Title: Effect of Temperature on the Rate of Coagulation of Rennin

Aim: To determine the effect of temperature on the rate of coagulation of rennin.

Literature Review
Cells coordinate their activities through intracellular signalling pathways and are able to
communicate with neighbouring cells. (Alberts et al, 2014), which includes responding to
changes in their inside environment, and external environment. A critical role is played by
enzymes in the processes that catalyse biochemical reactions and involved in signalling
pathways, which allow cells to carry out metabolic processes quickly and efficiently. ( Nelson
and Cox, 2017).Enzymes are proteins that are biological catalysts , which help fasten the
process of biochemical reactions in all living organisms. Enzymes are built up of amino acids
mostly. Enzymes are known to be similar or same to catalysts. A catalyst can be used
numerous times in any chemical fraction and does not reduce. Enzymes, on the other hand, can
easily lose their forms and might not function properly (Richard Willstätter, 1920). Enzymes are
proteins and are reusable. Enzyme activity means the catalytic properties of an enzyme. To test
how fast an enzymatic reaction happens, is the measure of the enzyme activity. Enzymes have
two significant core properties, they speed up the rate of chemical reactions, but in the process
they do not permanently change the reaction, or have themselves consumed. Secondly
enzymes can increase the reaction rate without changing the chemical equilibrium between
products and reactants (JBS Haldane, 1930). Another property of enzymes is specificity.
Enzymes tend to be very specific in their action. Enzymes have various specificities like: bond,
group, substrate, optical, cofactor and geometric. Furthermore, the third property is reversibility.
Enzymes' catalysed reversibility reaction depends upon the requirements of the cell. Lastly,
enzymes have a property known as sensitivity. Enzymes are very sensitive to changes in the
environments, and variations in factors like temperature, pH and substrate concentration.
These factors can have significant effects on an enzyme’s activity due to their impact on
enzyme structure and function..Higher temperatures cause more collisions among the
molecules and changes in pH will lead to the loss of activity in most enzymes. Temperatures
and pH in the environments, and the subtract concentration can all affect the rate in which an
enzyme catalyses a reaction. This experiment displays the effect of temperature on an enzyme
activity. It is known that as temperatures increase, the rate of enzyme activity also increases,
because when the temperature rises so does the kinetic energy of the molecules of the
substrate and enzymes.

This experiment investigated Rennin as its enzyme. Rennin is also known as chymosin, which is
an enzyme produced in the stomach of young mammals. It is a protein-digesting enzyme that
coagulates milk by transforming caseinogen into insoluble casein. The effect of temperature on
the rate of coagulation rennin has been investigated in this experiment. Coagulation in milk is
when the casein micelles destabilise and flocculate and cluster to create a form of a gel
enclosing the soluble milk components.(Gregory 2006).The rate of coagulation of rennin shows
the speed in which rennin makes milk proteins to coagulate. The rate of coagulation can be
affected by numerous factors such as temperature and different concentrations of rennin. Past
studies have highlighted that higher temperatures increase enzymatic activity and reduce
coagulation time, but if temperatures are above 36 degree celsius the coagulation time will
increase because of the heat-induced inactivation of rennin.( Dalgleish 1983). Various studies
have investigated the effect of temperature rate of coagulation of rennin. Studies have shown
that the optimal temperature above was around 40 degree celsius and showed a sharp decline
in activity around 45 celsius 9Kilara et al.1982). More studies show important focus on the
thermal stability of rennin at different temperatures, which have results showing that the enzyme
had been semi-stable at temperatures below 40 degree celsius, and had minimal activity loss.
Contrastingly, the enzyme quickly lost its activity at high temperatures, with no activity present
above 55 celsius. Sugimoto et al.( 2004). A study investigated the effect of temperature on
rennet coagulation activity on various milk types. Furthermore, the study discovered that the
optimal temperature for rennin activity varied based on the type of milk, thus higher
temperatures are required for coagulation of goat milk, but not for cow milk. ( Pintadol et al
2019).

This investigation of how temperature has affected the rate of coagulation of rennin has been
placed into action through an experiment.This is done by setting an experiment, to prepare
rennin solution, crush a junket tablet and add 10 ml of water. Prepare 6 water baths at
temperatures ranging from 0 to 100 degrees Celsius. Label 6 test tubes A and B, add 3 ml of
milk to each, and wait 10 minutes for the milk to reach the temperature of the water bath. Add
10 drops of rennin solution to the A tubes, shake, and record clotting time every 15 minutes.
Repeat 2 more times. Limitations in this experiment was, limited scope. The experiment will not
be able to cover all possible temperatures that could affect the rate of coagulation of rennin.
Thus, the results wouldn’t have been entirely representative of all temperature ranges.
Furthermore another limitation would be the small sample size. The experiment has used a
small sample size which could affect the statistical power of the study and lessen the
generalizability of the results. To sum up, the experiment can provide some insights into the
effect of temperature on the rate of coagulation of rennin. It is important to have these limitations
in mind so the finding can be interpreted accurately.

Hypothesis: Higher temperatures will cause the rate of coagulation of rennin to

increase.
Risk Assessment:
Risk Cause Identify

Risk 1 Heat Hot water Will cause burns on

skin.

Risk 2 Breakage of Glassware used in Will cause physical

equipment experiments. wounds.

Risk 3 Slipping, tripping, Liquid spilling on the Will cause injuries.

 falling. ground.

Methodology:
Firstly, one junket tablet was crushed using a mortar and a pestle. Once the rennin solution was
prepared, then ten ml of water was added and it was stirred. Afterwards, the mixture was
poured into a small beaker. Then six water baths were prepared at the following temperatures:
0, 20, 40, 60a and 80 degrees celsius.12 test tubes were labelled A and B half were A and half
were B and three ml of milk was added to each test tube using a pipette.Then, one test tube
labelled A and one test tube labelled B was placed in the water baths and were left for 10
minute. This was because it allowed the milk to rise or fall to the temperatures of the water bath.

To continue, the temperature of each of the water baths. Afterwards, ten drops of rennin
solution were added to the test tubes labelled A then, the timer was started. The test tubes were
gently tilted every 15 minutes to examine. The time was a record of the speed for the milk to
clot, if the milk didn’t clot after 15 minutes it was recorded as a desh. Lastly, these steps were
repeated two more times.

Results

Time (seconds) Time (seconds) Time (seconds)

Temperatures (°C) Trail 1 Trail 2 Trail 3

80 213 190 170

60 230 302 181

40 253 400 207

20 360 515 530

0 - - -

Figure 1: the time at which the milk clotted for each temperature.
Legend: X-axis: Represents Temperature
Y- axis: Represents the Time in seconds
Line: Represents the average rate of reaction
Figure 2: A graph showing the average rate of reactions of the experiment.