Ph testing of protein:

 

Introduction: Pepsin is a proteolytic enzyme that plays a critical role in the digestion of proteins in the stomach. It is produced in the chief cells of the gastric glands as an inactive precursor called pepsinogen, which is then activated by the low pH of the stomach.1      The pepsin will have Albumin added to it, to catalyze the reaction.  In this lab, Biuret is used as reagent for the reaction to see how well Pepsin works. A similar study published in the Journal of physiology conducted a similar experiment, but they did it on a much larger scale, used three different types of Albumin, and examined the bond types.2  During this experiment, we looked at one type of albumin and one type of Pepsin, and we tested it against how it would react with NaOH, we used six total samples, but one sample was just albumin and pepsin.

 

Methods:

1. put 3 ml of 2% albumin into six test tubes

2. put 3 ml of 1% pepsin into five of the test tubes, the last one should be labeled the “blank” sample.

3. Take the current pH of each sample.

4. Add NaOH to the first five tubes until there is a significant change in PH, about 25 drops each.

5. Create a table that lists the tube number, and the PH before the pepsin was added, and then a column for after the NaOH was added.

6. Put the samples in an incubator set to 37 °C

7. Wait one hour for the reaction to occur.

8. Take the measurements of pH after the time is up.

9. Record data in the table.

10. Add 1 mL of Biuret into the test tube. Biuret will change colors with the detection of either proteins or peptides. If protein is detected, it will change the mixture to violet in color. If a peptide is detected, pink or light violet in color.

 

Results:

Ph levels before and after the reaction occurred:

the first ph  level of ph after

blank 6

1) 5.38

2) 5.33       6.9

3) 4.95      6.4

4)  5.1       4.66

5) 5.2       3.84

6) 5.23       3.36

 

The samples were placed in test tubes, and the initial Ph was measured by a Ph sensor. Then, the samples were put in the incubator for the reaction to occur. Afterward, the Ph was measured again.

 

Color changing for evidence of proteins or peptides.

 

 

​

​

 

​

​

​

​

​

​

The samples were taken from the incubator after one hour, and the Biuret drops were added, this created the samples to change to the colors observed in these images. Samples one through five had evidence of peptides, from the color of the solution changed to violet, but the blank sample remained a full protein because the solution appeared dark purple.

 

Discussion: By the way the Ph and the color of the mixtures changed in the incubator, it seems that digestion occurred in all the samples. By observing the tubes it seems that the five tubes with Pepsin, NaOH, and Albumin had the highest amount of digestion. Tubes one through five became light violet in color suggesting that there is evidence of the breakdown of protein into peptides, but the blank sample appears to still be dark violet, so nothing reacted in this sample.  When someone has Achlorhydria, their digestion process would take much longer than the average person's because they lack the enzymes to catalyze and expedite the facilitation of digestion. Based on the data, the lack of Pepsin makes digestion improbable.

 

 

​

References:

Britannica, T. Editors of Encyclopaedia (2020, July 3). pepsin. Encyclopedia Britannica. https://www.britannica.com/science/pepsin

 

LOKEN, M. K., TERRILL, K. D., MARVIN, J. F., & MOSSER, D. G. (1958). Comparative studies of three methods for measuring pepsin activity. , (2), 251–258. https://doi.org/10.1085/jgp.42.2.251

​