The invention discloses an environment-friendly biological oil extraction method, comprises the steps of injecting a biological activator into an oil field test area by using a high-pressure plunger pump through a water injection well, wherein the injection amount of the biological activator is 0.05 to 0.1 PV, the injection speed is 80-150 m<3> / d, the air injection amount is 2-3*10<5> Nm<3> / d, theinjection mode is continuous injection, the biological activator is prepared by mixing glycerol, molasses, NaNO3, yeast extract, K2HPO4, MgSO4, KCl, ethanol, FeSO4, CaCO3 and Na2MoO4, and the components and the weight ratio of the biological activator are determined through a screening experiment and an analysis experiment. The crude oil recovery rate can be improved, beneficial activities and metabolites of microorganisms are utilized, through the effects of degrading crude oil, reducing viscosity, increasing local pressure, increasing fluidity, changing oil-water interfacial tension, improving rock wettability and the like, remaining oil is further extracted, the technological content is high, economic benefits are good, and environmental protection and no pollution are achieved.