Bromelain enzyme lab

[NastiaSawka]

My group wanted to experiment the effect that bromelain mixed in with different pH levels had on gelatin. We are supposed to find out the pH level that our enzyme will denature in.

We had 5 different pH levels: 1, 4, 7, 10, and 13. We did 5 trials for each pH level. Before we distributed the pH buffers into the test tubes, we poured in 100ml of the pH buffer and mixed in 10g of meat tenderizer, we poured 20ml of the solution in each test tube. We measured the weights of the gelatin before the experiment, and we used pure gelatin not flavoured jell-o. We measured the weights of the gelatin after leaving them in their solution. The measurements showed that the gelatin weighed more after the experiment, so we left the gelatin over the weekend to drain and we measured them again, the measurements were different but some still weighed more than the original. We found out that the gelatin weighed more because the molecules were being digested so it was breaking down and expanding, but we were not sure if that was reliable information. We found out that bromelain's ideal pH level is 4-4.5 but we are not sure how to tell when the enzyme denatures.

We are not sure if this was the right way to perform the experiment, or which pH level the enzyme denatures in.

[SciB]

Hi,

Bromelain is an enzyme that breaks down proteins by cleaving some of the bonds between amino acids. Its activity is maximal at pH 4.5. Your hypothesis would have been—at a pH other than 4, the digestion of gelatin will be reduced. The experiment to test this was to place weighed pieces of solid gelatin into a solution containing meat tenderizer in a buffer solution at various pH’s and leave them there for some period of time then weigh them again. How long did you leave the gelatin in the solution? What was the temperature of the solution? Enzymes take some time to work and work best at warm temperatures—that’s why humans are warm-blooded!

You said that you used meat tenderizer for the experiment. There are several different kinds of meat tenderizer available and some contain bromelain while others contain papain or both. They also have a LOT of sodium chloride, NaCl. I checked the label of McCormick’s unseasoned tenderizer and it contains 400 mg of sodium per ‘serving’ which is 1.1 g. if you are using a comparable tenderizer, that would mean you added 3.64 g of salt to 100 mL of buffer.

If my calculations are right that works out to 0.62M which may be a high enough concentration of NaCl to disrupt the ionic bonds in the enzyme—bromelain or papain. Thus the salt from the meat tenderizer could be denaturing the bromelain at every pH and it would not be able to digest the gelatin. The apparent increase in weight of the piece of gelatin may be due to its absorbing water.

I think the best way to have done this experiment would have been to use fresh pineapple juice which contains natural bromelain. Did you ever try making jello with fresh pineapple? It won’t gel because the bromelain digests the gelatin.

Why don’t you try a quick experiment using smaller amounts of tenderizer [2-4 g per 100 mL] or low-sodium tenderizer and see if the gelatin IS being digested. Once you are sure the enzyme is working then you can test the effect of using a different pH.

I hope this helps to answer your question. If you need more information just repost to this thread.

Sybee

[NastiaSawka]

Thanks Sybee,

I wanted to add in that we left the gelatin in the pH solutions for 24 and our meat tenderizer had only bromelain, no papain. Though I don't think this would make my experiment any more right.

I was also confused if you meant to try mixing the gelatin powder with meat tenderizer and then submerging it into the pH solution would work better?

But I guess we will need to start the experiment all over anyways.

So thank you for helping and giving another idea for the experiment!

[SciB]

Hi,

I’m glad my post was helpful. I think your enzyme was denatured by the NaCl in the tenderizer so try a smaller amount of tenderizer and use the low-sodium version. Since you are doing the experiment over, why don’t you get a pineapple and try some fresh juice along with the tenderizer? This would make a great comparison and kick your project up a notch.

As to exposing the gelatin, I did not mean to add tenderizer to gelatin powder. In all cases you have to follow the directions on the gelatin package to make it into a solidified gel that you can cut up into small cubes and add to the tenderizer plus buffer solutions.

Good luck and please let us know how this turns out. Don’t hesitate to post again if you have more questions.

Sybee