Enzymes

A catalyst is a substance that increases the rate of chemical reaction and is not
changed by the reaction itself.
An enzyme is a biological catalyst that catalyzes many important reactions
inside an organism (such as respiration) and therefore necessary to sustain life.
How enzymes work?
A substrate is a substance that an enzyme acts on. It is important to understand
that enzymes are very specific, and the reason for their specificity lies in their
active sites – a region of an enzyme that binds to a particular substrate. The
shape of the active site of an enzyme is complementary to only one specific
substrate. As the enzyme binds with the substrate, an enzyme-substrate complex
is formed. The reaction then occurs on the enzyme and the enzyme-product
complex is formed. The products eventually leave the enzyme.
Temperature and enzyme activity
Low temperatures reduce the rate of chemical reactions in general. This is
because molecules need to collide with one another and have enough energy for
a reaction to occur. In low temperatures, molecules are traveling at lower speeds
(less energy) and therefore the rate of successful collisions are lower. Moreover,
even when collisions do occur, the molecules may have insufficient kinetic
energies to begin with, and therefore the reaction may not occur. Enzyme
activity is therefore low in low temperatures. It is important to note however,
that low temperatures do not denature enzymes.
Higher temperatures generally increase the rate of chemical reactions.
Molecules are faster and have more kinetic energy. This means that rate of
successful molecular collisions are higher,and most molecules will have
sufficient energy required for the reaction.
However, temperatures that are far beyond the optimum temperature of the
enzymes can start to denature it, and reduce enzyme activity as a result. Most
enzymes have an optimum temperature of approximately 37 degrees in the
human body, and start getting denatured at above 50 degrees.
pH and enzyme activity
The optimum pH of an enzyme can vary. Pepsin is an enzyme found in the
stomach’s acidic conditions and therefore made to work best in a pH of
approximately 2. Amylase on the other hand, is found in saliva (more neutral
conditions) and therefore has an optimum pH of 7. Very high or very low pH’s
can denature these enzymes if it deviates too much from their optimum.
Practical: Enzymes & Temperature
 Amylase is an enzyme that digests starch (a polysaccharide of
glucose) into maltose (a disaccharide of glucose)
 The effect of temperature on the activity of amylase can be
investigated
Apparatus
 Spotting tile
 Measuring cylinder
 Test tube
 Syringe
 Pipette
 Stopwatch
 Water
 Thermometer
 Water bath
 Iodine
 Starch solution
 Amylase solution
Method
 Add 5cm3 starch solution to a test tube and heat to a set temperature
using beaker of water with a Bunsen burner
 Add a drop of Iodine to each of the wells of a spotting tile
 Use a syringe to add 2cm3 amylase to the starch solution and mix well
 Every minute, transfer a droplet of solution to a new well of iodine
solution (which should turn blue-black)
 Repeat this transfer process until the iodine solution stops turning
blue-black (this means the amylase has broken down all the starch)
 Record the time taken for the reaction to be completed
 Repeat the investigation for a range of temperatures (from 20°C to
60°C)
Results and Analysis
 Amylase is an enzyme which breaks down starch
 The quicker the reaction is completed, the faster the enzyme is working
 This investigation shows:
o At the optimum temperature, the iodine stopped turning blue-
black the fastest
 This is because the enzyme is working at its fastest rate and
has digested all the starch
o At colder temperatures (below optimum), the iodine took a longer
time to stop turning blue-black
 This is because the amylase enzyme is working slowly due
to low kinetic energy and few collisions between the
amylase and the starch
o At hotter temperatures (above optimum) the iodine turned blue-
black throughout the whole investigation
 This is because the amylase enzyme has become denatured
and so can no longer bind with the starch or break it down
Limitations
 Note that there are several different ways in which the temperature could
be controlled. The method described above is not very precise, an
improvement would be to use water baths kept at each temperature
 The starch and amylase solutions that need to be used should be placed in
a water bath and allowed to reach the temperature (using a thermometer
to check) before being used
 A colorimeter can be used to measure the progress of the reaction more
accurately
o A solution containing starch will be darker than a solution
containing glucose (as a result of the colour change of iodine)
o The absorbance or transmission of light through the coloured
solution can be measured using a colorimeter
Applying CORMS to practical work
 When working with practical investigations, remember to consider your
CORMS evaluation

CORMS evaluation
 In this investigation, your evaluation should look something like this:
o C - We are changing the temperature in each repeat
o O - This is not relevant to this investigation as we aren't using an
organism
o R - We will repeat the investigation several times to make sure our
results are reliable
o M1 - We will measure the time taken
o M2 - for the iodine to stop turning black
o S - We will control the concentration and volume of starch
solution, iodine and amylase used in the investigation