The invention relates to the technical field of textile and specifically relates to a preparation method of toughened antimicrobial mulberry fiber. According to the preparation method disclosed by theinvention, mulberry bark is utilized as a raw material; after acid leaching treatment, the mulberry bark is put into a mixed solution; soda boiled mulberry bark can be obtained after the mulberry bark is boiled; the soda boiled mulberry bark is acid leached again to be neutralized; degummed mulberry fibers can be obtained after treatment; the degummed mulberry fibers are mixed with an N-methylmorpholine water solution to be put into a rotary evaporator; propyl gallate is added; an obtained colloidal spinning solution is treated to finally obtain toughened antimicrobial mulberry fibers. According to the preparation method disclosed by the invention, when the mulberry bark is subjected to soda boil treatment, metal ions around cellulose can be combined with hydroxyl in the cellulose in themulberry bark, a sodium silicate solution can form sol to permeate into the fibers, and fiber net structures in the mulberry fibers are prevented from being damaged; after the mulberry fibers are treated by the N-methylmorpholine water solution, adhesive force between the fibers and flavonoid matters is improved; thus, antimicrobial activity of the mulberry fibers is enhanced, toughness of the mulberry fibers is improved, and an application prospect is wide.