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Thermal Characterization of Purified Glucose Oxidase from A Newly Isolated Aspergillus Niger UAF-1.

Anjum Zia M - J Clin Biochem Nutr (2007)

Bottom Line: Temperature optimum for glucose oxidase, catalyzed D-glucose oxidation was 40 degrees C.The enzyme showed a high thermostability having a half-life 30 min, enthalpy of denaturation 99.66 kJ mol(-1) and free energy of denaturation 103.63 kJ mol(-1).These characteristics suggest the use of glucose oxidase from Aspergillus niger UAF-1 as an analytical reagent and in the design of biosensors for clinical, biochemical and diagnostic assays.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry (Biochemistry), University of Agriculture, Faisalabad, Pakistan.

ABSTRACT
An intracellular glucose oxidase was isolated from the mycelium extract of a locally isolated strain of Aspergillus niger UAF-1. The enzyme was purified to a yield of 28.43% and specific activity of 135 U mg(-1) through ammonium sulfate precipitation, anion exchange and gel filtration chromatography. The enzyme showed high affinity for D-glucose with a Km value of 2.56 mM. The enzyme exhibited optimum catalytic activity at pH 5.5. Temperature optimum for glucose oxidase, catalyzed D-glucose oxidation was 40 degrees C. The enzyme showed a high thermostability having a half-life 30 min, enthalpy of denaturation 99.66 kJ mol(-1) and free energy of denaturation 103.63 kJ mol(-1). These characteristics suggest the use of glucose oxidase from Aspergillus niger UAF-1 as an analytical reagent and in the design of biosensors for clinical, biochemical and diagnostic assays.

No MeSH data available.


Related in: MedlinePlus

Optimized conditions for the production of glucose oxidase from Aspergillus niger UAF-1.
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Figure 1: Optimized conditions for the production of glucose oxidase from Aspergillus niger UAF-1.

Mentions: In the present studies, corn steep liquor (CSL) was used for growth and glucose oxidase production by the Aspergillus niger UAF-1. Culture media containing 2% substrate was subjected to fermentation for 36 hours at pH 5.5, 30°C temperature and 5% inoculum size. The results indicated enzyme activity (6.06 U mL−1) in a medium containing 2% CSL. The findings of present study are in accordance with most of the research workers [21, 22]. For the optimization of substrate, growth media containing 2% corn steep liquor was found to be the best by Zareen [23]. Furthermore, no studies have been found about the utilization of corn steep liquor as a substrate/inducer for glucose oxidase production. So, we utilized CSL as a substrate for the said enzyme production to optimize the conditions of better production and to explore the application of CSL as substrate for glucose oxidase. Submerged fermentation was carried out for 24, 36, 48, 60 and 72 hours with 2% CSL. Enzyme obtained high activity i.e. 7.88 U mL−1, after 36 hours incubation, thereafter biosynthesis of enzyme decreased. Willis [24] optimized the fermentation media for the production of glucose oxidase by Aspergillus niger and obtained highest glucose oxidase yield after 48 h. While our results indicate the maximum production of the enzyme after 36 h. Media was supplemented with glucose as carbon additive and fermentation carried out under optimum conditions. Addition of 4% glucose to fermentation flasks (in triplicate) resulted in better glucose oxidase production with 23.74 U mL−1 activity. Kona et al. [25] showed that 6% sucrose resulted in the highest enzyme activity. It was observed after optimization of media that 2% substrate, 36 h of fermentation, pH 5.5, temperature 30°C, urea 0.3%, KH2PO4 0.6%, CaCO3 0.04% and glucose 4% proved to be best, while with addition of MgSO4.7H2O production of enzyme decreased [26, 27] (Fig. 1).


Thermal Characterization of Purified Glucose Oxidase from A Newly Isolated Aspergillus Niger UAF-1.

Anjum Zia M - J Clin Biochem Nutr (2007)

Optimized conditions for the production of glucose oxidase from Aspergillus niger UAF-1.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2170954&req=5

Figure 1: Optimized conditions for the production of glucose oxidase from Aspergillus niger UAF-1.
Mentions: In the present studies, corn steep liquor (CSL) was used for growth and glucose oxidase production by the Aspergillus niger UAF-1. Culture media containing 2% substrate was subjected to fermentation for 36 hours at pH 5.5, 30°C temperature and 5% inoculum size. The results indicated enzyme activity (6.06 U mL−1) in a medium containing 2% CSL. The findings of present study are in accordance with most of the research workers [21, 22]. For the optimization of substrate, growth media containing 2% corn steep liquor was found to be the best by Zareen [23]. Furthermore, no studies have been found about the utilization of corn steep liquor as a substrate/inducer for glucose oxidase production. So, we utilized CSL as a substrate for the said enzyme production to optimize the conditions of better production and to explore the application of CSL as substrate for glucose oxidase. Submerged fermentation was carried out for 24, 36, 48, 60 and 72 hours with 2% CSL. Enzyme obtained high activity i.e. 7.88 U mL−1, after 36 hours incubation, thereafter biosynthesis of enzyme decreased. Willis [24] optimized the fermentation media for the production of glucose oxidase by Aspergillus niger and obtained highest glucose oxidase yield after 48 h. While our results indicate the maximum production of the enzyme after 36 h. Media was supplemented with glucose as carbon additive and fermentation carried out under optimum conditions. Addition of 4% glucose to fermentation flasks (in triplicate) resulted in better glucose oxidase production with 23.74 U mL−1 activity. Kona et al. [25] showed that 6% sucrose resulted in the highest enzyme activity. It was observed after optimization of media that 2% substrate, 36 h of fermentation, pH 5.5, temperature 30°C, urea 0.3%, KH2PO4 0.6%, CaCO3 0.04% and glucose 4% proved to be best, while with addition of MgSO4.7H2O production of enzyme decreased [26, 27] (Fig. 1).

Bottom Line: Temperature optimum for glucose oxidase, catalyzed D-glucose oxidation was 40 degrees C.The enzyme showed a high thermostability having a half-life 30 min, enthalpy of denaturation 99.66 kJ mol(-1) and free energy of denaturation 103.63 kJ mol(-1).These characteristics suggest the use of glucose oxidase from Aspergillus niger UAF-1 as an analytical reagent and in the design of biosensors for clinical, biochemical and diagnostic assays.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry (Biochemistry), University of Agriculture, Faisalabad, Pakistan.

ABSTRACT
An intracellular glucose oxidase was isolated from the mycelium extract of a locally isolated strain of Aspergillus niger UAF-1. The enzyme was purified to a yield of 28.43% and specific activity of 135 U mg(-1) through ammonium sulfate precipitation, anion exchange and gel filtration chromatography. The enzyme showed high affinity for D-glucose with a Km value of 2.56 mM. The enzyme exhibited optimum catalytic activity at pH 5.5. Temperature optimum for glucose oxidase, catalyzed D-glucose oxidation was 40 degrees C. The enzyme showed a high thermostability having a half-life 30 min, enthalpy of denaturation 99.66 kJ mol(-1) and free energy of denaturation 103.63 kJ mol(-1). These characteristics suggest the use of glucose oxidase from Aspergillus niger UAF-1 as an analytical reagent and in the design of biosensors for clinical, biochemical and diagnostic assays.

No MeSH data available.


Related in: MedlinePlus