The invention belongs to the technical field of microbial fermentation, and particularly relates to a preparation method of a probiotic inoculant. In the invention, a high-yield gamma-aminobutyric acid inoculant is obtained from the separation and screening of yogurt and pickled vegetables. The gamma-aminobutyric acid inoculant can increase organism appetites, promote growth, regulate intestinal flora and improve the absorlption effect of alimentary canals, and has health-care effects of promoting nutrient absorption and promoting growth. In the invention, flavonoids in paper mulberry leaves and saponin substances in stauntonia chinensis are extracted, and the flavonoids and the saponin substances are important growth factors for probiotics, and thorough the cooperation with the probiotics, the flavonoids and the saponin substances can improve the survival rate of the probiotics, prolong the action time of the probiotics, lead the probiotics to reach the body in the largest quantity, and stimulate the colonization and growth of endogenous and exogenous probiotics of intestinal tracts, and can also enhance the metabolic activity of the probiotics, enhance the immune functions of theorganisms, directly antagonize pathogens, and stimulate the production of endogenous antibiotic substances of the organisms.