29 Bio 100 Cellular Transport Worksheet and Answers PDF

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Bio 100 Cellular Transport Worksheet and Answers...

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Exercise # 3

Cellular Transport Studies with the electron microscope reveal that the plasma membrane consists essentially of lipid material with randomly dispersed protein molecules. The lipoprotein complex is not a rigid assemblage but a “sea” of lipid molecules among which are interspersed protein molecules that may “float” only on one or both sides of the membrane or traverse the whole width of the lipid bilayer. This is known as the “fluid mosaic” model of the plasma membrane structure. Among its functions, one of the most important feature of the cell membrane, would be to regulate the flow of materials into and out of the cell. The membrane possesses a certain selectivity with which it maintains a chemical balance between the cell and its environment. Another important feature exhibited by the plasma membrane would be permeability, which allows the passage of substances into and out of the cell and which is dependent upon the activities going on within the cell. Water-soluble substances that can pass through the membrane always diffuse from a region of high concentration to one of low concentration until a state of equilibrium (equal concentration) results. This accounts for the movement of oxygen into a cell and carbon dioxide out of a cell. However, in a living cell, equilibrium is never attained. A special case of the diffusion of water molecules through a membrane is called osmosis. A membrane is said to be differentially permeable when it allows the passage of small molecules like water but blocks the passage of larger molecules. When such a membrane separates two solutions containing varying concentrations of dissolved particles, water will diffuse from the less concentrated solution (hypotonic) to the more concentrated (hypertonic) until there is an equal concentration of water (isotonic) for a given volume of space on both sides of the membrane. Diffusion alone is not enough to explain the passage of substances across a living membrane. There are substances that accumulate inside the cell while others are excluded. This process, called active transport, is an energy-consuming process and can move substances from a region of lower to higher concentration and also through membranes that would otherwise block the passage of such substances.

MATERIALS • • • • • • • • •

Light microscope 5 Test tubes Test tube rack Beaker (any size) 1 Petri dish 100-ml Erlenmeyer flask Funnel Pipette Aspirator

CHEMICAL REAGENTS • Cooking oil • Milk • Egg albumen • Starch solution • Aqueous yeast suspension • Alkaline yeast suspension • Congo red • Collodion • Distilled water • Neutral red solution

• • • • • • • • •

Dissecting needle Filter paper Knife blade Cotton Sterile disposable blood lancet Slide Cover slip Filter paper Hot pan

• • • • • • • •

Potassium permanganate (𝐾𝑀𝑛𝑂4) Methylene blue crystals 40% formalin 2% iodine solution 0.5% NaCl 0.9% NaCl 1.0% NaCl 10% NaCl 50% glucose solution

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PROCEDURE

A. Structure of the plasma membrane A1. 1. 2. 3. 4. 5.

Into a test tube, put 5 ml of oil and 5 ml of water then shake vigorously. Let it stand for a few minutes and observe. a. What happens to the oil globules? Shake again the test tube without allowing the oil and water layers to settle. Transfer several drops of the mixture to a clean glass slide. Examine under LPO and look for the interphase between the oil and water. a. How does a living cell like the amoeba remain intact in an aquatic medium?

A2. 1. 2. 3. 4. 5.

Fill the beaker, of any size, with milk. Heat it to almost boiling point then allow it to cool. Observe the formation of a “skin” on the surface. Remove the skin and heat the milk again. Repeat this process several times. a. What similarities can you observe between the milk and the plasma membrane? b. Does the removal of the skin remove any components from the milk, preventing further skin formation? c. How does the organization of the plasma membrane differ from that of the milk?

A3. 1. 2. 3. 4. 5. 6.

Pour oil into a petri dish until it is half full. Put a drop of egg albumen solution into the oil solution. Watch for the formation of a delicate membrane at the protein-lipid interphase and notice that it becomes heavier after sometime. Try to wrinkle the membrane by gently probing it with a dissecting needle. Try to rupture the membrane with the needle. a. Does it heal itself? Place a small particle of nigrosine on top of the membrane and gently push it with the needle through the membrane. Observe closely. a. Does the dye dissolve and diffuse within the protein solution? b. Is there any resemblance to the manner by which particulate material might pass from the outside into the cell’s interior? Explain.

B. Selective action of cell membranes B1. 1. 2. 3. 4. 5. 6.

Place 1 ml of aqueous yeast suspension in each of the three test tubes labelled 1, 2, and 3. To each tube add three drops of Congo red solution. To test tube 1, add four drops of 40% formalin and shake gently. Heat the contents of test tube 2 until near boiling point. Let test tube 3 stand as it is. Place a drop of each suspension on a glass slide separately and observe under the microscope. a. In which of the test tubes did most of the cells become stained? b. Which cells did not accept the dye? c. What is the effect of formalin on the selectivity of the membrane? d. What is the effect of heat? e. If the dye has stained a few cells from test tube 3, think of the possible reasons for this.

B2. 1. 2. 3. 4. 5.

Place 25 ml of alkaline yeast suspension in a 100 ml Erlenmeyer flask or beaker. Add 25 ml of neutral red solution. Observe the color immediately and note any changes within a five-minute period. Filter 10 ml of the mixture through the fine filter paper to separate the yeast cells from the liquid (this can be done via centrifugation). Observe the color of the cells and the clear supernatant fluid.

Note: The internal pH of the yeast cells is about 5.5 to 6.0 even in an alkaline suspension. Neutral red turns deep red in acid solutions and yellowish-orange in basic solutions. a. What is the function of sodium-bicarbonate? b. What is the function of neutral red in this experiment?

C. Diffusion C1. 1. 2.

Place a pinch of potassium permanganate (𝐾𝑀𝑛𝑂4) and methylene blue crystals of about equal size 3-4 cm apart on the surface of a film of agar in petri dish. Observe the spread of color at regular time intervals as the molecules diffuse throughout the jelly-like agar. a. Since the molecular weight of 𝐾𝑀𝑛𝑂4 is 158 while methylene blue is 374, is there any correlation between the molecular weight and speed of diffusion?

C2. 1. 2. 3. 4.

5. 6. 7. 8. 9.

Prepare a collodion tube by placing 3 ml of collodion in a clean, dry 10 ml test tube. Fill the tube slowly and rotate it so that a thin film forms on the entire inner surface. Drain out any excess solution and place the tube in an inverted position in a test tube rack for 10 minutes or until the film dries. Using a knife blade, loosen the edge of the film at the mouth of the test tube and put water between the film and the tube to help in removing the collodion tube and knot closing it. Rinse the outside of the tube. Place the collodion tube in a beaker half full of water. Add about 5 ml of 2% iodine solution. After a few minutes look at the contents of the tube and record any color changes. Test the outer solution for the presence of starch and sugar at ten-minute intervals for one hour. a. On the basis of this experiment, would you say that molecules can diffuse through a differentially permeable membrane in opposite directions at the same time? b. What happened to the molecules of starch, glucose, and iodine?

D. Osmosis 1. 2. 3. 4. 5. 6. 7.

Prepare a water mount of a Boat of moses using distilled water. Observe the size of the cell. After viewing under the microscope, drain out the distilled water with the use of a tissue or filter paper. Place a drop of 0.5% NaCl on the side of the cover slip and allow the solution to penetrate the specimen. Observe changes on the size of the cell. Prepare a new water mount of a Boat of moses using distilled water. After viewing under the microscope, drain out the distilled water with the use of a tissue or filter paper. Placed a drop of 1.0% NaCl on the side of the cover slip and allow the solution to penetrate the specimen. Observe changes on the size of the cell.

E. Hemolysis and crenation of the red blood cell 1. 2. 3. 4. 5. 6. 7. 8. 9.

Clean the tip of your finger with sterile cotton soaked in 70% alcohol. Let it air dry and then prick it with sterile disposable blood lancet. Add a drop of your blood to a drop of 0.9% NaCl on a clean glass slide. Add another drop of blood to a drop of distilled water on another slide. Place a cover slip and examine under the microscope. Observe the size and shape of the cells. Now add a drop of 10% NaCl to one edge of the cover glass of the slide with sodium chloride and a piece of blotting paper to the opposite side. Note what happens to the cells as the concentrated salt solution reaches them. Compare results with those obtained in distilled water.

How does a living cell like the amoeba remain intact in an aquatic medium? Contractile vacuole What similarities can you observe between the milk and the plasma membrane? Both have regenerative abilities Does removal of the skin remove any components from the milk preventing further skin formation? No How does the organization of a plasma membrane differ from that of the milk? Milk is made up of proteins. Plasma membrane has a phospholipid bilayer Does the dye dissolve and diffuse within the protein solution? Yes Is there any resemblance to the manner by which particulate material might pass from the outside into the cell’s interior? Explain. Egg albumen and oil served the same function as the plasma membrane in the cell as a lipid-protein membrane allowing nigrosine to diffuse from a region of high concentration to low. However, in a living cell, equilibrium is never attained. Explain this phenomenon. Equilibrium is a steady state achieved when there are no further net movements of moleculesvia any process. Thus, it is only achieved when a cell is dead. Compare results with those obtained in distilled water. Distilled water – hemolysis – hypotonic solution 10% NaCl – crenation – hypertonic solution 0.9% NaCl – normal – isotonic solution...