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Xylanase enzyme is extracellular enzyme that plays a role in hidrolyzing xylan into short chains of xylose and xylooligosaccharides. The xylanase enzyme is one of the most important enzymes in various industrial sectors including the paper, textile and xylitol industries. Xylanase enzyme are produced by microorganisms such as bacteria, yeast and fungi with the help of xylan as an xylanase enzyme induction medium. The use of commercial xylan in industrial scale xylanase production is not economical because the price is too expensive $ 180-200/kg, so that an alternative is needed to use xylan from agricultural waste such as bagasse and wheat bran. Bagasse and wheat bran contain higher xylan compared to other agricultural wastes are 29.2% and 43.2%. This study aims to determine the optimal concentration in mixing bagasse and wheat bran substrates to the xylanase enzyme activity produced by Aspergillus niger and Aspergillus oryzae through the 2,4 dinitrosalicylic acid method. The test results of the xylanase enzyme activity of Aspergillus niger and Aspergillus oryzae showed that xylanase production increased with increasing concentration of wheat bran and decreased by adding concentration of bagasse. Based on the results of the enzyme activity test it can be concluded that the substrate which produces high xylanase enzyme at 100% wheat bran is 6.6 U / mL, while mixing the wheat bran and bagasse substrate which produces high xylanase activity at a concentration ratio of 75% wheat bran added 25% bagasse that is equal to 5.4 U / mL. The xylanase enzyme activity produced by Aspergillus oryzae was higher than Aspergillus niger which was 4.49 U / mL.