Application of Used Tea as Solid Matrix for Immobilization …
223
2.10 Optimal Temperature and pH for Enzyme Activity
The temperature activity profile of the immobilized enzyme was studied by incubating the immobilized enzyme in 100 mM Tris Hcl buffer (pH 8.0) for 30 min with
casein as substrate. The temperature range at temperatures (30, 35, 40, 45, 50, 55
and 60 °C), and enzyme activity was measured at each temperature.
Alternatively, the pH activity profile was studied incubating the enzyme substrate
solution at 37 °C and at different pH from 7.5–10. Then enzyme activity was
determined for various pH values.
2.11 Thermal Stability and pH Stability
Thermal stability of the free and immobilized enzyme was determined by 2 h of
incubation enzyme over the temperature range (30, 35, 40, 45, 50, 55, and 60) °C at
(pH 8.0). Residual activity was measured under standard conditions and expressed
as percentage of the relative protease activity.
The pH activity profile for partially purified free and immobilized enzyme was
determined at 37 °C for 2 h by incubating the enzyme with substrate at various pH
ranges from 3 to 8. The pH of the reaction mixture was varied using different buffers
(pH 7.5–9 and 10) and enzyme activity was calculated.
2.12 Storage Stability and Reusability
The immobilized alkaline protease was stored at Tris Hcl buffer (100 mM, pH 8.0)
at 4 °C for 30 days. The activity was measured every 6 days.
The immobilized alkaline protease was repeatedly used for hydrolysis in each
experiment. After each runs the immobilized enzyme was washed with distilled
water followed by Tris Hcl buffer and stored in the same buffer.
3 Results and Discussion
3.1 Effect of Amount of Solid Matrix on Immobilization
of Alkaline Protease
The amount of solid matrix for immobilization was optimized on the basis of immobilization yield and efficiency. Out of different amount of solid carrier used for
immobilization of protease, maximum enzyme yield (34%) and efficiency (45%)
was observed with 300 mg of matrix (Fig. 1). On increasing the amount of solid
223
2.10 Optimal Temperature and pH for Enzyme Activity
The temperature activity profile of the immobilized enzyme was studied by incubating the immobilized enzyme in 100 mM Tris Hcl buffer (pH 8.0) for 30 min with
casein as substrate. The temperature range at temperatures (30, 35, 40, 45, 50, 55
and 60 °C), and enzyme activity was measured at each temperature.
Alternatively, the pH activity profile was studied incubating the enzyme substrate
solution at 37 °C and at different pH from 7.5–10. Then enzyme activity was
determined for various pH values.
2.11 Thermal Stability and pH Stability
Thermal stability of the free and immobilized enzyme was determined by 2 h of
incubation enzyme over the temperature range (30, 35, 40, 45, 50, 55, and 60) °C at
(pH 8.0). Residual activity was measured under standard conditions and expressed
as percentage of the relative protease activity.
The pH activity profile for partially purified free and immobilized enzyme was
determined at 37 °C for 2 h by incubating the enzyme with substrate at various pH
ranges from 3 to 8. The pH of the reaction mixture was varied using different buffers
(pH 7.5–9 and 10) and enzyme activity was calculated.
2.12 Storage Stability and Reusability
The immobilized alkaline protease was stored at Tris Hcl buffer (100 mM, pH 8.0)
at 4 °C for 30 days. The activity was measured every 6 days.
The immobilized alkaline protease was repeatedly used for hydrolysis in each
experiment. After each runs the immobilized enzyme was washed with distilled
water followed by Tris Hcl buffer and stored in the same buffer.
3 Results and Discussion
3.1 Effect of Amount of Solid Matrix on Immobilization
of Alkaline Protease
The amount of solid matrix for immobilization was optimized on the basis of immobilization yield and efficiency. Out of different amount of solid carrier used for
immobilization of protease, maximum enzyme yield (34%) and efficiency (45%)
was observed with 300 mg of matrix (Fig. 1). On increasing the amount of solid