The invention belongs to the technical field of air purification, and particularly relates to a biological enzyme air purifying agent as well as a preparation method and application thereof. The preparation method comprises: respectively dissolving specific enzyme species, extracting supernatants, mixing and compounding the supernatants, concentrating the mixture at a low temperature, and purifying the mixture to obtain an enzyme mixed solution; adding tea polyphenols, citric acid and an activator, stirring the components evenly, and then adding a specific microbial agent to prepare a biological enzyme air purifying agent. The biological enzyme air purifying agent can remove formaldehyde, benzene, xylene, TVOC and organic substances of other odor sources in the air for a long time throughdecomposition, and also has a sterilization effect, and the whole process is environment-friendly and nontoxic and does not produce secondary pollution.