Exercise 6. Permeability Properties I.

Artificial Membranes Introduction It is important to recognize that body fluids contain solutes, such as proteins, that are charged and non-permeating. The very presence of such proteins can influence movement and concentrations of inorganic solutes inside and outside of cells. This experiment deals with how biomolecules affects the permeability properties in artificial membranes and how such effects relate to membrane potentials in living cells.

Objectives At the end of this exercise, the student should be able to: 1. Discuss the concept of Gibbs-Donna n equilibrium 2. Relate the above concept to the membrane potential of living cells. Materials Required 2 droppers 1 250-ml beaker 2 50-ml beakers 2 10-ml test tubes 1 10 microliters micropipettor with tips Stove or hot plate Wire gauze Stirring rod 50-ml graduated cylinder 1-ml or 5-ml measuring pipette Longganisa skin 3 to 5 grams gelatin powder distilled water 2.9% silver nitrate solution 2 g sodium chloride string triple beam balance

Procedure 1. Prepare the 5% gelatin solution by dissolving 3 grams of gelatin powder in approximately 60 to 80 ml water. Boil the mixture for 5 minutes while stirring. Allow the solution to cool. If the mixture should solidify, add some water and reheat. 2. Prepare the sodium chloride solution by dissolving 0.20 g NaCl in 50 ml distilled water. Label this as Solution A. In another beaker dissolve 0.20 g NaCl in 50 ml of gelatin solution. Label this as Solution B. 3. Form a watertight diffusion bag using the longganisa skin. Make sure that there are no leaks. Fill one of the diffusion bags with 25 ml solution B. Tie the end of the diffusion bag tightly with a piece of string. As much as possible, do not allow any airspace inside the bag for the fluid to move around. 4. Immerse the diffusion bag filled with solution B into the beaker containing solution A. Make sure that the tie ends of the bag are not immersed to prevent any fluid entry into the bag. Leave this set up for about half an hour.

5. After 30 minutes, remove the diffusion bag and empty the contents into a clean beaker and label this as solution BB. 6. Pipette 0.5 ml of solution A into a clean test tube. A micropippetor may be used to add 50 microliters of potassium chromate solution. 7. Add 25 microliters portions of 2.9% silver nitrate solution, shaking the solution continuously until the bright yellow solution changes to permanent brown hue. Record the total amount of silver nitrate added, and compute for the moles of chloride ions present in the solution. 8. Repeat step 7 for solution BB. Make a diagram of your set up in your data sheet and show your results in tabulated form.