report2 daphnia
TRANSCRIPT
INTERNATIONAL EDUCATION CENTRE (INTEC)UiTM Section 17 Campus 40200 Shah Alam Selangor Darul Ehsan.
TEL: 603-55227000
TITLE: THE EFFECT OF CAFFEINE ON HEART RATE
NAME: NUR AMALINA BT ZOLKEFLEE
SID NUMBER: 2010843164
CLASS: 11M2
DATE OF EXPERIMENT: 16TH AUGUST 2010
LECTURER’S NAME: DR.NORAINI BINTI FADZILLAH
PARTNER’S NAME: FATIN NABILAH BT MOHD NASIR
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INTRODUCTION
BACKGROUND INFORMATION
Human have consumed caffeine since Stone Age. Early people found that eating seed,
bark, and leaves of certain plants had increase their awareness, decrease their fatigue and
stimulate their mood. So what is actually caffeine ? Caffeine is a white crystalline
xanthine alkaloid, a phsychoactive stimulant drug. It is discovered by a German chemist,
Friedrich Ferdinand Runge in 1819. Caffeine can be found in beans, leaves and fruit of
some plants where it acts as natural pesticide that paralyse and kill certain insects feeding
on the plants. Caffeine is also consumed by human in infusions extracted from bean of
coffee plants and leaves of tea bush. In humans, caffeine is said to give a big impact on
human central nervous system(CNS). It is said can ward off drowsiness, restore alertness,
and has diuretic( a substance that cause increase in flow of urine) properties. But studies
did not show that caffeine can cause dehydration because most regular caffeine
consumers had developed a very strong tolerance in it. Basically, the normal function of
caffeine is it act as natural pesticide and inhibit seed germination of nearby coffee
seedlings, giving it a better chance of survival. The common source of caffeine is tea,
coffee and chocolate derived from cocoa bean. Caffeine content in coffee varies
depending on type of coffee bean and method used to derive the caffeine. But in general,
in 30ml of espresso, there are 40mg of caffeine, in 120ml of drip coffee, there are 100mg
of caffeine. Factually, dark-roast coffee has less caffeine than the lighter roast because
roasting process reduce the bean’s caffeine content.Is it true tea has more caffeine than
coffee? Well, although tea contains more caffeine than coffee (by dry weight), a typical
serving contains is less, as tea is normally brewed much weaker. Besides strength of the
brew, growing conditions, processing techniques and other variables also affect caffeine
content. Certain types of tea may contain somewhat more caffeine than other teas. Tea
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contains small amounts of theobromine and slightly higher levels of theophylline than
coffee. Recently, many manufacturers begun putting caffeine in their products especially
the shower products like shampoo and soap. Studies show that caffeine in shampoo
applied for two minutes is absorbed through the skin and reaches blood stream. They
claimed it can improve mental alertness. Caffeine is being used pharmacologically in
order to treat apnea in premature newborns. In 1819, Ferdinand isolated relatively pure
caffeine for the first time..
Diagram 1.1 shows the caffeine.
The nitrogen atoms which are planar( sp2 orbital hybridization) in caffeine resulting
caffeine’s molecule having aromatic character. Moving to effect of caffeine on human’s
body. As a CNS and metabolic stimulant, it helps to reduce fatigue and restore mental
alertness. It also helps in correcting irregular heart beats, make it faster and clearer flow
of thought, increase our focus, improve body coordination and act as competitive
inhibitor to adenosine, a neurotransmitter. Caffeine from coffee is absorbed by stomach
and small intestine within 45 minutes and then distributed to all body tissues. If we take
caffeine in moderation, it take less than hour for caffeine to begin affecting the body and
it will be ergogenic (increase a person’s ability for mental and physical labour) to us. A
study carried out show that trained runners showed 44% increase ‘race-pace’ endurance
as well as a 51% increase in cycling endurance, after a dosage of 9 milligrams of caffeine
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per kilogram of body weight. Caffeine will also increase the effectiveness of drug. For an
example, it makes pain reliever 40% more effective in relieving headache if being taken
with caffeine. Nowadays, more and more headache drugs include caffeine in their
formulae. But, there’s still bad effect if we take it excessively. This can lead to
caffeinism. Caffenism together with caffeine dependency can cause nervousness, anxiety,
irritability, muscle twitching, insomnia, headaches, heart palpitation and peptic ulcers. In
heart, caffeine bind to receptors on surface of heart muscle cells, which leads to an
increase in the level of cAMP ( cyclicAMP- function to increase activity of funny current
which directly increase the heart rate) inside the cells by blocking the enzyme that
degrades cAMP and mimicking the effects of epinephrine (which binds to receptors on
the cell that activate cAMP production). cAMP acts as a "second messenger," and
activates a large number of protein kinase A (PKA~cAMP-dependent protein kinase).
This has the overall effect of increasing the rate of glycolysis and increases the amount of
ATP available for muscle contraction and relaxation. According to one study, caffeine in
the form of coffee helps to reduce the risk of heart disease. However, the protective effect
was found only in participants who were not severely hypertensive ( patients that are not
suffering from a very high blood pressure). Research suggests that drinking caffeinated
coffee can cause a temporary increase in the stiffening of arterial walls. In order to
determine effect of caffeine in human life, we have to take a substitute of human being.
Daphnia has been chosen because it have sort of heart that we can see in magnification
due to its transparent body. Daphnia or commonly known as water flea is a small,
planktonic crustaceans. It is 0.2~0.5mm in length which can been seen by naked eyes. It
live in various aquatic environment from acidic swamps to freshwater lakes, ponds,
streams and rivers.
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Diagram 1.2 shows daphnia and where the heart locate.
In many Daphnia species, the skin is translucent. Therefore, it is an excellent subject as
one can observe the beating heart on top of the back, just behind the head. Average heart
rate for Daphnia is 180 beats per minute(BPM). Daphnia is prone to alcohol intoxication,
and therefore making it a good subject to study the effect of depressant on nervous
system. Not only alcohol, Daphnia can also become an indicator to study the effect of
caffeine, nicotine and adrenaline on heart beat. Daphnia is belong to clodoceran family
Daphiniidae. It reproduce parthenogetically usually in spring until end of summer. The
newly hatched Daphnia must moult several times before they are fully grown into an
adult usually after about two weeks. The young are small copies of the adult; there are no
true nymphal or instar stages. A fully mature female able to produce young about every
10 days under normal condition. The Daphnia’s lifespan are not more than 1 year.
Usually 108 days at 30C while 29 days at 280C. They eat tiny crustaceans but usually
ingest unicellular algae, protists, bacteria, and yeast(normally in lab). The beating of legs
produces a constant current through the skin which bring the material into digestive tract.
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The trapped food particles are formed into food bolus. The food bolus moves down the
digestive tract until is emptied through the anus. Basically, Daphnia is being used to test
the effect of toxins on ecosystem also known as indicator species. This is due to their
short lifespan and reproductive capabilities. In order to count the heart beat of Daphnia,
we need to use a microscope ; either compound microscope with low power objective or
dissecting microscope. A depression slide is used instead of plain slide. This is due to the
depression slide have polished, concave depression at the centre of slide where it can hold
large drop of water.
Diagram 1.3 shows microscope Light source, a light or a mirror (7) Diaphragm and condenser lens (8) Objective (3)
Ocular lens (eyepiece) (1) Objective turret (to hold multiple objective lenses) (2) Stage (to hold the sample) (9) Focus wheel to move the stage (4 - coarse adjustment, 5 - fine adjustment)
RATIONALE OF REPORT
Nowadays, due to the fast developing world, people tend not to care about the food or
the beverages the take each day. As long as it tastes nice then everyone would prefer to
have it. Did you ever know that the drink that we often have during our breakfast is an
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insecticide? Sounds horrible right? Yeah, caffeine acts as natural insecticide to the plants
it comes from. It can paralyse or even kill the insects feeding on its leaves, barks, fruits
etc. Cocoa in South Africa, and tea in Asia have all been used for hundred years to
produce drinks containing caffeine. Even these days, caffeine is also used as flavour
enhancer in soft drinks. In humans, caffeine acts as a stimulant drug, causing increased
amounts of stimulatory neurotransmitters to be released. At high levels of consumption
caffeine has been linked to restlessness, insomnia, and anxiety, causing raised blood
pressure. This can lead to heart and circulation problems. That is why it is important for
us to carry out this experiment. To see by ourselves how the heart beat increase when the
Daphnia is put into caffeine of 0.5% concentration. From this experiment, we can analyse
the data and use it to compare with human being, But we have to be careful because
Daphnia is different from human. Things happen to Daphnia might not be the same as
taken place in human being.
OBJECTIVES
Basically, our experiment is done to investigate the effect of caffeine on heart rate of
Daphnia. This experiment also is done to find the average heart beat of Daphnia at
normal condition( without being caffeinated) . Next, we would like to create awareness
among society how caffeine give such great impact on our heart. It can lead to
cardiovascular diseases because as our heart rate increase, this will at the same time
increase our blood pressure and thus give tense to our arteries, which is very harmful. By
measuring the heart rate of Daphnia in normal condition and caffeinated condition, we
can compare the heart rate and thus making it as an indicator for human heart rate.
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EXPERIMENTAL HYPOTHESIS
Presence of caffeine on Daphnia will increase the heart rate. This is because caffeine
is a stimulant. It means that it will increase the heart rate of living organism which
consume or absorb it( For this case, Daphnia absorb). At very high consumption, caffeine
linked to raised blood pressure. As blood pressure increase, the alertness (which is linked
to neurones in brain) also increase. The stimulated pituitary gland in brain send impulse
to adrenal gland to release adrenaline( a fight or flight hormone). This hormone
eventually increase the heart rate. That is how caffeine can increase our heart rate.
NULL HYPOTHESIS
There is no difference in rate of heart beat between Daphnia in culture solution and
caffeine solution. This is because Daphnia is highly resistant to caffeine. Furthermore,
caffeine does not affect anything to Daphnia’s body meaning that it will not affect human
body too.
VARIABLES
MANIPULATED VARIABLE: Presence of caffeine in solution where Daphnia is put in
CONSTANT VARIABLE: Species of Daphnia
RESPONDING VARIABLE: Heart beat per minute(BPM)
MATERIALS AND APPARATUS
MATERIALS: Culture of Daphnia, 0.5% caffeine solution, tissue paper, muslin cloth,
paper.
APPARATUS : plastic transfer pipette, beaker, stopwatch, compound microscope, pencil,
dissecting slide,
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METHOD
Measuring normal heart rate of Daphnia1. By using a plastic pipette, a Daphnia is withdrawn from the beaker
containing culture solution.
2. The Daphnia is placed on the cavity slide together with a few drops of
pond water( culture solution).
3. Excess pond water is wiped using muslin cloth.
4. By using a microscope , the Daphnia is focussed under the low-powered
objective lens. Then, the heart is focussed.
5. The heart rate is counted by tapping a piece of scrap paper eveytime the
heart beats and marks are counted afterwards. The heart rate is counted for
20 seconds. The count is repeated three times to get the average value.
6. Values are times with three to get the heart beat per minute.
7. Readings are recorded and tabulated.
Mesuring the heart rate of Daphnia in caffeine solution
1. The culture solution is drew off. By using a dropper, a drop of
caffeine solution is added onto the Daphnia.
2. By using a microscope , the Daphnia is focussed under the low-
powered objective lens. Then, the heart is focussed.
3. The heart rate is counted by tapping a piece of scrap paper
eveytime the heart beats and marks are counted afterwards. The
heart rate is counted for 20 seconds. The count is repeated three
times to get the average value.
4. Values are times with three to get the heart beat per minute.
5. Readings are recorded and tabulated
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SAFETY MEASURES AND RISK ASSESSMENT
There are a few safety measures that must be taken into consideration in order to make
sure that the experiment is going on smoothly without any accidents. But this experiment
is quite simple because it does not involve hazardous substances. The materials used are
apparently not harmful but still there are a number of safety measures that we must obey.
First, we must wear the lab coat to prevent any stain onto our cloth. We have to make
sure the concentration of caffeine used is not more than 1% because Daphnia does not get
enough oxygen in such concentrated solution. Do not eat and drink throughout the
experiment because the biology lab still keep the hazardous chemical substances even we
are not using it. We must also be very careful when handling the lab apparatus like
beakers and cavity slide which tend to break easily and will harm our safety. The
microscope are fragile and light bulbs can get so hot, so we have to be careful. We have
to take care of handling any chemicals that might be affected the heart rate of Daphnia.
And always keep the laboratory in clean state.
RESULTS AND DATA
Table 1: Daphnia in culture solution(normal condition)
Heart rate of Daphnia in
20s
Heart rate of Daphnia in one minute
10 30
9 27
8 24
Therefore, the average value of Daphnia’s heart rate= (30+27+24)/3
= 27 BPM
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Table 2: Daphnia in 0.5% caffeine solution
Heart rate of Daphnia in
20s
Heart rate of Daphnia in one minute
14 42
12 36
11 33
Therefore, the average value of Daphnia’s heart rate= (42+36+33)/3
= 37 BPM
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DISCUSSION
Based on our result, we found that the heart rate of Daphnia increase when it is put
in caffeine solution. In normal condition, the heart rate of Daphnia is 27 beats per minute,
while in caffeine solution, it is 37 beats per minute.The heart rate increase by 37%. Our
result meets our hypothesis. Caffeine solution is indeed a stimulant. Upon comparing our
result with other groups, they got the same as us that is an increment in heart rate in
caffeine solution. As we look on the sequence of our result, the heart rate decrease with
time, means that the first data is higher than the second, and so did the second data, it is
higher than the third one making the first data to be the highest and the last data is the
lowest. We conclude that as time goes on, the Daphnia is stressful enough to be in little
of water. It is dying actually.
CONCLUSION
After we have done this experiment, finally we came into conclusion heart rate of
Daphnia do increase in caffeine solution compared to its normal heart rate. Our
experimental hypothesis is accepted. The heart rate increase by 37% is because caffeine is
a stimulant. Caffeine belong to a group of compound called methylxanthines which affect
the heart rate by increasing beats per minute. It blocks adenosine receptor on surface of
heart muscle which produce adenosine(function to slow down heart rate in human causing
sleepiness). If this receptor is blocked, then fewer enzyme is produced and hence making
the heart rate increase. Daphnia is an unique organism because it absorbs chemicals that
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are present in the water surround it without selection. We conclude that our experiment
is successful because all the objectives aimed before experiment have been achieved.
LIMITATIONS AND ANOMALAOUS RESULT
There are a few limitations that prevent us from getting a very acurate and precise
result.This is due to lack in time. Most of our time is wasted to catch the tiny Daphnia in
pond water. It is so hard to see such small creature with naked eyes. We are also lack in
Daphnia which only 2 Daphnia provided for us. This is because Daphnia has short
lifespan. They died easily.. The actual value for Daphnia’s normal heart rate is 172 beats
per minute, but ours is only 27 beats per minute. While for in caffeine,it is 173 BPM, but
ours is 37 BPM only. We conclude that measurement that we took is not accurate due to
Daphnia become stressed in small volume of water. Therefore, the heart beats irregularly
and the measurements taken is not accurate enough. The illuminating light from
microscope have create a high temperature surrounding which is not suitable for Daphnia,
a temperature-sensitive organism. While taking the measurements, the Daphnia keep
moving and swimming around. With the beating leg and its active movement are
confusing us to see the heart beats
SUGGESTION FOR IMPROVEMENT
In order to get a more accurate result, a few suggestions had came across. The first one
is instead of using a plastic pipette to withdraw the Daphnia, it is better for us to use a
small fishing net. Using pipette took a lot of time because we tend to catch the
algae(bigger in size) rather than Daphnia which is small and transparent. The experiments
also should be repeated thoroughly (from beginning till end) to get a more valid result.
When the Daphnia moves around, we should wipe the solution so that it is hard for it to
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swim around with much less solution. To prevent the Daphnia from overheating, while
observing the heart rate, we should turn off the microscope and use the heat sink- a cavity
slide filled with iced-water was placed under different slide and was sealed by a
coverslip. This slide was then placed below the slide containing Daphnia.
REFERENCES AND BIBLIOGRAPHY
http://www.naturalhub.com/natural_food_guide_fruit_vitamin_c.htm; 4th August 2010
http://en.wikipedia.org/wiki/Vitamin_C; 4th August 2010 http://www.practicalbiology.org/areas/advanced/health-and-disease/whats-in-our-
food/measuring-the-vitamin-c-content-of-foods-and-fruit-juices,53,EXP.html; 4th August 2010
http://www.coursework.info/GCSE/Biology/Living_Things_in_their_Environment/Measurement_of_the_vitamin_C_content_of_L10763.html; 4th August 2010
http://en.wikipedia.org/wiki/Dichlorophenolindophenol; 4th August 2010
APPENDIX
Scientific classification Kingdom: AnimaliaPhylum: ArthropodaSubphylum: CrustaceaClass: BranchiopodaOrder: CladoceraFamily: DaphniidaeGenus: DaphniaMüller, 1785 Species
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Subgenus Daphnia
D. ambiguaD. arenata
D. catawba
D. cheraphila
D. latispina
D. melanica
D. middendorffiana
D. minnehaha
D. neo-obtusa
D. obtusa
D. oregonensis
D. parvula
D. pileata
D. prolata
D. pulex
D. pulicaria
D. retrocurva
D. tanakai
D. tenebrosa
D. villosa
Subgenus Hyalodaphnia
D. curvirostrisD. dentifera
D. dubia
D. galeata
D. lacustris
D. laevis
D. longiremis
D. longispina
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D. mendotae
D. thorata
D. umbra
Subgenus Ctenodaphnia
D. barbataD. brooksi
D. ephemeralis
D. exilis
D. lumholtzi
D. magna
D. salina
D. similis
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