Study of the protein adsorption on the catechol functionalized polyelectrolyte multilayer film on titanium surface

Objective To provide a theoretical basis for surface modification of titanium implants, the protein adsorption of the catechol functionalized polyelectrolyte multilayer film on titanium surface was explored. Methods According to the methods established in our previous study, catechol functionalized lipopolysaccharide-amine nanopolymersomes (cNPs) grafting rate of 40% was prepared by the reaction of lipopolysaccharide-amine amino nanopolymersomes (NPs) and 3, 4-dihydroxy phenyl propionic acid. The catechol functionalized hyaluronic acid (cHA) grafting rate of 10% was prepared by the reaction of hyaluronic acid (HA) and dopamine. Via layer-by-layer technique and initiated by cNPs, catechol functionalized polyelectrolyte multilayer film (cPEM) was constructed on titanium or quartz surface by using cHA and NPs, containing 3 (cHA/NPs) catechol double polyelectrolyte membrane. Similarly, initiated by NPs layer, polyelectrolyte multilayer film (PEM) without catechol was constructed on titanium or quartz surface by using hyaluronic acid (HA) and NPs. The chemical composition of the film surface was analyzed by infrared spectrum, and the surface roughness was measured by laser scanning confocal microscope (LSCM) . Zeta potential of film surface was recorded with a Zeta potential analyzer. Four kinds of proteins, namely Bovine serum albumin (BSA, pI = 4.7) , Fibronectin (Fn, pI = 5.8) , Bovine hemoglobin (BHb, pI = 6.8-7.0) and Polylysine (PLL, pI = 9.74) , were selected, as their isoelectric points (pI) were less than, equal to and greater than physiological pH 7.4, respectively. Solutions with proteins above (1 mg/mL) were prepared with 0.15 mol/L NaCl. The real-time protein adsorption process was monitored by using quartz crystal microbalance (QCM) . Atomic force microscopy was used to observe the morphology of protein before and after adsorption. The adsorption of fluorescently labeled proteins on the membrane surface was analyzed by LSCM and fluorescence microplate reader, respectively. The data were analyzed by One-Way ANOVA, SNK and LSD tests with SPSS 20.0 software package. The difference was statistically significant with P<0.05. Results The results of LSCM showed that the surface roughness of quartz was (301 ± 12) nm, and the surface roughness increased after assembling into cPEM and PEM, which were (656 ± 88) nm and (446 ± 25) nm, respectively. The three groups showed statistical difference (F = 66.974, P PEM > Ti. Atomic force microscopy showed that the surface of cPEM and PEM groups was uniform distribution of water-drop and island-like structure. After adsorption of BSA, the disk structure was observed on the surface, and the amount of cPEM was larger than that of PEM, suggesting that the adsorption amount of BSA on cPEM surface was more than that on PEM. Observation under LSCM and the results from fluorescence microplate reader showed that the adsorption amount of protein on quartz surface increased after being catechol functionalized. On the surface of the same kind of membrane, different proteins showed the capacity of BSA > Fn > BHb. Conclusions Protein adsorption on titanium surface can be improved by the modification of polyelectrolyte multilayer film, which can be further promoted by catechol functionalized modification. The driving force of protein adsorption may be mainly due to electrostatic interaction and the catechol catching influence on protein coupling. Key words: Titanium; Protein, adsorption; Catecholed polyelectrolyte multilayers