Several reports (including one by the Medical Daily

Magazine as noted in the bibliography) have shown that

coffee is beneficial to liver function and aids in preventing
liver disease. However, no studies have ever isolated
which specific ingredient(s) of coffee cause the positive
effects to liver function. This experiment tests the effects of
three main chemical ingredients of coffee (caffeine, niacin
and theobromine) on the activity of liver catalase, an
enzyme in the liver that breaks down hydrogen peroxide to
prevent it from harming the body. During this process,
hydrogen peroxide is converted to water and oxygen. The
quantity of oxygen produced is directly correlated with the
level of catalase activity. This is why the experimenter
uses gas pressure as an indicator of the livers activity.

Which ingredient in coffee (caffeine, theobromine, and

niacin) will increase the level of catalase activity of the
liver the most?

Among caffeine, niacin and theobromine in coffee, caffeine

will affect the activity of the liver catalase the most.

Independent Variable: Chemical ingredient

Dependent Variable: Amount of oxygen produced by liver
catalase (measured in kPa)

Part 1: Preparation
1. Put on an apron, gloves and safety goggles.
2. Measure 50ml of -4oC distilled water, and place into
the blender along with the 50 grams of liver.
3. Blend, and pour blended contents into the beaker,
and put the beaker into a container of ice to preserve
liver cells until use.
4. Prepare 3 groups of test tubes with 3 test tubes for
each group. Each group represents one ingredient;
there are 3 ingredients being tested, hence the 3
groups. Label the 3 tubes of each group as Solution
1 through Solution 3.
5. Weigh using the scale and add 0.1 grams of the first
ingredient to be tested into the tube labeled as
Solution 1, add 10 ml distilled water, mix using the
glass stirrer to dissolve the powder evenly.
6. Take 3.3 ml of the solution from Solution 1, add into
the tube labeled Solution 2, add 6.6 ml water, mix
well using the glass stirrer to produce a half-log, or
3.3 times, more diluted solution.
7. Repeat step 4, adding from Solution 2 into the tubes
labeled Solution 3. This will create a strong, undiluted
amount of ingredient as well as a 3.3 times and 10
times more diluted solution, allowing a greater range
of amounts of each ingredient to improve the
accuracy of the experiment.
8. Repeat steps 6 through 7 using the other 2
ingredients to make two more groups of solutions.
(The other 2 ingredients of coffee that we are using in
this experiment.)
9. Measure and place 100 microliters of the blended
liver into each of 3 empty test tubes using the
micropipette.
 10. Add 0.5ml of Solution 1 of the first ingredient into
the first test tube, which should now contain blended
liver.
11. Repeat steps 5-10 using another ingredient.
12. Prepare the laptop and oxygen sensor, but do not
start measuring yet.

Part 2: Experimentation
1. Measure and place 500 microliters of hydrogen
peroxide into the first test tube of the first ingredient
using the micropipette, .
2. Quickly cover the opening of the test tube with the
oxygen sensor and continuously record results using
the computer for the next minute. Save the file on the
computer, and record key data, such as the peak air
pressure (in kPa) of a trial and how long it took to
reach that peak.
3. Repeat steps 1 and 2 with all of the different test
tubes, using solutions 2, and 3 for 3 trials each until
all of the solutions (of the first ingredient) were tested
3 times.
4. Repeat steps 1-3 using the next ingredient.
5. Clean the blender and the test tubes with water and
(optionally) detergent. Dry them and place them back
where they belong.
6. Clean the tip of the oxygen sensor, verify that the data
was saved on the laptop, and place both back in their
proper locations.

Note: In the beginning of the procedure, 5 groups of

varying amounts of each ingredient were created. All 5
groups represent 1 ingredient, but in different amounts to
improve the accuracy of the experiment. After all, different
amounts of an ingredient will impact its effectiveness. Too
much or not enough of an ingredient will limit the
experiments success and precision. Later, the procedure
instructed the experimenter to measure and place 0.5ml of
each solution from its original, stock test tube into an
empty test tube. The purpose of the whole moving-around
thing is to save time, as re-measuring and re-mixing the
proper amounts for every amount of every ingredient will
unnecessarily waste a lot of time. The experiment will be
testing various amounts of each ingredient, with 3 trials for
each amount.

Conclusion:

The results have supported the hypothesis: out of the

three different chemical ingredients of coffee tested,
caffeine provides the most powerful stimulation for
catalase activity. In addition, the study has shown that
Niacin stimulated activity of the catalase the least. The
experiment proved another interesting point: all 3 of the
main chemical ingredients of coffee tested had beneficial
effects to the catalase activity, as the gas pressure for all 3
ingredients at varying amounts surpassed those of the
control tests. This is significant, because it gives people an
excuse for a morning cup of joe and proves that a limited
amount of caffeine in ones diet will improve liver function.
Future studies could include testing a larger range of
quantities to try to find out how much caffeine is most
beneficial. Other chemical ingredients of coffee could also
be tested to widen the scope of this experiment. Overall,
the experiment was conclusive in that all 3 ingredients
increased the activity of catalase, and that caffeine was
the most effective ingredient.

Olson, S. (2015, March 30). Coffee Might Protect The

Liver From Alcohol's Damaging Effects. Retrieved October
25, 2015.

Liver Facts. (n.d.). Retrieved October 25, 2015, from

http://www.liver.ca/liver-health/liver-facts.aspx

Goodsell, D. (2004, September 1). Catalase. Retrieved

November 15, 2015, from
http://www.rcsb.org/pdb/101/motm.do?momID=57

Emily, W., & David, Z. (2015, February 2). Niacin:

MedlinePlus Medical Encyclopedia. Retrieved November
15, 2015, from
https://www.nlm.nih.gov/medlineplus/ency/article/002409.h
tm

Liver Health and Wellness. (2015, January 7). Retrieved

November 13, 2015, from
http://www.liverfoundation.org/abouttheliver/liverhealth/

Adrian, J., & Fragne, R. (n.d.). Synthesis and availability of

niacin in roasted coffee. Retrieved November 13, 2015,
from
http://www.ncbi.nlm.nih.gov/pubmed/1897406
Electronic Scale Picture from Ohaus Corp. on Amazon

Gas Pressure Sensor Picture from Vernier Software &

Technology Website

Graduated Cylinder Picture from MSNucleus.org

Test Tube Rack Clip Art from clker.com

(shared by ocal 6-22-09)
 Scale (1)

Laptop (1)

Oxygen Sensor (1)

Graduated Cylinder (1)

Beaker (w/ labels) (1)

Blender (1)

Glass Stirrer (1)

Test Tubes (32)

Micropipette (1)

Distilled Water (1 liter)

Cows Liver (10g)

Hydrogen peroxide (50ml)

Caffeine (200mg)

Theobromine (200mg)

Niacin (200mg)

*Photo sources cited in bibliography