Influence of spacer length on heparin coupling efficiency and fibrinogen adsorption of modified titanium surfaces

Abstract

Abstract Background Chemical bonding of the drug onto surfaces by means of spacer molecules is accompanied with a reduction of the biological activity of the drug due to a constricted mobility since normally only short spacer molecule like aminopropyltrimethoxysilane (APMS) are used for drug coupling. This work aimed to study covalent attachment of heparin to titanium(oxide) surfaces by varying the length of the silane coupling agent, which should affect the biological potency of the drug due to a higher mobility with longer spacer chains. Methods Covalent attachment of heparin to titanium metal and TiO2 powder was carried out using the coupling agents 3-(Trimethoxysilyl)-propylamine (APMS), N- [3-(Trimethoxysilyl)propyl]ethylenediamine (Diamino-APMS) and N1- [3-(Trimethoxy-silyl)-propyl]diethylenetriamine (Triamino-APMS). The amount of bound coupling agent and heparin was quantified photometrically by the ninhydrin reaction and the tolidine-blue test. The biological potency of heparin was determined photometrically by the chromogenic substrate Chromozym TH and fibrinogen adsorption to the modified surfaces was researched using the QCM-D (Quartz Crystal Microbalance with Dissipation Monitoring) technique. Results Zeta-potential measurements confirmed the successful coupling reaction; the potential of the unmodified anatase surface (approx. -26 mV) shifted into the positive range (> + 40 mV) after silanisation. Binding of heparin results in a strongly negatively charged surface with zeta-potentials of approx. -39 mV. The retaining biological activity of heparin was highest for the spacer molecule Triamino-APMS. QCM-D measurements showed a lower viscosity for adsorbed fibrinogen films on heparinised surfaces by means of Triamino-APMS. Conclusion The remaining activity of heparin was found to be highest for the covalent attachment with Triamino-APMS as coupling agent due to the long chain of this spacer molecule and therefore the highest mobility of the drug. Furthermore, the adsorption of fibrinogen on the differently heparinised surfaces in real time demonstrated that with longer spacer chains the ΔD/Δf ratios became higher, which is also associated with better biocompatible properties of the substrates in contact with a biosystem.