Ultrahighly Charged Amphiphilic Polymer Brushes with Super

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Interfaces: Adsorption, Reactions, Films, Forces, Measurement Techniques, Charge Transfer, Electrochemistry, Electrocatalysis, Energy Production and Storage

Ultrahighly Charged Amphiphilic Polymer Brushes with Super-Antibacterial and Self-Cleaning Capabilities Ting Chen, Hui Yang, Xu Wu, Danfeng Yu, Aiqing Ma, Xu He, Keji Sun, and Jinben Wang Langmuir, Just Accepted Manuscript • DOI: 10.1021/acs.langmuir.8b04187 • Publication Date (Web): 05 Feb 2019 Downloaded from http://pubs.acs.org on February 8, 2019

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Ultrahighly Charged Amphiphilic Polymer Brushes with Super-Antibacterial and Self-Cleaning Capabilities Ting Chena,b, Hui Yanga,*, Xu Wuc, Danfeng Yuc, Aiqing Ma,d Xu He,d Keji Sun,d Jinben Wanga aCAS

Key Lab of Colloid, Interface and Chemical Thermodynamics, Institute of

Chemistry, Chinese Academy of Sciences, No. 2, 1st North Street, Zhongguancun, Beijing 100190, P. R. China bUniversity

of Chinese Academy of Sciences, No. 19, Yuquan Road, Shijingshan

District, Beijing 100049, P. R. China cDepartment

of Chemistry and Chemical Engineering, Guangzhou University, No.

230, Outer Ring Road, Panyu District, Guangzhou, Guangdong 510006, P.R.China dOil

Production Technology Research Institute, Shengli Oilfield Branch Company,

Sinopec, No. 306, Xisan Road, Dongying District, Dongying, Shandong 257000, P. R. China

ACS Paragon Plus Environment

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ABSTRACT Bacterial infection on biomaterial devices and the subsequent medical risks pose a serious problem in both human healthcare and industrial applications, resulting in a prevalence of various antimicrobial materials. Cationic amphiphilic polymer has been proposed to be a new generation of efficient antibacterial material, but the surface modified by such kind of polymer still shows incomplete bactericidal ability and easily contaminated performance. With this in mind, a novel kind of geminized cationic amphiphilic polymer brush surface has been developed in this study, presenting a complete antibacterial activity, due to the synergistic biocidal effect of electrostatic and hydrophobic interactions as well as the minimized contact area between bacteria and polymer surface. A structure self-adjustment process of polymer brush construction has been proposed, in which the mutual interference among cationic head groups can be avoided and the electrostatic repulsion and hydrophobic attraction can be balanced, in the formation of a smooth and tight surface. A self-cleaning capability of polymer surface has been carried out via hydrolysis and degradation, maintaining a high antibacterial activity. Therefore, we provide a facile and possible manipulation strategy to fabricate super-antibacterial and self-cleaning surfaces in a wide range of biomedical and industrial applications.


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INTRODUCTION Antibacterial materials are widely used to prevent invasive infections in various applications, such as medical implants or devices, water purification systems, and ship hulls.1-4 Cationic antibacterial polymer brushes attract considerable attention to replace traditional materials nowadays, due to the ability of interacting with microbial membranes through electrostatic interactions and the broad spectrum activity against both gram-positive and negative bacteria.5-8 A significant problem with the most current cationic polymer surfaces is that they can be easily masked by biomolecules or residues of dead cells, resulting in a lack of further interactions with pathogens and a trigger of adverse effects.9-10 Therefore, the development of antibacterial materials possessing both contact-active biocidal and antifouling properties is a promising approach to combat microbial contamination.11-12 For example, a dual functional coating was developed, in which the antibacterial upper-layer was consisted of gemini quaternary ammonium salt and the antifouling sub-layer was consisted of polyethylene glycol, showing a perfect antifouling activity against surface-attached bacteria and an incomplete bactericidal performance.13 Moreover, it is found that the antibacterial coatings weakly interact with substrates by casting method, which is easily detached from the surfaces, bringing in the loss of bactericidal and antifouling ability. To

overcome

such

issues,

here

poly{1,3-bis(N,N-dimethyl-N-octylammonium)-2-propyl

we

firstly

acrylate

prepared

dibromide},

a

geminized amphiphilic brush containing double cationic head groups and


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hydrophobic tails in each structural unit (denoted as PAGC8), and covalently tethered on solid surface in a simple one-pot method. It is shown in our previous work that the geminized amphiphilic polymer not only has ultrahighly positive charge density contributing to a strong attraction with bacteria, but also has hydrophobic structures interacting with lipid layers destroying the cell membrane of bacteria, potentially being more efficient due to the synergetic effect of both kinds of interactions.14-15 More interestingly, such antibacterial polymer brush may be endowed with self-cleaning capability via a strategy of hydrolysis and degradation. For comparison, poly{2-(methacryloyloxy)ethyl-N,N-dimethyl-N-octylammonium

bromide},

an

amphiphilic polymer brush comprising a single cationic head group and hydrophobic moiety

in

each

structural

unit

(denoted

as

PASC8)

and

poly{2-(methacryloyloxy)ethyltrimethylammonium chloride}, a hydrophilic polymer brush comprising a single cationic head group in each unit (denoted as PASC1) were prepared. We found that the geminized polymer brush PAGC8 presents a complete antibacterial activity due to the synergistic effect of electrostatic and hydrophobic interactions between quaternary ammonium groups or hydrophobic tails of polymers and the lipid layer of the bacterial cell membrane. After the hydrolysis and degradation, PAGC8 still maintains an excellent antibacterial activity. Therefore, we demonstrate that the geminized amphiphilic cationic polymer brushes can be a promising candidate material with super-antibacterial and self-cleaning capabilities in a wide range of biomedical and industrial applications. EXPERIMENTAL SECTION


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Materials.

N,N-dimethylaminoethyl

2-(methacryloyloxy)ethyltrimethylammonium 1-bromooctane,

methacrylate,

chloride

(80%

1,1,4,7,7-pentamethyldiethylenetriamine

2-bromoisobutylate,

copper

(I)

in

(PMDETA),

bromide

water), ethyl (CuBr),

(2-bromo-2-methyl)propionyloxypropyltrimethoxysilane, methanol, acetonitrile and N,N-dimethylformamide (DMF) were purchased from J & K Chemical Technology. Deuterium oxide (99.9%), acryloyl chloride, ammonium persulfate and ammonium iron[II] sulfate hexahydrate were purchased from Beijing Chemical Co. (A.R. Grade). The ampicillin resistant gram-negative bacteria Escherichia coli (E. coli) and gram-positive bacteria Staphylococcus aureus (S. aureus) were purchased from Beijing Bio-Med Technology Development Co., Ltd. Phosphate buffer saline (PBS, 10 mM, pH = 7.4) was purchased from Sigma Chemical Co., Ltd. and used in antibacterial tests. All of the reagents except those especially mentioned were used without further purification and all of the solutions were prepared using Millipore Milli-Q grade water in this work. Synthesis of monomers. 2-(methacryloyloxy)ethyltrimethylammonium chloride (for convenience, we called ASC1) was used without further treatment. Quaternized 2-(methacryloyloxy)ethyl-N,N-dimethyl-N-octylammonium

bromide

(we

called

ASC8) was prepared by blending N,N-dimethylaminoethyl methacrylate and 1-bromooctane overnight in acetonitrile with a molar ratio of 1:2. Geminized cationic monomers of 1,3-bis(N,N-dimethyl-N-octylammonium)-2-propyl acrylate dibromide, we called AGC8, were prepared through two-step reaction (Figure S1a). Firstly,


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1,3-dimethylamino-2-propanol were synthesized by blending epoxypropane and dimethylamine in ethanol solution at a molar ratio of 1:5, and quaternized with 1-bromine octane to obtain the monomer precursor, 1,3-bis(N,N-dimethyl-N-octyl ammonium)-2-hydroxylpropane dibromide, abbreviated as BHD-C8 (see step (1)). Secondly, acryloyl chloride was added dropwise to an anhydrous chloroform solution of BHD-C8 with a molar ratio of 1.2:1 (see step (2)). After recrystallization with acetone/diethyl ether repeatedly, the white solid powder of AGC8 was obtained. The monomer structures of ASC1, ASC8, and AGC8 were further confirmed by 1H NMR spectra (Figure S1 (b~d)).

O

O

Br

O O Si

Si

(1)

N Cl

O O

O

Si

Si

O O O

Si-ATRP

(2)

Br

O O

N Br

O O O

Si-PASC1

(a)

O O

or

O O

O

O O O

OH OH OH

Br

O

n

Br

n

Br

n

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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Si

N Br

O

or

Si

O O O

O O O

Si-PASC8

Si-PAGC8

(b)

Br N

(c)

Figure 1. Synthetic routes of different polymer brushes. Preparation of polymer brushes. PASC1 polymer brush was constructed through surface-initiated atom transfer radical polymerization (SI-ATRP), as illustrated in Figure 1a. The single crystal silicon wafers (1 cm ×1 cm) were firstly treated with piranha solution (H2SO4: H2O2 = 3:1(v/v)) to remove the organic residues on the surface, and then washed with deionized water and dried with pure nitrogen, followed by further treatment with plasma for 10 min. As step 1 in Figure 1, the freshly prepared hydroxyl terminated silicon wafers were immediately immersed in a 5% initiator solution of (2-bromo-2-methyl)propionyloxypropyltrimethoxysilane with


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triethylamine (0.1% v/v) for 16 h at room temperature, to obtain an ATRP initiator-coated surface, and the contact angle increases from ~12.1° to ~79.3°, indicating the successful modification of an ATRP initiator-coated surface (Figure S2). In step 2, ASC1 monomers (5.0 mL, 20 mmol), PMDETA (100 μL, 0.48 mmol), and ethyl-2-bromoisobutanoate (40 μL, 0.27 mmol) were dissolved in methanol (25 mL) and water (25 mL), and the mixture was degassed. Cu(I)Br (50 mg, 0.35 mmol) and ATRP initiator-coated silicon surfaces were then added to the flask for the polymerization and three freeze-pump-thaw cycles were conducted to remove oxygen. After 48 h, the solution was exposed to air to terminate the reaction. The silicon wafers were extracted, washed with methanol solution for 3 h to remove free polymers from the layer and dried through nitrogen blowing. Differently, ASC8 monomers (3.0 g, 8.50 mmol) and AGC8 monomers (5.0 g, 8.50 mmol) was used for preparation of PASC8 and PAGC8 polymer brushes respectively, in a same way, as shown in Figure 1 (b and c). Structure and property characterization. 1H NMR measurements were carried out on a Bruker AV500 FT-NMR spectrometer (500.1 MHz). XPS analysis was performed by a multi-functional X-ray photoelectron spectrometer system (ESCALab250Xi, VG, England) equipped with a 200 W monochromatic Al Kα X-ray source. The 500 μm X-ray spot was used for XPS analysis and the take-off angle was equal to 90°. The background pressure in the analysis chamber was about 3×10-10 mbar. All XPS spectra were referenced to the aliphatic hydrocarbon component of the C 1s signal at 284.8 eV. The polymers were dissolved in DMF and were filtered with


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0.45 μm organic membrane aperture prior to analysis and deuterium oxide was used to prepare the sample solutions. The polymer molecular weight and polydispersity were

determined

using

a

Waters

Breeze

1515

GPC

analysis

system

(dimethylformamide, DMF). Spectroscopic ellipsometry (M-2000 DITM, J. A. Woollam) was used to measure the thickness changes during the process of ATRP initiator immobilization and polymer brushes modification. A continuous wavelength scan from 370.1 to 999.1 nm and incidence angles of 70° was used in the measurements. After the establishment of an optical constant of bare silicon surface, the thickness was calculated by a Cauchy model with a resumed refractive index of 1.45 for ATRP initiator-coated surfaces and 1.47 for polymer brushes.16-17 Based on the thickness and molecular weight of the polymer brushes, the grafting densities of polymer chains on surface were estimated via σ = hρNA/Mn.12, 18 Where, σ is the graft density of dry polymer layer, h is the layer thickness determined by ellipsometry, ρ is the density of dry polymer layer (0.93 g/cm3 for PASC1; 1.00 g/cm3 for PASC8 and PAGC8), NA is Avogadro’s number, and Mn is the number-average molecular weight of polymer chains on surfaces. Surface ζ potentials of polymer surfaces were measured with streaming current mode by SurPASS electrokinetic analyzer from Anton Paar. Two slides of silicon wafers with the polymer brushes in 1 × 2 cm were used and the measurement was conducted in 0.1 mmol KCl aqueous solution with pH = 7.4. The surface topography was characterized in aqueous solution through AFM method in Peakforce tapping mode (Nano IIIA, Bruker Instruments, USA). The images with area of 1 × 1 μm were scanned using silicon cantilever (Fastscan-B,


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Bruker Instruments, Germany) with a nominal spring constant of 4 N/m and at a scan rate of 1.0 Hz. The roughness of different polymer surfaces was evaluated by analysis software for the whole images. Water contact angles were obtained on an optical contact angle meter (Krüss DSA CA goniometer, Germany) with 3 μL of distilled water. S. aureus (ATCC 6538) and E. coli (MG1655) were selected, and both of them exhibited negative charges on cell membrane surfaces.19-20 Single colony of S. aureus and E. coli bacteria on a solid Luria−Bertani (LB) agar plate was transferred to 10 mL of liquid LB culture medium in the presence of 50 μg/mL ampicillin and cultured overnight at 37 °C with shaking. Bacteria were collected by centrifugation and washed by PBS three times, then subsequently suspended in PBS. The optical density (OD600) of the bacterial suspension was adjusted to 1.0 to get a concentration of 5×108 cells·mL−1 and subsequently was diluted 5-fold by PBS. The sterilized polymer surfaces were placed in a 24-well plate and incubated with 10 μL of bacterial suspension. After 30 min of incubation at 37 °C, bacterial suspensions were harvested and then serially diluted (10 000-fold) in PBS. Each dilution of 100 μL was plated in triplicate on a solid LB agar plate and incubated at 37 °C for 16 h. The number of colony forming units (CFU) at each plate was counted after incubation and the average CFU/ml was determined. Each test was carried out in three replicates. The killing efficiency (KE) was calculated by equation (1):14, 21 KE (%)  1-

the colony forming units of the sample  100% the colony forming units of the controlled sample

(1)

The polymer brush surfaces incubated with bacteria were gently washed with PBS and fixed with 0.1 vol% glutaraldehyde in PBS overnight at 4 °C. After


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dehydrating with a serial of ethanol/water mixtures (40%, 70%, 90%, and 100%) and coating with platinum, the surface morphology was observed through a S4800 scanning electron microscopy (SEM, Hitachi, Japan). RESULTS AND DISCUSSION Characterization of polymer brush surfaces. O 1s

N 1s

C 1s

Si-ATRP

Si 2p

C-C/C-H

Si-ATRP

C 1s

Si Surface

+

PASC1

Br 3d

Si-ATRP PASC1

C-N

+

PASC1

C-N /C-O C-C/C-H

O=C-O

Cl 2p

PASC8

+

C-N /C-O

PASC8

C-C/C-H

O=C-O

600

C-N

C-N +

C-N

PAGC8

PAGC8

700

C-N +

PASC8

(a)

N 1s

C-O

500 400 300 200 Binding Energy (eV)

100

(b)

0

PAGC8

+

C-N /C-O

O=C-O

C-C/C-H

(c)

C-N

295 290 285 280 410 405 400 395 Binding Energy (eV) Binding Energy (eV)

Figure 2. (a) Wide-scan XPS spectra for different surfaces; (b) C1s, (c) N1s spectra and the related peak-fitting curves for different surfaces. Compared with bare silicon surface, the atom transfer radical polymerization initiator-coated surface (denoted as Si-ATRP) displays a strong chlorine signal for PASC1 polymer brush and bromine signal for PASC8 and PAGC8 polymer brushes (Figure 2a). After polymerization, the intensity of Si2p and O1s peaks of polymer brushes decreases, while C1s signal increases compared with that of the bare silicon and Si-ATRP surfaces. Integrated area of N1s peak at 402.1 eV (C–N+), referring to the quaternary ammonium groups, increases to 5.55, 6.14, and 6.73% for PASC1, PASC8, and PAGC8 coated surface (Table S1), respectively. As shown in the high-resolution C 1s spectra (Figure 2b), Si-ATRP surface exhibits two predominant peaks centered at 284.6 and 286.3 eV, corresponding to C–C and C–O bonds,


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respectively.22 In the case of polymer brushes, C 1s peak can be decomposed into three contributions: the peak at 284.6 eV is attributed to C–C and C–H of the alkyl chain, and the one at 286.3 eV is attributed to C–N+ or C–O. The presence of a characteristic peak at 288.6 eV (O=C–O species) provides a supporting evidence that the polymer brushes are constructed.23-24 The primary peaks at 402.1 eV (C–N+) and the weak peaks at 399.3 eV (C–N) can be observed after the introduction of polymer brushes (Figure 2c).25 The above results confirm the presence of desired elements and chemical groups in the polymer brushes and indicate the successful immobilization on the silicon surfaces.

(a) 0.5

2

Grafting density, chains/nm Surface potentials, mV

0.4 0.3 0.2 0.1 0.0 Si

(c)

PASC1 PASC8 PAGC8 47.9 °

PASC1

10 nm

(b)

80 60 40 20 0 -20 -40 -60 -80

Si Surface

0 0

0.5

0.5

1 μm

(d) PASC8

58.5 °

10 nm

0

0 0.5

1 μm

Rq = 3.34 nm

Rq = 0.37 nm

(e)

63.8 °

PAGC8

10 nm 0

0

0.5

12.1°

10 nm

0

0.5

0.5

1 μm

Rq = 2.82 nm

0

0.5

0.5

1 μm

Rq = 1.75 nm

Figure 3. (a) Grafting density and surface potential of different surfaces; (b) AFM topography and contact angle of silicon surface; AFM topography, contact angle, and schematic diagram of polymer brush surfaces: (c) PASC1; (d) PASC8; (e) PAGC8. After obtaining the molecular weight and thickness of different polymer brushes (Table S2 and S3), the grafting density of polymer chains on surface is estimated, with a value of 0.27, 0.28 and 0.37 chains/nm2 for PASC1, PASC8, and PAGC8, respectively (Figure 3a and Table S3). Surface potential shifts from negative potential


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of bare silicon surface to positive potential of polymer surfaces, in which the value of ζ potential increases from ~43.7 mV for PASC1 to ~66.2 mV for PAGC8. The roughness value (Rq) of the series of polymer brush surfaces, is bigger than that of bare silicon surface, ranging from 3.34 to 1.75 nm, as shown in Figure 3(b~e). In addition, the contact angle for the polymer brush surfaces is higher than that of the bare silicon surface, increasing from ∼47.9° for PASC1 to ∼63.8° for PAGC8. Electrostatic repulsion between cationic head groups is proposed to make a major negative contribution to the grafting modification of PASC1 system, resulting in a lower grafting density and surface potential, as well as the most fluctuant and heterogeneous surface morphology among all three polymer brush surfaces. The hydrophobic chains of PASC8 polymers contribute to the hydrophobic interaction between two adjacent backbones and therefore weaken the electrostatic repulsion between cationic head groups. Such a self-adjustment process is benefit for the structure optimization, leading to a higher grafting density of polymer chains and a higher charge density of polymer surface, and eventually forming a relative smooth surface with a higher hydrophobicity than that in the case of PASC1. Notably, PAGC8 polymer brushes, with double cationic head groups and hydrophobic chains in every repeat unit, balance the electrostatic repulsion and the hydrophobic attraction and avoid the mutual interference among cationic head groups, resulting in a smooth and tight surface layer with a relatively high hydrophobic characteristic and, therefore, possessing the highest surface grafting and charge density among all the three polymer brush surfaces.


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Antibacterial properties of polymer brush surfaces.

(a)

(e)

(b)

(f)

PASC1

PASC8

(c)

(g)

PAGC8

(d)

(h)

(i)

Killing Percentage (%)

Si Surface

E.coil S.aureus

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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100 80

S. aureus E. coli

60 40 20 0

Si PASC1 PASC8 PAGC8

Figure 4. Colony forming units of S. aureus (a~d) and E. coli (e~h) after incubation on different surfaces for 30 min; (i) Antibacterial activity against S. aureus and E. coli of different surfaces. Symbol of “×” represents no antibacterial activity. The antibacterial activity of different surfaces against S. aureus and E. coli was evaluated by the surface plating method, as shown in Figure 4. Dense bacterial colonies are found on the bare silicon surfaces (Figure 4a, e) and sporadic ones are observed on the polymer surfaces (Figure 4b, c, f, g), and especially for PAGC8 polymer surfaces keeping empty and clean features (Figure 4d, h and Figure S3). Furthermore, the bare silicon surfaces present a non-antibacterial activity after calculation (Figure 4i) and exhibit a surface morphology with intact cell walls (Figure 5a, e). PASC1 surface kills ∼71% of S. aureus and ∼88% of E. coli, in which some bacteria go with collapsed and merged cell membranes (Figure 5b, f). For PASC8 surface, the killing efficiency (KE) increases to ∼92% and ∼93% for S. aureus and E. coli, respectively, inducing the leakage of cytoplasm and the death of pathogens (Figure 5c, g). Notably, KE of PAGC8 brush surfaces reaches to nearly 100% for both kinds of bacteria, in agreement with the result that almost all the bacteria on the surfaces are severely damaged and broken (Figure 5d, h).


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S.aureus

Si Surface

E.coil

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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PASC8

PASC1

PAGC8

(a)

(b)

(c)

(d)

(e)

(f)

(g)

(h) 3.0 μm

Figure 5. SEM images of S. aureus and E. coli on different surfaces: (a) and (e) silicon surfaces; (b) and (f) PASC1 polymer surfaces; (c) and (g) PASC8 polymer surfaces; (d) and (h) PAGC8 polymer surfaces. Yellow arrows indicate lesions and collapses of bacterial membrane. Antibacterial mechanisms of polymer brush surfaces.

Intact membrane

Destroyed membrane

Live E.coil Live S.aureus Dead Bacteria

PASC1

PASC8

PAGC8

Higher

Lower

Controllable antibacterial activity, charge density, and grafting density

Figure 6. Schematic illustration of antibacterial mechanisms of different polymer brush surfaces. PASC1, as a cationic polymer brush surface, presents a limited antibacterial activity owning to the fluctuant surface morphology and the electrostatic interaction


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between the quaternary ammonium salts of the polymer branches and the negatively charged cell wall of both kinds of bacteria (Figure 6). For PASC8, as an amphiphilic cationic polymer brush surface, an improvement of killing efficiency can be obtained due to the synergistic effect of electrostatic and hydrophobic interactions on a relative smooth and tight polymer surface. The amphiphilic polymer brushes not only interact with the cell membrane surface of bacteria via electrostatic attraction, but also insert their hydrophobic chains into the bacterial cell membranes, resulting in the disruption of the lipid bilayer and the damage of bacteria. In the case of PAGC8, the geminized amphiphilic cationic polymer brush surface has an ultrahigh antibacterial activity for both kinds of bacteria, attributed to the enhanced synergistic effect of electrostatic and hydrophobic interactions, as well as the extremely smooth and tight polymer surface in favor of minimizing the contact area between bacteria and solid surface. Self-cleaning property of PAGC8 polymer brush surface. PAGC8

(a1)

(c1)

First Hydrolysis

(a2)

(c2)

Second Hydrolysis

(a3)

(c3)

Killing Percentage (%)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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120

E. coli

Hydrolysis

100 80 60 40 20

(b)

0

PAGC8 First

Second

Figure 7. (a1~a3) Colony forming units; (b) antibacterial activity; (c1~c3) SEM mages of E. coli after incubation on the hydrolyzed PAGC8 polymer brush surface. The self-renewed and sustained antibacterial ability of PAGC8 polymer brush surface was performed via hydrolysis in the presence of 0.6 mol L-1 NaOH solution at 37 °C for 30 min. Compared with the PAGC8 polymer surface with a superhigh


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antibacterial activity (Figure 7a1, b, c1), the degraded polymer surface provides sustained bactericidal ability after the first hydrolysis. There are still few bacterial colonies attached to the polymer surface (Figure 7a2), showing an antimicrobial activity of ~100% and nearly all of cell membranes being damaged (Figure 7c2). The surface competency for antibacterial ability maintain after partial degradation, due to the high coverage of quaternary ammonium salt groups and hydrophobic tails of polymer brushes.26-27 Even after the second hydrolysis, the antibacterial activity of the polymer brush surface still reach up to above 90% (Figure 7a3), and bacterial cells on the hydrolyzed polymer surface are severely collapsed as well (Figure 7c3), demonstrating the excellent sustained antibacterial ability of PAGC8 polymer brush surface. The content of C and N of the brushes decrease after hydrolysis compared with that of the PAGC8 polymer surface before hydrolysis (Table S4). In addition, the thickness of the polymer brush decreases from 39.6 nm to 34.3 nm and 27.1 nm, and the contact angle decreases from ~63.8° to ~36.3° and ~31.9° after the first and second hydrolysis. The results indicate that the side chains of polymer brushes were partially taken off, bringing in the slight collapse of the polymer brush and exposure of carboxylic groups on the surface (Figure 8). Therefore, most of dead bacteria attached to the surfaces can be released via hydrolysis and the high antibacterial ability can be retained by the fresh polymer surface, showing a promising material in combating challenging pathogenic infection.


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O O

Br

n

Br

(a)

n

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N

O OH

N O

O

O

O

HO

Hydrolysis Si

Si

O O O

O O O

N N

O C O

Si-PAGC8

Si-PAGC8

N N

O C OH

(b) NaOH 30 min

NaOH 30 min

First hydrolysis

PAGC8

(c)

NaOH 30 min

PAGC8

Second hydrolysis NaOH 30 min

First hydrolysis

Second hydrolysis

Figure 8. (a) Hydrolysis routes of PAGC8 polymer brush; (b) schematic illustration of hydrolysis process of PAGC8 polymer brush; (c) sustained antibacterial process of PAGC8 polymer brush. CONCLUSION A novel kind of ultrahigh charged amphiphilic polymer brush surface with super-antibacterial and self-cleaning capabilities has been developed in this study. Such polymer surface displays a killing efficiency of about 100% against both S. aureus and E. coli bacteria, on the one hand attributed to the contribution of ultrahigh charge density and hydrophobicity to the structure adjustment, and resulting in a smooth and tight surface and a small contact area with bacteria. On the other hand, the electrostatic and hydrophobic interactions between the geminized amphiphilic polymer brushes and the bacterial cell membranes are significantly strengthened with the increase of surface charge density and hydrophobicity. Moreover, the superhigh


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antibacterial polymer surface can reduce the formation of a biofilm on the surface via hydrolysis and degradation, and further kill bacterial cells effectively. Therefore, we demonstrate that the geminized amphiphilic cationic polymer brushes can be a promising candidate material with super-antibacterial and self-cleaning capabilities in a wide range of biomedical and industrial applications. ASSOCIATED CONTENT Supporting Information Figure S1 showing the synthetic process of AGC8 monomers as well as 1H NMR spectra of ASC1, ASC8, and AGC8 monomers, Figure S2 showing the contact angles of bare silicon surface and ATRP initiator-coated surface, Figure S3 showing the antibacterial activity of different surfaces against S. aureus and E. coli obtained from Confocal laser scanning microscopy (CLSM) images, Table S1 exhibiting the component analysis of polymer brushes calculated through XPS, Table S2 exhibiting the polydispersity and molecular weights of different polymers measured by GPC, Table S3 exhibiting the thickness, grafting density and surface potential of different polymer brush surfaces, and Table S4 exhibiting the surface composition, thickness and contact angle of PAGC8 polymer brush after hydrolysis. AUTHOR INFORMATION Corresponding Author *E-mail: [email protected] (H.Y.). Notes The authors declare no competing financial interest.


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ACKNOWLEDGMENTS This work was funded by the National Natural Science Foundation of China (21872152 and 21603240), the Important National Science and Technology Specific Project of China (2017ZX05013-003 and 2016ZX05025-003-009), and the Strategic Priority Research Program of CAS (XDB22030102). We thank Prof. Shu Wang from Institute of Chemistry, Chinese Academy of Sciences, Beijing, China, for the help in antibacterial tests. We thank Ms Xiaoyu Zhang and Ms Zhijuan Zhao from Institute of Chemistry, Chinese Academy of Sciences, Beijing, China, for the help in XPS measurements. We thank Ms Yeping Li from Anton Paar (Shanghai) Trading Co., Ltd for the help in surface potential measurements. We thank Dr Zhenwen Huang from Bruker (Beijing) Technology Co., Ltd for the help in AFM morphology measurements in aqueous solution. REFERENCES 1.

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TOC

Super-antibacterial and self-cleaning surface


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Ultrahighly Charged Amphiphilic Polymer Brushes with Super

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