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Various pretreatment methods such as dilute sulfuric acid and aqueous ammonia have been employed for biomass pretreatment, but they do not remove much of lignin portion and the effect is limited. Those methods work biomass to be exposed polysaccharide portion but at that step, xylan and glucan portion also could be removed by the reagent strength. Peracetic acid is promised reagent to remove the lignin portion selectively unlike the other reagents remove some of the sugars and are therefore undesirable. Peracetic acid could be converted from ethyl acetate and hydrogen peroxide by enzymatic biosynthesis using acetyl xylan esterase. In this study, recombinant acetyl xylan esterase was produced from Aspergillus ficuum expressed in Pichia pastoris [1]and immobilized on graphite supporter, and peracetic acid production was performed using immobilized acetyl xylan esterase. Immobilization was conducted at various conditions of temperature, reaction time and acidity. The condition ranges were employed to be 4-60°C of temperature, 3-48 h of reaction time and pH 4-9 of acidity. Optimized immobilization condition was determined to be 4°C of temperature, 24 h of time and pH 7 of acidity. Enzyme activity assay was also performed after immobilization, and the activity was found to be 170 U/g-matrix. Finally peracetic acid was produced using the immobilized acetyl xylan esterase, and approximately 4.91 mM of peracetic acid production was confirmed.