title: Immobilization of an anticoagulant benzamidine derivative: effect of spacer arms and carrier hydrophobicity on thrombin binding.
abstract: Prevention of blood coagulation is very often a prerequisite for successful medical devices. For that purpose, passivation of the key coagulation enzyme thrombin through the derivatization of the material's surface with an amidine-based molecule has been found to be promising. To further enhance the efficiency of this approach, thin layers of maleic anhydride copolymers offering different physico-chemical characteristics were tethered with carboxyl terminated polyethylene glycol to covalently immobilize a benzamidine-type derivative. The free carboxyl surface groups produced by the attachment of polyethylene glycol (PEG) were quantified by Ag(+) labeling and subsequent XPS detection. The film thickness as well as the carboxyl group content were found to be clearly dependent on the copolymer hydrophobicity and the nature of the PEG molecule. For the assessment of the anchorage of the thrombin to the benzamidine-derivative functionalized surfaces, the substrates were immersed in a buffered thrombin solution and the enzyme adsorption was studied using immunostaining/confocal laser scanning microscopy. Higher degrees of thrombin binding were observed for substrates configured with the hydrophilic compared to the more hydrophobic copolymer. Moreover, surface-bound spacers based on alpha,omega-heterobifunctional PEG amino acids (alphaAm,omegaAc-PEG) also enhanced the benzamidine surface density in comparison to homofunctional PEG diacids (alphaAc,omegaAc-PEG) because of a lower degree of carboxyl inactivation due to PEG 'bridging'. Altogether, the choice of copolymer coatings and the type of PEG spacers were demonstrated to enhance the efficiency of the thrombin scavenging by the covalently immobilized coagulation inhibitor.
authors.affiliation: Department of Biomaterials, Leibniz Institute of Polymer Research Dresden eV and the Max Bergmann Center of Biomaterials Dresden, Germany.
authors.forename: Katrin
authors.initials: K
authors.lastname: Salchert
authors.forename: Marie-Françoise
authors.initials: MF
authors.lastname: Gouzy
authors.forename: Maja
authors.initials: M
authors.lastname: Glorius
authors.forename: Anett
authors.initials: A
authors.lastname: Kühn
authors.forename: Mirko
authors.initials: M
authors.lastname: Nitschke
authors.forename: Carsten
authors.initials: C
authors.lastname: Werner
mesh_terms: D000327:Adsorption; D001550:Benzamidines / Q000737:chemistry*; D001665:Binding Sites; D020099:Coated Materials, Biocompatible / Q000737:chemistry*; D003432:Cross-Linking Reagents / Q000737:chemistry; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D008422:Materials Testing; D011092:Polyethylene Glycols / Q000737:chemistry*; D011485:Protein Binding; D013917:Thrombin / Q000737:chemistry*; D013927:Thrombosis / Q000517:prevention & control
chemical_list: D001550:Benzamidines; D020099:Coated Materials, Biocompatible; D003432:Cross-Linking Reagents; D011092:Polyethylene Glycols; D013917:Thrombin; C032157:benzamidine