The invention relates to a method of extracting natural anti-oxidation active substance from plant materials, which discloses a method of extracting natural anti-oxidation active substance from oil orange tree bark. The method includes the following steps: (1) crushing; (2) extracting: the water is the solvent, the mass of the bark materials which have been crushed to the volume of water is 1 : 5g/ml to1: 10g/ml, the bark materials crushed are immersed in the water, the extracting solution is gained under the temperature ranging from 50 DEG C to 80 DEG C and under the effect of ultrasonic; (3) flocculating settling: the chitosan acetic acid solution is the flocculating agent; and the chitosan acetic acid solution is added into the extracting solution and stirred for 10 minutes to 30 minutes; (4) subzero fractionating: the extracting solution which is added with the flocculating agent is cooled and kept statically, the clear solution is extracted and the sediment is threw away; (5) theclear solution is added with 15 percent to 20 percent active carbon and 0.4 percent to 0.6 percent EDTA-2Na which are fixedly contained in the clear solution, the clear solution is filtered for the cleaning of the carbon after the clear solution is stirred for 0.5 hour to 1 hour, the filtrate is used as the upper-column liquid for the fractionation by adsorption of the resin; (6) fractionation byadsorption of the resin, by using desolvation, the powdered natural anti-oxidation active substance is gained by the vacuum freeze drying.