Investigation of the effect of increasing amount of lead nitrate on hydrolysis of starch was done using the enzyme amylase

Methods:

1. Thermostatically controlled water bath was set up at 400c.
2. Six boiling tube were labelled  A-F  and place  in a test tube rack.
3. 10cm3 starch solution was added in a test tube.
4. Following quantities  of lead nitrate were added  to 5cm3  distilled water  in six test tubes labelled  1-6,

• 0gram
• 0.1 gram
• 0.2 gram
• 0.3 gram
• 0.4 gram
• 0.5 gram

and was shaken  it to ensure  it dissolves.

5. Contents of tube 1 was poured  into boiling tube A , tube 2 into  boiling tube B  etc.
6. 5cm3  amylase solution was added  to each of the boiling tubes , agitated  well and   the timer was started  after placing the tubes into the water bath.
7. The reaction was allowed to proceed for 20 minutes.
8. During this time the colorimeter  was adjusted  using a solution of 1cm3 iodine solution in a boiling tube  containing 10cm3 starch solution and 5cm3 distilled water. The colorimeter was set to 0% transmission with this solution.
9. After 20 minutes  1cm3 iodine solution was added to each tube.
10. Test each of this tubes  A-F  in the colorimeter noting down the absorbance  for each test tube and the data were recorded in the  table.

Results

Raw data

Table 2

Results for control set up:

 Table 3

Processed data

Table 4

 

Discussion

There   is clear dose dependent  inhibition of amylase activity and the concentration of  lead nitrate. As the concentration of lead nitrate increases there are decreasing amylase activity. Amylase activity in this case has been calculated  by the  conversion of  starch iodide complex measured by colorimeter.

Evaluating the experiment   it can be said that  there are  improvements that can be made to increase the accuracy of the results. Some of them are as follows:

 

1. Repetition of   this experiment for more times for accuracy of the results.

2. Wider range of concentrations of the lead nitrate inhibitor for prediction of  the reaction rates.

3. Larger volume of substrate to be used  so that it does not limit the  results. This would enable to tell if the lead nitrate sulphate was a  reversible or non-reversible inhibitor. If the inhibitor was  reversible then the reaction rate would be slow, but not stop, whereas

if it was non-reversible the reaction rate would slow very quickly and  stop altogether.

With the apparatus there was some degree of inaccuracy:

1) There were limitations with the accuracy of the

measurements that can be taken.  The results were rounded off to the nearest whole numbers.  When transferring these results to a graph, the

inaccuracy of the measurement could show either a higher or lower rate

of reaction depending on whether the measurement was rounded up or

down and would show an untrue picture of what was happening.

2. If the cuvettes are not properly washed there will little change in the results depend on the impurities of the cuvettes.

Conclusion

The experiment produced results shows the general trend as the known facts about the effects of inhibitors on enzyme activity.

These results support known facts and as such can be used to

prove that inhibitors such as lead  do have an effect the

reaction rates of enzyme activity.

Bibliography

• Wynn, C. H. (1973) The Structure and Function of Enzymes (Studies in
  Biology No. 42), Edward Arnold, p43
• P. Keusch (2000) Kinetics: Enzymatic Decomposition of Hydrogen  Peroxide, University of Regensburg Visited March 2003.
• Addison Ault (2002) Understanding Enzyme Inhibition J. Chem. Educ., 2002, 79 (3), p 311