Concentration of substrate:
If the concentration of the substrate is low then the particles will float about so there will be less collision in the particles and they wouldn’t take so long to react that means there are less collision taking place which means the rate of reaction decreases however if there are more concentration of the substrate then there will be more collision taking place that means that the rate of reaction increases. However the volume of substrate should be constant.
PH of enzymes:
All enzymes have a pH whether it is alkali or acid. The enzymes work best on certain pH. So if the pH is optimum then all the enzymes work best and the rate of reaction will increase. However if the more than the optimum pH increases then the enzyme denatures. By using a buffer solution would control the pH in the solution.
Gelatine contains eighteen amino acid those are insoluble but Bromelain in the fresh pineapple degrades the Gelatin to form amino acid, which are soluble.
So Bromelain hydrolysis down protein in liquid solution to form amino acid.
Preliminary investigation
Gelatin Experiment
Materials/Apparatus:
Gelatine (jelly)
Different concentrations of enzyme (pepsin) e.g. 1%, 3%
Petri dishes
Cork borers
Syringes
Method:
- Placed the gelatine in each Petri dish. There are three Petri dish altogether.
- Used the same diameter of cork borers to make a hole in the gelatine of same measurements throughout on all three.
- Measured the diameter of the hole before adding the enzymes.
- Made the pineapple into small pieces and placed it in the mixer machine to mix the pineapple into liquid juice then extracted the juice by adding the juice to the cloth and then squeezing the liquid into a clarified juice.
-
Used a syringe to put 5cm3 of the enzyme concentration of 1% and added 5cm3 of HCL in the gelatine hole in one Petri dish. Then placed 5cm3 of the 3% enzyme concentration and added 5cm3 of HCL in the gelatine hole in the second Petri dish and 5cm3 of the pineapple concentration and added 5cm3 of HCL in the third Petri dish containing the last gelatine hole.
- Cover the Petri dishes and leave the enzyme in contact with the gelatine for 45mins.
- At the end of this period, decant the enzyme, then measure the diameter of the holes again.
Table of results:
Experiment with photographic film
Materials/Apparatus:
Photographic film
Scissors
Boiling tubes and corks
Enzyme of appropriate concentration e.g.1% and 3%
Syringes
Method:
-
Used a scissor to cut the unused photographic film into strips of 3cm2.
- Weighed each strip, and then placed 1 strip in each clearly on labelled boiling tubes.
-
Added 20cm3 of the 1% pepsin in a boiling tube and then added 20cm3 of the 3% pepsin in a boiling tube and then finally added 20cm3 of the pineapple concentration in last boiling tube plus 5cm3 of HCL in every boiling tubes.
- Placed the cork on the mouth of the conical flask and started the stop watch.
- Then left the enzyme in contact with the photographic film and waited until the photographic film was clarified, checked the water bath every minute to prevent from getting any anomalous results.
- Then recorded the time it was clarified, after all the film had been clarified, got rid of the solution in the boiling tubes.
- Placed the film in a small bowl and placed it inside the oven to dry for 5 minutes.
- Then weighed them and recorded them in a table.
Table of results:
Carried out the preliminary work so that we could get knowledge of seeing if the method works and also determined what needs to be kept constant and what need to be changed during the experiment. If the experiment didn’t work then is there anything that needs to improve in actual method and also to see the effect of proteases on gelatin and photographic film.
The experiment with the jelly didn’t work because the jelly kept wobbling when making holes so the measurement of the hole is not constant and when adding the enzymes and HCL some spilled out and some was not inside the hole but ran outside the hole because the amount of the enzyme was excess. The size of the jelly was not right because when taking the jelly out from the container some was spoiled by breaking. It was hard to measure the hole because it kept wobbling. So I won’t be using this method in my final. The Petri dish used was so small that the solution came running out.
The photographic film experiment went well but for the time it took to clarify varies so each and every minute the solution need to be checked so the hot water bath had to be opened as a result the hot vapour force into the face so in the actual method the time it takes for film to clarify is known so the hot water bath could be opened after getting close to the actual time it took to clarify. The temperature can be increased a bit because the time it took for all the films to get clarify is more than 22.50minutes but if increased too much the enzymes could get denatured.
Secondary sources/references
- Did the preliminary work in class so that knowledge and experience of the method would be known.
-
Got some information about gelatin in . Some of the details that I got in my prediction are from this website and also for the risk assessment for gelatin.
- Got hydrochloric fact sheet from Google where I used to get information about the risk assessment.
Actual Method:
I would be using the same type of method that had been carried out in the preliminary experiment because the photographic film in the enzyme solution had worked well but I just want to improve it but increasing the temperature from 50°C to 55°C s that the rate of reaction will increases
Materials/ Apparatus/Equipment:
- Used photographic film
- Scissors
- 3 boiling tubes and cork
- Buffer solution (pH 6.5)
- Water bath 55°C
- Stop watch
- Syringes
- Extracted juice from stem
- Extracted juice from leaves
- Extracted juice from the flesh
- Ruler
The stem, leaves and the flesh of the pineapple is extracted by cutting each of the part into small pieces than placing it into the blender machine to make a solid part into liquid. Then a muslin cloth is used to squeeze the liquid to juice where the juice has been clarified. Then that juice would be used.
Method:
- Wear lab coat, safety goggles and gloves.
- Get all the equipment out and handle all the equipment with care.
-
Use a scissor to cut 3 strips of developed photographic film into strips of 3cm2.
- Weigh each strip, and then place 1 strip in each clearly on labelled boiling tubes.
-
Add 20cm3 of the juice extracted from the stem and 5cm3 of buffer solution in a boiling tube and then add 20cm3 of the juice extracted form the leaves and 5cm3 of buffer solution into another boiling tube and then finally add 20cm3 of the juice extracted from the flesh and 5cm3 of buffer solution into the last boiling tube plus 5cm3 of buffer solution in every boiling tubes.
- Place the cork on the mouth of the boiling tube.
- Then leave the enzyme in contact and wait 30minutes.
- After 30 minutes get rid of the solution in the boiling tubes and take the film out.
- Place the film into the crucial and place it inside the oven to dry for 5 minutes.
- Then weigh them and record them in a table below.
- Then calculate the change in mass and percentage change in mass to plot a graph.
- Percentage change in mass is calculated by using the formula below.
Change in mass/mass at the start *100%
Table of results:
Then present the results in histogram which is easier than the graph:
Which ever parts of the pineapple had the highest peak then it will have the highest efficient, as a result it will tells me that that certain part will have the highest concentration of Bromelain.
Variables:
Evaluation
Even though the experiment seems to be easier but there are some negative things about this experiment. They are:
- The leaves of the pineapple long pointed leaves with needle tipped and up curved spines on the margins, which will prick the fingers if not carefully picked. So there is a difficulty when cutting it to put inside the blender to extract the juice.
- When separating the stem and flesh because they both are attached to each other so when trying to extract the juice some part of flesh or stem can be in different places.
- Maybe increase the temperature more at 5°C because the Bromelain denatures at 70°C when kept more than 5 minutes.
I don’t think anything else that needs to be changed or improved and I think that my method would produce fair results.