How Do Protease in Some Fruit Juices Help Softening Meat?

Hypothesis:

The protease in some fresh juices digests collagen, which gives meat a tough texture by holding meat fibres together.

Design of investigation:

In order to test the hypothesis, the presence of protease in those fresh juices was tested at first. Then the amount of them was needed to be compared for finding their difference in effectiveness in tenderizing meat. Control experiments were set up for proving protease was taking part in softening meat. According to the hypothesis, protease is in charge of breaking down protein and softening meat.

The independent variable is the concentration of protease present in some fruit juices. To manipulate the independent variables, different kinds of fruit extracts which representing different source of protease should be tested in order to get a more comprehensive result. In this experiment, pineapple, papaya and kiwifruit were used because of their high amounts of protease. After all we can have a set of samples of different concentration of protease.

In this experiment, a plate of milk agar is needed for comparing the result of each sample. Circular wells were dug on the milk agar plates by the use of cork borer, and extracts could be put into the wells by dropper. One must note that milk-agar is white in color due to casein. If protease is present, casein will be digested by the enzyme giving a clear zone in the milk-agar plate. The size of the clear zone can help us to determine whether fruits differ in their in concentration of protease and effectiveness in softening meat.

The dependent variables are the size of the clear zone. To measure these variables, we should use a ruler to measure the diameter of the clear zones caused by different fruit juices. The amount of protease presented can be compared by performing the above test and then knowing their effectiveness in softening meat.

The results which were expected to obtain were that protease is in charge of digesting protein in order to soften meat. And higher concentration of protease, more efficiency in softening meat.

Controlled variables should be temperature, pH value and volume of the hole on the milk agar plate. The experiment should be done under room temperature and pressure. Use the same tooth-pick to make wells on milk agar so the size of them will be the same. Moreover, control experiment should be done by adding water and boiled fruit juices into the hole of agar plate and put it in the same environment as the setup. It can proved protease takes place in breaking down protein since protease will denature if it’s under a high temperature .

Assumptions:

The area of the clean zone is proportional to the amount of protease present in a sample. The protease present in the fruits ’s tissue can be extracted by water.

Apparatus：

Plastic vials and glass rods, measuring cylinder, petri dishes, boiling tues, dropper, mortar and pestle, spatula, beam balance, marker pen, forcep, toothpick, cork borer, knife, muslin cloth, beaker, bunsen burner, ruler

Materials：

Distilled water, a plate of milk agar, pineapple, papaya, kiwifruits

Procedure：

• Label seven boiling tubes for holding different juices: FA for fresh pineapple juice, FP for fresh papaya juice, FK for fresh kiwifruit juice, BA for boiled pineapple juice, BP for boiled papaya juice, BK for boiled kiwifruit juice and W for distilled water.
• Remove the skin of fruits by using a knife.
• Cut each fruit into smaller pieces by using a knife. Put each kind of fruit into different Petri dishes
• By using the beam balance, having same volume of the fruit from each sample. Put the other of them into plastic vials by using a spatula for making sure there are spare materials for further experiment.
• Grind the tissue by means of mortar and pestle to obtain about mL of juice, which is measured by the measuring cylinder.
• Filter the extract of each sample through the muslin cloth into a boiling tube. And stir them with glass rods.
• Transfer mL of each extract into a boiling tube which is measured by the measuring cylinder. Each fruit juice should be having two set of it.
• Transfer  mL of distilled water which is measured by the measuring cylinder to another unused boiling tube to be a control experiment.
• Circular wells are dug on the milk agar plate by using a cork borer. The circular agar blocks can be taken off by using a tooth-pick. Name the wells with marker pen.
• A forcep is used to hold the milk agar plate above the fire of Bunsen burner for about  minutes.
• Prepare hot water in a beaker. ( may be do it while the other group members are preparing the fruits.) Then put three tubes of juices into the beaker.
• After minutes, take out those tubes from the beaker gingerly.
• All liquid of boiling tubes could be put into the wells by dropper.
• Measure and record the size of each of the clear zone by a rular, compare the result.

Precautions:

• For preventing present of bacteria, using non-luminous to disinfect all apparatus.
• Wear safety goggles and gloves when handling the fruit juices, as they may be slightly corrosive if contacted.
• It is also important to wear safety goggles and gloves when using Bunsen burner, as it provides a layer of protection from being burned.

Discussion:

The results show that with increasing concentration of protease, the area of the clear zone larger, which represents a higher efficiency in breaking down protein and thus softening meat. This supports the hypothesis that the softening of meat is caused by the protease present in some fruit juices.

The following are the sources of error of this experiment:

• the milk-agar in the Petri dish might have contaminated when the cover was removed to drop some fruit juices into the wells.
• the measuring of the result might not be accurate enough.
• There are some protease may sink on the boiling tube. Their volume would be changed.
• Since a constant room temperature environment is needed, some of the protease may not work under its optimum temperature, which may affect the result.

To increase the accuracy of the experiment, we can do the followings:

• to drop the juices faster and more accurate, and then cover the lid of the Petri dish immediately
• To ensure contaminations are not present, using non-luminous to disinfect apparatus.
• use a smaller interval ruler to measure the results to increase the accuracy of the result of the experiment
• Stir the extracts properly to ensure there isn’t any protease sinks at the bottem of tube.
• filtration is conducted before dropping the fruit juices into the agar plate; this helps to prevent seeds and other impurities to affect the result of the experiment.
• the fruit juices are labeled clearly using labels. This helps to identify the diameter of the clear zones of each fruit
• Gring the tissue into fine pieces in a mortar before adding water.

We found that proteolytic enzymes are very important in digestion. Body needs them as they can breakdown the protein foods liberate the amino acids . In addition, proteolytic enzymes always use in various forms of therapy. Their use in medicine is notable based on several clinical studies , for examples, blood control or inflammatory conditions.

As for further investigation, i am intereste to find out what type of meat also be soften by fruit juice. And are there still have different liqiud can digest the protein of meat.

Conclusion:

The protease in some fresh juices digests protein, thus softening meat.