By: Pat Ebbert
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Amylase is an enzyme found in the body that
breaks down starch into simple sugars.
Amylase is found in the saliva and in the
pancreas of humans, breaking down the starch
from the foods we consume into simpler
sugars, which are later converted into glucose
needed for energy.
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Alpha-Amylase is the structure of amylase
found in human saliva and pancreas.
It breaks down long strands of carbohydrates
into simpler sugars such as maltose or glucose.
This form of amylase is considered superior
because it can act on any portion of the
substrate , unlike Beta-Amylase, making
reactions occur much faster.
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To the right is a
diagram showing the
structure of the AlphaAmylase enzyme.
The green dot
represents a chloride
ion, the pale yellow a
calcium ion. Both are
used to help the enzyme
bind to the substrate.
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An iodine atom
Iodine has the property of
binding to starch to create a
dark blue color.
If starch has been broken down
into simpler sugars, the iodine
cannot bind and therefore
leaves an area of no color
visible.
If amylase has broken down
surrounding starch into simpler
sugars, this area’s diameter can
be measured to give a relative
value of how much starch has
been broken down.
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Three variables are used in this experiment to
test their effect on amylase activity. All
variables are liquids that could be found in the
oral cavity, because that is where salivary
amylase is most prominent.
Listerine is used to see if mouthwash effects
amylase just as it kills bacteria.
Diet Pepsi is used to see if carbonation has any
effect on the amylase.
At last, Mylanta antacid drink is used to see if a
change in pH has an effect on amylase activity.
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The purpose of this experiment is the
determine whether or not the presence of
various liquids that may be found in the mouth
has any effect on amylase activity.
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Null Hypothesis: It is hypothesized that the
presence of Mylanta, Diet Pepsi, or Listerine
will have NO significant effect on the activity
of enzyme alpha-amylase when compared to
the controls.
Alternate Hypothesis: It is hypothesized that
the presence of Mylanta, Diet Pepsi, or
Listerine WILL have a significant effect on the
activity of enzyme alpha-amylase when
compared to the controls.
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2% stock solution of 1,4α-D-glucan
glucanohydrolase
(alpha-amylase) in
distilled water
3% stock solution of
iodine in distilled water
5ml macro-pipette
50-200µl micro-pipette
Micro-pipette tips
16 starch-agar plates
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8 test tubes (5ml)
45ml mixing tube
Squirt bottle
Sharpie marker
Drinking straws
Diet Pepsi
Mylanta antacid drink
Listerine mouthwash
1.
2.
3.
4.
Create eight different test tube solutions to be used in
experiment (explained on following slide).
Fill five wells in each of 16 starch agar plates with 50µl
of amylase for eight different data sets (10 trials of
each set, 80 total wells).
Two data sets are positive and negative control sets,
three are 10% variable solutions, and three are 50%
variable solutions (all of these come from the test
tubes).
After one hour, spray iodine solution onto each plate
and measure zone of starch-breakdown in millimeters.
Neg. Control
Pos. Control
10% Var.*
50% Var.*
Amylase
solution
0ml
1ml
1ml
1ml
Distilled
water
5ml
4ml
3.5ml
1.5ml
Variable
0ml
0ml
0.5ml
2.5ml
Total
5ml
5ml
5ml
5ml
This table represents what will be present in the eight test tubes used in
the experiment. After the solutions are prepared, 50µl of each tube are
pipetted into ten different starch-agar plate wells.
*The variable columns are repeated three times (Mylanta, Diet Pepsi,
Listerine)