The impact of different physicochemical parameters of fermentation on extracellular cellulolytic enzyme production by Trichoderma harzianum

Abstract

This study was undertaken to find out the optimum physicochemical parameters of fermentation, i.e. pH, incubation temperature and incubation time for the cellulase enzyme production of Trichoderma harzianum. The extracellular protein content was estimated by the dye binding method of Bradford. Endo-glucanase (EG), exoglucanase (or Cellobiohydrolase; CBH), β-glucosidase and total cellulase activity were investigated. The molecular weight of cellulase enzymes was studied using SDS-PAGE. To identify the predominant catalytic components in optimum conditions of enzyme production, cellulases were separated by an adapted two-dimensional electrophoresis technique. Estimated optimum conditions for cellulase enzyme were found as: pH 6.5, incubation temperature 28°C and incubation time 72 h. The SDS-PAGE profiles showed several enzyme bonds such as CBHs, EGs and BGLs. The T. harzianum had both enzyme bonds of Cel7A (CBHI) and Cel7B (EG). Finally, the results of the 2D PAGE analysis showed that the profile of protein in optimium conditions of enzyme production had several enzymes such as CBHs, EGs and the high values of cellulose activity due to synergism that occurred between the CBH and EG.

Creating a novel genetic diversity of Trichoderma afroharzianum by γ-radiation for xylanase-cellulase production

“…The growing conditions were described above previously, and triplicate flasks were harvested after 72 h of incubation at 28 °C. The supernatant obtained from the centrifugation of TFM (4500× g , 7 min, and 4 °C) was used for the extracellular protein content (μg/mL) and enzyme (cellulase and xylanase) activity assay [ 22 ].…”

“…The bovine serum albumin (BSA) was used as a standard (0–500 μg/mL) for the protein content calculation. Also, cellulase and xylanase activity (U/mL) were determined by the dinitrosalicylic acid (DNS) method [ 22 , 24 ] by UV/Vis spectrophotometer (Jenway, USA) at 540 nm. The amount of enzyme that liberates 1 μmol of glucose or xylose per hour in a standard assay was defined as a unit (U) of cellulase or xylanase activity, respectively.…”

“…Estimation of the Mw of protein bands was performed using Gel-Pro Analyzer 6.0 densitometry software (Media Cybernetics, Inc.) [ 8 ]. The Mw range of the protein marker (Sinaclon Company, Iran) was 11–245 kDa (kDa) [ 22 , 25 ].…”