effects of pepsin on milk

[kgaur]

Hi Science Buddies!
I'm doing a science project on milk and pepsin this year, and I don't know how I'm supposed to replicate digestion! I have to do this in order to demonstrate how pepsin breaks down the protein in milk, and which kinds of milk have their protein broken down the best for digestion. I want to replicate it up until the point after protein is broken down by pepsin. I don't know how to replicate everything! I've done some research, but I still don't know how I'm supposed to do it. I think I could demonstrate the mixing of the milk and saliva in the mouth by mixing milk and water ("saliva") in a blender, but I don't know what to do after that. HELP! I DON'T KNOW WHAT TO DO! I'M DESPERATE! I'M SORRY I SOUND SO FRANTIC, BUT I'M PARANOID! PLEASE HELP! THANK YOU!
yours in paranoia,
Kriti Gaur

P.S. I also wanted to know which forms I could test this in, because I'm using Vitamin D milk, 2% reduced fat milk, whole milk, chocolate milk,and soy milk in my experiment. I wanted to do my experiment so that the milk (IV) would be in its liquid state. Is that possible? Because if there are suggestions with dry milk, that's OK too, I guess.

[Walker]

Hello,

Since you want to learn about how pepsin effects milk proteins in normal digestion, you'll want to develop an experimental protocol that replicates the normal "working environment" of pepsin pretty well. Pepsin is produced in the stomach, so you should think about what it's like in there. By the time food in general gets to the stomach, it is well mixed (so your blender idea sounds like a good one), acidic (the stomach has special cells that produce natural acids to keep the pH very low) and warm (near body temperature of about 37 degrees Celsius), so it's reasonable to predict that pepsin will work best when it's added to a solution that's well-mixed, acidic, and warm (these are predictions that you could easily test if you want!)

If you add pepsin to a warm, acidic, well-mixed solution of milk proteins (nonfat dry milk, casein, or whey protein could be good choices for milk protein sources), you can expect the pepsin to start chopping up the proteins into smaller pieces. An important decision you'll need to make is how you are going to measure this activity of the pepsin. A fairly simple way, if you have access to electrophoresis equipment, would be to separate the milk proteins by size by using electricity to push them through a polyacrylamide gel. You could stain the proteins in the gel with a dye like Coomassie Blue, so then you'd actually be able to see the protein bands in the gel shift down to smaller sizes as the pepsin works. There are also biochemical tests that you could use to measure the activity of the pepsin, for example by tracking the change in spectrophotometric absorbance of a protein solution over time, or performing a biochemical test that produces a color change in the test tube based on the presence of more small peptides in the digested samples. If you post back with some information about the equipment you have to work with, the experts here can help you more with suggestions about what a good assay may be for you to use.

Once you have a good pepsin activity assay working, you'll need to decide what question you're most interested in asking about the effect of pepsin on the milk proteins. For example, you could find out how well pepsin works on different types of milk proteins, such as casein vs. whey proteins (they're both available in purified forms at nutritional supplement stores), or you could study the effectiveness of pepsin in different environments, such as at different pH values or temperatures, or you could find out whether the pepsin digestibility of milk is different when the milk is organic, or lactose-reduced, or ultra-pasteurized... your curiosity is the limit!

Hope this helps. Good luck with your project!

-Will

[deleted-71536]

Hi Kriti,

You have a very interesting project, and Will has given you some solid advice.

I just wanted to point out a few more things about digestion that may help you. You may already know that pepsin is a protease, which means that its job is to break down proteins. Protein digestion actually begins in the stomach, so you don't have to worry about what happens in the saliva. (Carbohydrate digestion does begin in the mouth, but you are focusing on proteins here.) As Will pointed out, the stomach is very acidic - the pH is about 1-2. It gets that way because of hydrochloric acid (HCl), so it's perfectly legitimate for you to use HCl to replicate the conditions of the stomach. You can still go with a blender here, since the stomach churns its contents to make sure they are well mixed.

You will need to think about how you will test which milk had its proteins "best" broken down by pepsin. Will has given you some good ideas to get started.

Let us know if you have more questions!

Good luck,
Heather