Albumin How do different methods of denaturation, including temperature changes of 30ºC and 60ºC, as well as 5g of Sodium Chloride

(NaCl), 5g of Sodium Hydrogen Carbonate (NaHCO3), 5 ml of Citric Acid (C₆H₈O) in Lemon juice, and 5 ml of Silver Nitrate (AgNO3)
Protein affect albumin in 40 ml of Gallus gallus domesticus (chicken) egg white after a xed duration of 5 minutes, and hence which
Lab method is the most effective at denaturing albumin?

Scienti c Background Research: Environmental/Ethical Considerations and Safety Issues:

Proteins are large molecules made up of amino acids, and are held in a natural shape from interaction
(Hydrogen or disul de bonds) of side groups on the amino acids between different areas of molecules.
Disruption of H-bonds causes the protein to change shape and denature, altering properties such as
solubility, gel formation, and enzyme activity. The globular protein albumin found in egg whites change
from a yellow tinted color to a white color when denatured.
Temperature: Albumin is hypothesized to denature over an optimum temperature of 62ºC when at a pH
of 7.4 (Baler et al., 2014). A temperature above will change the tertiary structure and shape (Clark, 1943)
Sodium Chloride Salt: High salt concentrations creates competition between the protein and salt ions to
hydrate. The salt will remove water molecules from protein surfaces which eventually denatures the
protein (Sinha & Khare, 2014).
Sodium Hydrogen Carbonate (aka. Sodium Bicarbonate) and Citric acid: Changes in pH alter electrostatic
changes and positions of hydrophobic and hydrophobic regions of the protein (Baler et al., 2014),
leading to a change in solubility which denatures the protein. Albumin has an average pH of around or Methodology of the Experiment
just above 7.  Overall set up of the experiment:
Silver Nitrate (heavy metal): Silver Nitrate has a pH of just above 3. Heavy metals can form a complex 1. Crack ope n10 eggs (Fig 1F). pour the egg white into a 600 ml
with functional side chain groups of proteins, which interfere with the proteins’ physiological activities, beaker (Fig 1B), and the egg yolk into another beaker.
therefore changing their functionality (Tamás et al., 2014).  2. Prepare 6 test tubes (Fig 1D) each with 40 ml of egg white
using a syringe (Fig 1A). Label each of the test tubes (should
look similar to gure 2). Prepare a timer set to 5 minutes.
Variable 1: Temperature
Independent and Dependent Variables: 3. Prepare and 2 sous vide water baths at ideally 30ºC and 60ºC.
Place a 600 ml beaker into each water bath and place a test
tube in each. Start the timer. When the timer ends, use the
tongs to place the test tubes in from of a black background.
Make any observations. 
Variable 2: Sodium Chloride Salt (NaCl)
4. Prepare 5 grams of Sodium Chloride by adding the salt to a
weighing boat (Fig 1C) on top of a weight scale (Fig 1G). Add
the Sodium Chloride into one test tube, and start the timer. 
Figure 1: An image of the apparatus used in this experiment
5. Stir the solution using a glass stirring rod. Start the timer and
write down observations. When the timer ends, place the test
tube in front of a black background, and make any nal
observations.
Variable 3: Sodium Bicarbonate (NaHCO3)
6.  Prepare 5 grams of Sodium Bicarbonate using a weighing boat on top of a weight scale (as done for Sodium Chloride).
Controlled Variables: Add the Sodium Hydrogen Carbonate into one test tube, and immediately start to timer. Repeat step 5.
Variable 4: Citric Acid (C₆H₈O) 
7. Prepare 3 lemons (Fig 1E), and extract lemon juice by squeezing them with a lemon squeezer.
Pour the solution into a 100 ml beaker. Dispose of the leftover lemons. Use a pipette to
transfer 5 ml of the juice into one test tube, and immediately start the timer. Repeat step 5.
Variable 5: Silver Nitrate (AgNO3)
8.  Measure 5 ml of Silver Nitrate using a pipette and a graduated cylinder. Pour the Silver
nitrate from the graduated cylinder into one test tube, and immediately start the
timer. Repeat step 5.
9. After all the experiments are complete, wash and clean any apparatus including glassware,
and safely dispose of any left over solutions. Return any apparatus to their original locations.

Raw Data: Qualitative observations of the egg whites containing albumin after being denatured through varying
methods of denaturation. These observations are seen in Figure 3, and explanations are referenced back to the
background research.

Uncontrolled Variables:

Apparatus and Materials:

Evaluation:
Conclusion and Analysis of Collected Data:
Strengths of the experiment:

The data collected in this investigation shows the effectiveness of 5 This investigation reveals the overall effect of
different protein denaturation methods on albumin proteins found in temperature, salt, pH, and heavy metals on the
egg white. The temperature was adjusted using sous vide water baths, denaturation of albumin. The methodology and
and solutions and compounds were simply added to the egg whites. results are not limited to Sodium Chloride salt,
The data collected is purely observational qualitative data. Using the or Silver Nitrate as the heavy metal variable.
Figure 2: Initial appearance of albumin egg observation and
white in test tubes. theorized explanations, Limitations of the experiment:
the followed conclusion
can be made:
References:
1. AGC Chemicals. (n.d.). Sodium bicarbonate. Sodium
Bicarbonate. Retrieved December 6, 2022, from:
https://www.agc-chemicals.com/jp/en/products/detail/
index.html?pCode=JP-
ENC010#:~:text=Sodium%20bicarbonate%20is%20a
Although the
%20white,substances%20with%20the%20lowest%20
alkalinity  Hydrogen bicarbonate
2. Baler, K., Martin, O. A., Carignano, M. A., Ameer, G.
A., Vila, J. A., & Szleifer, I. (2014, January 30). seems to have a
Electrostatic unfolding and interactions of albumin
driven by ph changes: A molecular dynamics study.
The journal of physical chemistry. B. Retrieved signi cant effect on
December 6, 2022, from https://www.ncbi.nlm.nih.gov/
pmc/articles/PMC3983335/  the albumin based
3. Clark, J. H. (1943, November 20). Denaturation
changes in egg albumin with urea, radiation, and heat.
The Journal of general physiology. Retrieved
only on observations,
December 6, 2022, from https://www.ncbi.nlm.nih.gov/
pmc/articles/PMC2142588/  the explanation behind
4. Marengo, K. (Ed.). (2019, May 21). Citric acid:
Structure, uses, intolerance, and more. Medical News the reaction suggests
Today. Retrieved December 6, 2022, from https://
www.medicalnewstoday.com/articles/citric-acid  that the foam was Further Investigations:
5. Sinha, R., & Khare, S. K. (2014, April 9). Protective
role of salt in catalysis and maintaining structure of
halophilic proteins against denaturation. Frontiers in unlikely to be purely • Expand the investigation to use a more varied range of temperatures and pH to
microbiology. Retrieved December 6, 2022, from
https://www.ncbi.nlm.nih.gov/pmc/articles/ the denaturation of albumin. However, the other reactions do not make additional observations.
PMC3988381/ 
6. Tamás, M. J., Sharma, S. K., Ibstedt, S., Jacobson, T.,
& Christen, P. (2014, February 25). Heavy metals and
have notable chemical reactions that would interfere with results, • Investigate the effect of the denaturation methods on other proteins such as
metalloids as a cause for protein misfolding and
which therefore determines the reactivity as shown in the diagram. meat based proteins, or gelatin as examples.
aggregation. Biomolecules. Retrieved December 6,
2022, from https://www.ncbi.nlm.nih.gov/pmc/articles/
Figure 3: Final image of results
PMC4030994/
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