Studies on the production of Glucose oxidase by Aspergillus terreus UniMAP AA-1
Glucose oxidase (GOx) has found a wide range of applications in chemical, food, beverage, biotechnology and other industries. It is commonly extracted from Aspergillus niger and selected strains of Penicllium sp. Currently there is a growing need to find alternative sources of this enzyme due to...
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| Format: | Thesis |
| Language: | English |
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| Online Access: | http://dspace.unimap.edu.my:80/xmlui/bitstream/123456789/21605/1/Full%20text.pdf http://dspace.unimap.edu.my:80/xmlui/bitstream/123456789/21605/2/p.%201-24.pdf |
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| Summary: | Glucose oxidase (GOx) has found a wide range of applications in chemical, food,
beverage, biotechnology and other industries. It is commonly extracted from
Aspergillus niger and selected strains of Penicllium sp. Currently there is a growing
need to find alternative sources of this enzyme due to some drawbacks associated with
A.niger and Penicllium sp. In this work, a novel GOx-producing strain, Aspergillus
terreus UniMAP AA-1, was isolated from soil of Agrotech Research Centre, Sg
Chucuh, Perlis.The screening tests for the GOx-producing strain were carried out on the
basis of color development test by using agar plate containing o-anisidine and
horseradish peroxidase. The screened strain was identified morphologically using light
microscope and Scanning Electron Microscope (SEM) and further verified by molecular
level identification. The strain was identified as a predominant extracellular GOx
producer and exhibits a pelleted morphology in fermentation culture. These findings
offer a new alternative to the existing GOx-producing strains which are known to be
associated with few drawbacks. Subsequently, a sequential optimization based on
statistical design and one-factor-at-a-time (OFAT) method was employed to optimize
the production of extracellular GOx from the potential strain. Plackett-Burman design
indicated glucose as the most influential variable followed by NaNO3, CaCO3, and
peptone on the GOx activity; while KH2PO4, MgSO4.7H2O and FeSO4.7H2O showed
negative main effect on the enzyme activity. Based on the result, glucose, NaNO3 and
CaCO3 were selected for further optimization studies. The influences of the three
medium components were investigated with one-factor-at-a-time (OFAT) and these
variables were subsequently optimized using a face centered central composite design
(FCCCD). The optimum conditions were found to be 10.64% (w/v), 1.21% (w/v) and
5.22% (w/v) for glucose, NaNO3 and CaCO3 respectively and the enzyme activity was
found to be 6.72 U/ml, which was about seven fold higher than that obtained in media
before optimization. The oxygen and glucose consumption as well as hydrogen
peroxide and gluconic acid production profiles of the crude enzyme are all in-line with
typical GOx properties. The kinetic constant, Km of the crude enzyme for its substrate,
determined by direct fits of Michaelis–Menten equation through nonlinear regression
(with correlation value or R2 =0.98) using solver function in Microsoft Excel software,
gave the value of within the range of 7.5-15 mM. The result indicates substrate
specificity of the crude enzyme towards β-D glucose (substrate) and demonstrated the
tight binding of the crude enzyme with its substrate. |
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