Investigating the Effect of Lipase Concentration on the Breakdown of Fat in Milk

INTRODUCTION:

Enzymes are proteins which can catalyse chemical reactions without changing themselves. The enzyme lipase breaks down the fat in dairy products such as full-cream milk for people who are lactose intolerant. Lipase acts on its specific substrate, lipids produces fatty acids. If enzyme concentration increases, random collisions between the substrates and active sites of enzyme increase due to the increasing amount of active sites which allow more collisions to happen, so the rate of breakdown of lipids to simpler substances will increase. During the experiment, sodium carbonate solution and pH indicator phenolphthalein will be added ahead of lipase enzyme; when the pH is below 8.2, phenolphthalein will turn from pink to colourless. Sodium carbonate solution is added in order to turn the solution alkaline and the indicator pink then back to colourless after lipase is added to catalyse the chemical reaction and speed up the breakdown of lipids into fatty acid in the milk. Fatty acid produced then is going to neutralise the solution with sodium carbonate solution added previously; phenolphthalein will finally turn from pink back to colourless.

AIM:

In this experiment I will investigate the effect of increasing enzyme concentration on the activity of the enzyme lipase that will be allowed to act on the same amount of milk and the time taken for the phenolphthalein to turn from pink to white compared.

HYPOPTHESIS:

With higher lipase concentrations, there will be more active sites in the solution to be fit in by the substrates, which means more collisions between the active sites and the substrates and quicker breakdown of lipids in the milk within shorter period of time. At enzyme concentration of 1% or 2%,

reactions can happen at a relatively slower pace, with lipids broken down and phenolphthalein turning from pink to white over a relatively longer time period. At 3%, enzyme activity will speed up with lipids broken down and phenolphthalein turning from pink to white over a relatively shorter time period. At 4% or 5%, enzyme activity will happen significantly faster.

APPARATUS:

1. Lipase solution at concentration 5%
2. 1 test tube
3. 5 dropping pipettes
4. full-fat milk
5. 4 measuring cylinders of 10ml (0.5ml)
6. 1 stopwatch (1s)
7. 1 stirrer
8. phenolphthalein
9. sodium carbonate solution at 0.5 mol dm3
10. Marker pen
11. distilled water
12. 4 beakers of 200ml (5ml)

VARIABLES:

Independent Variables:

1. Enzyme concentration- effectiveness of enzyme concentration will be tested at 1%, 2%, 3%, 4%, and 5%. Enzyme lipase at the concentrations of 1%, 2%, 3%, 4% will be made by mixing lipase at 5% and distilled water; all these solutions will be contained in different beakers to keep them pure. In order to make 1% lipase, for each 1ml lipase, 4ml distilled water will be added and poured into a beaker; to make 2% lipase, for each 1ml lipase, 3ml distilled water will be added; to make 3% lipase, for each 1ml lipase, 2ml distilled water will be added; in order to make 4% lipase, for each 1ml lipase, 1ml distilled water will be added.

Controlled Variables:

1. Volume of the milk used for every test will be kept constantly at 5ml; substrate concentration will be kept the same since full-cream milk will be used. Different volumes of milk will allow different amount of substrates to be contained in the milk. Therefore, more substrates contained in the milk will take it more time for the enzyme to break all the fats while the amounts of lipase and sodium carbonate solution stay the same.

2. pH of the substrate the identical amount of 7ml sodium carbonate solution, which is alkaline, will be added in the test tube; if not, more or less fatty acids will be produced and needed to neutralise the solution and for the phenolphthalein to turn from pink to colourless.

3. Volume of lipase solution 3ml lipase will be added to for each test. By keeping this the same, the amount of active sites of enzyme will be equal in all the cases, therefore allowing results to be fair. If more were placed in one of the test tubes, then the rate of activity of the enzyme at different concentrations will be wrongly judged, leading to incorrect conclusion.

4. Temperature will be kept