Phytase enzyme now becomes more important commercially. Presence of phytate in food
and feed make them less nutritive due to phytate complexes mainly with mineral ions and
proteins. Phytase in monogastric animals and human stomach either produced in small amount or
not. This leads to phosphorous Pi deficiency. Supplementation of food and feed with phytase
enzyme full fill this deficiency through degradation of phytate complexes and release of Pi.
Degradation of phytate complexes makes phosphorous other mineral ions and amino acids
available for growth and development. It was proved that feed conversion rate in poultry
increased due to supplementation of phytase in poultry feed. Feed of monogastric animals mostly
at industrial level pelleted to give it a shape or to kill microorganisms (sterility). At industrial
level enzyme production and processing cost about 2 billion. So this demands a thermostable
phytase to use at industrial level or its cost effective production.
Aspergillus niger have been used industrially for production of beneficial enzymes. A.
niger isolates procured from department of microbiology were confirmed through macro and
microscopic characteristics as A. niger. These isolate were screened for phytase production on
phytase screening medium PSM agar. Positive isolates identified through noval staining using
2% cobalt chloride, 6.25% ammonium molybdate and 0.42% ammonium vanadate for contrast.
Positive isolates next proceeded for phytase enzyme production in broth media (pH 5.6) using
0.5% sodium phytate as substrate. Incubation was done at 30oC for 5-7 days in shaking incubator
150rpm. After production quantification of enzyme was carried out through enzyme activity
assay. There maximum (274.99±10.14 FTU/ml) and minimum (68.88±2.55 FTU/mL) activity of
phytases from isolate PASN01 and PASN08 was observed. Phytases characterized through sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS PAGE) to know protein
molecular weights. Highest molecular weight 107.82kDa was PASN06 and lowest was 35.21kDa
of PASN 01.
Aspergillus niger spores subjected to steam heat treatment at 30oC, 45oC, 60oC, 75oC and
90oC for 15, 30, 45, 60minutes to identify thermostability. At 30oC and 45oC temperature, spores
of A. niger isolates found to be thermostable. But at 60oC, 75oC, or 90oC treatment spores
become inactivated or there 6.0 logarithmic reduction in spore count was observed.
Thermostability of phytases was found at 60oC, 75oC, 90oC for 15, 30, 45, and 60
minutes treatments. Enzyme from A. niger PASN01 and PASN08 observed as thermostable at
60, 75 and 90oC. Phytases from PASN01 and PASN08 showed 160.55±42.96 and 00±.00
FTU/mL decreased in activity after 45 minutes of treatment at 60oC temperature, respectively.
PASN01 phytase displayed 163.88±23.35, 172.77±7.52 and 171.66±7.26 FTU/mL decreased in
activity after 60minutes treatment at 60, 75 and 90oC. In case of PASN08 phytase at 60, 75 and
90oC temperature after 60minutes treatment, 13.33±10.41, 16.66±6.00 and 23.88±41.37 FTU/mL
decreased in activities were observed, respectively. PASN08 phytase observed more
thermostable than other phytases of A. niger isolates. Enzyme can bear pelleting and pre
pelleting temperatures. Enzyme from PASN08 also observed stable during storage at room
temperature.
Conclusion:
A. niger PASN08 spores inactivated or killed and phytase observed stable at 60oC
temperature, after 60mins treatment. Temperature 60oC may be used industrially for cost
effective thermostable phytase production from indigenous A. niger isolate PASN08.