IA Lab One: Enzymes
Design:
Background: The reaction between lactase (an enzyme) and lactose (a sugar in milk)
produces glucose and galactose. A Benedicts test is when Benedicts solution is added to a
mixture. If and/or when the mixture contains glucose the Benedicts solution will change
colors, becoming orange. In this way, performing a Benedicts test on just-mixed lactase
and lactose will indicate the end of the reaction (when the mixture changes colors) and it
will allow me to time the length of the reaction.
Research Question: How will increasing the amount of lactase added to milk affect the
rate of the reaction between lactose and lactase?
Hypothesis: I think that increasing the amount of lactase added to milk will decrease the
length of the reaction because increasing substrate concentration increases enzyme
activity.
Method for Collecting Data: I will record the length of the reaction in seconds and the
amount of lactase used in grams in a data table.
Independent Variable: Amount of lactase added to the reaction
Dependent Variable: Length of the reaction
Constants: Brand of lactase pill, type/brand of milk, amount of milk used, amount of
Benedicts solution used, temperature of water
Method for Controlling Variables: I will consistently use Lactaid Fast Act brand lactase
pills. I will consistently use 15 (± 1) mL of Schroeder brand 1% milk. I will consistently
use 5 (± .5) mL of Benedicts solution. To control the temperature of the boiling water
bath, I will keep the hot plate on the high setting.
Materials: Lactaid Fast Act lactase pills, Schroeder 1% milk, graduated cylinder, digital
scale, pestle and mortar, Benedicts solution, timers, funnel, hot plate, graduated dropper,
test tubes, test tube rack, large beaker, water, masking tape, marker, Petri dish
Procedure: 1. Gather all materials.
2. Fill a large beaker (big enough to hold four, 150 mL test tubes) and fill it with water.
Set it on the hot plate and turn the hot plate to high. Let it heat.
3. Place four test tubes in a test tube rack. Using the marker and tape, label each test tube
with the amount of lactase (beginning at 0.50 g) and the trial number (1-4).
4. Crush 10 or more lactase pills in with the pestle and mortar. (You will need more
lactase for the next concentration levels of lactase, so make plenty.) Crush until pills are
consistently a fine powder.
5. Fill each test tube with 15 (± 1) mL of milk.
6. Using the graduated dropper, add 5 (± .5) mL of Benedicts solution to each test tube of
milk.
7. Weigh out 0.50 (± 0.01) g of lactase pills (now in powder form) in a Petri dish.
8. Place the funnel in the test tube labeled trial one. Pour the measured lactase into the
milk. Swirl the test tube to blend. Put into the beaker of hot water. Start a timer. Watch
the solution. Once it changes colors (from purple to an orange), record the time in a data
table. Set the completed trial aside.
9. Clean out materials. Add water to the hot water beaker if needed. Crush more lactase
pills if needed. Again, place four test tubes in a rack and measure 15 (±1) mL into each
tube.
10. Repeat steps 7-9 with 1.00 (± 0.01) g of lactase powder.
11. Repeat steps 7-8 with 2.00 (± 0.01) g of lactase powder.
12. Clean and put away all materials.
Data Processing/Presentation:
Reaction Time of Lactase and Lactose
Amount of Lactase Added Trial
(± 0.01g)
Number
0.50
1.00
2.00
1
2
3
4
1
2
3
4
1
2
3
4
Length of Reaction
(± .01 seconds)
357.00
351.00
318.00
298.00
224.00
222.00
208.00
217.00
182.00
178.00
172.00
181.00
Average Reaction Times:
[(Trial one reaction time) + (trial two reaction time) + (trial three reaction time) + (trial
four reaction time)] / 4 = Average reaction time
0.50 g Lactase: [(357.00 ± .01s) + (351.00 ± .01s) + (318.00 ± .01s) +
(298.00 ± .01s)] / 4 = 331.00 ± .01s
1.00g Lactase:
= 217.75 ± .01s
2.00g Lactase:
= 178.25 ± .01s
Conclusion/Evaluation:
My results were conclusive and support my hypothesis. I conclude that increasing
the amount of lactase added to milk decreases the length of the reaction. My graph shows
a decrease in reaction time as the concentration of lactase increases. My conclusion is
supported with information from the CCBC Student Server. “Assuming a sufficient
concentration of substrate is available, increasing enzyme concentration will increase the
enzyme reaction rate.” (Kaiser, Gary E. "Enzymes." Student.ccbcmd.edu. 27 June 2001.
Web. 24 Nov. 2010.
<http://student.ccbcmd.edu/~gkaiser/biotutorials/proteins/enzyme.html>.) This explains
why the second decrease in reaction time is less than the first. As more active sites are
being introduced, there are less available substrates to occupy them and enzyme activity
decreases. If I did more trials with larger amounts of lactase, it’s probable that there
would be further decrease in enzyme activity.
There were possible sources of error in this lab. One of the most significant was
the temperature of the hot water bath I put the test tubes in (step eight). It was hard to
know if the temperature was consistent, especially after adding water if it was all boiling
away. Higher temperatures could have lead to a fasted Benedicts test. Another significant
error was the Benedicts test. To declare the end of the reaction, the mixture needed to
turn yellow. However, there were many color gradients of purple into yellow. It was hard
to know if I was keeping the indicating color consistent. Also, the crushed pills were not
crushed completely evenly. Some chunks were bigger than others. Finally, there was
some delay in beginning the timers at the start of the reaction. I had to pour in the lactase,
swirl the tube, put it in the hot water beaker, and then I could start the timer. This error is
less significant because it happened for every trial of my lab. All the data is similarly off.
I could make many improvements on this lab. I could keep a thermometer in the
hot water beaker to maintain a constant temperature. I could find a different way of
heating the water that would ensure a more even temperature. To fix the issue of test
color, I could keep the first test I complete and use that as a model for the color that
indicates the end of the reaction. I could make a strip (like a pH strip) to use while doing
the lab. As for the consistency in pill form, I could spend longer crushing the pills and
concentrate on getting an even mix. I could use a different method to get more consistent
results (a food processor, for example). Finally, I could get a partner to help me start the
timers sooner. I could swirl the tube while it’s in the hot water, after starting the timer.
Any of these changes could lead to more accurate results.