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Use of a redox indicator to show dehydrogenase activity.

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Introduction

Use of a redox indicator to show dehydrogenase activity Introduction: Triphenyl tetrazolium chloride (TTC) is an example of an artificial hydrogen acceptor, or redox indicator. TTC is colourless when oxidised, but forms red, insoluble compounds called formazans when reduced. TTC can therefore be used to show the presence of active dehydrogenase enzymes by a colour change. This experiment investigates the effect of temperature on the activity of dehydrogenases in yeast cells. Materials: * Actively respiring yeast suspension. This should be prepared by adding 100g of dried yeast to 1dm3 of water and mixing in 50g glucose. This mixture should be allowed to stand in a large beaker for about 2 hours before the experiment. * Triphenyl tetrazolium chloride solution, 0.5% * Distilled water * Test tubes and rack * Graduated pipettes, or syringes * Glass rods * Crushed ice * Beakers or water baths * Thermometer * Stopwatch Method: 1. Set up water bath at 30?C. 2. Pipette 10cm3 of yeast suspension into one test tube and 1cm3 of TTC solution into another test tube and stand them both in the water bath. ...read more.

Middle

The effect of a 10?C rise in temperature on the rate of a reaction can be expressed as the Q10 value. This is a ratio of the rate of a reaction between two temperatures. These are the Q10 values for the temperatures used in this experiment. Between the range 20?C and 30?C = 0.002176 ? 0.001147 = 1.897 Between the range 30?C and 40?C = 0.003205 ? 0.002176 = 1.473 Between the range 40?C and 50?C = 0.004782 ? 0.003205 = 1.492 Between the range 50?C and 60?C = 0.006359 ? 0.004782 = 1.330 Between the range 60?C and 70?C = 0.007937 ? 0.006359 = 1.248 Between the range 70?C and 80?C = 0.007622 ? 0.007937 = 0.960 Between the range 80?C and 90?C = 0.007308 ? 0.007622 = 0.959 Between the range 90?C and 100?C = 0.006993 ? 0.007308 = 0.957 One source of error in this experiment was that not all the yeast suspension was always transferred into the test tube of TTC solution. This meant that different amounts of yeast suspension could have been actually used for each of the temperatures. ...read more.

Conclusion

The overall accuracy of the experiment could have been improved if the whole experiment was repeated two or three times and an average for each temperature taken. In organic molecules oxidation involves the removal of both electrons and protons. This process is called dehydrogenation. Dehydrogenases are enzymes that catalyse these reactions. In cell respiration, dehydrogenases are used in the first part of the breakdown of glucose (glycolysis). The two trioses produced after the splitting of fructose-1,6-biphosphate undergo dehydrogenation. The hydrogen atoms produced by this reaction are taken up by NAD+ to from NADH + H+. The electrons then pass along a series of electron carriers, making up the electron transport system. This electron chain then results in ATP synthesis, which is of course the point of respiration. The role of dehydrogenases is also seen in the Krebs cycle. Four of the steps in the Krebs cycle involve the removal of pairs of hydrogen atoms, which are catalysed by NADH dehydrogenase or succinate dehydrogenase. In the same way, the NADH + H+ goes on to the synthesis of ATP from ADP and Pi. Neal Desai Mr. Cooksey U6AA Biology ...read more.

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4 star(s)

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Good biochemical knowledge demonstrated but a little more attention to detail is needed for a 5 star rating. Some discussion of the effects of temperature on enzyme activity would be relevant and helpful.

Marked by teacher Adam Roberts 18/07/2013

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