Studies on laccase from Pleurotus ostreatus NCIM 1200 Production optimization under solid state fermentation characterization and application

Abstract

newlineThe organism Pleurotus ostreatus NCIM1200 obtained from NCIM, newlinePune was used for the laccase production and characterization study under newlinesolid state fermentation condition using pineapple leaves as the substrate. newlineEleven different process parameters were selected to study their influence on newlinemaximal enzyme production through one-factor-at-a-time method. Further newlinestatistical optimization was performed using Design Expert 8.0 package which newlineresulted in the selection of five significant parameters viz. inoculum newlineconcentration, substrate size, substrate concentration, initial pH of the newlinemedium, additional carbon source, for laccase production. An overall 7.1 fold newlineincrease in laccase production was obtained after experimental validation of newlineRSM model. The laccase was purified by three steps including salt-out with newline(NH4)2SO4, ion-exchange chromatography with Q-Sepharose and gel filtration newlinechromatography with Sephadex G-100. The complete purification procedure newlineof extracellular laccase having a molecular mass of and#8776; 64.2 kDa, resulted in newline25% of yield and 2.5-fold of purification. The purified laccase showed a pI of newline3.7. Kinetic studies on the purified enzyme suggested the higher affinity of newlinelaccase enzyme towards phenolics rather than aromatic compounds. UVvisible newlinespectral studies confirmed the enzyme as white laccase lacking T1 newlinecopper site. The laccase gene specific sequence was isolated from Pleurotus newlineostreatus NCIM 1200 using primers corresponding to the highly conserved newlinecopper-binding regions of laccase enzyme and it was subjected to newlinecomputational analysis using various bioinformatic tools. The organism newlinePleurotus ostreatus NCIM 1200 and its extracellular enzyme laccase was newlinesuccessfully used for the decolourization of various synthetic textile dyes newlineincluding indigo carmine, amido black, evans blue, brilliant green, orange G newlineand acid yellow. Although this organism appears to lack Lignin Peroxidase, newlinethought to be necessary for lignin degradation, was effectively used for the newlinedegradation of lignin in coir retting field effluent.