Question

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PRE-LAB 6 The Hill Reaction Pre-laboratory Assignment Directions: is not complete at the beginning of lab, the student will be unable to participate in Questions: TYPE the answers to the following questions and turm in BEFORE lab begins. If pre-lab the lab session. 1.(2pts) While doing this experiment, it will be important to keep everything on ice. Why is it important to keep everything cold? What might happen if the proteins are warm temperature? ed to room 2. (2pts) While doing this experiment, you will be centrifuging ("spinning") your samples at 1000xg or 10,000 rpm. What does 1000xg really mean? What does 10,0000rpm really mean? Can you convert xg to rpm? (Is rps ALWAYS 10 times the xg?) 3. (2pts) Photosynthesis is a redox reaction. In the light-reaction of photosynthesis, what molecule is oxidized? In the dark-reaction (Calvin Cycle) of photosynthesis, what molecule is reduced? 4. (2pts) In the Hill Reaction (not photosynthesis), what would you expect to happen to absorbance when the test tube is exposed to light? Would it increase, decrease, or stay the same? Why? 5. (2pts) In the Hill Reaction (not photosynthesis), what would you expect to happen to absorbance if the test tube is NOT exposed to light? Would it increase, decrease, or stay the same? Why?

Answer 1

1.Keeping the temperature reduced will first of all reduce the chances of contamination by bacteria or other microbes.

Protein and enzyme denaturation starts when proteins are warmed.Because enzymes or proteins work at certain tempertaure and pH .Disturbing that range would disrupt their activitiy.

2.Relative Centrifugal Force(RCF) is measured in multiples of the standard acceleration due to gravity at the Earth's surface (x g).RPM on the other hand is revolutions per minute .RPM and RCF have following relationship-

g = (1.118 x 10^-5)R S^2.

where g is the relative centrifugal force ,R is the radius of the rotor in cm and S is the speed of the centrifuge in revolutions per minute .

3.In the light reaction  Twhich takes place in the thylakoid discs. Here, water (H₂0) is oxidized, and oxygen (O₂) is released. In the dark reaction CO2 is reduced to glucose.

4.The Hill reaction is formally defined as the photoreduction of an electron acceptor by the hydrogens of water, with the evolution of oxygen. In vivo, or in the organism the final electron acceptor is NADP⁺. We can measure the rate of the Hill reaction in isolated chloroplasts. This procedure uses a dye as an artificial electron acceptor that changes color as it is reduced. DCIP (2,6-dichlorophenolindophenol) is a dye which is blue in its oxidized form and colorless in its reduced form.   Since the DCIP will begin to revert to its oxidized (blue) state as soon as the chloroplasts in the reaction vessel are removed from the light path it is essential that all absorbance readings be taken as quickly as possible.  

Now when the test tubes are exposed to light oxidation of chlorophyll takes place.Since DCIP is a deep blue color, it should absorb red wavelengths .Since we are measuring absorption of a deep blue substance that will lose color during the assay, absorption will decrease with time. Color change will begin immediately when chloroplasts are added to the DPIP solution.