A preparation method of biological fracturing fluid for coal bed methane wells comprises the following steps: (1) screening hydrogenogens and methanogens; (2) acclimatizing related bacteria; (3) enriching the bacteria to prepare A liquid and B liquid, that is, adding the hydrogenogens separated and extracted in laboratory into a culture medium for enrichment for 3-6 days, adjusting the pH to 6.5 to obtain the A liquid, adding the methanogens into another culture medium for enrichment for 20-30 days, adjusting the pH to 7 to obtain the B liquid; (4) adding the A liquid and the B liquid in order according to on-site parameters. The mixed-bacteria fermented biological fracturing fluid prepared in the invention has certain sand-carrying capability; the bacteria in the fluid have hydrogen-producing and methane-producing capability, can not only increase the coal reservoir permeability, increase the coal bed methane resource amount, but also realize automatic viscosity reduction, and facilitate flow back. Residual hydrogenogens and methanogens after flow back can transform coal reservoirs continuedly, and generate biogenic coal bed methane continuously.