A study on Enzymes substance reactions

Enzymes were uncovered with a German chemist Eduard Buchner nearby the end of the 19th century. He previously been wanting to extract a fluid for medicinal use from yeast, however, the yeast extract retained going bad. He then decided to add sweets to the yeast, however, the yeast turned the sugar into alcohol, which is also known as fermentation. Buchner looked into into this and soon found out that living cells were not in charge of this fermentation which it was triggered by the fluid that was wanting to be extracted from the yeast. The term enzyme was coined for the substances in the drink that advertised fermentation. Although enzyme practically means "in yeast", it is now however being used as the collective noun for several hundreds of materials that have shown to have a catalytic action on specific chemical substance reactions.

Enzymes are biological or organic and natural catalysts composed of necessary protein. They catalyse (increase/decrease the rate of) chemical substance reactions without themselves being chemically altered at the end of the effect. It could therefore be utilized repeatedly therefore works well in smaller amounts. They essentially work by lowering the activation energy of the reactions and therefore allowing the reaction to place at a quicker rate. In enzymatic reactions, the molecules are the start of the process are called substrates, and the transformed molecules, the products.

Properties of enzymes
Enzymes have the following properties

Enzymes modify the rate of chemical reactions without themselves being chemically transformed by the end of the effect.

Enzymes are extremely powerful. Since enzymes are very specific, a tiny amount of the enzyme is with the capacity of catalysing an enormous chemical effect.

Enzymes are influenced by temperature. Enzymes are inactive at low conditions. Increasing the temp increases the activity of the enzymes. There can be an optimum working temperatures of which certain enzymes work best. This is normally between 37-42 degree centigrades. However, a higher temperatures, anything above 45 level centigrades normally destroys the dynamic sites of the enzymes and causes it to denature. This entirely damages the enzyme and they become functionless.

Enzymes are afflicted by pH. Certain enzymes work best in acidic conditions whereas certain enzymes work better in alkaline conditions. For example, pepsin works best in the tummy where the pH is below 7, however intestinal enzymes work better in coditions of pH of above 7.

Some enzymes may require a substance to be bound to them before they can catalyse chemical reactions. These substances are called co-enzymes.

Enzymes can work in either directions. Metabolic reactions are reversible and the way where the reaction goes is determined by the levels of substrate and products present. The effect will continue from left to right until an equilibrium is reached between the substrates and products. Also, when there is a sizable amount of products, then your reverse reaction starts and hence causes the product to be split up until again equilibrium is made.

Lock and Key & Induced Fit Hypothesis

Although enzymes have a large size, however, they have only a little region that is functional. That is known as the dynamic site. Energetic sites serves as a depressions on the surface of the enzyme. Just a few of the amino acids of the enzyme molecule make up this effective site; the rest are used to maintain its overall three-dimensional form. The energetic site is the website where in fact the substrate binds onto the enzyme in support of substrates with a particular molecular condition will have any chance to bind effectively with the particular enzyme. This is the reason why enzymes are specific in their actions as they can only bind to specific substrate substances.

Another idea which arose from the lock and key hypothesis is the induced fit hypothesis that shows that the enzyme alters its form just a little to ensure that the enzyme substances bind tightly with the substrate molecule. However, after the product or substrate leaves the effective dynamic site, the active view realigns itself to its original form.

Now, having talked about enzymes generally, we shall move onto talking about the enzymes more specific to the test. The enzyme being used in this investigation is Neutrase. Neutrase is a bacterial protease which is produced from a bacterial pressure called Bacillus Amyloliquefaciens. Protease can be an enzyme which catayses the hydrolysis of protein into polypeptides/amino acids. In humans, the digestion of proteins chiefly starts in the belly with Pepsin in the gastric juice where the protein are changed into polypeptides. Necessary protein digestion is displayed in this test by the actual fact that whenever the protein in the dairy are digested, the solution becomes from opaque cloudy white to a see-through solution.

In an enzyme handled effect such as that of necessary protein digestion in dairy, an increase in the attention of enzyme will lead to an elevated reaction rate. That is because of the fact that whenever there are definitely more enzyme substances present, you will see a greater potential for more of the enzyme molecules colliding with the substrate and therefore increasing the occurrence of the collisions. This increased occurrence of collisions will form an enzyme-substrate organic more rapidly.


The aim of this experiment is to see when there is a negative relationship between your enzyme attention and the rate of digestion of the necessary protein content in the milk. Upsurge in enzyme concentration resulting in decrease in the speed of digestion of proteins content.


H1 - There is a positive correlation between the enzyme focus and the speed at which the protein content in the milk digests. (Increase in rate of response).

H0 - There is no correlation between the enzyme awareness and the rate at which the proteins content in the dairy digests. (No impact on rate of effect).



Justification For Use

Marvel Powdered Milk

The way to obtain protein upon which the enzyme to work on.

Enzyme (Neutrase )

The enzyme which digests the proteins content in the dairy.

Sodium Phosphate Buffer (pH 6. 4)

In order to guarantee the pH of the perfect solution is remains regular for optimum working of the enzyme.

Test Tubes And Test Tube Rack

Test Tube: To hold the enzyme, buffer and dairy powder solution.

Test Tube Rack: To hold the test tubes.

Measuring Cylinder

To measure out the quantity of distilled normal water.

Measuring Scale

To solution out the mass of the dairy powder.

Pipette And Pipette Filler

To solution out accurately the volume of the enzyme.


To hold the normal water and the test tubes formulated with the enzyme, buffer and milk powder solution.


To time the duration it takes for the enzyme to completely digest the health proteins content in the dairy.


To add the answer of the enzyme, buffer and milk powder in to the curvettes.


To measure the light absorbancy principles.

Preliminary Test:

A preliminary experiment was conducted initially in order to check if the method designed for use was flawless or not. However, through the conduction of the experiment, a number of flaws were observed and hence taken care of to make a more flawless and acoustics experimental method.

In the primary experiment, it was chose that the heat range of the perfect solution is filled with the enzyme would be stored constant by by using a bunsen burner. However, fluctuation in temeperature were going to be evident and hence causing inaccuracies. As a result, it was decided to use a thermostatically controlled water bathroom to keep the temperature constant.

Also, when using the colorimeter, I decided to place the perfect solution is inside the curvette and then place it onto the colorimeter and record the prices as time passes until it reached 0. However, this might give inaccurate results predicated on the fact that when the answer was removed from the water bathtub and positioned inside the curvette, the heat would decrease over time and hence have an effect on enzyme activity. Because of this, it was chosen that the perfect solution is be taken off the water bath and placed into the curvette at regular intervals. After each 20 seconds, a new curvette would be utilized with solution removed from the water shower and then placed onto the colorimeter to track record the light absorbancy values.


Measure out 10. 00 grams of milk powder using an electronic mass balance into a beaker. Then assess out 200 cm^3 of distilled drinking water by using a measuring cylinder. Add this inflatable water into the beaker filled with the dairy powder and use a spatula to stir the perfect solution is well.

Now, different concentrations of enzymes are to be prepared.

To make a 1% amount solution of enzyme, add 1cm^3 of the Neutrase into a test tube utilizing a pipette. Then add 99cm^3 of distilled normal water measured out by using a measuring cylinder. Tremble the test tube including the perfect solution is (100cm^3) well.

To make a 2% attentiveness solution of enzyme, add 1cm^3 of the Neutrase into a test tube utilizing a pipette. Then add 98cm^3 of distilled water measured out by using a measuring cylinder. Tremble the test tube made up of the perfect solution is (100cm^3) well.

Use the same method as above to make 3%, 4% and 5% solutions. Place each of the test tubes into the test tube rack.

Add Sodium Phosphate Buffer ( 6. 4 pH ) to each test tube.

Place one of the test tubes containing the enzyme solution and anybody of the dairy solution test tubes into the water shower for 5 minutes.

After five minutes, immediately pour the enzyme solution into the test tube made up of the milk solution.

Use a colorimiter to check the quantity of light passing through the perfect solution is as time passes. Press 'R' to reset the colorimeter (use the red filter) as red light is transmitted the best which will give accurate readings.

After every 10 seconds, place some solution from the test tube into the a curvette and the curvette onto the colorimeter and record the effect.


The sole indie adjustable in this test is the attentiveness of the Neutrase solution which varies from (1-5)%.

Other variables which could affect the research have been tabulated the following


How may a big change in this changing affect the info?

How will it be controlled?

Type of milk used.

Some milks will have significantly more or less protein molecules present than others which alters the amount of substrate molecules being available to form enzyme-substrate complexes.

This will be controlled by ensuring that the same dairy powder is used throughout the test.

pH of the perfect solution is.

Enzymes work better in alternatives of different pH. The experience of the enzyme depends after the pH of the solution; if optimal pH solution can be used, the enzyme would work better than if non-optimum pH was used.

This will be controlled by ensuring that a pH buffer ( Sodium Phosphate - pH 6. 4) is used. This can minimise any changes in the pH of the solution and ensure that the most effective pH is maintained constant for reliable working of the enzyme.

Volume of Neutrase solution used.

A larger level would of enzyme ( Neutrase ) soltuion would bring about a larger amount of enzyme-substrate complexes and therefore increasing the speed of the effect.

Neutrase solution level will be stored constant by precisely measuring the quantity required by using a pipette and pipette filler.

Temperature of the perfect solution is.

Temperature alters the speed of enzyme activity and an increased temperature would lead to better enzyme activity. However, anything normally above 45 degree centigrades would lead to denaturing of the enzyme.

Using a thermostatically controlled water shower will maintain a frequent temperature. The heat range would be stored regular at the maximum working heat for the enzyme.

Volume of dairy used.

A larger level of milk would lead to a greater amount of enzyme-substrate complexes being made as there is a greater variety of protein present and so increasing the speed of the reaction.

Milk level will be maintained constant by precisely measuring the quantity required utilizing a pipette and pipette filler.

Health & Security Regulations:

Wearing a lab overcoat inside the laboratory.

Wearing safety goggles to protect the eyes from chemicals.

Wearing plastic material gloves when controlling the enzyme and milk solution and to prevent contaminants.

Keeping the test tubes in a rack to avoid unintentional breakage or spilling.

Bunsen burners will be guaranteed they are kept on yellowish flame when not used.


The results of the test demonstrate that the H1 hypothesis is accurate. There is a positive correlation between the enzyme (Neutrase) attention and the pace of which the health proteins in the dairy digests. In other words, increasing the focus of the enzyme escalates the rate of reaction.

The results show that an increase in the awareness of the enzyme contributes to a descrease in enough time taken for the solution to look clear. That is because of the fact that when the focus of the enzyme is increased, there are more active-sites present on the enzymes onto that your substrate binds onto. This creates more and more enzyme-substrate complexes. The higher concentration of enzyme increases the kinetic energy of the molecules and increases the consistency of the collisions between your enzyme and substrate molecules. A lesser concentration of enzymes means there would be fewer lively sites and hence, a slower rate of response.

However, if we stored increasing the attention of the enzyme, you might observe that the brand would start to level off because every one of the effective sites on the enzymes would be occupied and therefore, increasing the concentration would have no effect on the rate of the effect.


Observing the results, one may easily generate the final outcome that there surely is a positive correlation between the enzyme attentiveness and the digestion of the necessary protein content. Even though, the group of results achieved are not fully accurate and hence, there are anomalies. However, advancements can be made to minimise inaccuracies and unreliability in the test.

One of the factors that could have damaged the experiment might have been human reaction error in timing. Also, the starting of the stop-clock for each and every of the tubes could have been different and this could have resulted in unreliable results. To avoid this, however, an automatic stop-clock could be used.

Another factor that can have influenced the experiment might have been the movements of the reacting mixture constant. The mixing between the substrate ( Health proteins in the milk ) and the Enzyme ( Neutrase ) cannot have been same in each tube. This could have led to inaccuracies in the result as the molecules in the tubes with the better combining would have better kinetic energy and hence, the effect rates would be quicker as more and more enzyme-substrates would be formed quicker. In order to avoid the scope of mixing in each tube from influencing the results, I will ensure that the next time, there will be no stirring of the mixtures when alternatives are added at all so that way, it'll be regular ( no combining ) for each and every tube.

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