In this paper, chitin nano-whiskers (CNWs) were prepared through acid hydrolysis. The surface deacetylation of the prepared CNW in alkali conditions was carried out. The morphologies, crystallinity, degree of deacetylation and antibacterial property of the nano-whiskers were studied by transmission electron microscopy (TEM), Fourier transform infrared spectra, X-ray diffraction, antibacterial testing and so on. The results confirmed that the amino groups were successfully introduced onto the surface of the CNW, so that the nano-rod-like morphology of the whisker was still maintained. The degree of deacetylation of CNW increased from 4.4% up to a maximum of 55.2% with the increase in NaOH concentration and reaction temperature. Furthermore, deacetylation kinetics of CNW was also investigated. It was found that the deacetylation process followed the pseudo-first-order kinetics. The apparent rate constant of the reaction increased with the increase in NaOH concentration and reaction temperature. The apparent activation energy of the reaction was estimated as 27.71 kJ/mol. The deacetylated CNW exhibited much improved antibacterial property, and the percentage bacterial reduction of the sample reached 85%.