The effects of pH and temperature on proteins
Farshad Sedaghat
James Van Zoeren
Monta Vista High
School
November7, 1995
ABSTRACT
In this
experimet, our purpose was to determine how temperature and harsh chemical
treatments ( 1 M HCl ) affect the denaturing of the proteins in egg whites. Each
time, 1 ml. of egg white was taken and tested in different environments to see
when the proteins in the egg white would be denatured. In general, as the
temperature of the egg white was increased, the amount of HCl needed to denature
the proteins in the egg whites was decreased.
Introduction
What happens when you heat up an egg? It clumps
together and turns white. What happens when you mix egg with strong acids,
bases, or harsh chemicals? They curdle up and change shape and color. Gentle
heating or certain chemical treatments can loosen the linkages between proteins,
but many proteins will re-form their original structure once normal conditions
arc restored. Higher temperatures and stronger chemicals make the polypeptides
that form the protein permanently lose their shape and make them denatured. [1].
This is what happens when eggs arc heated up. Their proteins have permanently
lost their shape and have changed their structure and color.
In our
experiment, we wanted to find out at what temperatures and degree of chemical
treatment causes proteins to be denatured. We used chicken egg whites as the
protein and tested them under difererent temperatures and chemical treatments (
1 M HCl). We also used temperature and acid together to see how temperature
would affect the amount of acid needed to denature the egg white. We believe
that as the temperature is increased, Iess HCl needs to be added in order to
denature the egg white. This experimen could have health related purposes for
determining under what conditions proteins can he denatured and in which
environments they could survive safely.
Materials and
Method
Two chicken eggs were cracked and the egg whites were separated
from the egg yolks and put in a beaker. Separating the egg yolk from the egg
white was done very carefully to make sure that no egg yolk was present in our
egg whites.
First, we took 1 ml. of the egg white using the large
pipette and placed it in a test tube. This first test tube was tested using
temperature(°C) only. We took a large beaker, filled it half'way with tap water
and placed it on the hot plate which was turned to high heat or the #10 knob. (
For different hot plates, turn the heat up to as high as possible. )
When the temperature of the water reached to about 50 °C, the test tube
containing the egg white was placed in the water. We observed the egg white
until it turned Lo a cloudy white color. We knew that the proteins had denatured
therefore changing color and form. We recorded the temperature at which the egg
white denatured The pH of the egg white was taken and recorded.
Next, 1
ml, of the egg white was tested to see how much 1 M HCl was needed to denature
it at room temperature thing the 1 mL pipette, HCl was slowly added to the egg
white little by little. When the egg white clumped together and had a white
color, we knew the proteins had been denatured. The amount of HCl used to
denature the protein was recorded along with the pH. ( HCl was added very slowly
to the egg white and the test tube was constantly swirled to allow the HCl to
mix thoroughly with the egg whites. The measurement of HCl used was recorded as
precisely as possible.)
Next, temperature and HCl were both used to
determine at what temperature and how much acid was needed to denature the
proteins of the egg white. 1 ml, of egg white was placed in a test tube. The
test tube was placed on the hot plate, in water about 4() °C warm. The egg white
was allowed to rest in the water for about 30 seconds in order to get warm.
Then, HCI was slowly added to the egg white. As soon as the egg white started to
clump together, both the temperature of the egg white and the amount of HCl used
were recorded. The pH of the solution was also recorded. The same procedure was
repeated for temperatures around 50°C , 60°C , and 70 °C. The same steps were
done at 0°C except ice was used to cool the waler down. ( NOTE: The temperature
at which the denaturing, occurred had to be recorded right away because as soon
as the egg whites wore taken out of the water, they changed temperature very
rapidly).
Results
We lound that as the temperature of the egg
whites was increased, the amount of HCl needed to denature the proteins was
decreased. The graph shows the amount of HCl needed to denature the egg proteins
at a certain temperature. It is obvious that as the temperature is increased,
the amount of HCl needed is decreased.
Discussion
When we started this experiment we expected that as
the temperature of the egg white was increased. the amount old HCl needed to
denature the protein would be decreased. Our hypothesis was supported by our
data. B comparing, the temperature and amount of I M HCl added, we can see that
the amount of HCl needed to denature the protein is decreased as the temperature
is increased. Based on our results, we hypothesize that the graph of amount of
HCl vs. temperature is almost an inverse variation. As the temperature gets
larger. the amount of HCl gets smaller.
A protein's overall
three-dimensional shape is determined by the interactions between its amino
acids. There are four different structures (primary, secondary, tertiary, and
quaternary) which affect the shape of proteins. The shape of the proteins are
determined by the different kinds of hording between the amino acids (hydrogen
bonds, ionic bonds, disulfide bonds, and hydrophobic interactions). It was
determined that the egg, white alone (without any HCl) was denatured at 66.0 °C.
At high temperatures, the protein's atoms, and the associated water molceules,
have so much energy that their motions disrupts the hydrophobic, hydrogen, and
ionic bonds that give the protein its normal shape 121 It was also determined
that the egg white alone (at room temperature, 22.8°C was denatured when 0.92
ml. of HCl was added to it.
The pH of each denatured egg, white was
taken to see at what pH that certain temperature denatured the egg: proteins. In
the higher temperature test tube, the pH of the denatured egg whites was lower
which further proves that as temperature rises, less harsher chemical treatments
will denature the egg white. For example, the pH of the denatured egg white at
2.0°C was a very low 3, but the pH of the denatured egg white at 59.0 °C was 5
Our calculations and measurements were handled very accurately and
precisely to assure that no mistake would affect the data of our lab. However,
some factors might have resulted the outcome of our experiment and there are
also many opportunities for human errors. For example, it was difficult to keep
the temperature of the water at a constant. The temperature would either quickly
rise or decrease. Therefore, the temperatures were determined as carefully as
possible. Also, if we had diluted the 1 M of HCl to 0.1 M, our experiment would
have had more accurate results because 1 M HCl causes the egg white to denature
very quickly. No matter how careful we were, there was always the chance that we
added 0.1 or 0.3 ml. of HCl alter the egg whites had already been denatured.
If it was possible to experiment the denaturing of the egg white at
intervals of 5 °C( From 0 to 65 °C), the experiment would have produced a very
accurate range of the denaturing, of the proteins in eggs. However, keeping the
temperature al constant temperatures is very difficult to do with the materials
we have.
To improve upon our experiments, experiments can be done to
test the denaturing of eggs under different environments such as using NaCH or
other basic solutions. Tests can also be done to see if denatured proteins
harmed by acid could be renatured by titrating it with a base.
References
[1 and 2]Arms, Karen; Camp, Pamela S; Jenner Jannan
V; Zalisko, Edward J.: Biology, A Journey Into Life, Third Edition, pg 53-55.
Harcourt Brace College Publisher, New York, 1994
[3] Janet Van Zoeren, De
Anza College, personal communicaton