2. Properties of enzymes Experiment



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2. Properties of enzymes
Experiment 2.1. The effect of temperature on enzyme activity
Enzymes, like all proteins, are susceptible to denaturation that may be induced by chemical or physical factors. Each enzyme has a temperature optimum – the value of temperature at which this enzyme has maximum of catalytic activity, i.e. provides proceeding of catalyzed chemical reaction with maximum velocity. The maximum rate of major enzymatic reactions in human organism is at temperatures in the range of 38C−40C. Enzymes will be denatured and lose their biological properties if the temperature is higher. It should be noted that some microorganisms have enzymes with high thermostability. When the temperature decreases to a level below the optimum the activity of enzymes also decreases. Dependence of enzyme activity on temperature can be demonstrated on human salivary amylase.

Salivary amylase hydrolyses starch commonly to dextrins. During the reaction of iodine with intact starch a blue coloration is developed. Products of starch hydrolysis give different colours with iodine in dependence on their molecular weight. Amylodextrins give violet coloration with iodine, erythrodextrins – reddish brown, maltose and achrodextrins do not change the yellow colour of iodine.


Procedure
Place 3 ml of undiluted saliva into a test tube and boil it for 5 minutes, cool it by tap water. Then take 3 test tubes and, according to the table, pour 10 drops of 1% starch solution into each tube. Next, add 10 drops of non-boiled saliva into the first test tube, 10 drops of boiled saliva into the second test tube and 10 drops of distilled water into the third test tube as a reference. All of them are to be placed in a thermostat at 38C. In 10 minutes, perform the qualitative test on starch (iodine reaction). Add 1–2 drops of 0.1% iodine solution into each test tube and compare the colorations that will develop in these test tubes. Write down your observations in the table below, explain the results and make a conclusion:


Reagents

The first test tube

The second test tube

The third test tube

1% starch solution

10 drops

10 drops

10 drops

Non-boiled saliva

10 drops





Boiled saliva



10 drops



Distilled water





10 drops




Incubation 10 min, 38C

0.1% iodine solution

1–2 drops

1–2 drops

1–2 drops

Coloration











Explanation of the results, conclusion: ____________________________________
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