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Chemical Grafting of Poly(ethylene glycol) Methyl Ether Methacrylate onto Polymer Surfaces by Atmospheric Pressure Plasma Processing Raechelle A. D’Sa and Brian J. Meenan* Nanotechnology and Integrated Bio-Engineering Centre (NIBEC), University of Ulster, Shore Road, Newtownabbey, BT37 0QB, Northern Ireland Received July 20, 2009. Revised Manuscript Received September 4, 2009 This article reports the use of atmospheric pressure plasma processing to induce chemical grafting of poly(ethylene glycol) methyl ether methacrylate (PEGMA) onto polystyrene (PS) and poly(methyl methacrylate) (PMMA) surfaces with the aim of attaining an adlayer conformation which is resistant to protein adsorption. The plasma treatment was carried out using a dielectric barrier discharge (DBD) reactor with PEGMA of molecular weights (MW) 1000 and 2000, PEGMA1000 and PEGMA2000, being grafted in a two step procedure: (1) reactive groups are generated on the polymer surface followed by (2) radical addition reactions with the PEGMA. The surface chemistry, coherency, and topography of the resulting PEGMA grafted surfaces were characterized by X-ray photoelectron spectroscopy (XPS), time-of-flight secondary ion mass spectrometry (ToF-SIMS), and atomic force microscopy (AFM), respectively. The most coherently grafted PEGMA layers were observed for the 2000 MW PEGMA macromolecule, DBD processed at an energy dose of 105.0 J/cm2 as indicated by ToF-SIMS images. The effect of the chemisorbed PEGMA layer on protein adsorption was assessed by evaluating the surface response to bovine serum albumin (BSA) using XPS. BSA was used as a model protein to determine the grafted macromolecular conformation of the PEGMA layer. Whereas the PEGMA1000 surfaces showed some protein adsorption, the PEGMA2000 surfaces appeared to absorb no measurable amount of protein, confirming the optimum surface conformation for a nonfouling surface.
1. Introduction The in vivo response of a biomaterial (or implant) is influenced by surface chemistry and topography. It is these properties that then determine the initial stages of biocompatibility when contact is made between the material/device and a biological system (cells or tissue). An important aspect of some biomaterials that come into contact with bodily fluids is the prevention of nonspecific adsorption of biomolecules such as proteins, platelets, and bacteria.1-3 Generally, most polymeric biomaterials used in the medical device industry are highly hydrophobic and interact strongly with proteins in most biological environments.4,5 In some cases, this can result in severe clinical complications such as coagulation, complement activation, thrombus formation, fibrous encapsulation, and immunological reactions.6-8 It is well accepted that grafting of hydrophilic molecules such as poly(ethylene glycol) (PEG) and its structural analogues onto a polymer surface results in a reduction of the nonspecific adhesion of such biomolecules to polymeric surfaces.9,10 Whereas the
mechanism by which PEG and related materials repel proteins is not fully understood, surface density and molecular weight (MW) have been recognized as key factors in the effectiveness of this effect.8,11-15 Many of the polymeric biomaterials that are used clinically have proven mechanical and physicochemical properties but can lack the functionality to allow for direct covalent tethering of hydrophilic coatings such as PEG. Plasma surface modification has been shown to be a useful means for creating the surface conditions that allow for covalent tethering of such coatings. Although atmospheric pressure plasma processing offers a costeffective means of achieving the desired surface functionality, the facility offered by normal “corona” based methods is very limited due to the transient nature of the surface effects induced. The advent of dielectric barrier discharge (DBD) atmospheric pressure processing holds out more promise in this regard, but its ability has yet to be fully realized.16-20 Plasma surface modification of a polymer activates the surface to create a so-called “bonding layer” which can then be utilized
*Corresponding author. Address: Nanotechnology and Integrated BioEngineering Centre (NIBEC), University of Ulster, Shore Road, Newtownabbey, Co Antrim, BT37 0QB, Northern Ireland. Telephone: þ44(0) 28 90368939. E-mail:
[email protected].
(11) Emoto, K.; Harris, J. M.; Van Alstine, J. M. Anal. Chem. 1996, 68, 3751– 3757. (12) Bergstrom, K.; Osterberg, E.; Holmberg, K.; Hoffman, A. S.; Schuman, T. P.; Kozlowski, A.; Harris, J. H. J. Biomater. Sci., Polym. Ed. 1994, 6, 123–132. (13) Sofia, S. J.; Premnath, V.; Merrill, E. W. Macromolecules 1998, 31, 5059– 5070. (14) Irvine, D. J.; Mayes, A. M.; Satija, S. K.; Barker, J. G.; Sofia-Allgor, S. J.; Griffith, L. G. J. Biomed. Mater. Res. 1998, 40, 498–509. (15) Michel, R.; Pasche, S.; Textor, M.; Castner, D. G. Langmuir 2005, 21, 12327–12332. (16) Borcia, G.; Anderson, C. A.; Brown, N. M. D. Appl. Surf. Sci. 2004, 221, 203–214. (17) Borcia, G.; Anderson, C. A.; Brown, N. M. D. Appl. Surf. Sci. 2004, 225, 186–197. (18) Liu, C.; Brown, N. M. D.; Meenan, B. J. Surf. Sci. 2005, 575, 273–286. (19) Liu, C.; Cui, N.; Brown, N. M. D.; Meenan, B. J. Surf. Coat. Technol. 2004, 185, 311–320. (20) Liu, C.; Brown, N. M. D.; Meenan, B. J. Surf. Coat. Technol. 2006, 201, 2341–2350.
(1) Chapman, R. G.; Ostuni, E.; Liang, M. N.; Meluleni, G.; Kim, E.; Yan, L.; Pier, G.; Warren, H. S.; Whitesides, G. M. Langmuir 2001, 17, 1225–1233. (2) Ostuni, E.; Chapman, R. G.; Holmlin, R. E.; Takayama, S.; Whitesides, G. M. A. Langmuir 2001, 17, 5605–5620. (3) Kingshott, P.; Griesser, H. J. Curr. Opin. Solid State Mater. Sci. 1999, 4, 403– 412. (4) Ratner, B. D.; Bryant, S. J. Annu. Rev. Biomed. Eng. 2004, 6, 41–75. (5) Castner, D. G.; Ratner, B. D. Surf. Sci. 2002, 500, 28–60. (6) Anderson, J. M.; Rodriguez, A.; Chang, D. T. Semin. Immunol. 2008, 20, 86– 100. (7) Anderson, J. M. Annu. Rev. Mater. Res. 2001, 31, 81–110. (8) Morra, M. J. Biomater. Sci., Polym. Ed. 2000, 11, 547–569. (9) Lee, J. H.; Lee, H. B.; Andrade, J. D. Prog. Polym. Sci. 1995, 20, 1043–1079. (10) Harris, J. M. In Poly(ethylene glycol) Chemistry: Biotechnical and Biomedical Applications; Plenum Press: New York, 1992.
1894 DOI: 10.1021/la902654y
Published on Web 10/01/2009
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for the immobilization of macromolecules such as PEG; that is, grafting is achieved by chemically tethering molecules onto the plasma-activated surface.21-24 This is generally considered in a two-step process, where (1) reactive groups are generated on the polymer surface followed by (2) heterogeneous reactions between the macromolecules of interest and the active surface groups. Whereas plasma processing has been used to graft molecules onto substrates, it is not widely applied due to the high engineering costs associated with vacuum based plasmas that are normally required to carry out these reactions.22-27 In this regard, DBD processing offers an attractive alternative since it allows for rapid, continuous in-line processing and lower operational costs. However, the utility of a DBD reactor to induce effective plasma grafting of macromolecules has not been proven. This article reports a study of the use of DBD processing to induce plasma grafting of methacrylate terminated PEG (PEGMA) onto poly(methyl methacrylate) (PMMA) and polystyrene (PS) polymer substrates. The pendant vinyl group of the PEGMA species is targeted to interact with the oxidative groups generated by the DBD treatment of the polymeric surfaces. The nature of the grafted polymer chains has been characterized by X-ray photoelectron spectroscopy (XPS) and time-of-flight secondary ion mass spectrometry (ToF-SIMS). XPS indicates the oxidation of the surface in stage 1 and the grafting of PEGMA in stage 2. ToF-SIMS has been used to corroborate the XPS findings and to determine the coherency of the surface coatings. Changes in surface morphology that result from the grafting have been monitored by atomic force microscopy (AFM). The ability of the grafted PEGMA to act as a nonfouling layer has been measured by exposing the surfaces to a bovine serum albumin (BSA) solution. XPS analysis was used to determine the presence or otherwise of adsorbed protein by monitoring the attendant nitrogen signal. It should be noted that the albumin adsorption experiments were undertaken as a means of further characterizing the actual structure of the PEGMA created on the polymer surfaces by DBD induced grafting, and as such they do not represent a protein adsorption study per se.
2. Materials and Methods 2.1. Materials. Commercial grade PMMA (1 mm thick) and PS (1.2 mm thick) sheets (Goodfellow, Cambridge U.K.) were used as the polymer substrates for these studies. Sheets were cut into 1.5 1.5 cm2 sized squares and cleaned by sonication in ethanol for 15 min and dried in air overnight prior to use. Poly(ethylene glycol) methyl ether methacrylate (PEGMA, Sigma-Aldrich, U.K.) of two different molecular weights (MW), namely, (1) 1000 Da (PEGMA1000) and (2) 2000 Da (PEGMA2000), was used as the source material for grafting onto the polymer surfaces using the DBD processing method. PEGMA1000 was used as received, while the PEGMA2000 was supplied as a 50% w/v solution in water and was subsequently dehydrated under reduced pressure to retrieve the solid polymer from which the required solutions were produced as described later. Bovine serum albumin (BSA) produced from fatty acid and globulin-free lyophilized power (Sigma-Aldrich, U.K.) was used for the protein adsorption studies. (21) Goddard, J. M.; Hotchkiss, J. H. Prog. Polym. Sci. 2007, 32, 698–725. (22) Uyama, Y.; Kato, K.; Ikada, Y. Adv. Polym. Sci. 1998, 137, 1–39. (23) Uchida, E.; Uyama, Y.; Ikada, Y. Langmuir 1994, 10, 481–485. (24) Kato, K.; Uchida, E.; Kang, E.; Uyama, Y.; Ikada, Y. Prog. Polym. Sci. 2003, 28, 209–259. (25) Zhao, B.; Brittain, W. J. Prog. Polym. Sci. 2000, 25, 677–710. (26) Wang, P.; Tan, K. L.; Kang, E. T.; Neoh, K. G. J. Membr. Sci. 2002, 195, 103–114. (27) Zou, X. P.; Kang, E. T.; Neoh, K. G. Surf. Coat. Technol. 2002, 149, 119– 128.
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2.2. Atmospheric Pressure Plasma Treatment. The operational characteristics of the DBD reactor (Arcotec GmbH, M€ onsheim, Germany) have been described elsewhere.28-30 In brief, the DBD working electrode is driven by a sinusoidal high voltage AC power supply at a frequency in the range 40-80 kHz, to control the output power and the impedance of the operational working load. The discharge was generated in a gap (2 mm wide) created between three stationary tubular metal electrodes (diameter of 11.8 mm) arranged in parallel, and a 2 mm thick rubber covered aluminum platen electrode that slides under the metal electrodes to create the active discharge zone. The distance between the adjacent electrodes is 52 mm. The power dissipated between the electrode and plate system was measured by using a Tektronix TDS3052B (Tektronix Inc., Beaverton) digital oscilloscope coupled directly to the working electrode via a high voltage probe. Exposure of the sample surfaces to the atmospheric pressure plasma discharge in a given experimental condition can be described quantitatively by two parameters, namely, the power density (Pd) and the residence time of the substrate in the plasma (R) as described in eqs 1 and 2 below: p Pd ¼ ð1Þ ðwxlÞ R ¼ ð2xNÞ
w v
ð2Þ
where w is the total width of the discharge strip region, that is, the product of number of electrodes (3 in this case) and width of the discharge strip generated by each electrode on the sample surface (average value 1.5 mm), l is the length of the discharge region along the electrodes (in this case 210 mm), and N is the number of cycles. Each cycle consists of the plate sliding back and forth through the discharge zone with a transit speed denoted v and hence is equivalent to two periods of exposure (i.e., forward and backward). The energy dose delivered by the plasma, that is, the total energy imparted to the surface of the sample per unit area in J/cm2, is denoted in eq 3: D ¼
2NPd vl
ð3Þ
The values of Pd and R describe the total energy that the surface of a sample receives during exposure to the discharge in a given experimental run. Although Pd and R can be combined in a single experimental parameter of energy dose (D), it has been shown that changes in the discharge power can generate different effects to those obtained for the same energy dose but different processing times.30 For the purposes of this study, the power is fixed at 500 W and so the value of D is reported, with the relevant values of Pd and R provided for completeness.
2.3. Plasma Induced Grafting of PEGMA onto PS and PMMA. The MW 1000 and 2000 Da PEGMA materials were dissolved in methanol (Sigma-Aldrich, U.K.) at a concentration of 10% w/v. The polymer surfaces of interest were DBD treated as per the pretreatment conditions listed in Table 1, and samples were immediately dip coated in the two PEGMA-methanol solutions for 10 s. The coated polymer specimens were allowed to dry in an oven at 50 °C for 2 h and subsequently DBD treated again, as per the grafting treatment parameters given in Table 1. The grafted samples were washed repeatedly with methanol and submersed in Milli-Q ultrapure water for 16 h on a rocker table to remove any noncovalently bound PEGMA. Samples were dried under nitrogen and stored in sterile Petri dishes prior to analysis. (28) Upadhyay, D. J.; Cui, N.; Anderson, C. A.; Brown, N. M. D. App. Surf. Sci. 2004, 229, 352–364. (29) Upadhyay, D. J.; Cui, N.; Anderson, C. A.; Brown, N. M. D. Colloids Surf., A 2004, 248, 47–56. (30) Cui, N.; Upadhyay, D. J.; Anderson, C. A.; Brown, N. M. D. Surf. Coat. Technol. 2005, 192, 94–100.
DOI: 10.1021/la902654y
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D’Sa and Meenan Table 1. Experimental Conditions for DBD Treated and PEGMA Grafted Polymer Substrates DBD modification/ pretreatment
substrate nomenclature PS PSox0 PSox00 PS-g-PEGMA1000-(II) PS-g-PEGMA1000-(III) PS-g-PEGMA1000-(IV) PS-g-PEGMA2000-(I) PS-g-PEGMA2000-(II) PS-g-PEGMA2000-(III) PMMA PMMAox0 PMMAox00 PMMA-g-PEGMA1000-(II) PMMA-g-PEGMA1000-(III) PMMA-g-PEGMA1000-(IV) PMMA-g-PEGMA2000-(I) PMMA-g-PEGMA2000-(II)
2
2
D (J/cm )
Pd (W/cm )
R (s)
grafting molecule MW
D (J/cm2)
Pd (W/cm2)
R (s)
314.9 105.0 105.0 210.0 314.9 52.5 105.0 210.0
5.3 5.3 5.3 5.3 5.3 5.3 5.3 5.3
56.6 18.8 18.8 37.5 56.6 9.4 18.8 37.5
1000 1000 1000 2000 2000 2000
105.0 210.0 314.9 52.5 105.0 210.0
5.3 5.3 5.3 5.3 5.3 5.3
18.8 37.5 56.6 9.4 18.8 37.5
210.0 105.0 105.0 210.0 314.9 52.5 105.0
5.3 5.3 5.3 5.3 5.3 5.3 5.3
37.5 18.8 18.8 37.5 55.0 9.4 18.8
1000 1000 1000 2000 2000
105.0 210.0 314.9 52.5 105.0
5.3 5.3 5.3 5.3 5.3
18.8 37.5 55.0 9.4 18.8
2.4. XPS Analysis. X-ray photoelectron spectroscopy (XPS) was carried out using a Kratos Axis Ultra DLD spectrometer (Kratos, U.K.) at
