The invention provides a vitamin B1 modified montmorillonite composite mycotoxin adsorbent, a preparation method and application thereof, and belongs to the technical field of non-metallic mineral materials. The adsorbent comprises montmorillonite and vitamin B1 between layers of the montmorillonite. The content of the vitamin B1 is 15-45% by mass of the montmorillonite. With the vitamin B1 as a modifier, the montmorillonite is modified since the vitamin B1 simultaneously has the polar hydrophilic group and the non-polar hydrophobic carbon chain, such that the obtained composite mycotoxin adsorbent has the amphiphilic property, can simultaneously adsorb the strong polar mycotoxin and the weak polar mycotoxin, has characteristics of high adsorption efficiency, no toxic-side effect on animals, and environmental protection; the invention also provides a preparation method of the vitamin B1 modified montmorillonite composite mycotoxin adsorbent, and the method is simple to operate and suitable for industrial mass production.