L. Edirisinghe – Year 9
The Effect of Heat on Enzyme Activity
The Effect of Heat on Enzyme Activity
Scientific Investigation – Report
BHP Billiton Science and Engineering Awards
Lakmali Edirisinghe
Gosford High School, Year 9
1
L. Edirisinghe – Year 9
The Effect of Heat on Enzyme Activity
Table of Contents
Abstract
3
Aim
3
Hypothesis
3
Introduction/ Background Research
3
Materials
5
Method
6

Gelatine Preparation
6

Papaya Puree Preparation
6

Conducting the Experiment
7
Safety Precautions/ Risk Assessment
8
Variables
8
Fair Test Justification
8
Results
9

Experiment 1
9

Experiment 2 (Repeat 1)
10

Experiment 3 (Repeat 2)
11

Photographs
12

Qualitative Observations
12
Discussion
13

Analysis and Comparison with Previous Studies
13

Explanation
13

Explanation for the Unexpected Qualitative Observations
16

Limitations and Improvements
16
Conclusion
17
Acknowledgements
18
Bibliography
18
2
L. Edirisinghe – Year 9
The Effect of Heat on Enzyme Activity
Abstract
For this experiment, I investigated the effect of temperature on papain’s (an enzyme
found in papaya) ability to break down proteins in gelatine, in an effort to identify its
optimal temperature. I set up 9x100mL measuring cylinders with an equal amount of
set gelatine, and poured a controlled amount of papain of varying temperatures into
each cylinder; namely 20oC, 30oC, 40oC, 50oC, 60oC, 70oC, 80oC, 90oC and 100oC. I
recorded the bottom level of a marble, used as a marker inside the cylinders, at
regular intervals of 5 minutes over a one-hour period and tabulated the results into
my logbook. After 3 trials, I learnt that papain is most active at temperatures between
80oC and 90oC, averaging 11.67 mL and 12.33 mL broken down in an hour
respectively – equating to 18% of original value. This disproves my hypothesis, which
expected the optimal performance temperature to be 65oC; backed by previous
studies. However, I have identified multiple limitations with my experiment, which
may have influenced these results.
My research did conclude that the surrounding temperature of the papaya before
preparation had a significant impact on its enzyme’s performance. The most effective
fruit was subjected to only slight temperature differences. This finding may improve
the lives of many in developing nations, who rely on papain heavily to aid digestion
and heal skin conditions including inflammation and wounds.
Aim
To identify papain’s optimal performance temperature by investigating the effect of
temperature on its ability to break down proteins in gelatine.
Hypothesis
I predict this experiment will prove that papain will reach an optimal performance
temperature at 65oC, and reduce its speed of breaking down proteins from any
temperature higher than that, forming an approximate bell-curve shaped graph.
Introduction/ Background Research
Before I conducted my experiment, I did
some research to familiarise myself with new
terms and concepts. A full explanation of my
3
Figure 1 – The Induced-fit Model of Enzymes
L. Edirisinghe – Year 9
The Effect of Heat on Enzyme Activity
experiment can be found in the discussion section of this report.
An enzyme is a (biological) protein that acts as a catalyst.
A catalyst changes (usually increases) the
rate of a chemical reaction by altering the
pathway of reaction. This new pathway is
usually more energy-efficient, allowing more
reactions to be made faster.
One of the most prominent theories of how
enzymes work is the ‘Induced fit model’,
shown in Figure 1. It is very similar to a wellFigure 2 - Energy efficiency of catalysts
used ‘lock and key’ analogy, but the enzyme
(lock) has to change its shape slightly to
accommodate the substrate (key). A substrate is a substance an enzyme reacts with.
The enzyme collides with one or more appropriate substrate (reactant) molecules
and binds to them at the active site, to form an enzyme-substrate complex. The
enzyme then goes through a process called catalysis, which ‘transforms’ the
substrates. This usually means breaking them down to form smaller ones. Once the
transformation is complete, the enzyme ejects the new substrate(s) (product/s), and
returns to its normal shape, ready collide into the next reactant. It is important to
note that enzymes do not get ‘used up’, and can therefore be reused to transform
other substrates. A typical enzyme molecule can transform 1000 individual substrate
molecules per second, but this depends on the temperature, and many other factors.
Figure 3 - Enzyme process, using sucrose and sucrase as examples.
4
L. Edirisinghe – Year 9
The Effect of Heat on Enzyme Activity
Papain (the main enzyme in papaya fruit) is a protease, which specifically breaks
down proteins into amino acids, consequently breaking peptide bonds (between the
amino and carboxyl groups of two different amino acids). Likewise, other enzymes
have different jobs, and cannot do anything else. Enzymes are prevalent in most
fruits, but of different intensity. This is why apples and strawberries are able to set in
gelatine (a protein), but stronger enzymes such as those in raw pineapple and
papaya are not (tinned fruit has their enzymes specifically denatured for preservation
purposes, so tinned pineapple and other such fruit will set). Enzymes are affected by
four major variables; temperature, pH, activators and inhibitors (activators increase
the speed of an enzyme while inhibitors slow it down). Each enzyme has a specific
value of temperature and pH level where it can reach optimal performance.
Previous studies have shown papain to have an optimal temperature of 65 oC, and an
optimal pH range of 6.0 – 7.0. On either side of the optimal temperature, the enzyme
activity decreases, in a pattern shown in Figure 4 and 5.
Figure 4 - Optimum PH level of papain from previous studies
Figure 5 - Optimum temperature of papain from previous studies
Materials
For this experiment (without repeats), I needed;

25g (4 leaves) Pure Gelatine (1 sachet of Dr Oetker’s gelatine powder)

Water (585mL at 60oC – a kettle is helpful in achieving this temperature)

Paper, pen and tape for labelling

Papaya Puree
o Water (230mL)
o 1 Papaya fruit (1kg)
o Blender
o Knife (+ vegetable peeler, spoon and cling-wrap)
5
L. Edirisinghe – Year 9

9x 100mL Measuring Cylinders

A saucepan (a small, heatproof pot)

Refrigerator (at around 2oC)

Stove (with range hood)

Thermometer (suitable for cooking)

9x Marbles
The Effect of Heat on Enzyme Activity
Method
*There are 3 stages to this experimental procedure – gelatine preparation, papaya
puree preparation and the actual experiment – which uses the previously prepared
gelatine and papaya. * A hand drawn scientific diagram can be found in my logbook*
Gelatine Preparation
1. First, I labelled 9 cylinders each as: 20oC, 30oC,
40oC, 50oC, 60oC, 70oC, 80oC, 90oC, 100oC
respectively.
2. Then, I poured 585mL of hot water (60oC) into
a saucepan.
3. I opened one sachet (25g) of powdered
gelatine and sprinkled it onto the water,
stirring it through.
Stirring Gelatine in a Saucepan
4. When the gelatine had been fully dissolved (the water was transparent,
and granule-free), I poured 65mL of the liquid into each of the 9
measuring cylinders – ensuring the bottom of the concave meniscus was in
line with the marking.
5. I then refrigerated all the measuring cylinders overnight.
For the Papaya Puree
1. First, I cut the papaya into segments
lengthwise, and spooned out the seeds. I
cling-wrapped 3/4 of the fruit, and put it
in the fridge for later.
2. Then, I peeled the skin off the remaining
1/4 of fruit, and diced the flesh into small
chucks of about 2.5cm3.
6
Preparing Papaya Fruit
L. Edirisinghe – Year 9
The Effect of Heat on Enzyme Activity
3. I put the chunks and 230mL of water (at room temperature) into a blender
and pureed the fruit, until it reached a smooth, even consistency.
For the Experiment
1. I first poured 25mL of papaya puree into a small, heatproof pot, and
ensured it was 20oC. I then retrieved the measuring cylinder marked ‘20oC’
from the refrigerator.
2. After that, I dropped a marble onto the set gelatine inside the cylinder, and
poured the 25mL of puree into the cylinder. It served as my ‘control’ in the
experiment, as the room temperature was 20oC at the time.
3. I quickly recorded the level where the bottom of the marble was on the
measuring cylinder, using its markings. I also
recorded the time, both values in my logbook.
4. Every 5 minutes, I recorded the time, and level
of the marble. This was tabulated in my
logbook.
5. While this was happening, I took another 25mL
of papaya puree, and poured it into the pot.
However, this time I heated it to 30oC, using a
stove (on low heat) and a thermometer to check.
6. I took the 30oC labelled measuring cylinder from
the fridge, put a marble inside it, and poured the
Heating Papaya Puree
heated puree on top. I recorded the time, and the level of the marble, like
the control. These two variables were also recorded at 5 minute intervals.
7. Steps 5-6 were repeated for all 7 other cylinders. I stopped recording each
cylinder 1 hour after they first came into contact with the papaya puree.

I managed to synchronise all the tests, so at a specific time (e.g. 7:05
pm) I could record the levels of all the measuring cylinders at once
<see my logbook for case in point>.
8. I then repeated steps 1-7 twice, to ensure consistency cleaning all
equipment thoroughly between repeats. My first repeat used leftover
papaya from the previous experiment; my 2nd required a new papaya (an
explanation for this can be found in the ‘limitations and improvements’
section of this report).
7
L. Edirisinghe – Year 9
The Effect of Heat on Enzyme Activity
Safety Precautions/ Risk Assessment
There were a number of safety hazards that I had to address in this experiment.

I poured hot gelatine into thin measuring cylinders, exercising high levels of
care; including standing the cylinder on a level benchtop, and pouring as
slowly as possible, with a steady hand.

I prepared papaya puree by cutting, skinning and dicing a papaya fruit. This
was done with clean, dry hands on a stable cutting surface, keeping fingers
away from the knife and peeler blades. Cleaning the blender blade after the
experiment was done with similar caution.

I heated papaya puree on a stove. Before I did, I ensured the stovetop and
equipment was clean and dry, and the range hood was turned on. I kept the
stove on a low heat, and waited until the puree reached the required
temperature, stirring as it was heating to prevent the bottom from burning.
Variables
Independent Variable
Temperature of papaya
puree (papain
enzymes)
Dependent Variable
Amount of gelatine
broken down (or
inversely, level of
remaining set
gelatine, in mL)
Controlled Variables
 Amount and concentration of
substrate and enzyme
 Room and fridge temperatures
 Equipment used in both preparation
and experimental procedures.
 Type of enzyme (papain) and
 Type of substrate (gelatine)
 Time intervals for recording (5 mins)
For a Fair Test:
I had to experiment with each cylinder in the same period of time. I could not finish
recording one cylinder, then start the next one, as each gelatine sample would be in
the fridge for drastically different times, which may alter the results of the
experiment. I had to multitask with 9 different samples, ensuring that all measuring
cylinders were out of the fridge in about an hour since the commencement of the
experiment.
Whenever I was pouring something, I ensured the bottom of the concave meniscus
lined up exactly with the appropriate marking. I also controlled all variables that were
not independent or dependent (see sections ‘Variables’ and ‘Method’). I used pure
gelatine instead of jelly because any flavouring or additives may affect the results.
8
L. Edirisinghe – Year 9
The Effect of Heat on Enzyme Activity
Results
For Experiment 1:
Temp of
o
Level of marble in
measuringcylinder (mL)
Enzyme ( C)
9
20
30
40
50
60
70
80
90
100
0
65
65
65
65
65
65
65
65
65
5
64
64
64
63
63
59
58
58
58
10
64
63
62
61
60
58
57
55
57
15
63
63
62
59
59
57
55
54
56
Time after enzyme added (mins)
20
25
30
35
40
63
62
62
62
61
62
62
61
61
61
62
61
60
60
60
59
58
58
57
57
58
57
57
56
55
56
55
55
55
54
54
54
53
53
52
54
53
53
52
52
56
55
55
55
55
45
61
61
60
56
55
53
52
51
55
50
61
61
60
55
54
52
52
51
55
55
61
61
60
55
54
52
51
51
54
60
61
61
60
55
54
52
51
51
54
L. Edirisinghe – Year 9
The Effect of Heat on Enzyme Activity
For Experiment 2 (repeat 1):
Temp of
Enzyme
o
( C)
Level of marble in
measuringcylinder (mL)
20
30
40
50
60
70
80
90
100
0
65
65
65
65
65
65
65
65
65
5
64
63
63
63
64
63
62
61
63
10
64
63
63
63
63
62
62
59
63
15
63
62
62
62
63
62
61
59
62
Time after enzyme added (mins)
20
25
30
35
40
63
63
63
63
63
62
62
62
62
62
62
62
62
62
62
62
62
62
62
62
63
63
63
62
62
62
61
61
61
61
61
60
60
60
60
58
57
57
57
57
62
62
62
62
62
45
63
62
62
62
62
61
60
57
62
50
63
62
62
62
62
61
60
57
62
Timeline - Lines for temperatures 30, 40, 50 are behind lines 100 and 60 – see table
10
55
63
62
62
62
62
60
60
57
61
60
63
62
62
62
62
60
59
57
61
L. Edirisinghe – Year 9
The Effect of Heat on Enzyme Activity
For Experiment 3 (repeat 2):
Temp of
Enzyme
o
Level of marble in
measuringcylinder (mL)
( C)
11
20
30
40
50
60
70
80
90
100
0
65
65
65
65
65
65
65
65
65
5
64
63
63
63
62
62
61
62
62
10
64
63
63
63
61
61
60
61
62
15
63
63
62
62
61
60
59
60
61
Time after enzyme added (mins)
20
25
30
35
40
63
63
62
62
62
62
62
61
61
60
62
61
60
60
59
61
60
59
59
57
60
60
59
58
57
59
58
57
56
54
57
56
55
54
53
60
58
57
55
54
60
59
59
59
58
45
62
59
58
56
55
53
53
53
58
50
61
59
58
56
53
53
51
52
56
55
61
59
56
54
53
52
50
51
55
60
61
58
55
53
52
51
50
50
54
L. Edirisinghe – Year 9
The Effect of Heat on Enzyme Activity
Photographs
Experiment 3’s “80-degree measuring cylinder” at 3:20 PM, 3:40 PM and 4:00 PM respectively.
Experiment 1; all 9 cylinders lined up in order. The cylinder on the right (100 degrees) has its water evaporated,
and is therefore a darker shade of orange. The experiment is still in progress.
Qualitative Observations
Some qualitative observations (temperatures are averages only) were:

Papaya puree was always 15mLs high, and did not mix with the liquid gelatine.

The marble makes a small dip in the gelatine, and the nature of the liquefied
gelatine itself distorts the shape of the marble to assume a ‘magnified’
appearance (which was taken into account when recording).

Bubbles form at 50oC, and proper evaporation begins at 70oC, when clumping
of solids starts to form and the build-up of some bright orange, powderylooking residue. Puree is significantly darker at 100oC, which I assume is due
to slight burning as a result of significant evaporation.
12
L. Edirisinghe – Year 9
The Effect of Heat on Enzyme Activity
* I investigated the reasons of some unexpected observations in my discussion.
Discussion
Analysis + Comparison with Previous Studies
All three trials have produced very similar trends, all proving that temperature has a
huge effect on the functionality of enzymes. In my experiment, Papain enzymes
broke gelatine down the least at room temperature (20oC), and progressively
liquefied more gelatine until the optimal temperature of either 80 or 90 degrees is
reached (average of 86.67oC). This equates to 11.67 mL and 12.33 mL broken down in
an hour respectively – liquefying 18% of original value to the nearest percent. After
that, papain did not break gelatine as quickly. One important difference between the
tests was the actual repeats, as although they proved the same thing, they had a
huge disparity between the lowest levels reached. Repeat 1 found the lowest level
(the highest amount) of liquefied gelatine to be 57mL, while repeat 2 saw it as 50mL.
The experiment’s final result consequently contradicts both my hypothesis and
previous studies, which say the optimal temperature is 65oC (some sources provide a
range between 60oC and 80oC). Despite this, the general shape of the final graph
agrees with a portion of my hypothesis, and previous studies (see ‘Introduction’). I
am rather surprised at this result; but I have outlined some external factors which
may be falsely increasing the ‘optimal temperature’, in a section headed ‘Limitations
and improvements’.
Explanation
As
explained
in
the
introduction,
enzymes are proteins. A protein is a
‘string’ of amino acids, which vary from
protein to protein. Amino acids consist
of an amino group and carboxyl group;
formed on a nitrogen and carbon
backbone. Specific chemical groups
(side chains – noted ‘R’ in Fig 6), that
Figure 6 - The structure of a protein. NOTE: this protein is DENATURED
identify the enzyme and allow it to
perform specific functions. There are
20 main types of Amino acids, but plenty more exist. Strands of amino acids form a
helix, which bunches and ‘tangles’ itself up.
13
L. Edirisinghe – Year 9
The Effect of Heat on Enzyme Activity
Figure 7 - Taking apart a protein: From Left to Right: 1) The protein is made up of a string of amino acids. 2) These strands
make an alpha-helix. 3) These helixes fold, and assemble with other folded helixes (4) to form a complex protein structure.
The proteins are tangled due to a process
called
‘folding’.
Folding
is
important
in
proteins, as it ‘activates’ them; usually making
new, non-covalent bonds to different parts of
the same strand which increases the protein’s
functionality. A covalent bond occurs when
two molecules or compounds share their
electrons in order to achieve a stable atomic
charge of 8. Consequently, non-covalent
Figure 8 - A simple diagram showing the non-covalent
bonds of an active (folded) protein
bonds do not involve the sharing of electrons. The protein takes anywhere between 1
nanosecond and 1 millisecond to fold, in a way still unknown to scientists.
Figure 9 – Papain in its raw, enzyme form. The pink part is hydrophobic – it fails to mix or dissolve in water.
14
L. Edirisinghe – Year 9
The Effect of Heat on Enzyme Activity
A single papain enzyme looks like this:
Figure 10 - The structure of a papain enzyme. Each circle is an amino acid, and the -s-s- figures represent non-covalent
bonds. The numbers are markers; there are 212 amino acids in 1 enzyme alone. -SH is the main active site of the enzyme.
Papain is fairly small in size, compared to other enzymes.
Before reaching optimum temperature, increasing heat makes enzymes and
substrates move faster, and more likely to collide. However, increasing the enzyme’s
temperature beyond its optimum value breaks these non-covalent bonds, hence
‘untangling’ the enzyme. This process is called denaturing. Untangling the enzyme
makes it inactive, as its shape changes. The substrate can no longer ‘fit into’ the
enzyme. The absence of active catalysts means the process of breaking down
proteins is slowed.
15
Figure 11 - When the enzyme is denatured, the perfect substrate is unable to 'fit'
15
L. Edirisinghe – Year 9
The Effect of Heat on Enzyme Activity
Gelatine, the substrate of the experiment, is a protein, a product of collagen which is
sourced from the horns, hooves, bones and connective tissue of animals,
predominantly cattle. Gelatine ‘melts’ (i.e. loses its solidity) at 35oC, but this process
speeds up with more temperature. Gelatine unravels its folded proteins when in
contact with hot water.
Explanation for the Unexpected Qualitative Observations
I believe the reason why the papaya did not sink as the gelatine liquefied (contrary to
the rule of density) is that there was no room for any space exchanging to occur. The
papaya, taking the shape of its container, did not allow the liquefied gelatine to slip
up past it. The papaya is a very thick suspension, and the liquefied gelatine is a sol (a
colloid with solids [proteins] dispersed in a continuous liquid medium). This may
have caused some errors with the results, as the papaya never really touched solid
gelatine. The puree was always 15 cm high as it was an enzyme – it doesn’t get used
up in the reaction.
Set gelatine has a refraction index (R.I.) of about 1.4, while its liquefied form is slightly
less. As both values are larger than 1, the R.I. of air, and the measuring cylinder itself
was curved like a biconvex lens, the gelatine-filled measuring cylinder acted very
similar to a magnifying glass, distorting the size and shape of the marble.
Limitations and Improvements
One of the biggest shortcomings of my experimental procedure was that I used both
water and papaya to make a ‘puree’. The problem with that is I do not have proper
evidence to prove fully that enzymes were the cause of the gelatine to liquefy. As hot
water also liquefies gelatine, it is hard to tell what component of the puree allowed
some levels to drop more than others. Although water does not denature (with
regard to the 100oC measuring cylinder), it evaporates, which along with
aforementioned fact that the enzyme did not touch solid gelatine for a considerable
period of time, makes water and enzymes equally possible contributing factors to the
results of my experiment. Another thought is that the cold gelatine (from the fridge)
cooled down the temperature of the enzyme on contact.
Further research suggests that the skin, not the flesh, of the papaya fruit has the
highest concentration of enzymes. Raw papayas have more enzymes than when ripe.
However, the varying factor between experiments 2 and 3 in their final lowest mark
was definitely a product of enzyme environment prior to experimentation, as all other
variables were kept the same. For experiment 1, I bought a fresh papaya, left it in a
16
L. Edirisinghe – Year 9
The Effect of Heat on Enzyme Activity
hot car for an hour, then placed it in the fridge for 10 minutes before cutting it up.
For experiment 2, I used the remainder of papaya from experiment 1; which was cut,
cling-wrapped and stored in the fridge (at 2oC) for a week. After a poor result, I
decided to purchase a new, fresh papaya. I cut it 20 minutes later, with minimal
temperature fluctuations during that period, to conduct my 2nd repeat.
Another limitation was the amount of accuracy of recording. I was forced to
approximate all values off to the nearest mL, due to time constraints, meniscus levels,
lack of markings smaller than millilitres, the bottom of the marble making a dip in the
gelatine and the gelatine magnifying the marble, warping depth perception.
To make the experiment more successful if I were to do it again, I would use less
water and more fruit to make the puree, or find a purer or more concentrated form
of papain to use instead of a puree. I would also attempt to actively keep the papain
in contact with set gelatine, using a paddle pop stick or similar apparatus. For each
repeat, I would also treat each papaya to the same temperatures before preparation.
With regard to the recording accuracy, I could examine each cylinder more carefully
before recording, or acquire larger measuring cylinders, so a 1mL limit of accuracy
will not affect the results as much.
If I was to do this experiment again, I would also start the process with frozen puree,
and have 11 experiments, ranging from 0oC to 100oC (and not just from 20oC), to
give me a better and broader understanding of the effect of temperature on papain.
Conclusion
In conclusion, I found that temperature has an enormous effect on the productivity
of enzymes (suspended in water), but so did the environment of the papaya before
its preparation. My experiment found the optimal temperature for papain was
between 80oC and 90oC (86.67oC on average), becoming less active on either side of
the figure. This disproved my hypothesis, backed by scientific evidence, which was
65oC, but proved that either too big an increase or decrease in temperature will stunt
activity. If I had used pure papain instead of a papaya and water puree, and been
stricter on controlling the environments of the papaya prior to its preparation, the
results would have changed; becoming less ambiguous and more reliable.
Provided that papain has extensive therapeutic properties, my research concludes
that keeping the papaya at consistent room temperatures before preparation has the
best chance of speeding up the healing process of wounds as well as easing inflamed
areas with topical application, and increasing digestion upon consumption. The
17
L. Edirisinghe – Year 9
The Effect of Heat on Enzyme Activity
practical aspects of this experiment include increasing the effectiveness of naturally,
enzyme-driven remedies, which may benefit the lives of many in developing nations,
the citizens of which rely heavily on papain as natural medicine.
Acknowledgements
I would like to thank my father for purchasing/ providing the materials and apparatus
used in this experiment (except the measuring cylinders), outlined in the ‘Materials’
section of this report. I would also like to thank my science teacher (Mrs Fletcher) for
her approval and referral of my experiment to the STANSW Young Scientist Awards.
My acknowledgements extend to Gosford High School’s science faculty (Ms Day), for
lending me the 9x 100 mL measuring cylinders used in my experiment.
All idea generation, research, and experimental proceedings, including writing this
report and logbook, was done solely by me. All research sources are cited below:
Bibliography (24 sources)

HILL, Graham (1986) Chemistry Counts London: Hodder and Stoughton ISBN: 0-340-37631-7
<first accessed 07/05/16>

Australian Institute of Food Science and Technology (2009) [Australia] NSW Department of
Education and Training. URL:
http://www.curriculumsupport.education.nsw.gov.au/secondary/science/11_12/aifstresources/inde
x.htm - <first accessed 13/05/16>

C. Monrose-Forde Effect of Temperature on Enzyme Action URL:
http://brilliantbiologystudent.weebly.com/effect-of-temperature.html - <first accessed 22/05/16>

Marc Leger (2012) You Can’t Kill What’s Not Alive, and Other Thoughts About Boiling [United
States] Atoms and Numbers URL: http://www.atomsandnumbers.com/2012/you-cant-kill-whatsnot-alive-and-other-thoughts-about-boiling/ - <first accessed 22/05/16>

AJ Kerrigan (2014) Enzymes Lab [United States] Health Science Academy. URL:
http://academygenbioii.pbworks.com/w/page/70219744/Enzymes%20Lab%20 - <first accessed
22/05/16>

Sam Adam-Day (2015) Enzymes [United Kingdom] URL: http://alevelnotes.com/Enzymes/144 <first accessed 22/05/16>

Sigma-Aldrich (International LTD) (2016) Papain URL: http://www.sigmaaldrich.com/lifescience/metabolomics/enzyme-explorer/analytical-enzymes/papain.html - < accessed 22/05/16>

British Broadcasting Corporation (2010) Enzymes [United Kingdom] URL:
http://www.bbc.co.uk/schools/gcsebitesize/science/add_aqa_pre_2011/enzymes/enzymes1.shtml <first accessed 22/05/16>

McKinley, O’Loughlin (2016) McGraw-Hill Global Education Holdings, LLC Animation – How
Enzymes Work [United States] URL:
http://highered.mheducation.com/sites/0072495855/student_view0/chapter2/animation__how_en
zymes_work.html - <first accessed 22/05/16>
18
L. Edirisinghe – Year 9


The Effect of Heat on Enzyme Activity
Andrew Rader (2016) Enzymes Make the World Go 'Round Andrew Rader Studios [United States]
URL: http://www.chem4kids.com/files/bio_enzymes.html - <first accessed 22/05/16>
Lakshmi Santhosh (2016) The Effects of Temperature on Enzyme Activity and Biology Demand
Media [United States] URL: http://classroom.synonym.com/effects-temperature-enzyme-activitybiology-6049.html - <first accessed 29/05/16>

Michael Graw (2016) What Are the Effects of Boiling & Freezing on Enzyme Activity? Demand
Media [United States] URL: http://classroom.synonym.com/effects-boiling-freezing-enzymeactivity-23207.html - <first accessed 29/05/16>

Emily Updegraff (2016) Why Does Heating Interfere with the Activity of an Enzyme? Demand
Media [United States] URL: http://classroom.synonym.com/heating-interfere-activity-enzyme10251.html - <first accessed 30/05/16>

David Robson (2016) Protein 101: What it is and Why You Need Plenty of it For Muscle Growth
Allmax Nutrition [New Zealand] URL: http://www.allmaxnutrition.com/postarticles/supplements/protein-101-what-it-is-and-why-you-need-plenty-of-it-for-muscle-growth/
- <first accessed 30/05/16>

S. S Husain; G Lowe (1969) Completion of the Amino Acid Sequence of Papain University of
Oxford [United Kingdom] URL:
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1184853/pdf/biochemj00695-0126.pdf - <first
accessed 11/06/16>

Stuart Mason Dambrot (2014) Promising proteins: Scientists develop new drug discovery tool
using spectroscopy and simulation ScienceX Network [United States] URL:
http://medicalxpress.com/news/2014-07-proteins-scientists-drug-discovery-tool.html - <first
accessed 11/06/16>

Stanford University (2016) The Science (of Protein Folding) [United States] URL:
https://folding.stanford.edu/home/the-science/ - <first accessed 11/06/16>

Purvi Gosrani (2013) Heat Shock Proteins Linkedin Corp. (Slide Share) [United States] URL:
http://www.slideshare.net/poorvishah10/heat-shock-proteins - <first accessed 18/06/16>

Homaei AA; Sajedi RH; Sariri R; Seyfzadeh S; Stevanato R (2010) Cysteine enhances activity and
stability of immobilized papain. National Center for Biotechnology Information, U.S. National
Library of Medicine [United States] URL: www.ncbi.nlm.nih.gov/pubmed/19479190 - <first
accessed 11/06/16>

Dee Rawsthorne Jelly Vision – Jelly Facts Institute of Food Research [United Kingdom]
http://www.ifr.ac.uk/jellyvision/Jelly_facts.pdf - <first accessed 18/06/16>

Winsby, Roy Refractive Index – a Note for the beginner © Onview.net Ltd (Microscopy-UK)
[United Kingdom] URL: http://www.microscopyuk.org.uk/mag/indexmag.html?http://www.microscopy-uk.org.uk/mag/art97/winrind.html <first
accessed 18/06/16>

K. Eldredge, J. Koh, S. Kongkatong, K. Burch, M. Harel, E. Miller, S. Bray, D. Canner Papain
Weizmann Institute of Science [Israel] URL: http://www.proteopedia.org/wiki/index.php/Papain
<first accessed 18/06/16>

Dr. Edward Group (2011) The Health Benefits of Papain Global Healing Center [United States] URL:
http://www.globalhealingcenter.com/natural-health/papain/ <first accessed 18/06/16>

Edinformatics (author unknown) TYPES OF MIXTURES -- SOLUTIONS, SUSPENSIONS, COLLOIDS
(1999) Edinformatics [United States] URL:
http://www.edinformatics.com/math_science/mixtures.htm <first accessed 18/06/16>
19