Isolation of Thermotolerant Phytase Producing Fungi and Optimisation of Phytase Production by Aspergillus niger NRF9 in Solid State Fermentation Using Response Surface Methodology

다국어 초록 (Multilingual Abstract)

Phytase is an enzyme that hydrolyses phyticacid into myo-inositol and inorganic phosphate. Onehundred thirteen fungi were isolated from 58 soil samplescollected from different sites with high organic mattercontent. They were screened for phytate hydrolysing activityon phytase screening medium (PSM). Clear zones wereformed around the colonies of 72 isolates on Ca-containingPSM. Solid state fermentation was done for the selectedfungal isolates for production of the phytase enzyme.
Among the isolates, NRF9 showed maximum enzymeactivity (39.2 U/gds) upon quantitative screening after5 days of incubation at 30°C. The fungus grew vigorouslyand secreted enzyme at temperatures of 30, 37, and 45°C.
The maximum phytase production was obtained at 30°C;the next-best production was obtained at 37°C. The isolatewas identified and characterised as Aspergillus niger NRF9on the basis of 18S rDNA sequencing. Various processparameters were optimised using a one-variable-at-a-timeapproach’. The significant factors for phytase productionwere identified as incubation time, solid:liquid ratio andglucose concentration. These factors were further optimisedusing a Box-Behnken design. Maximum phytase production(112.1 U/gds) was observed at 5 days of incubation time,a 1:2 solid:liquid ratio and a 1% glucose concentration.
Validation of these optimised variables was done in shakeflasks. Optimisation resulted in a 2.9-fold increase in phytaseproduction compared to unoptimised cultivation conditions.

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