Question

1. design a follow-up experiment to investigate the enzyme peroxidase from plants.

2. investigate the effect of pH on enzyme activity.

Answer 1

1. Design a follow-up experiment to investigate the enzyme peroxide from plants.

Peroxidase enzyme is present in organisms such as plants, bacterias and humans. It cleaves Hydrogen peroxide by use of oxygen for respiration, and form a toxin by product.

Reaction:

2H₂O₂ + Peroxidase -----> 2H₂O + O₂ + peroxidase

Byproducts are water and oxygen, as they are not as much toxic as H₂O₂, there is no deterioration in the inner region of the cell.

Experimental design

The rich source of catalase(peroxidase enzyme) is in turnip, and it is abstracted by considering the factors such as Substrate concentration, enzyme concentration, pH and temperature which affects the speed of the enzymatic reaction.

Spectrometers are used for monitoring the reaction rate, by using a reagant, guiacol. Oxidation of guiacol takes place in the presence of oxygen, and the color changes from clear to brown. The color of guiacol becomes intense, as the amount of oxygen product increases. Reaction is proceeded by taking 10ml reaction mixture which consists of guiacol, turnip extract, pH7 buffer and hydrogen peroxide. Every second of a minute absorbance measurement is observed. The graph is against time(x) and graphed absorbance(y). The rate of the reaction is obtained from the slope.

Maintaining standard reaction as a reference, variable of interest is measured individually, by running three set of reactions and compared with the standard. Effect of substrate concentration is intented by taking three sets of tubes, which differ with each other and with the substrate: amount of hydrogen peroxide is used. Slopes of the resulting lines are recorded and the average reaction rates(dependent variable, y) against the test factor(independent variable, x) is graphed. The curves from the graphs gives information about the status of the reaction rate, when the conditions gets deviate from the optimum and the enzyme's optimum value for each factor.

O.300 O.250 O.200 O.IS0 O100. 0.000 25 37 loo TEMPERATURE Reachon

2. Effect of pH on enzyme activity

Alteration in the pH will disturb the activity of the enzyme. Here the Optimum pH is referred as the point at which the enzyme is extremely active and the pH is also a positive value.

OPTIMUM pH pH REACTION VELOCITY

Enzyme Activity is extremely low, when the pH value is higher or lower to the extent. The stability of the enzyme is based on the pH value. For an enzyme along with activity, a region of pH optimal stability is possible. Optimum pH value will differ for each and every enzyme.

Ex. Pepsin has 1.5 - 1.6 Optimum pH, and Trypsin has 7. - 8.7, Urease has 7, Maltase has 6.1 - 6.8, Catalase has 7.0 Optimum pH

Apart from pH and temperature, ionic strength also play a role in fixing the reaction of the enzyme. For exact and consistent enzyme, these physical and chemical parameters should be examined and optimised.