The invention provides a preparation method of zinc oxide affinity anti-ultraviolet and antibacterial silk fibers. The preparation method includes the steps: performing degumming on silk after impurities in the silk are removed, washing the degummed silk by the aid of deionized water, and drying the washed silk to obtain the degummed silk; performing activating treatment on the degummed silk by the aid of strong oxidizing agents, soaking the degummed silk after activating treatment into zinc nitrate water solution, filtering and washing the soaked degummed silk, soaking the filtered and washed degummed silk into ammonium hydroxide, adjusting temperature, and keeping the temperature to obtain pre-covering silk fibers; adding dichloromethane into the pre-covering silk fibers, uniformly mixing mixture, adding polyethylene glycol monomethyl ether, lactide and stannous iso caprylate, mixing mixture, performing microwave treatment, filtering and washing on the mixture under protection of nitrogen, standing in heating and humidifying steam, and washing and drying the mixture to obtain the zinc oxide affinity anti-ultraviolet and antibacterial silk fibers. The obtained silk fibers have excellent ultraviolet resistance, inhibit and resist bacteria, remove odor and are resistant to soak and wash, good in moisture absorption performance, and a user feels comfortable when wearing a product made of the silk fibers.