PRODUCTION OF XYLANASE FROM FUNGI ASSOCIATED WITH THE RHIZOSPHERE SOIL OF CASSAVA (Manihot esculenta Crantz) TUBER

Abstract

This research is aimed at screening fungi associated with rhizosphere soil of cassava tuber for xylanase production, optimization, purification and characterization. Soil samples were collected from the Federal University of Technology, Akure, Nigeria and transferred to the laboratory for physicochemical and microbiological analyses. Microorganisms associated with the samples were isolated and identified using standard microbiological techniques. The fungal isolates from the rhizosphere soil of cassava were identified as Debaryomyces nepalensis, Fusarium oxysporium, Microsporium persicolor, Aspergillus terreus, Aspergillus fumigatus Geotrichum candidum, Alternaria alternata and Penicillium polonicum. All the fungal isolates were screened positive for xylanase activity on mineral salt medium supplemented with araboxylan (1%) as sole carbon source. The process parameters were optimized using one factor at a time technique. The identities of the isolates authenticated as Debaryomyces nepalensis and Penicillium polonicum by molecular techniques were regarded as good xylanase producers and they were selected for optimization studies. The optimal incubation periods for maximal xylanase production by Penicillium polonicum and Debaryomyces nepalensis were 120 and 144 hours respectively. The two fungal isolates had the same optimal pH and temperature which were; 5.0 and 50oC. Maximum xylanase production was achieved from D. nepalensis with the use of xylan as sole source of carbon and peptone as sole source of nitrogen, while optimum xylanase production was achieved from P. polonicum with the use of xylan as sole source of carbon and sodium nitrate as sole source of nitrogen. The crude enzyme from the best xylanase-producing fungus was purified using ammonium sulphate precipitation, ion exchange and gel-filtration chromatography and the biochemical properties were thereafter characterized. The activity of purified enzyme from D. nepalensis was optimal at pH 8.0 and at temperature 50oC retaining more than 50% of its activity over a broad range of temperature and pH. The enzyme activity was mildly inhibited by Mg2+, Hg2+, Cu2+, Na+ and K3Fe and stimulated by Mn2+, Ca2+, Ni2+ and Co2+ metal ions. Also, chemical agent such as ethylenediamine tetra acetate (EDTA) and urea inhibited the enzyme activity. The molecular weight of the purified enzyme was estimated to be 47KDa. The Km and Vmax were found to be 0.180 mg/ml and 0.189 μmol/min/ml respectively. The studied xylanase from D. nepalensis had desirable biochemical properties such as a broad pH range, moderate thermostability and the ability to withstand enzyme inhibitors, which makes them a good choice for many industrial applications.