Enzymes are a group of polypeptides that have a function as a catalyst in biochemical processes that can occur inside or outside the cell. Enzymes function to lower the activation energy because a chemical reaction assisted by enzymes must have a lower energy.
Enzymes can be separated from their sources, which come from animal cells and plant cells. This process of separating the enzyme from its source is called an enzyme isolation.
An enzyme is isolated to determine the characteristics that are in an enzyme. One of the functions of enzyme isolation is to separate an enzyme. One of the enzymes that can be separated from the source is the bromelain enzyme.
Bromelain enzyme is a proteolytic enzyme that is only present in pineapple fruit and stems. This proteolytic enzyme is an enzyme that can break down small molecules from a protein.
This bromelain enzyme can be produced from the side of a fruit juice factory called Ananas Comosus. Ammonium Sulfate and alcohol are some of the compounds that can be used as a precipitate in this bromelain enzyme.
Bromelain enzyme turns out to have several beneficial uses for our bodies, some of those uses are reducing pain in swelling, especially swelling after surgery or also because of wounds including being able to heal burns.
This enzyme is also useful for reducing arthritis, can also add lung function in a respiratory infection and several other uses that cannot be mentioned.
Consumption of bromelain in an anticoagulant compound should not be taken together because it will cause increased bleeding. So, to improve digestion in humans so as not to cause bleeding, the bromelain enzyme content should be consumed at a dose of 500 mg in capsule form.
The anticoagulant in bromelain is a substance that can prevent blood clots from binding to calcium and inhibiting the formation of thrombin which converts fibrinogen to fibrin in a blood clotting process.
Examples of anticoagulants are heparin, oxalate, Trisodium citrate dihydrate and EDTA or ethylenediaminetetraacetic acid. Several bromelain enzyme researchers such as Peckolt in 1870, Chittenden in 1892 and Cadwell in 1905 conducted research on ways to isolate bromelain enzymes from pineapple juice.
Meanwhile, Henich and Gortner researched the isolation of the bromelain enzyme in pineapple juice in 1957. From several studies that have been carried out, it turns out that the molecular weight of a bromelain enzyme is 33,000.
In addition, there are some differences regarding the amino acid content in the bromelain enzyme derived from pineapple juice and that derived from pineapple stem juice.
Because of the many studies on the isolation of bromelain enzymes, this activity is often also used as a biochemical practice by several experts. To carry out a biochemistry practice there are several procedures that must be carried out and categories that are used as a benchmark.
Basically, there are two ways to isolate bromelain enzymes.
The method is as follows:
1. Bromelain enzyme isolation using acetone
Usually, acetone is a cosmetic to remove nail polish or nail polish. Acetone can also be used for a study to isolate enzymes. The ways to use acetone to isolate the bromelain enzyme are as follows:
Prepare the pineapple. Peel the pineapple from the skin and cut into small pieces as desired.
Blend the small pieces of pineapple by adding ice according to taste so that it becomes a juice. The addition of ice cubes in the content of pineapple juice is intended to make the enzymes in the pineapple not damaged.
Separate the pineapple juice pulp from the juice. Use a filter to make it easier.
Refrigerate the juice or filtrate for 3 hours.
After cooling the pineapple filtrate for 3 hours, add acetone with 3 different levels, namely 35%, 55% and 70%.
This mixture of acetone and pineapple filtrate was precipitated using a centrifuge for 15 to 30 minutes. Separate the precipitate that forms.
Add the filtrate with ammonium sulfate with a concentration of 40% and centrifuge again to obtain a second precipitate. After that, add 60% ammonium sulfate and then return to the centrifuge.
After producing the third precipitate, test the protein content of the pineapple juice. To determine the protein content of the enzyme from the precipitate formed, use a spectrophotometer by determining the wavelength.
2. Isolation of bromelain enzyme using Ammonium Sulfate
Making a practical experiment to isolate bromelain enzyme from Ammonium Sulfate is as follows:
Prepare pineapple and peel it from the skin.
Cut the pineapple according to taste and add phosphate buffer with a pH level of 7 then once mixed, blend the pineapple pieces.
After blending, take the filtrate and filter and then cool for 15 minutes.
Add ammonium sulfate with a concentration of 20% and then cool again for 15 minutes.
After the filtrate mixture was cooled, the solution was centrifuged for 15 minutes at a speed of 3500 rpm at 0˚ C. Separate the precipitate formed from the filtrate mixture and centrifuged.
Add more ammonium sulfate to the front of the filtrate and also increase the concentration to 40% and then centrifuge to produce a second precipitate.
Repeat the previous step and increase the ammonium sulfate content to 60% to obtain a third precipitate. After getting the third precipitate, test the protein content.
In conducting research on isolating the bromelain enzyme, the materials and tools that must be prepared are:
- Tool
- Blender
- Knife
- Cutting board
- 2 250 ml beakers
- Plastic
- Stirring rod
- 2 measuring cups
- Analytical balance
- Napkin
Materials needed:
- 250 grams of pineapple
- 153 ml of cold acetone
- Ice cubes to taste according to taste
- Aquades 100 ml
In this biochemical study, the isolation of the bromelain enzyme will use a category that uses acetone. But the steps described will be slightly different from the work steps mentioned above. The steps are as follows:
Clean the pineapple and cut it into small pieces. Can resemble dice and then weigh as much as 250 grams.
Add 100 ml of distilled water then puree with a blender. Before in the blender, the color of the pineapple is bright yellow, while after blending the color of the pineapple becomes light yellow.
Strain the pineapple pulp into gloves and the juice or filtrate is kept for 15 minutes.
Add the cold acetone little by little and stir continuously until the acetone content is 30%. Then after that let stand for 1 full day or 24 hours.
After being allowed to stand for 1 day, weigh the precipitate formed in the filtrate. Usually, the resulting precipitate will be yellow in color. This precipitate is called the bromelain enzyme.
Record any visible observations.
In recording the observations, a treatment table with the results of the observations must be needed so that we know more about the developments that occur.
So, making biochemical research on how to isolate the bromelain enzyme is not that difficult, we are just looking for some chemicals to produce the enzyme itself. Not too complicated or difficult. I have explained simple ways in the practicum easily and simply.
That's all about the bromelain enzyme, complete with the practical steps. Hopefully what I write can be useful for readers, good luck!