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BACKGROUND
In the fixed concentration of enzyme, a change in concentration of the substrate may either increase or decrease the reaction state. So, the basic objective of this test was to study the effect of changing amount of the substrate to the salivary amylase activity.
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REQUIREMENTS
Reagents: Starch solution
Stock enzyme solution
Iodine
Apparatus: Test tube rack
Colorimeter tubes
Four Erlenmeyer flasks
1 ml pipette
Calorimeter
Distilled water
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PROCEDURE
Arrange a test tube rack having total 4 rows with each row having 10 colorimeter tubes.1 In each row the first tubes should be labeled as 1:2, 1:4, 1:8 and 1:16 depending on the dilutions of the starch. Now take Erlenmeyer flask and label it depending on the respective dilutions and then to each flask add 50ml of distilled water. Now to each flask add 50 ml of starch solution. Mix it properly and from this transfer 50 ml of the mixed solution to the second flask and then again mix it properly. Repeat same with the third and fourth flask. Then from fourth flask discard 50 ml of the solution. Now tale 1 ml pipette and transfer 0.2 ml of the enzyme solution to all flask. Mix it properly and transfer 3 ml of the solution from each flask to colorimeter tube. To each tube add 3 drops of iodine and allow it to mix properly. Then place the tubes in the colorimeter for measuring absorbance at 540 nm. Calculate the transmittance percent. Do this for every flask with 2 minute gap.2
Transmittance percent=antilog (2- absorbance)
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CONCLUSION
With increase or decrease in substrate concentration the % transmittance and absorbance will increase or decrease.
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REFERENCES
- Briggs GE, Haldane JB. A Note on the Kinetics of Enzyme Action. Biochem J. 1925;19(2):338–339
- Quastel JH, Whetham MD. Dehydrogenations produced by resting Bacteria. I. Biochem J. 1925;19(3):520–531.
