Comparative analysis of optimal endoglucanase production by Trichoderma reesei and intergeneric fusants of Trichoderma reesei Saccharomyces cerevisiae

Abstract

Intergeneric fusants of Trichoderma reesei QM 9414/Saccharomyces cerevisiae NCIM 3288 developed in the authors' laboratory can convert cellulosic materials directly to ethanol in a single step process. The production of endoglucanase in this case is a key factor. The production pro®le of this enzyme by the intergeneric fusants is different from Trichoderma reesei QM 9414 (WT). The production of endoglucanase was studied seperately by Trichoderma reesei (WT) using optimal production medium which was designed as per the combined screening approach of Plackett-Burman followed by a central composite experimental plan and the intergeneric fusants using optimal production medium obtained by Box-Behnken optimization procedure. Dried grass was used as the cellulosic substance whose concentration was kept constant during the statistical optimization procedure. The concentration of dried grass was later varied keeping the other optimized medium constituents constant to ®nd the ®nal optimum medium composition for endoglucanase production. List of symbolsvalue of x i at the centre point of the investigated area Dx i step change IntroductionWith the reserves of non-replenishable energy sources slowly getting exhausted, efforts are on for tapping energy from the replenishable ones. Since ethanol can be obtained from renewable resources, it provides a promising alternative fuel source. Besides, ethanol is also used as a solvent, a germicide, an antifreeze and a chemical intermediate [1]. Ethanol is produced by fermentation by the S. cerevisiae-molasses technology. However, scanty and uneven supply of molasses and its higher consumption for more costly chemical and biochemical production renders severe handicap to this technology [2,3]. Ethanol is also produced from cellulose, an abundant biomass. So far only multistage approach of ethanol production has been taken up, which has its own limitations of being capital intensive, time consuming and low yielding [4]. Hence, in order to achieve a single stage conversion of cellulose to ethanol, Trichoderma reesei QM 9414, a hyper-cellulase producer, was fused with Saccharomyces cerevisiae NCIM 3288, a potent ethanol producer, in the authors' laboratory [5].The resultant fusant has been found to be stable and produce ethanol from cellulosic substances [6]. Production of endoglucanase, one of the enzymes responsible for the conversion of cellulosic materials to fermentable sugars, is a key factor in this case. Comparative analysis of endoglucanase production by the intergeneric fusants of T. reesei/S. cerevisiae and T. reesei (WT) under optimal production conditions using response-surface methodology has been presented. 2 Materials and methods 2.1 Microorganisms Trichoderma reesei QM 9414 was obtained from the National Chemical Laboratory, Pune, India. The intergeneric fusants of Trichoderma reesei QM 9414/Saccharomyces cerevisiae NCIM 3288 was developed in the authors' laboratory [5]. 2.2 Culture maintenance T. reesei QM 9414 was maintained on malt-agar slants, having (kg/m 3 ): malt-e...

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