Elastomer

Apr 13, 2018 - The polymer brush structure at the water interface was investigated using ... Highly Stretchable and Instantly Recoverable Slide-Ring G...
0 downloads 0 Views 565KB Size
Download PDF
Subscriber access provided by UNIV OF DURHAM

Interface Components: Nanoparticles, Colloids, Emulsions, Surfactants, Proteins, Polymers

Polyrotaxane brushes dynamically formed at a water/elastomer interface Kanta Yanagi, Norifumi L Yamada, Kazuaki Kato, Kohzo Ito, and Hideaki Yokoyama Langmuir, Just Accepted Manuscript • DOI: 10.1021/acs.langmuir.8b00649 • Publication Date (Web): 13 Apr 2018 Downloaded from http://pubs.acs.org on April 15, 2018

Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.

is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

Page 1 of 7 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

Langmuir

Polyrotaxane brushes dynamically formed at a water/elastomer interface Kanta Yanagi,† Norifumi L. Yamada,‡ Kazuaki Kato,† Kohzo Ito,† and Hideaki Yokoyama*,† † ‡

Graduate School of Frontier Sciences, The University of Tokyo, Chiba 277-8561, Japan High Energy Accelerator Research Organization, Ibaraki 319-1108, Japan

ABSTRACT: Dense polymer brushes with closely packed rotaxane structures were formed at the interface of water and a styrenebutadiene elastomer by spontaneous segregation of an amphiphilic polyrotaxane (PR), a mechanically interlocked polymer consisting of hydrophobic polybutadiene threading through multiple hydrophilic γ-cyclodextrin (γ-CD) derivatives. Segregation of PR at the water/elastomer interface was suggested by X-ray photoelectron spectroscopy. The polymer brush structure at the water interface was investigated using neutron reflectometry. Brush structures were found to depend on the number of CDs on the PRs; the PR with a small number of CDs formed a thinner and homogeneous brush, whereas the PR with a higher number of CDs formed a thicker and less ordered brush. These PR-brushes showed protein repulsion, resulting from the surface-hydrated brush layer preventing direct contact of proteins.

1. INTRODUCTION End-grafted polymer chains at interfaces are called polymer brushes.1 They exhibit unique properties such as anti-fouling behavior,2 colloid stabilization,3 and lubrication4 owing to steric hindrance of the dense polymer chains. Polymer brushes have mainly been fabricated by two methods.5,6 In the “grafting-to” method, polymer chains are grafted onto the interface either chemically or physically. In the “grafting-from” method, polymers are chemically synthesized from initiators present on the surfaces. Recently, our group reported a novel, easy method for fabricating polymer brushes.7–9 This third method utilizes spontaneous segregation of amphiphilic diblock copolymers at the water/elastomer interface from the elastomer side to form a so-called “dynamic polymer brush.” When a mixture of the hydrophobic polydimethylsiloxane (PDMS) elastomer and diblock copolymers consisting of PDMS and hydrophilic polyethyleneglycol (PEG) comes into contact with water, it leads to the spontaneous formation of dense, highly stretched PEG polymer brushes at the water/elastomer interface.7 In this system, the hydration energy of the hydrophilic PEG block is the driving force, and the hydrophobic PDMS block anchors the PEG brush to the elastomer surface. Dynamic polymer brushes can be generally regarded as an interfacial segregated layer of amphiphilic copolymers containing both hydrophobic and hydrophilic blocks. Amphiphilic polyrotaxanes (PRs) with hydrophobic linear chains and hydrophilic cyclic molecules are another type of amphiphilic copolymer candidates for surface segregation. PRs are composed of an axis polymer that threads through multiple cyclic molecules trapped by bulky end groups.10 Since the cyclic and linear molecules are not chemically connected but simply topologically constrained, the cyclic molecules can slide along the axis and rotate. Therefore, the cyclic molecules can either be dispersed randomly along the axis chain (similar to random copolymers) or aggregated in blocks (similar to block copoly-

mers) as shown in Figure 1. When amphiphilic PRs are used instead of block copolymers in the dynamic polymer brush architecture, unique PR brushes with cyclic molecules exposing themselves to water are expected to be formed (see upper part of Figure 4). Although many researchers have investigated potential applications of surface-immobilized rotaxanes as molecular devices11, there have been few studies on surface-immobilized PRs. Our method is a novel approach for realizing PR brushes on a surface with self-healing property, in which, if the brush is somehow lost, the remaining PRs embedded in the film can segregate to repair. Such self-repairing is advantageous compared to other conventional methods such as grafting-to brushes12–14 and inclusion complex brushes15,16 of grafted polymers and CDs. In this study, we employed amphiphilic polyrotaxane with hydrophilic rings and hydrophobic chains. The ring molecules in polyrotaxane can move along the linear polybutadiene and their distribution can be random or blocky and dynamically change. The interface between water and hydrophobic elastomer drives intramolecular phase separation of ring molecules within a polyrotaxane molecule to be blocky polyrotaxane, and such blocky polyrotaxane segregates to the water interface to form dynamic polymer brush. Furthermore, the effect of PR architecture on brush structure as well as its protein repulsion property was investigated.

Figure 1. Possible structures of a polybutadiene-based PR. The left and right images represent random copolymer-like PR and block copolymer-like PR. Two cyclodextrins were used as the end cappers.

ACS Paragon Plus Environment

Langmuir 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

2. EXPERIMENTAL SECTION 2.1 Materials. 2-(2-Methoxyethoxy)acetyl chloride was purchased from Alfa Aesar. Trimethylamine was obtained from Tokyo Chemical Industry Co., Ltd. Methoxy acetyl chloride, lithium chloride (LiCl), lithium bromide (LiBr), dimethyl sulfoxide (DMSO), toluene, chloroform, pyridine, N,Ndimethylformamide (DMF, super dehydrated), lysozyme from egg white, and phosphate buffer saline 10X (pH = 7.4) (PBS) were obtained from Wako Pure Chemical Industries, Ltd. Poly(styrene-b-butadiene-b-styrene), styrene 30 wt% (SBS; based on the information by the supplier Mw ~140,000), bovine serum albumin (BSA), and fibrinogen from bovine plasma were obtained from Sigma-Aldrich Co. LLC. Deuterium oxide (99.8 atom % D) was obtained from Kanto Chemical Co., Inc. All reagents were used without further purification. Two types of polybutadiene-based PRs with different coverage were prepared following previously published methods.17 Following this method, we obtained PRs whose both ends were capped with γ-cyclodextrins (γ-CDs). Note that only γ-CDs can form a complex with polybutadiene with 1,2-additional units. Molecular weight (Mn) of the axis polymers used for both PRs was 4200 and it contained about 20% of 1,2-addition units. According to the 1H NMR spectra (see Figure S3 in the Supporting Information), a single PR with lower coverage included about seven CDs in total (two at the ends and five on its axis) with a Mn of ~14000 and it was denoted as PR07. The coverage of the polybutadiene-based PR was calculated as defined previously18 and found to be about 11%. The other single PR with a higher coverage included about fifteen CDs in total (two at the ends and thirteen on its axis) with a Mn of ~23000 and it was denoted as PR15. Its coverage was 27%. 1H NMR spectra were measured using a JNM-AL400 spectrometer (JEOL Ltd.). Attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectra were recorded on a Nicolet 4700 (Thermo Electron Co., Ltd.) spectrometer. Gel permeation chromatography (GPC) was performed with DMSO/LiBr (10 mM LiBr) as the eluent on a Prominence UFLC (Shimadzu Corporation) using PEG standards for calibration. 2.2 Modification of PR07. PR07 (50 mg, 3.6×10–6 mol, previously dried in vacuum) and LiCl (60 mg, 1.4 mmol) were dissolved in dry DMF (2.8 mL) under argon and then trimethylamine (0.20 mL, 1.4 mmol) was added. This solution was cooled in an ice bath, followed by the dropwise addition of 2(2-methoxyethoxy)acetyl chloride (0.30 mL, 2.0 mmol). The resulting mixture was stirred overnight at 25 °C. The product was purified by dialysis using a cellulose membrane (whose molecular weight cut-off was 12000–14000) for 3 d followed by freeze-drying to yield the modified PR07 as a white powder (yield = 67 mg). As original PR07 was not used for study, the modified PR07 has been simply denoted as PR07 in this article. 2.3 Modification of PR15. PR15 (80 mg, 3.5×10–6 mol, previously dried in vacuum) and LiCl (80 mg, 1.9 mmol) were dissolved in dry DMF (4.0 mL) under argon and then trimethylamine (0.17 mL, 1.2 mmol) was added. This solution was cooled in an ice bath, followed by the dropwise addition of methoxyacetyl chloride (0.078 mL, 0.86 mmol). The resulting mixture was stirred at 20 °C overnight. The product was purified by dialysis using a cellulose membrane (whose molecular weight cut-off was 12000–14000) for 2 d followed by freezedrying to yield the modified PR15 as a white powder (106 mg). As original PR15 was not used in other parts of this paper, the

Page 2 of 7

modified PR15 has been simply denoted as PR15 in this article. For PR 15, methoxyacetyl chloride was used to modify the CDs instead of 2-(2-methoxyethoxy)acetyl chloride for PR07. By using those modifiers, the modification degrees of CDs (54% for PR07 and 51% for PR15) were maintained the similar values. 2.4 Sample preparation. SBS and 20 wt% of PR07 or PR15, were dissolved in chloroform (for PR07) or pyridine (for PR15). The total polymer concentration of the solution was 1 wt% (PR07) or 5 wt% (PR15). The solution was passed through a 0.45 µm filter and spin-coated at 2000 rpm onto silicon or quartz substrates. These films were annealed at 150 °C for 12 h under vacuum. Pure SBS films were also prepared as control samples from a 2 wt% toluene solution of SBS following the same procedure. Film thicknesses were estimated by ellipsometry (JASCO, M-150) and found to range from 200–500 nm. For the X-ray photoelectron spectroscopy measurements discussed in section 3.2, sample films were immersed in distilled water for 24 h at 25 °C and then dried overnight in vacuum. 2.5 X-ray photoelectron spectroscopy (XPS). Surface composition analysis in vacuum was investigated by XPS (JEOL, JPS-9000) using Al Kα as the X-ray source. Before the measurement, samples were dried overnight in vacuum. The take-off angle was 90° from the surface and the peak positions of the alkyl carbons were set to 285.0 eV as reference. After the background was subtracted from the raw data by the Shirley method, peak area was calculated, followed by conversion to atomic number fraction using the equipped software. 2.6 Neutron reflectometry (NR). NR experiments were conducted with the Soft Interface Analyzer (SOFIA)19,20 at JPARC, Ibaraki, Japan. Specular neutron reflectivity from the polymer/D2O interface was measured at 25 °C for more than 24 h after being in contact with water. We confirmed that the surface reconstruction was complete within 24 h by Quartz crystal microbalance (data are not shown). The scattering length density (SLD) depth profiles were computed by fitting the reflectivity curves using the Motofit package.21 The curves were fitted with a multilayer model consisting of a quartz substrate, SBS matrix film, swollen D2O brush layer, and ambient D2O. The SLDs of quartz, SBS matrix, PR brush, and D2O were assumed to be 4.2, 0.68, 1.9 (PR07), 2.1 (PR15), and 6.36×10–4 nm–2, respectively. Note that the hydrogendeuterium exchange at hydroxyl protons of the PR-brush is expected to occur in ambient D2O. The polymer layers (matrix and brush) were divided into some sublayers in order for the reflectivity to be fitted with the simplest model; a single layer for SBS film, two layers for the PR15-blended film, and twenty-nine layers for the PR07-blended film. In the SBS and PR15-blended films, the thickness of each layer and SLD of the brush layer were used as fitting variables. In the PR07blended film, the SLDs of the brush layers were fixed from 0.7–6.3×10–4 nm–2 in ascending order with 0.1–0.3×10–4 nm–2 increments and the thicknesses of these layers were used as fitting variables. Their values were optimized to obtain the SLD profiles around the interfaces between the film and D2O. Resolutions were set as 5% for SBS and 20% for PR-blended films. 2.6 Protein adsorption. Fibrinogen, BSA, and lysozyme having molecular weights of 170, 66, and 14 kDa, respectively, were dissolved in PBS with concentrations of 1, 4, and 5 mg mL–1, respectively. Sample films were immersed in distilled

ACS Paragon Plus Environment

Page 3 of 7 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

Langmuir water for 24 h for brush formation before they were immersed in the filtered (0.45 µm) protein solution for 24 h. After rinsing thrice with PBS for 15 min to remove the non-adsorbed and loosely adsorbed proteins, the films were rinsed with distilled water to remove PBS. Finally, the sample films were dried in vacuum overnight for XPS measurement. Experiments were carried out at 25 °C.

3. RESULTS AND DISCUSSION 3.1 Polybutadiene-based PR derivatives. Characteristics of modified PRs are summarized in Table 1. 1H NMR spectra, GPC traces, and IR spectra are shown in the Supporting Information. Two types of PRs with different number of CDs were prepared and modified with amphiphilic side chains to decrease the number of hydrogen bonds of CDs. These strong intermolecular hydrogen bonds typically lead to the aggregation of PRs in an elastomer, which may prevent brush formation. However, the hydroxyl groups are also important for brush formation because they lead to a large hydration energy gain, which is a driving force of brush formation in the dynamic polymer brush architecture. Therefore, an appropriate modification is required whereby the number of hydrogen bonds is reduced while still maintaining the hydrophilicity of CDs. After the modification, the reduction in the number of hydrogen bonds was confirmed by the shift of the OH vibration peak in the IR spectra and the hydrophilicity of CDs was confirmed by brush formation (discussed in section 3.3). Note that the SBS elastomer was used as a matrix to provide affinity for the polybutadiene-based PR and enough molecular mobility at room temperature. 3.2 Segregation of PR. To reveal the surface compositions of the films under a hydrophobic environment, XPS measurements were conducted. In the C1s spectra shown in Figure 2, large alkyl carbon (C–C, C=C) peaks, which mainly originated from the SBS matrix, were observed at ~285 eV in all samples. On comparing the spectra measured before (dashed line) and after (solid line) sinking in water, the peak for ether carbon (C–O) at ~287 eV became slightly larger in the PR07blended film and significantly larger in the PR15-blended film. Furthermore, there was a clear increase in the ester carbon (CO–O) peak at ~289 eV, while no difference was observed in the SBS film upon water contact. The hydrophobic SBS elastomer does not show any surface rearrangement on water sinking. On the other hand, in PR-blended films, the PRs existed slightly at the outermost surface even before the contact was made with water and increased further after sinking in water. This was because the C–O and CO–O photoelectrons could have only originated from the modified CDs on the PRs. This surface enrichment can be attributed to the brush formation in water and segregated PRs that somewhat remained at surface after drying; surface segregation of PRs is an indirect proof of the formation of PR-brushes. Because CDs with many hydroxyl groups generally increase the surface energy if they appear on the surface, they have a tendency to be embedded in the bulk. However, this is not the case for the CD derivatives with about half of their hydroxyl groups modified by methyl groups, which have a lower surface energy.22–24 When the brushes were dried, the segregated PRs could interact with one another on the surface and fail to bury themselves in the matrix again. In this situation, these CDs can cover the surface in vacuum because of the low surface energy, although

they seem to be hydrophilic enough to segregate on the water interface. 3.3 Structures of PR-brushes. In order to reveal the interfacial structure in water, neutron reflectivity was measured at the polymer/D2O interface. Note that D2O was used to enhance the contrast for neutron reflectometry. The reflectivity curves and SLD depth profiles are shown in Figure 3. The SLD profiles are primarily dominated by the depth profile of the D2O

Figure 2. C1s spectra of SBS and PR-blended (20 wt%) films before and after sinking in water for 24 h. In addition to large C–C peaks, C–O and CO–O peaks were confirmed after water treatment. The spectra were shifted vertically for clarity. volume fraction. The D2O volume fraction approaches from 0 to 1 as the SLD increases from the value of SBS (0.68×10–4 nm–2) to the value of D2O (6.36×10–4 nm–2). The interfacial structure of the SBS film does not show any noticeable characteristics except the fringes from the total film thickness as expected. The SLD changes sharply from the value of SBS to that of D2O (Figure 3b), indicating a flat interface (Figure 4c). Unlike the SBS film, the PR07-blended film showed a characteristic broad fringe with a minimum around q = 0.4 nm–1, which indicated the existence of an additional thin layer structure. From the result of fitting with a multilayer model, it was suggested that there is a D2Oenriched swollen polymer layer whose SLD is 2.3×10–4 nm–2 with a thickness of 6.6 nm at the interface (Figure 3b). The PR15-blended film also showed a broader fringe in the whole q-range, indicating the presence of additional thin layers. Based on fitting, it was suggested that there are D2O-swollen polymer layers with ~20 nm thickness and height distribution at the interface (Figure 3b). Considering that the XPS results indicate that the PRs segregate at the water/SBS interface, these D2O-swollen polymer layers are most likely segregated PR-brushes. In the “dynamic polymer brush” system, segregated diblock copolymers form a highly stretched dense polymer brush layer at the interface.7–9 Unlike the simple diblock copolymer, PR is expected to have several different possible interfacial structures owing to the availability of multiple CD locations along the polymer chain backbone. Among these, there are two important structures; (i) CDs aggregate to form a block at only one end of the PR to form a single brush (I-shape, Figure 4a) and (ii) CDs aggregate to form blocks at both ends of the PR, forming two brushes per molecule (U-shape, Figure 4b). The

ACS Paragon Plus Environment

Langmuir 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

I- and U-shapes are considered as the segregated structures of the diblock and triblock copolymers, respectively. The estimated interfacial structures based on NR data and molecular architecture of PRs are shown in Figure 4c–e. Brush structures are considered as follows. In PR07 the height of a single modified CD was estimated to be 1–1.5 nm in terms of the chemical structure and molecular volume, but due to the flexibility of side chains, the height of the modified CD could not be determined unambiguously. Hence, a 6.6 nmthick PR-brush would be described by 4–6 vertically stacked CDs in a single PR. From the SLD prolife, the volume fraction of PR in

ent. Brush structures can be described by a mixture of I and Ushapes. The maximum height is ~ 20 nm, which agrees with the height of ~15 modified CDs. This I-shape brush comprised only half of the brush. The thinner ~10 nm-thick layer can be considered as a U-shape; a single U-shaped PR forms two brush chains with half of the maximum brush height (~10 nm) (Figure 4e). From the SLD profile, the surface brush coverage was calculated to be 0.67. This density does not take the Ushape into account. The U-type brush formed by a single PR covers twice as large an interfacial area as that covered by the I-type brush. If this correction is taken into account, the surface coverage of PR15 becomes a considerably larger value of 0.77. The different characteristics of the brushes are summarized in Table 2. Comparing the brush structures of PR07 and PR15, PR15 with more CDs forms a thicker and less-ordered (lower surface coverage) brush structure. A thicker brush is natural because the length of the hydrophilic segment (number of CDs) is longer in PR15. However, when more CDs aggregate in the I-shape, the system has to pay a larger entropic penalty. Therefore, although the hydration energy becomes larger in the I-shape as described in brush structure of PR07, not all brushes adopt the I-shape in order to reduce the unfavorable entropic loss. In addition, when the PR chain is sufficiently long, an extended PR crystal is not likely to be observed, but a 10 nm-thick folded crystal of CDs is observed. Recently, it was reported that a 10 nm-thick well-ordered folded rotaxane structure was formed on the Au surface after the formation of an inclusion complex between the grafted PEG brushes and α–CDs regardless of the graft density.15 This thickness seems to be a characteristic stable persistent length of PR. There is a possibility that the metastable U-shape was formed and remained as well as the stable I-shape. Note that an upside-down U-shape brush with ~10 nm thick is also possible.

Figure 3. (a) Neutron reflectivity curves and (b) SLD depth profiles near the SBS/D2O interface. A D2O-swollen PR layer exists in SBS with PR. The reflectivity curves are shifted for clarity. the brush layer was calculated to be ~0.9.Therefore the surface brush coverage, which is defined by graft density/hexagonally close-packed maximum CD density, was calculated to be ~1.0, respectively (method of calculating the surface brush coverage is explained in the Supporting Information). In order to realize this situation, the PRs should be closely packed both vertically and in plane. This suggests that the number of CDs exposed to water ought to be maximized in order to obtain hydration energy gain of CDs, which is a driving force for brush formation in the dynamic polymer brush. Therefore, considering that each PR has 5 CDs on the axis in addition to the end-capping CDs, the brush structure of PR07 is comprised of ~6 vertically stacked CDs in a single PR to form a single thin PR-brush, which covered the surface almost entirely (Figure 4d). In the PR15-blended film, a height distribution was observed for the brush layer, indicating that the height (number of vertically stacked CDs) of the PR-brushes were different. The layer whose SLD was 6.0×10–4 nm–2 was not included in the brush layer because this layer is very similar to D2O ambi-

ACS Paragon Plus Environment

Page 4 of 7

Page 5 of 7 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

Langmuir Figure 4. Possible PR-brush structures similar to the (a) diblock copolymer (I-shape) and (b) triblock copolymer (Ushape). Estimated interfacial structures of the (c) SBS, (d) PR07-blended, and (e) PR15-blended films. Table 2. Characteristics of PR-brushes. Sample

Height / nm

Surface coveragea

PR07

6.6

1.0

PR15

10 and 21

0.67

a Defined as graft density/hexagonally close-packed maximum CD density. 3.4 Protein adsorption. In order to investigate the fundamental brush properties, protein adsorption tests were carried out. The adsorbed amount was roughly evaluated by comparing the N atomic concentrations of the sample surfaces (Figure 5) because the N signals originated solely from the adsorbed proteins, as confirmed by using control samples (see Figure S6 in Supporting Information). N concentrations were calculated from the peak areas of the C1s and N1s spectra. Compared with the SBS film, the PRblended films showed a reduction in the adsorption against all proteins. These results could be attributed to the PR-brushes at the water/elastomer interface. A number of mechanisms for the protein repellency of polymer brushes such as excluded volume,25 high mobility,25,26 structured (mediated) water,27,28 have been proposed and in this case, the following mechanism operates. The PR-brushes form a hydrated layer at the elastomer surface and this layer has a large excluded volume, which reduces the direct contact of the matrix surface with proteins. Moreover, in order for the proteins to be adsorbed on the surface, hydrated water must be released and brushes must be highly deformed, which are energetically unfavorable processes. The protein repellency of the PR-blended surface with other chemical structures of PRs have also been reported in other studies,14,29,30 but the interfacial structure was not revealed. The most significant difference was observed in the adsorption of fibrinogen. Similar behavior was observed in the graft PR system. Inoue et al.14 have reported the different protein repellency behaviors of a hydrophobic surface and hydrophilic PR grafted surface against albumin and fibrinogen. Fibrinogen showed a larger difference of adsorption on surfaces than albumin owing to the lesser intermolecular interactions of albumin and the coagulation ability of fibrinogen. Adsorption results were also explained by the size difference of the proteins.31,32 In the case of small proteins, adsorption occurs even in a tiny unprotected area. However, for large proteins, adsorption is effectively prevented by the smaller areal coverage of the brush. In our study, BSA and Lysozyme showed only weak adsorption onto bare SBS and hence the largest difference was observed for fibrinogen. PR07 with a distinct brush structure and higher surface coverage showed the best anti-fouling property. The importance of surface coverage (graft density) has also been suggested by other researchers.25,31–34

Figure 5. Calculated N concentrations at sample surfaces after protein adsorption. PR-blended films showed protein repulsion against all proteins having different MW (MW = 14, 66, and 170 kDa for lysozyme, BSA, and fibrinogen, respectively).

4. CONCLUSIONS Dense PR-brushes were successfully fabricated at a water/elastomer interface by spontaneous surface segregation of amphiphilic PRs, which consist of hydrophilic γ-CD derivatives and hydrophobic polybutadiene backbones. PRs with different number of CDs formed different brush structures; PRs with a larger number (~15) of CDs formed ~20 nm-thick brushes with inhomogeneous heights, including folded Ushaped brushes. On the other hand, PRs with fewer number (~7) of CDs formed ~7 nm-thick all extended I-shaped brushes with homogeneous heights. These PR-blended films showed significantly diminished adsorption of a variety of proteins. This indicated that PRs with fewer CDs formed dense extended brushes with higher surface coverage that prevented the adsorption of proteins most effectively.

ASSOCIATED CONTENT Supporting Information Characterization figures (GPC traces, ATR-FTIR spectra and 1H NMR spectra), calculation of surface brush coverage, XPS spectra of control samples for protein adsorption. This material is available free of charge via the Internet at http://pubs.acs.org.

AUTHOR INFORMATION Corresponding Author * (H.Y.) E-mail: [email protected]

Author Contributions The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript.

Funding Sources This work was supported by JSPS KAKENHI Grant Number 15H03862. This work was partially supported by ImPACT Program of Council for Science, Technology and Innovation (Cabinet Office, Government of Japan).

Notes The authors declare no competing financial interests.

ACS Paragon Plus Environment

Langmuir 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

ACKNOWLEDGMENT The neutron reflectometry experiments were performed using SOFIA at J-PARC with support from the S-type (2009S08, 2014S08) and general use (2015A0253) research project of KEK.

REFERENCES (1) Milner, S. T. Polymer Brushes. Science 1991, 251(4996), 905– 914. (2) Higaki, Y.; Kobayashi, M.; Murakami, D.; Takahara, A. Antifouling behavior of polymer brush immobilized surfaces. Polym. J. 2016, 48(4), 325–331. (3) Vincent, B. Effect of adsorbed polymers on dispersion stability. Adv. Colloid Interface Sci. 1974, 4(2–3), 193–277. (4) Klein, J.; Kumacheva, E.; Mahalu, D.; Perahla, D.; Fetters, L. J. Reduction of frictional forces between solid surfaces bearing polymer brushes. Nature 1994, 370 (6491), 634–636. (5) Zhao, B.; Brittain, W. J. Polymer brushes: surface-immobilized macromolecules. Prog. Polym. Sci. 2000, 25(5), 677–710. (6) Edmondson, S.; Osborne, V. L.; Huck, W. T. S. Polymer brushes via surface-initiated polymerizations. Chem. Soc. Rev. 2004, 33(1), 14–22. (7) Inutsuka, M.; Yamada, N. L.; Ito, K.; Yokoyama, H. High Density Polymer Brush Spontaneously formed by the Segregation of Amphiphilic Diblock Copolymers to the Polymer/Water Interface. ACS Macro Lett. 2013, 2(3), 265–268. (8) Tanoue, H.; Inutsuka, M.; Yamada, N. L.; Ito, K.; Yokoyama, H. Kinetics of Dynamic Polymer Brush Formation. Macromolecules, 2017, 50, 5549–5555. (9) Tanoue, H.; Inoue, K.; Yamada, N. L.; Ito, K.; Miyao, S.; Ishizne, T.; Yokoyama, H. Thermo-responsive Dynamic Polymer Brush Fabricated by Segregation of Amphiphilic Diblock Copolymers. Submitted. (10) Ito, K.; Kato, K.; Mayumi, K. Polyrotaxane and Slide-Ring Materials; The Royal Society of Chemistry: Cambridge, 2016; pp 1. (11) Coronado, E.; Gavina, P.; Tatay, S. Catenanes and threaded systems: from solution to surfaces. Chem. Soc. Rev. 2009, 38(6), 1674–1689. (12) Yang, D. H.; Katoono, R.; Yamaguchi, J.; Miura, Y.; Yui, N. Immobilization of Polyrotaxane on a Solid Substrate as the Design of Dynamic Surface. Polym. J. 2009, 41(11), 952–953. (13) Yamada, K.; Katoono, R.; Yui, N. Controlled loop and graft formations of water-soluble polymers on SAMs for the design of biomaterials surfaces. Polym. J. 2012, 44(3), 286–293. (14) Inoue, Y.; Ye, L.; Ishihara, K.; Yui, N. Preparation and surface properties of polyrotaxane-containing tri-block copolymers as a design for dynamic biomaterials surfaces. Colloids Surf. B. 2012, 89, 223–227. (15) Takahashi, S.; Yamada, L. N.; Ito, K.; Yokoyama, H. Inclusion Complex of α–Cyclodextrin with Poly(ethylene glycol) Brush. Macromolecules 2016, 49(18), 6947–6952. (16) Tawa, K.; Kuboyama, N.; Ahmed, S. a.; Tanaka, M.; Nakaoki, T. Sensitive Detection of a Pseudo-Polyrotaxane Ultrathin Film by SPR and QCM-D Methods. Sens. Actuators, B 2009, 138(1), 126−133. (17) Kato, K.; Komatsu, H.; Ito, K. A Versatile Synthesis of Diverse Polyrotaxanes with a Dual Role of Cyclodextrin as both the Cyclic and Capping Components. Macromolecules 2010, 43(21), 8799–8804. (18) Michishita, T.; Okada, M.; Harada, A. Complex formation of polybutadiene with cyclodextrins. Macromol. Rapid Commun. 2001, 22(10), 763–767.

(19) Mitamura, K.; Yamada, N. L.; Sagehashi, H.; Torikai, N.; Arita, H.; Terada, M.; Kobayashi, M.; Sato, S.; Seto, H.; Goko, S.; Furusaka, M.; Oda, T.; Hino, M.; Jinnai, H.; Takahara, A. Novel neutron reflectometer SOFIA at J-PARC/MLF for in-situ soft-interface characterization. Polym. J. 2013, 45(1), 100–108. (20) Yamada, N.L.; Torikai, N.; Mitamura, K.; Sagehashi, H.; Sato, S.; Seto, H.; Sugita, T.; Goko, S.; Furusaka, M.; Oda, T.; Hino, M.; Fujiwara, T.; Takahashi, H.; Takahara, A. Design and performance of horizontal-type neutron reflectometer SOFIA at J-PARC/MLF. Eur. Phys. J. Plus 2011, 126, 108. (21) Nelson, A. Co-Refinement of Multiple-Contrast neutron/XRay Reflectivity Data Using MOTOFIT. J. Appl. Crystallogr. 2006, 39 (2), 273−276. (22) Zhang, R.; Seki, A.; Ishizone, T.; Yokoyama, H. Reduced Hydrophobic Interaction of Polystyrene Surfaces by Spontaneous Segregation of Block Copolymers with Oligo (Ethylene Glycol) Methyl Ether Methacrylate Blocks: Force Measurements in Water Using Atomic Force Microscope with Hydrophobic Probe. Langmuir 2008, 24(10), 5527–5533. (23) Ishizone, T.; Han, S.; Hagiwara, M.; Yokoyama, H. Synthesis and Surface Characterization of Well-Defined Amphiphilic Block Copolymers Containing Poly[oligo(ethylene glycol) methacrylate] Segments. Macromolecules 2006, 39(3), 962–970. (24) Yokoyama, H.; Miyamae, T.; Han, S.; Ishizone, T.; Tanaka, K.; Takahara, A.; Torikai, N. Spontaneously Formed Hydrophilic Surfaces by Segregation of Block Copolymers with Water-Soluble Block. Macromolecules 2005, 38(12), 5180–5189. (25) Harris, J. M. Poly(ethylene glycol) Chemistry: Biotechnical and Biomedical Applications; Plenum: New York, 1992. (26) Hirata, T.; Matsuno, H.; Kawaguchi, D.; Hirai, T.; Yamada, N. L.; Tanaka, M.; Tanaka, K. Effect of Local Chain Dynamics on a Bioinert Interface. Langmuir 2015, 31(12), 3661–3667. (27) Lu, D. R.; Lee, S. J.; Park, K. Calculation of solvation interaction energies for protein adsorption on polymer surfaces. J. Biomater. Sci. Polym. Ed. 1992, 3(2), 127–147. (28) Tanaka, M.; Mochizuki, A. Clarification of the blood compatibility mechanism by controlling the water structure at the bloodpoly(meth)acrylate interface. J. Biomater. Sci. Polym. Ed. 2010, 21(14), 1849–1863. (29) Park, H. D.; Lee, W. K.; Ooya, T.; Park, K. D.; Kim, Y. H.; Yui, N. In vitro biocompatibility assessment of sulfonated polyrotaxane-immobilized polyurethane surfaces. J. Biomed. Mater. Res., Part A 2003, 66A(3), 596–604. (30) Park, H. D.; Bae, J. W.; Park, K. D.; Ooya, T.; Yui, N.; Jang, J. H.; Han, D. K.; Shin, J. W. Surface modification of polyurethane using sulfonated PEG crafted polyrotaxane for improved biocompatibility. Macromol. Res. 2006, 14(1), 73–80. (31) Jeon, S. I.; Andrade, J. D. Protein-surface interactions in the presence of polyethylene oxide. II. Effect of protein size. J. Colloid Interface Sci. 1991, 142(1), 159–166. (32) Sofia, Susan J.; Premnath, V.; Merrill, Edward W. Poly(ethylene oxide) Grafted to Silicon Surfaces: Grafting Density and Protein Adsorption. Macromolecules 1998, 31(15), 5059–5070. (33) Jeon, S. I.; Lee, J. H.; Andrade, J. D.; De Gennes, P. G. Protein-surface interactions in the presence of polyethylene oxide. I. Simplified theory. J. Colloid Interface Sci. 1991, 142(1), 149–158. (34) Szleifer, I. Protein Adsorption on Surfaces with Grafted Polymers: A Theoretical Approach. Biophys. J. 1997, 72(2, Pt. 1), 595– 612.

ACS Paragon Plus Environment

Page 6 of 7

Page 7 of 7 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

Langmuir Table 1. Characteristics of the synthesized PRs.

a

Sample

Number of CDs

PR07

7

PR15

15

Chemical structure of the modification group of CDs

Modification degree

Mna

Mw /Mnb

Yield

-OCOCH2OCH2CH2OCH3

0.54

2.5  10

1.23

76%

-OCOCH2OCH3

0.51

3.6  10

1.35

85%

Calculated from NMR spectra. b Obtained from GPC traces.

ACS Paragon Plus Environment

7