Adsorption of cellulase on fibers is a key factor in determining its efficiency on fiber treatment and microfibrillated cellulose (MFC) preparation. Different adsorption behavior, treatment efficiency and performance of MFC and MFC film were observed due to the different properties of cellulases. Herein, bleached eucalyptus kraft pulp (BHKP) was pretreated by complex cellulase (D cellulase) and endocellulase (R cellulase) with different dosages for MFC preparation. Enzyme activity, adsorption ratio, adsorption kinetics and adsorption thermodynamic of the two cellulases were comprehensively studied, and the impacts of the cellulase pretreatment on the properties of MFC and MFC film were investigated. The results showed that D cellulase possessed higher adsorption ratio than R cellulase, but R cellulase demonstrated higher adsorption rate than D cellulase. High temperature discouraged the adsorption of the two cellulases because of their exothermic natures. The crystallinity index (CrI), specific surface area (SSA) and morphology of MFC were tuned by the combination of two cellulases at different dosages. The CrI of MFC treated by D cellulase and R cellulase increased from 40.45% to 66.50% and 66.67% respectively when the cellulase dosage was 10 U/g. The elongation at break (E) and tensile strength (TS) of MFC film treated by D cellulase were decreased first and then increased slightly, but the MFC film treated by R cellulase decreased continuously. The MFC film prepared by D cellulase possessed the best barrier property at 20 U/g and the corresponding oxygen permeability coefficient was 4.37×10-14 cm3·cm/cm2·s·Pa. However, the oxygen permeability coefficient of MFC film pretreated by R cellulase at a dosage of 10 U/g was 4.13×10-14 cm3·cm/cm2·s·Pa. This work shows that R cellulase was more suitable than D cellulase for BHKP pretreatment to prepare MFC film.