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- Title
Purification and characterization of endo b-1,4-D-glucanase from Trichoderma harzianum strain HZN11 and its application in production of bioethanol from sweet sorghum bagasse.
- Authors
Bagewadi, Zabin K.; Mulla, Sikandar I.; Ninnekar, Harichandra Z.
- Abstract
An acidophilic-solvent-thermostable endo β-1,4-D-glucanase produced from a potential Trichoderma harzianum strain HZN11 was purified to homogeneity by DEAE-Sepharose and Sephadex G-100 chromatography with 33.12 fold purification with specific activity of 66.25 U/mg and molecular mass of ~55 kDa. The optimum temperature and pH were 60 °C and 5.5 retaining 76 and 85% of activity after 3 h, respectively. It showed stability between pH 4.5-6.0 and temperature between 50-70 °C indicating thermostability. Endo β-1,4-D-glucanase was activated by Ca2+ and Mg2+ but inhibited by Hg2+, Pb2+ and Cd2+. The effect of thiol reagents, metal chelators, oxidizing agents and surfactants on enzyme activity has been studied. Purified endo β-1,4-D-glucanase exhibited highest specificity towards carboxymethyl cellulose. Kinetic analysis showed the Km, Vmax and Ki (cellobiose inhibitor) of 2.5 mg/mL, 83.75 U/mg and 0.066 M, respectively. The storage stability of purified endo β-1,4-Dglucanase showed a loss of mere 13% over a period of 60 days. The hydrolysis efficiency of purified endo β-1,4- D-glucanase mixed with cocktail was demonstrated over commercial enzyme. Optimized enzymatic hydrolysis of sweet sorghum and sugarcane bagasse released 5.2 g/g (36 h) and 6.8 g/g (48 h) of reducing sugars, respectively. Separate hydrolysis and fermentation of sweet sorghum bagasse yielded 4.3 g/L bioethanol (16 h) confirmed by gas chromatography-mass spectrometry (GC-MS). Morphological and structural changes were assessed by scanning electron microscopy (SEM) and Fourier transform infrared spectroscopy. Elemental analysis was carried out by SEM equipped with energy dispersive X-ray technique. These unique properties prove the potentiality of enzyme for biomass conversion to biofuel and other industrial applications.
- Subjects
TRICHODERMA harzianum; CELLULASE; ETHANOL as fuel; SORGO; GAS chromatography/Mass spectrometry (GC-MS); ENERGY dispersive X-ray spectroscopy
- Publication
3 Biotech, 2016, Vol 6, Issue 1, p1
- ISSN
2190-572X
- Publication type
Article
- DOI
10.1007/s13205-016-0421-y