There are enzymes responsible for the ripening of fruit which break down the starch content to produce more sweet sugars and make the fruit softer, making it more edible. Therefore, altering the conditions which effect enzyme rate of reaction, will effect how quickly a pineapple will ripen. Other enzyme activity increases in the fruit during ripening, due to certain hormones (such as ethylene). Applying this rule to pineapple: the bromelain enzyme activity will increase as the pineapple ripens. If I were to put a pineapple in cold conditions, this would slow down the ripening process because the enzymes responsible would have less kinetic energy, and I am therefore indirectly reducing the activity of bromelain.
In this investigation scenario, when under-ripe pineapple was used in jelly, it set better than when ripe pineapple was used. Taking in to consideration the information I have found out above, I propose that this could have occurred due to a protease enzyme that breaks down the protein in the jelly, which is more active in the ripe pineapple than the under ripe pineapple. I will now plan a full investigation to prove my proposal by testing pineapples at different stages of ripeness.
Method
Apparatus
Jelly (gelatine)
Ruler
4 equal sized petri dishes
Borer
3 under ripe pineapples
Blender
Timer
Fridge and Fridge freezer
Pipette
Preparations
I will place one pineapple (A) in a freezer for two weeks to stop the ripening process. Another pineapple (B) will be placed in a freezer for one week and kept at room temperature for the second week. The third pineapple (C) will be kept at room temperature for two weeks. I will make sure that any pineapples kept at room temperature will not be placed near a window or radiator where the temperature may fluctuate. The pineapples in the freezer will be kept on the same shelf as each other. I will take the pineapples out of the freezer and place the in the fridge 24 hours before the experiment, to make sure they are all the same temperature at the start.
Method
- I will prepare the jelly, according to instructions on the packet. I will then pour it in to four petri dishes and put it in the fridge to set. Before pouring the jelly in to the dish, I will measure ¾ of the way up of the dish and make a mark. This mark is where I will pour the jelly up to to make sure that there is the same amount of jelly, which reaches the same height in each dish.
- To prepare the pineapple, I will remove the top and bottom, stand vertically and remove the skin, cut in to quarters and remove the core. I will not use the core because this is not usually eaten so does not apply to the scenario where the pineapple was being used in food. I will be as consistent as possible with each pineapple to make sure that I am using the same type of tissue. I will then place the quarters in the blender for ten seconds and place in a labelled beaker (labelled A, B or C). Blending the pineapple will break walls of the pineapple tissue, meaning that the enzymes will be more exposed and take effect more quickly than if the pineapple had not been blended.
- (To take place as soon as possible after step 2) I will take the petri dishes out of the fridge and with a borer make three holes, as far away from each other as possible in the jelly of each of the dishes. I will label the dishes A, B, and C, and measure the diameter of each of the holes made by the borer with a ruler. I will number each of the holes by writing the number on the lid and placing it underneath the dish with numbers in the same place as the corresponding hole.
- I will use a pipette to place the pineapple pulp in to the holes. Each type of pineapple will be in its own, labelled petri dish. One of the petri dishes will not have any pineapple put in it, and will be used as a control.
- I will then put the petri dishes back in the fridge and leave them there for seven days. This is enough time for the enzyme, of even the ripest pineapple to take effect. Any longer than this, and the liquefied gel from each hole may join up, making it difficult to take measurements. Putting the dishes back in the fridge will prevent any bacteria or foreign bodies attaching to the jelly which could effect the results. I will start the timer the minute that I have placed the pineapple into the holes, and have a different timer for each petri dish so I know they have had exactly the same amount of time in the fridge.
- After every 24 hours, I will remove them from the fridge again. From research I have done, I have found that enzymes in the pineapple will turn the gelatine from a gel to a liquid. I will measure the diameter of the area that is liquid and record my results in a table as below.
Taking away the diameter of the hole at the start, from the diameter of the liquid and the hole, gives the diameter of the gel which has turned to liquid. I will use one of these charts every 24hrs for seven days, leaving me with seven mean results for each dish. When these diameters are plotted in a time/diameter graph they will allow me to see the rates at which the diameters have changed. Adding them will give me the mean total diameter of gel that has turned to liquid.
As you can see from the table, I have used three holes for each type of pineapple so that I can work out an average. This reduces the effect of error and anomalous results.
Results
Table to show results and analysed data
At the start, the holes are different sizes. If the data were analysed with this being the case, then it would be impossible to see trends in the data. Therefore I have taken away the size of the hole at the start from the size of the hole after two days to give me the change in hole size. I then found the mean of the change in size for each day to represent the data and so that comparisons and trends can be found.
In this data the independent variable is the number of days after purchase, and the dependent variable is the size of the hole after two days.
A graph to show how the mean change in hole size changes with increasing age of the pineapple
It is possible to see from the table and graph that the mean change in size of the hole increased as the number of days after purchase increased. I have chosen to use a scatter graph to makes trends more visible and show the effect of altering the dependent variable has on the independent variable.
Conclusion
From this experiment it can be concluded that the longer the time after purchase of the pineapple the more the size of the hole increased. This is shown on the graph by the lines connecting each of the plotted points having positive gradients. The plotted points nearly form a straight line showing a steady increase in hole size, and suggesting that if the pineapple had been tested after a period longer than 3 days after purchase that the hole size would continue to increase.
The size of the hole increases because the protein molecules which make up the jelly are hydrolysed by a protease enzyme (Module 2) called bromelain which is found in pineapple. When a molecule of protein is hydrolysed, amino acids and water are formed (Module1) and this will occur wherever the enzyme meets the protein in the jelly. Therefore the enzyme takes effect from the initial hole, spreading outwards which creates the effect of the hole size increasing.
The independent variable is the number of days after purchase of the pineapple; this gives an indication of the ripeness of the pineapple, with the pineapple getting riper with each increasing day after purchase. The enzymes had a greater effect on the jelly the riper that the pineapple was, which shows the enzyme bromelain increases in activity as the pineapple ages.
Other enzymes that increase in activity as the pineapple ages are pectinase and cellulase. Cellulase breaks down cellulose and pectinase breaks down pectins. Cellulose and pectins are both polysaccharides. Cellulose strengthens cell walls, and pectins help to hold cells together. When these polysaccharides are broken down the pineapple becomes softer and more digestible (Module 1). It is hormones such as ethylene which are responsible for the increase in activity of these enzymes. I propose that it is the same hormones that cause the increase in activity of bromelain. Bromelain may play a part in the ripening process by helping to soften the pineapple tissues. It would do this by breaking down structural, fibrous proteins (as bromelain is a protease). This explains the increase in activity of bromelain, as the pineapple gets riper.
The ripening process makes the pineapple more appealing for animals to eat, and if not eaten, will cause the pineapple to decay to expose the seeds. These are methods of seeds dispersal which are necessary for aiding survival of the pineapple plant.
Evaluation
There are no points in the graph which appear to be out of place, suggesting that there are no anomalous results in the experiment. However, there are only 4 points on the graph, and the experiment was only carried out once, so it is possible that anomalous results will go unnoticed. For example, if the experiment included a pineapple that was tested 4 days after purchase then we may find that the mean change in holes size was the same as or less than the result of the pineapple that had been tested three days after purchase. This would suggest that something else was occurring in the pineapple that the original experiment had not shown or that the result for the pineapple tested 3 days after purchase was anomalous. With that in mind there must also be a limit as to how many days after purchase the pineapple may be used, it may be too decayed after a certain time.
One of the main sources of error in this experiment is that the holes started off at different sizes. This means they would all contain different volumes of the pineapple juice, and therefore the holes for the same day would contain a different amount of enzymes. If a hole contains more enzymes than another, then its capacity for a size increase is larger.
Having less enzymes would also result in a decreased rate of reaction but we must also take in to account of the fact the size of the hole will effect its surface area to volume ratio. Initially, the enzyme will react with the jelly only on the surface of the hole, where it meets the jelly. The smaller a hole is, the larger its surface area to volume ratio, and vice versa. The smaller holes will have a larger proportion of their enzymes exposed to the jelly than the larger holes.
With both of theses factors to consider, the results would have been fairer if the holes were the same size.
A pestle and mortar were used to break up the pineapple flesh so that the enzymes available in the juice. However, it is possible that this process was different for each pineapple. If it had been carried out more thoroughly then more enzymes would be available which would cause a greater increase in hole size.
If the petridishes were even slightly knocked when measuring the size of the hole then this mixed up the jelly with the liquid surrounding the hole and the contents of the hole. Mixing up these components would aid the reaction and make the measurements incorrect because the liquid will move further in the jelly as a results of being moved. It is possible that some of the measurements are incorrect because of this, which could cause anomalous results.
Measuring the number of days after purchase may not be an accurate measure of the age of the pineapple and subsequently how old and ripe the pineapple is. This is because it is not known how long each of the pineapples had been on the shelf, so its possible that the pineapple tested 3 days after purchase is less ripe than the pineapple tested two days after purchase. The pineapples may have also been treated with a spray to prevent them ripening too quickly, and this may have reached some of the pineapples more than others.
It may not be correct to make the conclusion that I have about the bromelain being associated with decay of the pineapples so that the seeds are exposed, because in this experiment the pineapples are commercially grown and seedless. For further work, I would like to use a seeded, naturally grown pineapple and compare the results.