Kinetic study of NTPDase immobilization and its effect of haemocompatibility on polyethylene terephthalate.

Poor haemocompatibility of material surfaces is a serious limitation that can lead to failure of blood-contact devices and implants. In this work, we have improved the haemocompatibility of polyethylene terephthalate (PET) surfaces by immobilizing apyrase/ecto-nucleoside triphosphate diphosphohydrolase (NTPDase) on to the carboxylated PET. NTPDase immobilized PET surfaces scavenge the ADP released by activated platelets, which prevents further platelet activation and aggregation. The surface properties of the modified PET were characterized by scanning electron microscope (SEM), energy dispersive X-ray spectroscopy (EDAX), and contact angle measurement. The enzyme attachment and stability on the modified PET surfaces were evaluated. The kinetics of free enzyme and immobilized enzyme were studied and fitted using the Michaelis-Menten kinetic model. Both free and immobilized NTPDase followed Michaelis-Menten kinetics with similar Michaelis-Menten constants (Km). This suggests that the activity of NTPDase was unchanged upon immobilization. Protein adsorption and %hemolysis was significantly reduced for carboxylated PET and NTPDase immobilized PET surfaces compared to unmodified PET. Lactate dehydrogenase assay showed that the number of adhered platelets reduced by more than an order of magnitude for the NTPDase immobilized PET surface compared to unmodified PET. These results clearly indicate that NTPDase immobilization significantly enhances the haemocompatibility of PET surfaces.