteins, and platelets to evaluate antibacterial and antifouling activities as well as protein/platelet adhesion by XTT (S. aureus) and Cell TiterBlue (E. coli) assays and confocal and scanning electron microscopic imaging.

2. EXPERIMENTAL SECTION 2.1. Materials. CH3O-PEG-OH (known as MPEG, Mn 2400 g mol−1, PDI 1.05) was purchased from Polymer Source, lyophilized, and transferred to a glovebox 1 day prior to use. N-(3,5Trifluoromethyl)phenyl-N′-cyclohexylthiourea (TU) was prepared according to a procedure reported previously.36 TU was dissolved in dry tetrahydrofuran and dried over CaH2 overnight. The mixture was filtered, and the solvent removed in vacuo. 1,8-Diazabicyclo[5,4,0]undec-7-ene (DBU) was dried over CaH2 overnight, and dried DBU was obtained after vacuum distillation. Both dried TU and DBU were transferred to a glovebox prior to use. FITC-conjugated bovine serum albumin (FITC-BSA), 3-mercaptopropyltrimethoxysilane, and all other chemicals were purchased from Sigma-Aldrich and used as received unless stated otherwise. Silicone Kit Sylgard 184 was bought from Dow Corning and used according to the manufacturer’s protocols. The LIVE/DEAD Baclight bacterial viability kit (L-7012) was obtained from Invitrogen. S. aureus (ATCC No. 6538) and E. coli (ATC No. 25922) were purchased from ATCC (USA). 2.2. Synthesis of MTC-OCH2BnCl Monomer.37,38 Briefly, in a dry two-neck 500 mL round-bottom flask equipped with a stir bar, MTC-OH (3.08 g, 19.3 mmol) was first dissolved in dry THF (50 mL) with 5−8 drops of dimethylformamide (DMF). Subsequently, oxalyl chloride (3.3 mL) was added in one shot (pure form), followed by an additional 20 mL of THF. The solution was stirred for 90 min, after which volatiles were blown dried under a strong flow of nitrogen to yield a pale yellow solid intermediate (5-chlorocarboxy-5-methyl1,3-dioxan-2-one, MTC-Cl). The solid was then subjected to heat at 60 °C for 2−3 min for the removal of residual solvent and was redissolved in dry dichloromethane (CH2Cl2, 50 mL), followed by immersing the flask in an ice bath at 0 °C. A mixture of pchloromethylbenzyl alcohol (2.79 g, 17.8 mmol) and pyridine (1.55 mL, 19.3 mmol) was dissolved in dry CH2Cl2 (50 mL), which was C

DOI: 10.1021/ma5022488 Macromolecules XXXX, XXX, XXX−XXX

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5.27−4.94 (m, 44H, −COOCH 2 −), 4.90−4.48 (m, 44H, −C6H4CH2Cl), 4.46−3.79 (m, 112H, −CH2OCOO−), 3.64−3.54 (m, 12H, −CH2CH2NR2), 3.53−3.39 (m, 217H, −OCH2CH2− from 2.4 kDa MPEG), 3.36−3.25 (m, 44H, −N+CH2CH2CH2−), 3.00− 2.96 (s, 132H, −N+[CH3]2), 2.24−2.08 (m, 12H, −OCH2CH2CH2−), 1.86−1.65 (m, 44H, −N+CH2CH2CH2−), 1.406−1.10 (m, 110H, −N+CH2CH2CH2− and −N+CH2CH2CH2CH3), 1.05−0.83 (m, 84H, −CH3). Polymer 2.4k-S with the tethering block as the end block was synthesized in similar fashion, with slight modification to the sequence of monomer addition. In a glovebox, 24.1 mg (0.010 mmol) of 2.4 kDa MPEG-OH initiator and 0.3 g (1.0 mmol) of MTC-OCH2BnCl were charged in a 20 mL glass vial equipped with a stir bar. Dichloromethane was added, and the monomer concentration was adjusted to 2 M. Once the initiator and monomer were completely dissolved, 7.5 μL (0.05 mmol) of DBU and 18.6 mg (0.050 mmol) of TU were added to initiate the polymerization. After 15 min, 36.7 mg (0.10 mmol) of MTC-FPM was added. The reaction pot was left to stir at room temperature for another 40 min before quenched with 30 μL of trifluoroacetic acid. Subsequently, the polymer intermediate was purified via precipitation twice in cold diethyl ether and was dried on a vacuum line until a constant weight was achieved. The protected polymer was then deprotected, followed by quaternization according to the procedures used for the synthesis of 2.4k-V. Polymer 2.4k-S (with Protected Maleimide). 1H NMR (400 MHz, CDCl3, 22 °C): 7.39−7.25 (m, 320H, −C6H4CH2Cl), 6.59−6.40 (m, 6H, −CHOC2H4CHO−), 5.26−5.21 (m, 6H, −R2CHOCHR2−), 5.18−5.02 (m, 160H, −COOCH 2 −), 4.81−4.63 (m, 160H, −C6H4CH2Cl), 4.62−4.48 (m, 6H, −COOCH2CH2−), 4.49−3.99 (m, 332H, −CH2OCOO−), 3.85−3.61 (m, 217H, −OCH2CH2− from 2.4 kDa MPEG), 3.59−3.53 (m, 6H, −CH2CH2NR2), 2.91−2.75 (m, 6H, −CC2HCC2H−), 1.92−1.87 (m, 6H, −OCH2CH2CH2−), 1.27−1.20 (m, 249H, −CH3). Polymer 2.4k-S (with Deprotected Maleimide). 1H NMR (400 MHz, CDCl3, 22 °C): 7.41−7.24 (m, 320H, −C6H4CH2Cl), 6.73− 6.63 (m, 6H, −COC2H4CO−), 5.25−5.03 (m, 160H, −COOCH2−), 4.65−4.46 (m, 160H, −C 6 H 4 CH 2 Cl), 4.44−4.40 (m, 6H, −COOCH2CH2−), 4.38−3.97 (m, 332H, −CH2OCOO−), 3.84− 3.61 (m, 217H, −OCH2CH2− from 2.4 kDa MPEG), 3.57−3.52 (m, 6H, −CH2CH2NR2), 1.91−1.88 (m, 6H, −OCH2CH2CH2−), 1.34− 1.14 (m, 249H, −CH3). Polymer 2.4k-S (Quaternized). 1H NMR (400 MHz, (CD3)2CO, 22 °C): 7.69−7.25 (m, 320H, −C6H4CH2Cl), 7.17−6.55 (m, 6H, −COC2H4CO−), 5.42−5.27 (m, 6H, −COOCH2CH2−), 5.26−4.92 (m, 160H, −COOCH2−), 4.89−4.47 (m, 160H, -C6H4CH2Cl), 4.45− 3.81 (m, 332H, −CH2OCOO−), 3.61−3.54 (m, 6H, −CH2CH2NR2), 3.54−3.40 (m, 217H, −OCH2CH2- from 2.4 kDa MPEG), 3.35−3.24 (m, 160H, −N+CH2CH2CH2−), 2.98 (s, 480H, −N+[CH3]2), 2.23− 2.00 (m, 6H, −OCH 2 CH 2 CH 2 −), 1.87−1.61 (m, 160H, −N+CH2CH2CH2−), 1.36−1.07 (m, 405H, −N+CH2CH2CH2− and −N+CH2CH2CH2CH3), 1.05−0.83 (m, 249H, −CH3). 2.5. Gel Permeation Chromatography (GPC). Polymer molecular weights were analyzed by GPC using a Waters HPLC system equipped with a 2690D separation module and two Styragel HR1 and HR4E (THF) 5 mm columns (size: 300 × 7.8 mm) in series arrangement, coupled with a Waters 410 differential refractometer detector. THF was employed as the mobile phase at a flow rate of 1 mL min−1. Number-average molecular weights and polydispersity indices of polymers were calculated from a calibration curve based on a series of polystyrene standards with molecular weights ranging from 1350 to 151 700. 2.6. 1H NMR Analysis. 1H NMR spectra of monomers and polymers were recorded on a Bruker Advance 400 NMR spectrometer, operated at 400 MHz and at room temperature. The 1H NMR measurements were performed using an acquisition time of 3.2 s, a pulse repetition time of 2.0 s, a 30° pulse width, 5208 Hz spectral width, and 32K data points. Chemical shifts were referred to solvent peaks (δ = 7.26 and 1.94 ppm for CDCl3 and CD3CN-d6, respectively).

−OCH 2 CH 2 − from 2.4 kDa MPEG), 3.56−3.51 (m, 14H, −CH2CH2NR2), 2.91−2.81 (m, 14H, −CC2HCC2H−), 2.17−1.98 (m, 14H, −OCH2CH2CH2−), 1.26−1.19 (m, 321H, −CH3). The protected polymer was then deprotected by dissolving in 10 mL of toluene and heated to 110 °C overnight. After that, the toluene was removed under vacuum, and the deprotected polymer was dissolved in 2 mL of dichloromethane and precipitated in cold diethyl ether. The polymer was subsequently dried on a vacuum line until a constant weight was achieved. 1H NMR (400 MHz, CDCl3, 22 °C): 7.40−7.24 (m, 396H, −C 6 H 4 CH 2 Cl), 6.72−6.65 (m, 12H, −COC2H4CO−), 5.21−5.02 (m, 198H, −COOCH2−), 4.59−4.48 (m, 198H, −C6H4CH2Cl), 4.45−4.40 (m, 12H, −COOCH2CH2−), 4.38−3.94 (m, 420H, −CH2OCOO−), 3.83−3.60 (m, 217H, −OCH 2 CH 2 − from 2.4 kDa MPEG), 3.59−3.54 (m, 12H, −CH2CH2NR2), 2.17−1.98 (m, 12H, −OCH2CH2CH2−), 1.27− 1.14 (m, 315H, −CH3). Finally, the polymer was dissolved in 20 mL of acetonitrile, and an excess (2 mL) of N,N-dimethylbutylamine was added to fully quaternize the OBnCl pendant groups. The reaction mixture was stirred overnight in a 50 mL round-bottom flask at room temperature, and the solvent was then removed in vacuo. The obtained product was dissolved in a mixture of acetonitrile and isopropanol (1:1 in volume) and dialyzed against the solvent mixture for 2 days. The solvent was removed under reduced pressure, and the final product was dried in a vacuum oven until a constant mass was achieved. Polymer 2.4k-V: 1H NMR (400 MHz, (CD3)2CO, 22 °C): 7.65−7.34 (m, 360H, −C6H4CH2Cl), 7.18−6.22 (m, 10H, −COC2H4CO−), 5.46−5.29 (m, 10H, −COOCH2CH2−), 5.25−5.04 (m, 180H, −COOCH2−), 4.80−4.52 (m, 180H, −C 6H 4 CH 2 Cl), 4.40−3.90 (m, 380H, −CH2OCOO−), 3.70−3.56 (m, 10H, −CH2CH2NR2), 3.54−3.38 (m, 217H, −OCH2CH2− from 2.4 kDa MPEG), 3.34−3.20 (m, 180H, −N+CH2CH2CH2−), 2.99 (s, 540H, −N+[CH3]2), 2.29−2.10 (m, 10H, −OCH2CH2CH2−), 1.81−1.70 (m, 180H, −N+CH2CH2CH2−), 1.34−1.25 (m, 180H, −N+CH2CH2CH2−), 1.23−1.10 (m, 270H, −N+CH2CH2CH2CH3), 1.05−0.84 (m, 285H, −CH3). Polymer 2.4k-Vshort was synthesized similarly to polymer 2.4k-V, with a shorter degree of polymerization of benzyl chloride moieties. In a glovebox, 24.1 mg (0.010 mmol) of 2.4 kDa MPEG-OH initiator and 36.7 mg (0.10 mmol) of MTC-FPM were charged in a 20 mL glass vial equipped with a stir bar. Dichloromethane was added, and the monomer concentration was adjusted to 2 M. Once the initiator and monomer were completely dissolved, 0.53 μL (0.01 mmol) of DBU was added to initiate the polymerization. After 45 min, 0.105 g (0.35 mmol) of MTC-OCH2BnCl was added to the reaction mixture and followed by addition of 2.1 μL (0.040 mmol) of DBU and 6.5 mg (0.050 mmol) of TU. The reaction proceeded at room temperature under stirring for another 40 min before it was quenched with 20 μL of trifluoroacetic acid. Subsequently, the polymer intermediate was purified via precipitation twice in cold diethyl ether and was dried on a vacuum line until a constant weight was achieved. Polymer 2.4k-Vshort (with Protected Maleimide). 1H NMR (400 MHz, CDCl3, 22 °C): 7.39−7.24 (m, 92H, −C6H4CH2Cl), 6.52−6.45 (m, 14H, −CHOC2H4CHO−), 5.26−5.21 (m, 14H, −R2CHOCHR2−), 5.18−5.08 (m, 46H, −COOCH2−), 4.65−4.52 (m, 46H, −C6H4CH2Cl), 4.42−4.19 (m, 120H, −CH2OCOO−), 4.09−4.40 (m, 14H, −COOCH2CH2−), 3.84−3.61 (m, 217H, −OCH 2 CH 2 − from 2.4 kDa MPEG), 3.60−3.52 (m, 14H, −CH2CH2NR2), 2.85−2.80 (m, 14H, −CC2HCC2H−), 1.96−1.85 (m, 14H, −OCH2CH2CH2−), 1.33−1.17 (m, 90H, −CH3). Polymer 2.4k-Vshort (with Deprotected Maleimide). 1H NMR (400 MHz, CDCl3, 22 °C): 7.40−7.24 (m, 88H, −C6H4CH2Cl), 6.73−6.64 (m, 12H, −COC2H4CO−), 5.19−5.08 (m, 44H, −COOCH2−), 4.61−4.51 (m, 44H, −C 6 H 4 CH 2 Cl), 4.46−4.18 (m, 112H, −CH2OCOO−), 4.12−4.03 (m, 12H, −COOCH2CH2−), 3.83− 3.61 (m, 217H, −OCH2CH2− from 2.4 kDa MPEG), 3.60−3.53 (m, 12H, −CH2CH2NR2), 1.97−1.87 (m, 12H, −OCH2CH2CH2−), 1.34−1.14 (m, 84H, −CH3). Polymer 2.4k-V short (Quaternized). 1 H NMR (400 MHz, (CD3)2CO, 22 °C): 7.87−7.29 (m, 88H, −C6H4CH2Cl), 7.17−6.53 (m, 12H, −COC2H4CO−), 5.43−5.29 (m, 12H, −COOCH2CH2−), D

DOI: 10.1021/ma5022488 Macromolecules XXXX, XXX, XXX−XXX

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μL, 3 × 105 CFU mL−1) was seeded onto uncoated and polymercoated PDMS surfaces, topped up with 60 μL of MHB, and cultured at 37 °C for 24 h. Each surface was washed thrice with sterile PBS and was carefully placed in individual 8 mL tubes containing 1.5 mL of PBS. Each tube was sonicated for 8 s to detach bacterial cells, and viable counts in the resulting suspensions were performed by plating on agar medium to enumerate bacteria that were attached to the disklike PDMS surface.42 2.14. Antifouling Analysis of Various PDMS Surfaces by XTT Assay. Another quantitative measurement of live bacteria cells attached onto the PDMS surface was performed by studying 2,3bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide (XTT) reduction.43 XTT reduction assay measures cell metabolic activity in live cells. Briefly, S. aureus in MHB (20 μL, 3 × 105 CFU mL−1) was seeded onto uncoated and polymer-coated PDMS surfaces, topped up with 60 μL of MHB, and cultured at 37 °C for 24 h. Each surface was washed thrice with sterile PBS, followed by incubation with 100 mL of PBS, 10 μL of XTT, and 2 μL of menadione at 37 °C for 2 h. The optical density (OD) of formazan dye produced by XTT reduction in each sample was recorded at 490 nm with a reference wavelength of 660 nm using a microplate reader (TECAN, Sweden). The experiment was conducted in triplicates. 2.15. Antifouling Analysis by Cell Titer Blue Assay. The Cell Titer-Blue cell viability assay provided quantitative analysis of live E. coli cells attached onto the PDMS surface, as XTT was unable to reproduce similar quantitative results. E. coli in MHB (20 μL, 3 × 105 CFU mL−1) was seeded onto the uncoated and polymer-coated PDMS surfaces, topped up with 60 μL of MHB, and cultured at 37 °C for 24 h. The surface was washed twice with sterile PBS, followed by incubation with 100 mL of PBS and 20 μL of Cell Titer Blue Reagent at 37 °C for 2 h. The fluorescence intensity of resorufin produced after reduction within mitochondrial enzymes of viable cells was determined at excitation wavelength of 560 nm and emission wavelength of 590 nm using the microplate reader. The experiment was conducted in triplicates. 2.16. LIVE/DEAD Baclight Bacterial Viability Assay. A LIVE/ DEAD Baclight bacterial viability kit (L-7012, Invitrogen), containing both propidium iodide and SYTO fluorescent nucleic acid staining agents, was used to label bacterial cells on the uncoated and polymercoated PDMS surfaces. Briefly, the red fluorescent dye propidium iodide, which only penetrates damaged cell membrane, was used to label dead bacterial cells. The green fluorescent dye SYTO 9, which can penetrate cells both with intact and damaged membranes, was used to label all bacterial cells. Bacteria solution (3 × 105 CFU mL−1, 20 μL) was seeded onto the uncoated and polymer-coated PDMS surfaces, followed by incubation at 37 °C for 24 h or 7 days. The surfaces were washed thrice with clean PBS after the bacteria solution was removed. Subsequently, each PDMS sample was placed individually into a 48-well plate with 200 μL of a dye solution, prepared from a mixture of 3 μL of SYTO (3.34 mM) and 3 μL of propidium iodide (20 mM) in 2 mL of PBS. The procedure was conducted at room temperature in the absence of light for 15 min. Eventually, the stained bacterial cells attached to the surfaces were examined under a Zeiss LSM 5 DUO laser scanning confocal microscope (Germany), and the images were obtained using an oil immersed 40× object lens at room temperature. 2.17. Analysis of Bacteria Attachment and Biofilm Formation by Field-Emission Scanning Electron Microscopy (FESEM). FE-SEM was employed to evaluate the attachment and biofilm formation of S. aureus or E. coli on the uncoated and polymer-coated PDMS surfaces. Bacteria solution (3 × 105 CFU mL−1, 20 μL) was seeded onto the uncoated and polymer-coated PDMS surfaces, followed by incubation at 37 °C for 1 or 7 days. An additional 20 μL of MHB was added after every 24 h to prevent the bacteria culture medium from drying out. At the predetermined time points, the PDMS surfaces were washed thrice with sterile PBS, followed by fixation with 2.5% glutaraldehyde in PBS overnight. The fixed bacteria were dehydrated with a series of graded ethanol solution (25%, 50%, 75%, 95%, and 100%, 10 min each) before the PDMS samples were mounted for platinum coating. A field emission scanning electron

2.7. Preparation of Polydimethylsiloxane (PDMS) Silicone Rubber. PDMS silicone rubber was prepared by mixing 10 base parts to 1 curing part thoroughly, followed by degassing under vacuum for 30 min. The mixture was spin-coated onto a Petri dish (for LIVE/ DEAD cell staining and SEM studies) using a SAWATECH AG Spin Module SM-180-BT, or it was cast into a 48-well plate for XTT, Titer Blue cell viability, and colony assays. Both the Petri dish and plate were placed overnight in a vacuum oven at 70 °C for curing. After curing, the PDMS sample formed in the Petri dish was cut into square pieces (0.5 cm × 0.5 cm with a thickness of about 1 mm). The disk-like PDMS samples were gently removed from the bottom of the 48-well plate with flat forceps. All PDMS samples were first sonicated with deionized (DI) water, followed by isopropanol and DI water. The samples were dried under a stream of nitrogen before use. 2.8. Vapor Deposition of PDMS Surface. Clean PDMS surface was exposed to ultraviolet/ozone (UVO) radiation for 1 h in a commercial PSD-UVT chamber (Novascan). The surface was then briefly exposed to humid air and dried under a stream of nitrogen. Subsequently, the dried PDMS surface was placed on a clean piece of weighing paper in a small vacuum desiccator, together with 1 mL of 3mercaptopropyltrimethoxysilane loaded in a clean vial. The vapor deposition process was carried out overnight with the desiccator sealed under vacuum at 70 °C to provide thiol-functionalized surface. The treated surface was dried under a stream of nitrogen and kept in a sealed desiccator at room temperature prior to use. 2.9. Polymer Coating. The polymers of different compositions (2 mg) were first dissolved in 400 μL of HPLC grade water, 500 μL of PBS (pH 7.4), and 100 μL of SDS solution (to prevent micellization of the polymers during the coating process). Subsequently, the clean PDMS surface treated with 3-mercaptopropyltrimethoxysilane was immersed in the polymer solution for 1 day at room temperature. The polymer-coated PDMS samples were sonicated in a mixture of isopropropanol and water (1:1 volume ratio) and dried under a stream of nitrogen before use. 2.10. X-ray Photoelectron Spectroscopy (XPS) Measurements. The difference in chemistry between uncoated and polymercoated PDMS surfaces was analyzed by XPS (Kratos Axis HSi, Kratos Analytical, Shimadzu, Japan) with Al Kα source (hν = 1486.71 eV). The angle between the surface of the sample and the detector was kept at 90°. The survey spectrum (from 1100 to 0 eV) was acquired with pass energy of 80 eV. All binding energies were calculated with reference to C 1s (C−C/C−H bonds) at 285.0 eV. 2.11. Static Contact Angle Measurements. The static contact angles of both uncoated and polymer-coated surfaces were measured by an OCA15 contact angle measuring device (Future Digital Scientific Corp.). DI water (20 μL) was used for all measurements. All samples were analyzed in triplicates, and the static contact angle data were presented as mean ± SD. 2.12. Killing Efficiency of Polymer-Coated Surfaces (Colony Assay). The concentration of S. aureus or E. coli in Mueller-Hinton broth (MHB, cation-adjusted) was adjusted to give an initial optical density (OD) reading of 0.07 at the wavelength of 600 nm on a microplate reader (TECAN, Switzerland), which correlates to a concentration of Mc Farland 1 solution (3 × 108 CFU mL−1). The bacterial solution was diluted by 1000 times to achieve a loading of 3 × 105 CFU mL−1. Subsequently, 20 μL of this bacterial solution was added to the surface of an uncoated or coated disk-like PDMS sample, which was placed in a 48-well plate. Additionally, 60 μL of MHB was added to the surface, and the 48-well plate was incubated at 37 °C for 24 h. The bacterial solution (10 μL) was then taken out from each well and diluted with an appropriate dilution factor. The bacterial solution was streaked onto an agar plate (LB Agar from first Base). The number of colony-forming units (CFUs) was tabulated and recorded after an incubation of about 18 h at 37 °C. Each test was conducted in triplicates. 2.13. Antifouling Analysis of Pristine, Thiol-Functionalized, and Polymer-Coated PDMS Surfaces by Surface Viable Colonies. Quantitative measurement of live bacterial cells attached onto PDMS surface was performed by directly enumerating the bacteria adhered to the surface. Briefly, S. aureus or E. coli in MHB (20 E

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Figure 2. Static water contact angles and high resolution N 1s spectra of pristine, thiol-functionalized, and 2.4k-V as well as 2.4k-S coated PDMS surfaces. of the negative control)] × 100. The data were analyzed and expressed as mean and standard deviation of three replicates for quantification of each type of PDMS surface.

microscope (FE-SEM, JEOL JSM-7400F, Japan) was used to observe the PDMS surfaces. 2.18. Analysis of Platelet Adhesion. Fresh rat blood was centrifuged at 1000 rpm min−1 and at room temperature for 10 min to obtain platelet rich plasma (PRP) in the supernatant. Uncoated and polymer-coated PDMS surfaces were immersed in PRP and incubated at 37 °C for 30 min. The samples were then washed thrice with PBS, followed by the same bacteria fixation and FE-SEM analysis procedures described above. 2.19. Fluorescence Analysis for Protein Fouling. Individual surfaces were incubated overnight with 20 μL of FITC-BSA solution (1 mg/mL) at 37 °C. The surfaces were then washed thrice with clean sterile PBS solution before they were observed under an inverted fluorescence microscope (Olympus IX71, U.S.A). Meanwhile, the FITC-BSA solutions were removed from the respective surfaces and dissolved in 1 mL of sterile PBS solution. The fluorescence intensity of the solution was investigated using a PerkinElmer-LS55 luminescence spectrometer with Jobin Yvon Fluorolog-3 at 495 and 525 nm excitation and emission wavelengths, respectively. 2.20. Hemolysis Test. Freshly obtained rat blood was diluted to 4 vol % with PBS buffer. The red blood cell suspension in PBS (500 μL) was added into a 2 mL Eppendorf tube, which contained uncoated or polymer-coated PDMS samples individually. The tube was incubated for 1 h at 37 °C for hemolysis to proceed. After incubation, the tube was centrifuged at 2200 rpm for 5 min at room temperature. Aliquots (100 mL) of the supernatant from each tube were transferred to a 96well plate, and hemoglobin release was measured at 576 nm using the microplate reader (TECAN, Sweden). In this procedure, the red blood cells in PBS were used as a negative control, while the red blood cells lysed with 0.2% Triton X were used as a positive control. The absorbance analysis for red blood cells lysed with 0.2% Triton X was taken as 100% hemolysis. The calculation for percentage of hemolysis was as follow: hemolysis (%) = [(OD576 nm of the sample − OD576 nm of the negative control)/(OD576 nm of the positive control − OD576 nm

3. RESULTS AND DISCUSSION 3.1. Polymer Synthesis. The polymers were synthesized via metal-free organocatalytic ring-opening polymerization of MTC-OCH2BnCl and MTC-FPM using MPEG as a macroinitiator in the presence of the cocatalysts DBU and TU (Figure 1a). The reaction was quenched with trifluoroacetic acid and left to stir for 5 min. 1H NMR integration values of monomers relative to the PEG initiator correlated well, confirming controlled polymerization and predictable molecular weights via initial monomer to initiator feed ratio. In addition, the proton NMR analysis displayed all the peaks associated with both initiator and monomers. Polymers 2.4k-V, 2.4k-Vshort, and 2.4k-S polymers had narrow molecular weight distribution with polydispersity index (PDI) ranging between 1.22 and 1.28 (Table 1 and Figure S1). Subsequently, after precipitating twice in cold diethyl ether, the two polymers were isolated and dried. The furan-protected maleimide groups in the polymers were subsequently deprotected by dissolving in toluene and heating to 110 °C overnight. The deprotected polymers were purified by reprecipitation in cold diethyl ether twice, and 1H NMR showed a downfield shift from 6.49 to 6.68 ppm, which correlates to the deprotected maleimide pendant groups (Figure S1a vs Figure S2b in the Supporting Information). There was negligible difference in PDIs before and after deprotection, indicating that the deprotection did not affect the polymer chain. Excess N,N-dimethylbutylamine was added to the polymers dissolved in 20 mL of acetonitrile to F

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Figure 3. Viable surface colonies analysis of S. aureus and E. coli at 1 and 7 days.

−SH appeared with an atomic content of 2.35% (Figure S3c). Moreover, the N (1s) peak was observed on the surface grafted with 2.4k-V or 2.4k-S, showing comparable nitrogen atomic contents for both polymer coatings (i.e., 0.61% and 0.45%, respectively). In addition, the binding energy of sulfhydryl (SH) from the thiol-functionalized surface at 166 eV shifted to 164 eV after polymer grafting due to the formation of thiolether bond between maleimide of the polymer and sulfhydryl group of the thiol-functionalized surface. In the high-resolution N 1s spectrum of the coated surface, there were two distinct peaks (Figure 2). The first peak at 400 eV represents the amine from the maleimide pendant group (Figure 1a), and the second peak at 403 eV is from N,N-dimethylbutylammonium functional groups. These findings demonstrated successful grafting of the polymers onto the PDMS surface. 3.3. Antimicrobial Activity of Polymer-Coated Surfaces. Pristine PDMS silicone and thiol-functionalized surfaces, and surfaces coated with the two polymers, were tested against Gram-positive S. aureus and Gram-negative E. coli. With the pristine surface serving as the control, killing efficiency for the thiol-functionalized surface and surfaces coated with the two copolymers was studied. The number of S. aureus in solution decreased slightly when the solution was incubated with the thiol-functionalized PDMS surface for 24 h as compared to the pristine surface, while the surfaces coated with the cationic polymers 2.4k-V and 2.4k-S killed 98.5% and 89.4% bacterial cells, respectively (Figure S4a). In addition, there was significant reduction in viable colonies of E. coli in solution incubated with the 2.4k-V and 2.4k-S coated surfaces with killing efficacies of 93.9% and 82.5%, respectively (Figure S4b). The 2.4k-V polymer coating had a greater killing effect against both S. aureus and E. coli in solution than the 2.4k-S polymer coating. Unlike 2.4k-S, where PEG shields the cationic antibacterial block, the configuration of 2.4k-V offered easier access of the cationic block to bacteria in solution, leading to more effective eradication of the bacteria (Figure 1b). In addition, 2.4k-Vshort with a shorter cationic block had comparable antibacterial activity in solution as compared to 2.4k-V (99.4% vs 98.5% against S. aureus; 96.9 vs 93.9% against E. coli), which was also stronger than 2.4k-S. As Gram-negative E. coli possess an additional lipopolysaccharide-containing

achieve complete quaternization. The quaternized polymers were purified via dialysis for 2 days. From 1H NMR analysis, the presence of a new distinct peak at 2.99 ppm confirmed quantitative quaternization of −OCH2BnCl pendant groups (Figure S2b vs Figure S2c). The two polymers 2.4k-V and 2.4kS were of similar cationic block length and a comparable number of maleimide groups (Table 1), while 2.4k-Vshort had a shorter degree of polymerization for the cationic block and comparable repeat units of maleimide groups as compared to 2.4k-V. 3.2. Polymer Coating and Characterization. Polymers were grafted onto PDMS surfaces, and their antibacterial and antifouling properties (Figure 1b) were studied. Clean samples of PDMS silicone rubber were exposed to ultraviolet/ozone (UVO) radiation to generate hydroxyl groups, and 3mercaptopropyltrimethoxysilane was deposited onto the surface at 70 °C to provide thiol functional groups via condensation reaction. The thiol functional groups on the PDMS surface reacted with the maleimide pendant groups on the polymer via Michael addition.39 The static water contact angles of treated and untreated PDMS surfaces were measured to study wettability change after coating. As shown in Figure 2 (insets), the contact angle of silicone rubber surface decreased drastically after UV/ozone passivation (108.6 ± 0.7° vs 22.3 ± 1.0°). After functionalizing with mercaptopropyltrimethoxysilane, the PDMS surface regained partial hydrophobicity (83.8 ± 2.4°). Cationic polymer coating led to increased wettability (2.4k-S polymercoated surface: 71.7 ± 0.8°; 2.4k-V polymer-coated surface: 77.2 ± 0.7°; 2.4k-Vshort polymer-coated surface: 74.2 ± 1.7° (Figure S3a). These findings demonstrate that the cationic polymer coatings increased the wettability of silicone rubber surface. The XPS spectra of silicone rubber before and after polymer coating were obtained and analyzed to affirm successful grafting of the polymers onto the thiol-functionalized PDMS surface. The atomic content of C 1s, O 1s, N 1s, and S 2p peaks were analyzed and compared among the pristine, thiol-functionalized, and 2.4k-V as well as 2.4k-S grafted surfaces (Figure S3b). After successful vapor deposition of 3-mercaptopropyltrimethoxysilane onto the pristine surface, the S 2p peak from G

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Figure 4. Live and dead S. aureus (a) and E. coli (b) cell staining on the uncoated silicone PDMS surface and surfaces coated with thiol, 2.4k-V, and 2.4k-S. The surfaces were imaged under confocal laser scanning microscopy after 1 and 7 days of incubation (green denotes live and dead cells; red denotes dead cells).

outermost membrane as compared to Gram-positive S. aureus, the polymer coatings had relatively lower killing efficiency against E. coli as compared to S. aureus. 3.4. Bacterial Antifouling Activities of PolymerCoated Surfaces. Antifouling activity is one of the most important properties that ideal catheters should possess to prevent catheters-associated infections. To quantitatively investigate bacteria fouling on the polymer-coated silicone rubber surfaces, the number of viable bacterial cells fouled on the surfaces was measured. After 1 day of incubation, there was already a high number of live S. aureus and E. coli cells fouled onto both pristine and thiol-functionalized surfaces (S. aureus: 8.8 log and 8.6 log CFUs, respectively; E. coli: 8.5 log and 8.2 log CFUs, respectively) (Figure 3). The CFU values increased significantly especially on the thiol-functionalized surface after 7 days. In contrast, the polymer-coated surfaces showed significant antifouling activity on both day 1 and day 7 with 2.4k-S being more effective. Higher antifouling efficacy offered by 2.4k-S coating indicates that the antifouling PEG block as the outmost layer is more effective to prevent bacteria from adhering onto the surface. A complementary XTT assay (Figure S5a), which measures bacterial cell viability, was performed to further evaluate antifouling activity of the polymer-coated uncoated surfaces,

and the results are well correlated to the viable surface colonies of S. aureus determined by agar plating (Figure 3). Cell TiterBlue cell viability assay was employed to quantify fouling of E. coli as XTT assay was unable to detect E. coli. Similar to S. aureus, the pristine and thiol-functionalized surfaces showed extensive E. coli fouling, while polymer coatings inhibited E. coli fouling with 2.4k-S coating being more promising (Figure S5b). LIVE/DEAD bacterial cell staining was performed to further confirm the antifouling property of polymer coatings against both S. aureus and E. coli. As shown in Figure 4, a large number of live cells were seen on the pristine and thiol-functionalized surfaces after 1 day and 7 days of incubation. The surface coated with 2.4k-S had significantly less fouling as compared to surface coated with polymer 2.4k-V, which is in agreement with both viable surface colonies (Figure 3) and XTT assay results (Figure S5a). Moreover, 2.4k-Vshort polymer coating performed similarly to 2.4k-V polymer coating, displaying greater fouling on the surface as compared to 2.4k-S polymer. Biofilm on surfaces consists of bacteria, their secreted extracellular matrix, and organic debris and is extremely difficult to remove.44,45 From SEM analysis, the control surfaces without polymer coating developed S. aureus and E. coli biofilm especially at 7 days (Figure 5). The 2.4k-V coating was unable to prevent biofilm formation, especially E. coli biofilm. In sharp H

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Figure 5. Prevention of biofilm formation. FE-SEM images of S. aureus (a) and E. coli (b) after 1 day/7 days of incubation on uncoated and coated PDMS surfaces. Size of the scale bars: 10 μm.

examined by SEM analysis. Platelet fouling was seen on the entire pristine surface (Figure S7). Interestingly, the surface coated with 2.4k-V was shown to attract a number of platelets. However, very few platelets were observed on the surface coated with 2.4k-S coated surface, implying that 2.4k-S coating successfully prevented platelet fouling. Hemolytic activity of the untreated and polymer-coated surfaces was evaluated using rat red blood cells. All surfaces exhibited almost no or minimal hemolysis after incubation with red blood cells (Figure S8), which is ideal for use as antifouling coatings, especially for intravenous catheters.

contrast, 2.4k-S coating with the optimal configuration inhibited bacteria fouling, effectively preventing biofilm formation of both S. aureus and E. coli. It is significant that the 2.4k-S coating inhibited the fouling of Gram-negative bacteria E. coli for a week as similar antifouling performance was not reported in most polymer coatings.46,47 3.5. Hemocompatibility of Polymer-Coated Surfaces. The uncoated and coated surfaces were examined for their protein adsorption, platelet adhesion, and hemolysis to study blood compatibility. Proteins are present in blood and adsorption of the proteins may mask the antifouling function of polymer coatings. FITC-labeled BSA was used as a model protein to evaluate protein adsorption on the pristine and polymer-coated surfaces. From the fluorescence microscopic images of the surfaces (Figure S6a), although thiol-functionalized surfaces inhibited protein fouling, the pristine surface showed the greatest degree of protein adsorption, followed by 2.4k-V coated surface. Protein adsorption was significantly reduced on the surface coated with 2.4k-S (Figure S6b). The antifouling PEG block was positioned at the topmost position within the covalently tethered triblock copolymer 2.4k-S on the surface (Figure 1b), effectively preventing proteins and bacteria from fouling onto the surface.48 Meanwhile, in the 2.4k-V coating the cationic block might be easily accessed by proteins, promoting protein fouling. Platelet adhesion may cause thrombus formation. Platelet adhesion on the pristine and copolymer-coated surfaces was

4. CONCLUSION Triblock copolymers rationally composed of three components including an antifouling PEG, antibacterial cationic polycarbonate, and maleimide-functionalized polycarbonate (for anchoring onto silicone rubber surface) were successfully synthesized via metal-free organocatalytic ring-opening polymerization as antibacterial and antifouling coating materials. The polymers were grafted onto thiol-functionalized PDMS silicone rubber surfaces through the maleimide via Michael addition. The effects of anchoring block placement in the polymers and cationic block length were studied. The 2.4k-S coating with PEG as the outermost layer was more effective against S. aureus and E. coli fouling over 1 week than 2.4k-V with the surface anchoring block as the middle block, preventing biolfilm formation over 7 days. The cationic length I

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did not affect the antibacterial and antifouling activity significantly. More importantly, the 2.4k-S coating was able to resist protein fouling and platelet adhesion and did not cause significant hemolysis. Therefore, this polymer coating holds potential for prevention of bacterial fouling, biofilm formation, and catheter-associated bloodstream infections.

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ASSOCIATED CONTENT

S Supporting Information *

GPC diagrams, 1H NMR spectra of polymers, antibacterial, protein adsorption, platelet adhesion, and hemolysis activities of polymer coatings. This material is available free of charge via the Internet at http://pubs.acs.org.

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AUTHOR INFORMATION

Corresponding Authors

*E-mail [email protected] (Y.Y.Y.). *E-mail [email protected] (J.L.H.). Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS This work was funded by Institute of Bioengineering and Nanotechnology (Biomedical Research Council, Agency for Science, Technology and Research, Singapore) and SERC Personal Care Programme Grant No. 1325400028.

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REFERENCES

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DOI: 10.1021/ma5022488 Macromolecules XXXX, XXX, XXX−XXX