Abstract: By incorporating organic-inorganic nano-hybridization into wet phase inversion coating-forming method, a novelantimicrobial polyurethane synthetic leather coating with in-situ generated nano-TiO 2 (PUT) was prepared. The antibacterialactivity against Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, Micrococcus luteus, antifungal activityagainst Aspergillus niger and toxicity of this PUT coating were investigated respectively. Experimental results showed thatthe antimicrobial activity of the PUT coating increased with increasing nano-TiO 2 concentration. Low nano-TiO 2 concentra-tion (≤0.50 wt%) resulted in negligible or limited antimicrobial activity. When the nano-TiO 2 concentration increased up to0.75 and 1.00 wt%, the antibacterial activity of the PUT coating exceeded 82 and 93 % respectively, and no Aspergillus nigergrowth was observed on the coating surface within 28 days. Cell culture assay indicated that the PUT coating had no detri-mental effect on the morphologies and proliferation rate of normal human dermal fibroblasts, which indicated a non-toxic andskin-friendly characteristic. According to these results, the PUT coating was capable of reducing the risk of microbial con-tamination while remained skin-friendly to wearers. More importantly, the organic-inorganic nano-hybridization techniquedeveloped in this study was carried out simultaneously during the established wet phase inversion coating-forming methodfor PU leather manufacture, which promised its possible application on an industrial scale.