Experiment 2: Concentration Gradients and Membrane Permeability
In this experiment, you will dialyze a solution of glucose and starch to observe: The directional movement of glucose and starch. The effect of a selectively permeable membrane on the diffusion of these molecules.
An indicator is a substance that changes color when in the presence of a specific substance. In this experiment, IKI will be used as an indicator to test for the presence of starch.
Materials
(5) 100 mL Beakers
10 mL 1% Glucose Solution, C6H12O6
4 Glucose Test Strips
(1) 100 mL Graduated Cylinder
4 mL 1% Iodine-Potassium Iodide, IKI
5 mL Liquid Starch, C6H10O5
3 Pipettes
4 Rubber Bands (Small; contain latex, handle with gloves on if allergic)
Permanent Marker
* Stopwatch
* Water
* Scissors
*15.0 cm Dialysis Tubing
*You Must Provide
*Be sure to measure and cut only the length you need for this experiment. Reserve the remainder for later experiments.
Attention!
Do not allow the open end of the dialysis tubing to fall into the beaker. If it does, remove the tube and rinse thoroughly with water before refilling it with the starch/glucose solution and replacing it in the beaker.
Note:
If you make a mistake, the dialysis tubing can be rinsed and used again.
Dialysis tubing must be soaked in water before you will be able to open it up to create the dialysis “bag.” Follow these directions for this experiment:
1. Soak the tubing in a beaker of water for ten minutes.
2. Place the dialysis tubing between your thumb and forefinger, and rub the two digits together in a shearing manner. This motion should open up the “tube” so that you can fill it with the different solutions.
Procedure
1. Measure and pour 50 mL of water into a 100 mL beaker using the 100 mL graduated cylinder. Cut a piece of dialysis tubing 15.0 cm long. Submerge the dialysis tubing in the water for at least ten minutes.
2. Measure and pour 82 mL of water into a second 100 mL beaker using the 100 mL graduated cylinder. This is the beaker you will put the filled dialysis bag into in Step 9.
3. Make the glucose/sucrose mixture. Use a graduated pipette to add 5 mL of glucose solution to a third 100 mL beaker and label it “dialysis bag solution.” Use a different graduated pipette to add 5 mL of starch solution to the same beaker. Mix by pipetting the solution up and down six times.
4. Using the same pipette that you used to mix the dialysis bag solution, remove 2 mL of the dialysis bag solution and place it in a clean beaker. This sample will serve as your positive control for glucose and starch.
a. Dip one of the glucose test strips into the 2 mL of glucose/starch solution in the third beaker. After one minute has passed, record the final color of the glucose test strip in Table 3. This is your positive control for glucose.
b. Use a pipette to transfer approximately 0.5 mL of IKI into the 2 mL of glucose/starch solution into the third beaker. After one minute has passed, record the final color of the glucose/starch solution in the beaker in Table 3. This is your positive control for starch.
5. Using a clean pipette, remove 2 mL of water from the 82 mL of water you placed in a beaker in Step 2, and place it in a clean beaker. This sample will serve as your negative controls for glucose and starch.
a. Dip one of the glucose test strips into the 2 mL of water in the beaker. After one minute has passed, record the final color of the glucose test strip in Table 3. This is your negative control for glucose.
b. Use a pipette to transfer approximately 0.5 mL of IKI into the 2 mL in the beaker. After one minute has passed, record the final color of the water in the beaker in Table 3. This is your negative control for starch.
Note : The color results of these controls determine the indicator reagent key. You must use these results to interpret the rest of your results.
6. After at least ten minutes have passed, remove the dialysis tube and close one end by folding over 3.0 cm of one end (bottom). Fold it again and secure with a rubber band (use two rubber bands if necessary).
7. Test to make sure the closed end of the dialysis tube will not allow solution to leak out. Dry off the outside of the dialysis tube bag with a cloth or paper towel. Then, add a small amount of water to the bag and examine the rubber band seal for leakage. Be sure to remove the water from the inside of the bag before continuing. Using the same pipette that was used to mix the solution in Step 3, transfer 8 mL of the dialysis bag solution to the prepared dialysis bag.
Figure 4: Step 9 reference.
9. Place the filled dialysis bag in the 100 mL beaker filled with 80 mL of water, leaving the open end draped over the edge of the beaker as shown in Figure 4.
10. Allow the solution to sit for 60 minutes. Clean and dry all materials except the beaker holding the dialysis bag.
11. After the solution has diffused for 60 minutes, remove the dialysis bag from the beaker and empty the contents of the bag into a clean, dry beaker. Label the beaker “final dialysis bag solution.”
12. Test the final dialysis bag solution for the presence of glucose by dipping one glucose test strip into the dialysis bag. Wait one minute before reading the results of the test strip. Record your results for the presence of glucose in Table 4.
13. Test for the presence of starch by adding 2 mL IKI. After one minute has passed, record the final color in Table 4.
14. Use a pipette to transfer 8 mL of the water in the beaker to a clean beaker. Test the beaker water for the presence of glucose by dipping one glucose test strip into the beaker. Wait one minute before reading the results of the test strip, and record the results in Table 4.
15. Test for the presence of starch by adding 2 mL of IKI to the beaker water. Record the final color of the beaker solution in Table 4.
Table 3: Indicator Reagent Data
Indicator
Starch Positive
Control (Color)
Starch Negative
Control (Color)
Glucose Positive
Control (Color)
Glucose Negative
Control (Color)
Glucose Test Strip
n/a
n/a
IKI Solution
n/a
n/a
Table 4: Diffusion of Starch and Glucose Over Time
Indicator
Dialysis Bag After 60 Minutes
Beaker Water After 60 Minutes
IKI Solution
Glucose Test Strip
Post-Lab Questions
1. Why is it necessary to have positive and negative controls in this experiment?
2. Draw a diagram of the experimental set-up. Use arrows to depict the movement of each substance in the dialysis bag and the beaker.
3. Which substance(s) crossed the dialysis membrane? Support your response with data-based evidence.
4. Which molecules remained inside of the dialysis bag?
5. Did all of the molecules diffuse out of the bag into the beaker? Why or why not?
Experiment 1: Diffusion through a Liquid
In this experiment, you will observe the effect that different molecular weights have on the ability of dye to travel through a viscous medium. Materials
1 60 mL Corn Syrup Bottle, C12H22O11
Red and Blue Dye Solutions (Blue molecular weight = 793 g/mole; red molecular weight = 496 g/mole)
(1) 9 cm Petri Dish (top and bottom halves)
Ruler
* Stopwatch
* Clear Tape
*You Must Provide
Procedure
1. Use clear tape to secure one-half of the petri dish (either the bottom or the top half) over a ruler. Make sure that you can read the measurement markings on the ruler through the petri dish. The dish should be positioned with the open end of the dish facing upwards.
Carefully fill the half of the petri dish with corn syrup until the entire surface is covered. Develop a hypothesis regarding which color dye you believe will diffuse faster across the corn syrup and why. Record this in the post-lab questions. Place a single drop of blue dye in the middle of the corn syrup. Note the position where the dye fell by reading the location of its outside edge on the ruler. Record the location of the outside edge of the dye (the distance it has traveled) every ten seconds for a total of two minutes. Record your data in Table 1 and use your results to perform the calculations in Table 2. Repeat the procedure using the red dye, the unused half of the petri dish, and fresh corn syrup.
Table 1: Rate of Diffusion in Corn Syrup
Time (sec)
Blue Dye
Red Dye
Time (sec)
Blue Dye
Red Dye
10
70
20
80
30
90
40
100
50
110
60
120
Table 2: Speed of Diffusion of Different Molecular Weight Dyes
Structure
Molecular Weight
Total Distance
Traveled (mm)
Speed of Diffusion
(mm/hr)*
Blue Dye
Red Dye
*Multiply the total distance diffused by 30 to get the hourly diffusion rate
Post-Lab Questions Record your hypothesis from Step 3 here. Be sure to validate your predictions with scientific reasoning.
Which dye diffused the fastest?
Does the rate of diffusion correspond with the molecular weight of the dye?
Does the rate of diffusion change over time? Why or why not?
Examine the graph below. Does it match the data you recorded in Table 2? Explain why, or why not. Submit your own plot if necessary.
https://nuonline.neu.edu/bbcswebdav/pid-9451339-dt-content-rid-14232100_1/courses/BIO1101.90155.201714/BIO1101.90155.201714_ImportedContent_20160930044714/CourseRoot/html/lab006s001.html
https://nuonline.neu.edu/bbcswebdav/pid-9451340-dt-content-rid-14232401_1/courses/BIO1101.90155.201714/BIO1101.90155.201714_ImportedContent_20160930044714/CourseRoot/html/lab006s002.html
https://nuonline.neu.edu/bbcswebdav/pid-9451341-dt-content-rid-14232402_1/courses/BIO1101.90155.201714/BIO1101.90155.201714_ImportedContent_20160930044714/CourseRoot/html/lab006s003.html
