Kocher, Gurvinder SinghLota, Pardeep2018-01-182018-01-182017http://krishikosh.egranth.ac.in/handle/1/5810039556The present study was conducted with the objective of standardization of a fungal consortium from selected ligninolytic cultures for biological pretreatment of paddy straw and its subsequent conversion to bioethnaol. The proximate analysis of paddy straw revealed that 30 mesh straw had highest proportion of cellulose (39.5%) besides, 23.3% hemicellulose and 11.5% lignin. Among different chemical pretreatment methods, acid (1.0% H2SO4) –autoclave (15 psi for 90 min) followed by alkali (2.0% NaOH) pretreatment resulted in 82.61% and 59.23% decrease in lignin and hemicellulose, respectively and 36.60% increase in relative proportion of cellulose. Four fungal strains (viz., LS1, LS2, LCu1 and LCu2) from a total of fifty nine strains isolated from diverse natural habitats, tested positive for the production of laccase (Lacc), lignin peroxidase (LiP) and manganese peroxidase (MnP). Out of three consortia developed on the basis of ligninolytic activities, consortium III comprising Pleurotus ostreatus and Phanerochaete chrysosporium, resulted in Lacc, LiP and MnP activities of 2.40, 37.92 and 62.50 nkat/gds, respectively, significantly higher than the activities of individual fungal strains. Biological pretreatment of paddy straw under Response surface methodology optimized conditions (moisture, 121.0 %; temperature, 31.3°C and log spore count, 8.0 spores/ml) resulted in cellulose, 43.0%; hemicellulose, 12.7%; lignin, 7.0%; total sugars,13.65 mg/gds and reducing sugars, 4.00 mg/gds. The pretreatment of paddy straw with concentrated (10x) ligninolytic enzyme under shake flask conditions using optimized physico-chemical parameters viz. paddy straw concentration, 2.5 g; enzyme volume, 8.0 ml; Ca2+ ions (10 mM) and incubation temperature, 45°C in 72 h of enzymatic action resulted in 80.87% and 47.64% decrease in lignin and hemicellulose, respectively and 29.96% increase in relative proportion of cellulose. For saccharification studies, from a total of forty seven fungal strains isolated from diverse natural sources, two strains (Aspergillus sp. CTS1 and Aspergillus sp. CTS2) tested positive for thermophilic and cellulolytic nature. The two strains were used for the development of fungal consortium which revealed filter paper, carboxymethyl cellulase and cellobiase activities of 10.2, 30.0 and 7.9 U/gds, respectively, significantly higher than the activities of individual fungal strains. Under the RSM optimized conditions [temperature, 55°C; pH,4.8 and substrate concentration, 5% (w/v)] and 30FPU/g dry substrate Arrowzyme yielded 476.0 mg/gds reducing sugars from biological pretreated paddy straw which were further increased to 492.0 mg/gds by the addition of Tween 20 at 0.2% (v/v). Enzymatic saccharification of biological pretreated straw by in-house concentrated cellulase (CC1) from thermophilic fungal consortium resulted in 229.0 mg/gds reducing sugars. The saccharification of chemical pretreated straw with Arrowzyme and CC1 resulted in 566.0 mg/gds and 250.0 mg/gds reducing sugar, respectively. Optimization of fermentation under separate hydrolysis and fermentation (SHF) conditions using RSM [S.cerevisiae inoculum size 10.0 % (v/v) and DAHP concentration, 0.2% (w/v)] resulted in 0.172 g/g and 0.193g/g ethanol from biological and chemical pretreated hydrolysate, respectively. Similarly, the CC1 saccharified hydrolysate resulted in an ethanol content of 0.076 g/g (biological pretreated) and 0.085 g/g (chemical pretreated). Fermentation of non-enzymatically saccharified biological and chemical hydrolysate by S.cerevisiae produced 0.013 g/g and 0.041 g/g ethanol, respectively, whereas sequential fermentation of chemical hydrolysate with S.cerevisiae and Pachysolen tannophilus produced 0.047g/g ethanol. The combined ethanol production from enzymatically and non-enzymatically saccharified biological and chemical pretreated paddy straw hydrolyaste was observed to be 0.185 g/g and 0.240 g/g, respectively.ennullThesis