Isolation, Molecular Identification, Mutation Induction of Egyptian Fungus Aspergillus niger NM-NRC and Medium Optimization Using Response Surface Methodology for Pectinase Hyper Production

Abstract

A class of enzymes called pectinases aids in the digestion of pectic substances. In the food business, it's commonly used to produce  and clarify juices and wines. The aim of this study was to isolate, assess, and examine pectinase enzymes from fungi derived from various soil specimens. A total of 15 fungal isolates were first screened from 10 distinct soil samples. Out of them, only one exhibited the most pronounced pectinolytic activity was isolate No. 1 exhibited the greatest enzyme output after a 3 days incubation period, with a substrate citrus pectin concentration of 1% and a temperature of 30◦C. The enzyme production was measured at 51.47 U/mg. Internal transcribed spacer (ITS) sequencing and PCR were used to confirm the molecular identity of isolate No. 1, which was determined to be Aspergillus niger NM-NRC. The results were deposited in the NCBI database under the accession numbers  OQ600201. In order to increase the efficiency of pectinase production in this strain, a series of mutagenesis techniques
were employed, including ultraviolet (UV) radiation, ethidium bromide (EtBr), and hydrogen peroxide (H2O2). A number of mutants were produced, and the mutant H-34 from Aspergillus niger NM-NRC was shown to be the most effective in breaking down pectin, with a pectinase activity of 113.43 U/mg. Response Surface Methodology (RSM) was used to establish the ideal conditions for pectinase
expression in the Aspergillus niger NM-NRC mutant H-34. The optimal culture conditions for achieving the greatest pectinase-specific activity of 164.45 U/mg were a temperature of 30oC, pH of 9, 48 hours of incubation, and a pectin concentration of 2%. Additionally, 1.5% glucose and 1.5% beef extract were used as carbon and nitrogen sources, respectively. In conclusion, the strain Aspergillus niger NM-NRC mutant H-34 has been determined to be highly effective in producing pectinase on a large scale for commercial use.