Diffusion Lab - Procedure.

Part One - Semipermeable Membranes.

We will lead off with control tests to check the validity of our experimental procedures. You will need 8 test tubes, numbered 1 - 8. To the first six test tubes, you will add six drops of the following test solutions: tubes 1 & 2, pure water; tubes 3 & 4, sugar solution; tubes 5 & 6, starch solution (make sure it's properly mixed). To tubes 7 & 8, add three drops of sugar solution and three drops of starch solution.

The odd-numbered tubes will be used for a Benedict's sugar test: add six drops of Benedict's reagent, then record the combined solution's starting color on the answer sheet. Place the test tubes into a boiling water bath (wear safety glasses!) for three minutes. Record and interpret the final color (a positive for presence of sugar is a range of colors, from green through red, orange, and copper-brown).

The even-numbered tubes will be used for a Lugol's starch test: record the solution's starting color on the answer sheet, then add two drops of Lugol's reagent. Record and interpret the final color (a positive for presence of starch is a blue-black color).

Our experimental test with semipermeable tubing membrane involves dialysis tubing, which comes flattened and dry on a roll. Cut about 15 centimeters and soak it until it is soft and pliable. Gently open the tubing completely with a glass rod. Twist one end, bend it and tie it tightly with thread. Leaving enough room to similarly tie off the other end, half fill the tubing with sugar solution and half fill it with starch solution. Check the tube for leaks, then rinse it and blot it dry. Set it aside for a minute or two on a dry paper towel to check for slow leaks.

Half fill a smaller beaker with water, the add 5 milliliters of Lugol's to the beaker and mix. Place the tubing into the beaker solution and let sit for at least 20 minutes (longer is fine; you can probably do Part Two in the meantime), then answer the first Part One questions on the answer sheet. Label two test tubes: "O" for the fluid outside the tubing, and "I" for the fluid inside the tubing. Using the labeled droppers available, place 6 drops of the appropriate fluids in the tubes, the run Benedict's tests on the solutions as was done in the control tests. Now answer the second Part One questions on the answer sheet. Wash and return all glassware.

Part Two - Osmosis in Living Cells.

Place a single Elodea leaf in a small drop of water and cover with a cover slip. Find a thin area near an edge of the leaf and work your way to high (40X lens, 400X magnification) power. Look for cells with several chloroplasts, but avoid cells that are really full of chloroplasts. On the answer sheet, draw and label a single cell.

Place a drop of concentrated salt solution (the osmolarity is much higher than that of the cell's interior fluid) next to the cover slip so that it "bleeds" underneath. You may need to draw water out from the other side of the cover slip with a small piece of paper towel. Keep the cells in focus (often the focus changes as you jostle the cover slip) and watch for significant changes - the reaction could happen immediately or take a few minutes. Draw and label (the nucleus often cannot be seen, but it may become visible as an oval gap in the cell) a changed cell (it does not necessarily have to be the cell you drew first) and make special note of just how the cells have changed. Answer the first Part Two questions.

"Bleed" water back under the cover slip and, with a piece of paper towel on the opposite side, remove salt solution until another significant cell change occurs, then answer the second Part Two questions. If you absolutely know what should happen (and more importantly, why the change should occur), you can answer the questions without getting the actual change to occur.

Clean and return your slide and cover slips. Rinse them well - a little salt contamination is invisible but can have significant effects on some experiments that they may be used for later.





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