Lotus-leaf-like structured chitosan–polyvinyl pyrrolidone films as an anti-adhesion barrier

Accepted Manuscript Title: Lotus-leaf-like structured chitosan-polyvinyl pyrrolidone films as an anti-adhesion barrier Author: Jin Ik Lim Min Ji Kang Woo-Kul Lee PII: DOI: Reference:

S0169-4332(14)02078-9 http://dx.doi.org/doi:10.1016/j.apsusc.2014.09.087 APSUSC 28743

To appear in:

APSUSC

Received date: Revised date: Accepted date:

1-6-2014 12-9-2014 15-9-2014

Please cite this article as: J.I. Lim, M.J. Kang, W.-K. Lee, Lotus-leaf-like structured chitosan-polyvinyl pyrrolidone films as an anti-adhesion barrier, Applied Surface Science (2014), http://dx.doi.org/10.1016/j.apsusc.2014.09.087 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

Lotus-leaf-like structured chitosan-polyvinyl pyrrolidone films as an anti-adhesion barrier

ip t

Jin Ik Lim · Min Ji Kang · Woo-Kul Lee* Laboratory of Biointerfaces/Tissue Engineering, Department of Chemical Engineering,

cr

Institute of Tissue Regeneration Engineering, College of Engineering, Dankook University,

an

M

For submission to Applied Surface Science

us

Jukjeon-dong, Yongin-si, Gyeonggi-do, Republic of Korea

Ac ce p

Woo-Kul Lee, Ph.D.

te

d

*To whom correspondence should be addressed

Tel: +82-31-8005-3540

FAX: +82-31-8021-7216

E-mail: [email protected]

Highlights

Page 1 of 27

Improved mechanical properties by hydrogen bond between chitosan and PVP chains.

-

Improved anti-adhesion effect by lotus-leaf-like structured chitosanPVP (L-Chitosan-PVP) film

-

L-Chitosan-PVP film as a blood/tissue anti-adhesion barrier for postsurgical treatment.

cr

ip t

-

us

Abstract

For postsurgical anti-adhesion barrier applications, lotus-leaf-like

an

structured chitosan-PVP films were prepared using a solution casting

M

method with dodecyltrichloro-immobilized SiO2 nanoparticles. We evaluated whether the lotus-leaf-like structured chitosan-PVP films (L-chitosan-PVP)

d

could be applied as postsurgical anti-adhesion barriers. A recovery test

te

using a tensile strength testing machine and measurement of crystallinity using X-ray diffraction indicated that films with 75% PVP were the optimal

Ac ce p

composition of the chitosan-PVP films. Also, dodecyltrichloro-immobilized SiO2 nanoparticles was synthesized and sprayed on the film after pretreatment with the instant bio-glue. Analysis of cell adhesion, proliferation, and anti-thrombus efficiency were performed via a WST assay, field emission scanning electron microscopy, and hemacytometry. The contact angle with the lotus-leaf-like surface was of approximately 150°. Furthermore, the L-chitosan-PVP film yielded a lower cell and platelet adhesion rate (around less than 4%) than that yielded by the untreated film. These results indicate that the lotus-leaf-like structure has a unique property and that this novel L-chitosan-PVP film can be applied as a

Page 2 of 27

blood/tissue-compatible, biodegradable material for implantable medical devices that need an anti-adhesion barrier.

Introduction

us

1.

cr

structure, Superhydrophobic surface, Surface modification

ip t

Keywords: Anti-adhesion barrier, Anti-thrombotic surface, Lotus-leaf-like

Adhesions are abnormal fibrous connections that develop between

an

tissues and organs as a result of the inflammatory processes, such as that due to infections, inflammations, and endometriosis, and they are a major

M

unsolved problem for surgery. Most frequently, however, they are a sequel to various accidents or diseases after an incision, cauterization, suturing,

te

other tissue trauma [1,2].

d

mechanical or thermal injury, ischemia, abrasion, foreign-body reaction, or

Ac ce p

Many studies have been undertaken to find an effective approach to prevent postsurgical adhesion. Pharmacological methods (aspirin, dexamethasone, and heparin) and barrier-based devices (polymer solutions, solid sheets, and hydrogels) are two important approaches to used to prevent adhesions [3–7]. Drugs have a low efficiency for local targeting and produce side effects, therefore film-type anti-adhesion barriers have been usually used in an attempt to prevent undesired contact between the damaged serosal tissues and other intra-abdominal surfaces during critical re-epithelialization after surgery [8–11]. Some of these methods have been developed as commercial products,

Page 3 of 27

such as hyaluronic acid solution (Sepracoat; Genzyme, Cambridge, MA), oxidized regenerated cellulose (Interceed; ETHICON Women’s Health and Urology, Somerville,NJ), and sodium hyaluronate carboxy methyl cellulose

ip t

films (Seprafilm; Genzyme, Cambridge, MA). However, these have drawbacks in that they have a short time of persistency and are breakable and sticky.

cr

[12–14]. To improve upon these shortcomings, many researchers have

us

attempted to design novel biomaterials that provide a more biocompatible matrix and an enhanced surface treatment for anti-adhesion that are

an

required with fewer complications and higher efficiencies [15]. Chitosan (CS) is a naturally derived crystalline polysaccharide. It has gained

M

considerable attention as a biomaterial and has diverse applications in tissue engineering as a result of its low cost, large-scale availability,

te

[16].

d

antimicrobial activity, minimal foreign body reaction, and biocompatibility

Ac ce p

In this study, the CS and polyvinyl pyrrolidone (PVP) were selected as polymers for the barrier materials due to their water solubility, biocompatibility, and enhanced strength. These materials have been extensively employed in biotechnical and biomedical fields. Blends of CS/PVP appeared clear in appearance, and the IR spectra indicated carbonyl–OH hydrogen bonding between chains in the blend which consequently gave them their enhanced strength [17]. Control over the interface properties through physical rather than chemical modification of biopolymer surface is desirable since the surface treatment is required to improve the anti-adhesion efficiency of the

Page 4 of 27

biopolymer. The antithrombotic effects of highly hydrophobic surfaces have already been reported by many researchers [18–20], and the development of various methods to fabricate superhydrophobic surfaces have been reported,

ip t

including the template tool, sol-gel technology, the solution method, layerby-layer self-assembly, the plasma treatment process, and the

cr

electrospinning method [18–26]. Furthermore, superhydrophobic surfaces,

us

such as the lotus-leaf-like structure, have been extended to an increasing number of applications such as contamination prevention (including self-

an

cleaning windows, windshields, exterior paints for buildings), micro fluidic devices, electromechanical systems, and enhanced lubrication [27–32].

M

The objectives of this study were as follows: (1) to select the optimal composition for an enhanced CS/PVP barrier; (2) to determine the surface

d

treatment conditions in order to achieve a lotus effect; and (3) to evaluate

Ac ce p

te

the anti-adhesion efficiency using fibroblasts.

Page 5 of 27

ip t cr

Materials and methods

us

2.

In this study, 86% deacetylated chitosan (Korea-Chitosan Co, Seoul,

an

Korea) of ~1,000 KDa was used as a basis material for the anti-adhesion barrier. Polyvinyl pyrrolidone (PVP) of 360KDa, ethanol, silica particles of

M

12-nm in size, dodecyltrichlorosilane, and toluene (99 wt.%) were purchased from Sigma-Aldrich (St. Louis, MO, USA). All other chemicals used were of

d

an analytical grade. Ultrapure water from a Milli-Q water system was used

te

to prepare the aqueous solutions.

Ac ce p

Two grams of PVP and two grams of CS were each dissolved in a 1% (v/v) acetic acid solution (100 ml) under magnetic stirring for 48 h at room temperature. The resulting solutions were filtered and stored at 4 °C until use. The solutions were then mixed into the chitosan acidic solution to attain PVP solution concentrations of 0, 25, 50, and 75% (v/v). The mixture was injected into a Teflon plate (200 × 200 × 3 mm3), dried at room temperature for 1 w, then vacuum dried for 24 h. The obtained chitosanPVP film was washed stage-by-stage in decreasing ethanol concentrations (100%, 90%, 80%, 70%, 50% and 0% v/v) for 2 h per stage. Then, the film was vacuum dried.

Page 6 of 27

The crystal intensity diagrams of the chitosan-PVP films were measured via X-ray diffraction (XRD: D8 advance; Bruker AXS, Karlsruhe, Germany) operating with Cu-Kα radiation (λ = 0.15406 nm) at 40 kV and

ip t

100 mA at a speed of 1°/min.

To examine the recovery from the bending stress, the chitosan-PVP

cr

films (200 × 200 × 3 mm3) were prepared at PVP concentrations of 0, 25, 50,

us

and 75%. The recovery test was carried out using a universal testing machine. A 5 N load cell with a crosshead speed of 1 mm/min (bending

an

angle=180°) was used for this purpose. The recovery was calculated as Recovery (%)=A1/A0×100, where A0 indicates the original angle, and A1

M

indicates the final angle for 1 h after stress release.

The FTIR spectra of chitosan, PVP, and their blend (75% PVP) were

te

PerkinElmer, USA).

d

scanned (4000–500 cm−1) using the FT-IR Spectrometer (Spectrum GX,

Ac ce p

The superhydrophobic surface was prepared by using the previously reported method [33, 34]. Two grams of SiO2 nanoparticles and 40 mL of dehydrated toluene were placed into a Schlenk flask. They were refluxed for 3 h after adding 1 mL of dodecyltrichlorosilane. The procedure imparts hydrophobicity to the SiO2 nanoparticles by immobilizing the dodecyltrichloro groups on their surface. After filtration, the obtained dodecyltrichloro group-immobilized SiO2 particles were dried at 353 K. The dodecyltrichloro group-immobilized SiO2 particles (0.1 g) were added to 5 mL of 1-butanol. After stirring (30 min) and sonicating (20 min), the suspension was manually sprayed over chitosan-PVP films (200 × 200 × 3 mm3) using a

Page 7 of 27

glass vaporizer. The resulting samples were then placed horizontally and were dried at room temperature overnight. To evaluate the stability of the SiO2 sprayed on the chitosan-PVP film

ip t

with 75% PVP, three types of films [one with 0.1 g SiO2 in 5 mL butanol

simply sprayed on the chitosan-PVP film (200 × 200 × 3 mm3), one with SiO2

cr

sprayed after pretreatment with 2 mL of Dermabond (octyl 2-cyanoacrylate)

us

as a binder on the chitosan-PVP film, and another with 0.1 g of SiO2 and 2 mL of Dermabond mixture sprayed on the film] were dried at room

an

temperature for 48 h, immersed in 5 ml PBS, and incubated at 37 °C and 20 rpm for 0, 4, 24, 48, and 72 h. After incubation, the films were dried at room

M

temperature and the time-based change of the water contact angle (Phoenix 150; Surface Electro Optics, Seoul, Korea) and the morphology were

d

observed using a field-emission scanning electron microscope (FE-SEM: S-

te

4700; Hitachi, Tokyo, Japan).

Ac ce p

Using the same method as in the previous recovery test, the same three types of films of three types [SiO2 simply sprayed on chitosan-PVP film, SiO2 sprayed after pretreatment with Dermabond (octyl 2-cyanoacrylate) as a binder on chitosan-PVP film, and SiO2 and Dermabond mixture sprayed on the film] were prepared, and the recovery test was carried out using a universal testing machine.

Chitosan-PVP films (one with a lotus-leaf-like-structured film and one prepared with untreated chitosan as control) were pre-wetted with medium [Dulbecco's modified eagle's medium (DMEM) supplemented with 2 mM L-glutamine and 10% fetal bovine serum] and incubated at 37 °C under

Page 8 of 27

a 5% CO2 environment. After 12 h, the medium was aspirated, and NIH-3T3 cells (ATCC-L929, Manassas, VA, USA) were plated directly on each film in a 200-µl media suspension. The cell density was set at 3 × 104 cells per well.

ip t

After another 1 h, 1,800 µl medium was added to each well, and the initial adhesion and proliferation were quantified using a WST-8 assay.

cr

The films were equilibrated overnight using 2 ml of PBS. Prior to the

us

adhesion studies, the buffer was removed from a syringe and 2 ml of human whole blood was introduced into the syringe. The syringes were tapped to

an

remove air bubbles, sealed, and rotated in a shaking incubator at 37°C for an adhesion time of 1 h. After each adhesion time, the films were quickly

M

removed from the syringe and were fixed in a formaldehyde solution for 24 h. The platelets that had adhered to the surface of the film were determined

3.

te

Ac ce p

a hemacytometer.

d

using a field emission scanning electron microscope and were counted using

3.1.

Results and discussion Measurement of crystal intensity of the chitosan-PVP films To characterize the chitosan-PVP films, XRD and mechanical tests

were performed. XRD crystallization intensity increased after the addition of PVP to chitosan. The main diffraction peak at around 2θ=19° and small peaks at 21.5° and 23.7° were observed in the small-angle region of the diffraction profiles for the chitosan-PVP films. The intensity increased as PVP concentration increased, and this could be attributed to the hydrogen bonding between chitosan and the PVP polymer chains in the mixtures (Fig.

Page 9 of 27

1). Also, we believe that the increase in crystallization intensity implies a change in the mechanical properties chitosan-PVP mixtures to become tough and strong as a result of the presence of PVP. Recovery test of the chitosan-PVP films with respect to various PVP

ip t

3.2.

concentrations

cr

The anti-adhesion barrier requires an optimal toughness or recovery

us

for application to surgical sites of various shapes, including curves, angles, waves, etc. The recovery of chitosan-PVP films in response to bending stress

an

was compared to that of chitosan, and the results indicated that the recovery of the chitosan-PVP films increased with increasing PVP

M

concentration, whereas chitosan and the film with 25% PVP were broken after bending one times. Table 1 indicates that a chitosan-PVP film with 75%

d

PVP showed the highest recovery efficiency. Also, we infer that an improved

te

recovery was a result of the increased hydrogen bonding between chitosan

Ac ce p

and PVP. Therefore, chitosan-PVP film with 75% PVP was determined to be the film composition with the optimal physical properties and was prepared for the following tests. 3.3.

FTIR analysis

Fig. 2 shows the FTIR spectra of the chitosan, PVP, and the blended

film. The spectrum of the pure chitosan (Fig. 2a) shows a broad peak at wavelengths in the region of 1570–1655 cm−1, indicating the presence of amides I and II. The peaks at 1639 and 3342 cm−1 are attributed to the amide carbonyl band and to free hydroxyl groups. PVP shows an amide carbonyl band at 1688 cm−1 (Fig. 2c), which is higher than that observed in

Page 10 of 27

pure chitosan. The chitosan/PVP blend (Fig. 2b) also shows a peak resulting from the amide carbonyl group of PVP at 1688 cm−1. As a result, the chitosan-PVP blend shows a single carbonyl band at 1660 cm−1, indicating

ip t

an interaction between the chitosan and the PVP that can occur between amine groups of both chitosan and carbonyl groups of PVP [35]. Morphological analysis of the L-chitosan-PVP films

cr

3.4.

us

The L-chitosan-PVP film surface was compared against films with different surface treatments, such as pure chitosan-PVP and L-chitosan-

an

PVPs. In contrast with the smooth-surfaced pure chitosan-PVP [Fig. 1(a)], Lchitosan-PVP films generally showed a rougher surface that was sprayed

M

with nanoparticles. As shown in Fig. 1(b) and (c), no difference was observed between the morphologies of the SiO2 simply-sprayed film and the SiO2

d

sprayed film after pretreatment with Dermabond. Also, in the case of the

te

SiO2 and Dermabond mixture sprayed film [Fig. 1(d)], particles of an uncertain

Ac ce p

boundary were observed because these were nanoparticles covered by Dermabond. Therefore, we anticipate a superhydrophobic surface of the SiO2 simply-sprayed film and SiO2 sprayed film after pretreatment with Dermabond. 3.5.

Stability of SiO2 particles on the L-chitosan-PVP film In general, the water contact angle by the superhydrophobic surface

is of over 150°. The anti-adhesion and anti-pollution effects of the superhydrophobic surface are a result of the trapped hydrophobic air layer between the particles on the film. The stability test of the SiO2 particles on the film was performed using SiO2 simply sprayed on the film, SiO2 sprayed

Page 11 of 27

on the film after pretreatment with Dermabond (octyl 2-cyanoacrylate), and SiO2 and Dermabond mixture sprayed on the film. Each were prepared and sprayed, and the change in water contact angle was measured as the

ip t

stability. As indicated in Fig 3, in the case of SiO2 simply sprayed on the film, a high water contact angle with a relatively lower stability was observed

cr

[time(hr)/contact angle, 0/165, 4/163, 24/165, 48/142, and 72/121],

us

whereas SiO2 sprayed on the film after pretreatment with Dermabond showed relatively high stability with lower water contact angle than the

an

SiO2-only sprayed film. We suggested that the low stability of the simply sprayed surface is due to the non-bonded interaction of the film surface with

M

only a weak hydrophobic interaction of between SiO2 particles. The good stability with a somewhat lower water contact angle by Dermabond is a

d

result of the fixation effect of the particles and the partially interrupted air

te

trapped by the reaction of Dermabond as a binder between the chitosan-PVP

Ac ce p

surface and the SiO2 particles. In the case of the SiO2 and Dermabond mixture, a superhydrophobic effect was not observed. As the cause, we suggest that the air area between the SiO2 particles was substituted by the Dermabond as a polymerizable solution. Therefore, the SiO2 sprayed film after pretreatment with Dermabond was selected to be the optimal surface treatment method with a water contact angle of near 150°. 3.6.

Recovery test of the L-chitosan-PVP films with respect to various surface treatment methods

Table 2 indicates that L-chitosan-PVP films with 75% PVP showed a somewhat lower recovery efficiency than chitosan-PVP films with 75% PVP

Page 12 of 27

(Table 1), except for the SiO2 simply-sprayed film. We suggest that the mechanical properties of the L-chitosan-PVP films were not affected by he only slightly sprayed SiO2 nanoparticles. Also, the decreased recovery of the

ip t

films, except for the SiO2 simply-sprayed film, is a result of the mechanical properties of the brittle and hard Dermabond layer on the L-chitosan-PVP films. However, the L-chitosan-PVP films showed a recovery of over 70%,

cr

and we expect that this is sufficient for application to surgical sites of

3.7.

Cell adhesion and proliferation test

us

various shapes.

an

For the cell affinity measurements, fibroblasts cells cultured on films. In the cases with untreated chitosan-PVP films, after a 3-h cell culture, a

M

more than 3-fold increase was observed in the cell adhesions on the film when compared to those with the L-chitosan-PVP films (Fig. 4). Moreover,

d

after 24, and 72 h of culture, increased cell proliferation was observed. We

te

suggest that the initial cell adhesion efficiency on the film was important for tissue anti-adhesion, and the low initial cell adhesion efficiency of L-

Ac ce p

chitosan-PVP film is a result of the superhydrophobic structure. Also, cell proliferation is depressed by a low initial cell adhesion efficiency of the Lchitosan-PVP film. Therefore, we confirmed that the L-chitosan-PVP film can be used in clinical applications as an optimal anti-adhesion barrier. 3.8.

In vitro platelet adhesion Various biomaterial films for postsurgical anti-adhesion are usually

used at a surgical site. However, when these biomaterials are in contact with blood from surgical site, proteins are first adsorbed instantaneously onto the surfaces and these are then deformed. Then, the platelets adhere to the surfaces and are activated and aggregated, and this process is thought to

Page 13 of 27

play a major role in thrombus formation. Furthermore, film encapsulation by side fibrous tissues is ultimately induced by thrombus formed on the film. Therefore, a platelet adhesion test on the films was conducted. Fig. 5 shows

ip t

the in vitro adhesion behavior of the platelets on the chitosan-PVP films.

Fewer platelets adhered to the L-chitosan-PVP film (approximately 4%) than

cr

to the control chitosan-PVP film. We suggest that the lower amount of

us

adhesion is due to non-wettable property of the superhydrophobic surface formed by the dodecyltrichloro-immobilized SiO2 particles on the chitosan-

Conclusion

M

4.

an

PVP film.

Our goal was to conduct a fundamental study of the chemo-physical

d

properties of materials synthesized for an anti-adhesion barrier to improve

te

anti-thrombotic and anti-adhesion efficiency. Therefore, a superhydrophobic

Ac ce p

chitosan-PVP film was prepared by spraying dodecyltrichloro-immobilized SiO2 particles on chitosan-PVP film. We conducted various tests to assess the adequacy of these films for cell anti-adhesion and as anti-thrombotic solutions. When we assessed the mechanical properties as postsurgical antiadhesion barriers, the chitosan-PVP film proved to be tougher and presented a higher recovery. The water contact angle of the sprayed dodecyltrichloroimmobilized SiO2 particles was near 150°. Furthermore, cell anti-adhesion and anti-thrombus effects were confirmed. Future studies are essential to understanding the immobilization of various hydrophobic chains on SiO2 nanoparticles so as to be able to further increase superhydrophobicity and

Page 14 of 27

increase their anti-adhesion efficiency in order to guarantee their suitability

ip t

for applications in various fields, including medicine.

cr

Figure legends

us

Fig. 1 XRD patterns of chitosan-PVP films at various PVP concentrations Fig. 2 FTIR spectra of (a) chitosan, (b) chitosan-75% PVP and (c) PVP.

an

Fig. 3 Scanning electron microscope (SEM) images of L-chitosan-PVP films. (a): pure chitosan-PVP film, (b): SiO2 simply-sprayed film, (c) SiO2 sprayed

M

film after pretreatment with Dermabond, (d) SiO2 and Dermabond mixture sprayed film, × 10,000.

te

stress

d

Fig. 4 Change in water contact angle on the chitosan-PVP film under water

Ac ce p

Fig. 5 Cell adhesion and proliferation on the lotus-leaf-like structured chitosan-PVP film and the untreated chitosan-PVP film Fig. 6 Platelet adhesion on the lotus-leaf-like structured chitosan-PVP film and the untreated chitosan-PVP film viewed with an electron micrograph [a: platelet on chitosan-PVP film as a control (×500), b: Lotus-leaf-like structured chitosan-PVP film (×500, ×20,000)].

Page 15 of 27

ip t cr us

References

[1] E. Shahrama, S.H. Sadraie, G. Kaka, H. Khoshmohabat, M.

an

Hosseinalipour, F. Panahi, M.R. Naimi-Jamal, Evaluation of chitosane gelatin films for use as postoperative adhesion barrier in rat cecum model,

M

Inter. J. Surg. 11 (2013) 1097-1102.

[2] L. Li, N. Wang, X. Jin, R. Deng, S. Nie, L. Sun, Q. Wu, Y. Wei, C. Gong,

d

Biodegradable and injectable in situ cross-linking chitosan-hyaluronic acid

Ac ce p

(2014) 3903-3917.

te

based hydrogels for postoperative adhesion prevention, Biomaterials 35

[3] J.L. Hill-West, R.C. Dunn, J.A. Hubbell, Local release of fibrinolytic agents for adhesion prevention, J. Surg. Res. 59 (1995) 759-763. [4] Z. Zhang, J. Ni, L. Chen, L. Yu, J. Xu, J. Ding, Biodegradable and thermoreversible PCLA-PEG-PCLA hydrogel as a barrier for prevention of post-operative adhesion, Biomaterials 32 (2011) 4725-4736. [5] E. Ersoy, V. Ozturk, A. Yazgan, M. Ozdogan, H. Gundogdu, Effect of polylactic acid film barrier on intra-abdominal adhesion formation, J. Surg. Res. 147 (2008) 148-152. [6] B.W. Hellebrekers, T. Trimbos-Kemper, J. Trimbos, J.J. Emeis, T.

Page 16 of 27

Kooistra, Use of fibrinolytic agents in the prevention of postoperative adhesion formation, Fertil. Steril. 74 (2000) 203-212. [7] L. Muzii, R. Marana, L. Brunetti, F. Margutti, M. Vacca, S. Mancuso,

ip t

Postoperative adhesion prevention with low-dose aspirin: effect through the selective inhibition of thromboxane production, Hum. Reprod. 13 (1998)

cr

1486-1489.

us

[8] D.A. Grainger, W.R. Meyer, A.H. DeCherney, M.P. Diamond, The use of hyaluronic acid polymers to reduce postoperative adhesions, J. Gynecol.

an

Surg. 7 (1991) 97-101.

[9] C.I. Lauder, G. Garcea, A. Strickland, G.J. Maddern, Use of a modified

M

chitosane dextran gel to prevent peritoneal adhesions in a rat model, J. Surg. Res. 171 (2011) 877-882.

d

[10] C. Weis, E.K. Odermatt, J. Kressler, Z. Funke, T. Wehner, D. Freytag,

te

Poly (vinyl alcohol) membranes for adhesion prevention, J. Biomed. Mater.

Ac ce p

Res. B Appl. Biomater. 70 (2004) 191-202. [11] T. Ito, Y. Yeo, C.B. Highley, E. Bellas, D.S. Kohane, Dextran-based in situ crosslinked injectable hydrogels to prevent peritoneal adhesions. Biomaterials 28 (2007) 3418-3426. [12] B.C. Ward, A. Panitch, Abdominal adhesions: Current and novel therapies, J. Surg. Res. 165 (2011) 91–111. [13] T.H.M. Abou-Aiad, K.N. Abd-El-Nour, I.K. Hakim, M.Z. Elsabee, Dielectric and interaction behavior of chitosan/polyvinyl alcohol and chitosan/polyvinyl pyrrolidone blends with some antimicrobial activities, Polymer 47 (2006) 379–389.

Page 17 of 27

[14] J.J. Chang, Y.H. Lee, M.H. Wu, M.C. Yang, C.T. Chien, Electrospun anti-adhesion barrier made of chitosan alginate for reducing peritoneal adhesions, Carbohydrate Polymers 88 (2012) 1304– 1312.

ip t

[15] N. Bölgen, I. Vargel, P. Korkusuz, Y.Z. Mencelo˘glu, E. Pis¸ kin, In vivo performance of antibiotic embedded electrospun PCL membranes for

cr

prevention of abdominal adhesions, J. Biomed. Mater. Res. B Appl. Biomater.

us

81B (2007) 530–543.

Biomaterials 24 (2003) 2339-2349.

an

[16] E. Khor, L.Y. Lim, Implantable applications of chitin and chitosan,

[17] M.V Risbuda, A.A. Hardikarb, S.V. Bhata, R.R. Bhonde, pH-sensitive

M

freeze-dried chitosan–polyvinyl pyrrolidone hydrogels as controlled release system for antibiotic delivery, J. Control. Release 31 (2000) 23-30.

d

[18] W. Lee, M.K. Jin, W.C. Yoo, J.K. Lee, Nanostructuring of a polymeric

te

substrate with well-defined nanometer-Scale topography and tailored

Ac ce p

surface wettability, Langmuir 20 (2004) 7665-7669. [19] K. Acatay, E. Simsek, C. Ow-Yang, Y.Z. Menceloglu, Tunable, superhydrophobically stable polymeric surfaces by electrospinning, Angew. Chem. Int. Ed. 43 (2004) 5210-5213. [20] J.I. Lim, S.I. Kim, Y. Jung, S.H. Kim, Fabrication and Medical Applications of Lotus-leaf-like structured superhydrophobic surfaces, Polym. korea 37 (2013) 411-419. [21] Daniel Ebert, Bharat Bhushan, Durable Lotus-effect surfaces with hierarchical structure using micro- and nanosized hydrophobic silica particles, J. Colloid Interf. Sci. 368 (2012) 584-591.

Page 18 of 27

[22] Z. Yoshimitsu, A. Nakajima, T. Watanabe, K. Hashimoto, Effects of Surface Structure on the Hydrophobicity and Sliding Behavior of Water Droplets, Langmuir 18 (2002) 5818-5822.

porous microsphere/nanofiber composite film prepared by

ip t

[23] L. Jiang, Y. Zhao, J. Zhai, A lotus-leaf-like superhydrophobic surface: a

cr

electrohydrodynamics, Angew. Chem. Int. Ed. 43 (2004) 4338-4341.

us

[24] Y. Yao, X. Dong, S. Hong, H. Ge, C.C. Han, Shear-Controlled

Micro/Nanometer-Scaled Super-Hydrophobic Surfaces with Tunable Sliding

an

Angles from Single Component Isotactic Poly(propylene), Macromol. Rapid Commun. 27 (2006) 1627-1631.

M

[25] K. Tsujii, T. Yamamoto, T. Onda, S. Shibuchi, Super Oil-Repellent Surfaces, Angew. Chem. Int. Ed. Engl. 36 (1997) 1011-1012.

d

[26] R. Buzio, C. Boragno, F. Biscarini, F.B.D. Mongeot, U. Valbusa, The

te

contact mechanics of fractal surfaces, Nat. Mater. 2 (2003) 233-236.

Ac ce p

[27] J.I. Lim, S.I. Kim, S.H. Kim, Lotus-leaf-like structured heparinconjugated poly(L-lactide-co-ɛ-caprolactone) as a blood compatible material, Colloid Surface B 103 (2013) 463-467. [28] P. Aussillous, D. Quere, Liquid marbles, Nature 411 (2001) 924-927. [29] T.L. Sun, L. Feng, X.F. Gao, L. Jiang, Bioinspired surfaces with special wettability, Acc. Chem. Res. 38 (2005) 644-652. [30] A. Nakajima, K. Hashimoto, T. Watanabe, Recent Studies on SuperHydrophobic Films, Monatsh. Chem. 132 (2001) 31-41. [31] C. Yu, X. Jiansheng, G. Zhiguang, Recent Advances in Application of Biomimetic Superhydrophobic Surfaces, Prog. Chem. 24 (2012) 696-708.

Page 19 of 27

[32] B. Zhao, J.S. Moore, D.J. Beebe, Surface-directed liquid flow inside microchannels, Science 291 (2001) 1023-1026. [33] S.A. Kulkarni, S.B. Ogale, K.P. Vijayamohanan, Tuning the hydrophobic

ip t

properties of silica particles by surface silanization using mixed selfassembled monolayers, J. Colloid Interf. Sci. 318 (2008) 372–379.

cr

[34] H. Ogihara, J. Xie, J. Okagaki, T. Saji, Simple Method for Preparing

us

Superhydrophobic Paper: Spray-Deposited Hydrophobic Silica Nanoparticle Coatings Exhibit High Water-Repellency and Transparency, Langmuir 28

an

(2012) 4605−4608.

[35] D. Anjali Devi, B. Smitha, S. Sridhar, T.M. Aminabhavi, Novel

M

crosslinked chitosan/poly(vinylpyrrolidone) blend membranes for

Ac ce p

te

280 (2006) 45–53.

d

dehydrating tetrahydrofuran by the pervaporation technique, J. Memb. Sci.

Page 20 of 27

Table 1 Recovery (%) of the bending stress of the chitosan-PVP films with respect to various PVP concentrations (n=5)

25

50

75

Recovery (%)

-

-

7±3

ip t

0

92 ± 5

cr

PVP content (%)

us

Table 2 Recovery (%) of the bending stress of the L-chitosan-PVP films with

SiO2 simply sprayed

Recovery (%)

91 ± 4

SiO2 sprayed after pretreatment with Dermabond

M

L-chitosanPVP films

an

respect to various surface treatment methods (n=5)

72 ± 3*

74 ± 4*

Ac ce p

te

d

*P > 0.05

SiO2 and Dermabond mixture sprayed

Page 21 of 27

Chitosan +25%PVP +50%PVP +75%PVP

6000

cr us an

4000 3000

M

Intensity

5000

ip t

Fig. 1

5

10

15

20

te

1000

d

2000

25

30

35

Ac ce p



Page 22 of 27

Ac ce p

te

d

M

an

us

cr

ip t

Fig. 2

Page 23 of 27

Ac ce p

te

d

M

an

us

cr

ip t

Fig. 3

Page 24 of 27

Ac ce p

te

d

M

an

us

cr

ip t

Fig. 4

Page 25 of 27

us

0.35

TCP Chitosan-PVP L-Chitosan-PVP

an

0.30 0.25

M

0.20 0.15 0.10

d

Absorbance (490 nm)

0.40

cr

ip t

Fig. 5

te

0.05

Ac ce p

0.00

1 3

2 24

3 72

Culture time (hr)

Page 26 of 27

Ac ce p

te

d

M

an

us

cr

ip t

Fig. 6

Page 27 of 27