Enzymatic hydrolysis of lignocellulose is generally considered as the major economic bottleneck of the bioethanol from lignocellulose production process. Among the strains that can produce cellulolytic enzymes, Trichoderma reesei is the fungal strain commonly used for the industrial production of enzymes for lignocellulose hydrolysis, especially because of its high capacity of enzyme secretion (>30g.L^-1 protein for industrial strains). Understanding the complexity of the synergistic mechanisms involved will result in defining the optimal ratios between processive and non-processive cellulolytic enzymes. The present approach allows a rapid investigation of multiple cellulase ratios not naturally found in fungal secretomes. This analysis is based on the separation of the main cellulolytic activities present in the secretome of the cellulolytic strain T. reesei. The fractionation is carried out by Fast Protein Liquid Chromatography (FPLC) and the separation method used leads to the purification of the main active enzymes, i.e. CBHI, CBHII, EGI and EGII, up to 94% without loss of activity. The separated enzymatic fractions are used in a miniaturized degradation test of pre-treated lignocellulosic substrates. The degradation capacities are determined by the final sugar production and hydrolysis yields using HPLC. These tests were used to evaluate the effects of these reconstituted pools on the degradation of a steam-exploded wheat straw. Multiple combinations of different purified enzymes simulated in vitro variations of each cellulase and  provided new information on their respective roles in lignocellulose degradation.