Cellulase Production by Penicillium Oxalicum Ti-11 with Traditional Chinese Medicine Residue as Substrate

PurposeThe study aims to search for efficient cellulase producer and explore the possibility of traditional Chinese medicine residue as a substrate for cellulase production, so as to realize the waste utilization of traditional Chinese medicine residue.MethodsThe cellulase-producing strain was identified through morphological and molecular biological methods. Scanning electron microscope (SEM) and Fourier transform infrared spectroscopy (FTIR) were used to characterize the structure of traditional Chinese medicine residues before and after fermentation. The enzyme activity was determined by DNS method, and the enzyme production conditions were optimized by single factor and response surface methodology.ResultThe strain grew well in forsythia leaf residue, and the highest FPA could reach 2.06 IU/mL. In addition, the structural characteristics of traditional Chinese medicine residue that before and after enzymatic hydrolysis were characterized by SEM and FTIR. The results showed that the structure of the residue was destroyed after enzymatic hydrolysis, the damage of forsythia leaf residue was the most serious, and enzymatic hydrolysis promoted the dissolution of cellulose, lignin and hemicellulose. The enzyme production conditions of the strain were optimized by Plackett-Burman design and response surface analysis. The FPA could reach 2.79 IU/mL under the optimal conditions of FLR concentration 24.84 g/L, (NH4)2SO4 concentration 2 g/L, temperature 34.44℃, pH 6.20, rotational speed 200rpm, inoculum 6%, which was 35.44% higher than that before optimization.ConclusionsThe results showed that traditional Chinese medicine residue could be used as the induced substrate for fungal cellulase production. This study provides an idea for the low-cost production of fungal cellulase and the waste utilization of traditional Chinese medicine residue.

Cellulase production and efficient saccharification of biomass by a new mutant Trichoderma afroharzianum MEA-12

Background Cellulase plays a key role in converting cellulosic biomass into fermentable sugar to produce chemicals and fuels, which is generally produced by filamentous fungi. However, most of the filamentous fungi obtained by natural breeding have low secretory capacity in cellulase production, which are far from meeting the requirements of industrial production. Random mutagenesis combined with adaptive laboratory evolution (ALE) strategy is an effective method to increase the production of fungal enzymes. Results This study obtained a mutant of Trichoderma afroharzianum by exposures to N-methyl-N’-nitro-N-nitrosoguanidine (MNNG), Ethyl Methanesulfonate (EMS), Atmospheric and Room Temperature Plasma (ARTP) and ALE with high sugar stress. The T. afroharzianum mutant MEA-12 produced 0.60, 5.47, 0.31 and 2.17 IU/mL FPase, CMCase, pNPCase and pNPGase, respectively. These levels were 4.33, 6.37, 4.92 and 4.15 times higher than those of the parental strain, respectively. Also, it was found that T. afroharzianum had the same carbon catabolite repression (CCR) effect as other Trichoderma in liquid submerged fermentation. In contrast, the mutant MEA-12 can tolerate the inhibition of glucose (up to 20 mM) without affecting enzyme production under inducing conditions. Interestingly, crude enzyme from MEA-12 showed high enzymatic hydrolysis efficiency against three different biomasses (cornstalk, bamboo and reed), when combined with cellulase from T. reesei Rut-C30. In addition, the factors that improved cellulase production by MEA-12 were clarified. Conclusions Overall, compound mutagenesis combined with ALE effectively increased the production of fungal cellulase. A super-producing mutant MEA-12 was obtained, and its cellulase could hydrolyze common biomasses efficiently, in combination with enzymes derived from model strain T. reesei, which provides a new choice for processing of bioresources in the future.

Enhancement of Indigenous Fungal Cellulase Production by Gamma Rays

The increasing demand for cellulases causes the need for a high cellulase-producing microbe. Mutagenesis is an efficient way to produce a high-titer cellulase-producing strain. Mutagenesis using gamma rays irradiation has the advantage that it can cause a double strand break of DNA. Repair of double-strand break tends to has an error-prone repair that leads to the alteration of DNA sequence. The aim of this study was to screen high cellulase-producing indigenous fungal mutants produced by mutagenesis. Trichoderma sp. PK1J2 was subjected to gamma irradiation at 300 Gy. The mutants produced were screened using a plate medium containing cellulose as a sole carbon source. After staining with congo red, colonies with wider clear zones were grown in a liquid medium for four days, and the cellulase activities were analyzed. Mutant M8 produces endoglucanase, FPase, and β-glucosidase at 0.46 U/ml, 0.18 U/ml, and 1.10 U/ml, respectively, which were 90%, 50%, and 30% higher than those of the parental strain.

Analytical study on Fungal Cellulase Produced by Penicillium Expansum grown on Malus Domestica (Apple Fruits)

The rise in world industrialization and the cost of importing enzyme by local industries have led to arise in the search for novel and native enzyme producing microorganisms. Cellulase is an enzyme that catalyzes the breaking down of carbon chains in cellulose and hemicellulose, this research therefore aimed at studying fungal cellulase produced by Penicillium expansum grown on malus domestica (apple fruits). Fresh apple fruit was allowed to deteriorate under laboratory condition until there was visible mould growth. The mould with desired features of the organism of interest was subcultured by direct plating on PDA plates to which 10 % streptomycin has been added to prevent bacterial contaminants. The plates were incubated at 28±2 0C for 7 days until a visible mass of blue mycelia appear. The isolate was further subcultured onto freshly prepared media until pure culture was obtained. Characterization and identification of isolate were done using macroscopy and microscopy techniques. The isolate was re-inoculated into healthy apple fruits and the fruits were incubated at temperature of 28±2 oC for 8 days. Cellulolytic activity was examined every day throughout the incubation period. Crude enzyme was extracted each day using standard methods. Carboxyl methyl cellulose was used as standard for the crude cellulase activity assay after extraction from the infected apple fruits using Dinitrosalicylic acid (DNSA). Culture parameters like pH and temperature were also optimized to determine their effect on cellulolytic activity of the fungus. Cellulase activity was defined as the amount of glucose produced in μmol/mg/min under the assay condition. The highest cellulase activity of 86.84±0.52 μmol/mg/min was observed on day 6 of incubation at 28±2 oC and at pH 7. In conclusion, it is evident from this research that P. expansum isolated could be used as potential novel organism for industrial production of cellulase under optimized fermentation conditions.

COMPLEX OF ENZYMES FOR EFFICIENT CONVERSION OF DRIED APRICOT BIOCOMPONENTS IN THE PRODUCTION OF FOOD INGREDIENTS FOR ALCOHOLIC BEVERAGES

The effectiveness of the use of pectolytic enzyme preparations - Fructocim P6L and fungal cellulase - Bruzaym BGX for biocatalysis of high-molecular substances of dried apricot pulp in the preparation of food ingredients (alcoholic juices and fruit drinks) for alcoholic beverages is shown.