Experimental study on the antibacterial property and cytocompatibility of medical biodegradable zinc alloy materials in vitro

Abstract

Objective To study the antibacterial property and cytocompatibility of medical biodegradable zinc alloy in vitro. Methods Escherichia coli and Staphylococcus aureus strains were respectively mixed with LB culture medium, the concentration of 1.0×108CFU/mL was regulated by the bacterial turbidimetric instrument. The bacteria was cultured in carbon dioxide incubator of 37℃ for 24 h as bacteria original liquid. The same size of zinc alloy and titanium alloy (Ti6Al4V) rods were prepared for experiment, the surface oxide layer was polished and removed, which was cleaned ultrasonically in 100% ethanol and distilled water, disinfected with ethylene oxide, standby. (1) Escherichia coli/Staphylococcus aureus strains were respectively co-cultured in LB culture medium with biodegradable zinc alloy, titanium alloy. The absorbance value at the time of 0、2、4、6、8、12、24、48、72 h after cultured at 600 nm wavelength was determined by enzyme-linked immunosorbent assay.(2) Zinc alloy and titanium alloy were respectively placed on Staphylococcus aureus strains solid culture plates, bacteriostatic circles were observed at 24、48 h after cultured. (3) L929 cells were cultured with different concentrations of zinc alloy extracts, cell morphology were observed at the time of 1、5 d after cultured, CCK8 method was used to detect the cell proliferation and cell relative rate of increment(RGR) was calculated. (4)The MC3T3-E1 cells were seeded respectively on zinc alloy surface and titanium alloy to observed adhesion and growth state 2 days after cultured. Single factor analysis of variance (ANOVA) and t test were used to compare the data between the groups. Results The absorbance values of Escherichia coli and zinc alloy co-culture group were significantly lower than Escherichia coli culture group, Escherichia coli and titanium alloy co-culture group at different phase points, the differences were statistical significant(P values were less than 0.05). The absorbance values of Staphylococcus aureus and zinc alloy co-culture group were significantly lower than Staphylococcus aureus culture group, Staphylococcus aureus and titanium alloy co-culture group at different phase points, the differences were statistical significant(P value was less than 0.05). Bacteriostatic circles were observed near biodegradable zinc alloy, no bacterial colony appeared around (4.38±0.40)mm near the zinc alloy at 24 hours after cultured, while no bacterial colony appeared around (4.75±0.44)mm at 48 hours after cultured, no significant difference existed between the diameter of bacteriostatic circles (t=-1.10, P=0.31), no bacteriostatic circle was observed near the titanium alloy. The growth of L929 cells were good in the culture and negative control group with different concentrations of zinc alloy. CCK8 assay suggested that cell RGR in the leaching solution groups were more than 75%, the cell toxicity was 0-1 grade at different time points, the cytotoxicity of zinc alloys was safe and acceptable. MC3T3-E1 cells could grow and adhere on the surface of zinc and titanium alloy. There was no abnormalities compared with the titanium alloy group in cell morphology. Conclusion Zinc alloy material has good antibacterial activity against Escherichia coli and Staphylococcus aureus, and has good cellular compatibility, the cell toxicity is safe and acceptable, and cells can grow and adhere on the surface of it. Key words: Zinc alloys; Biodegradable implants; Antibacterial property; Cytotoxicity