Wheat Germ Acid Phosphatase

Wheat Germ Acid Phosphatase - 4-1-05The Affects of Varying...

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Unformatted text preview: 4-1-05The Affects of Varying Conditions on the Enzyme Wheat Germ Acid PhosphataseObjectives:Enzymes are important for nearly all functions in the cells of both plants and animals. Reactions that normally would take hours or days can be catalyzed by an enzyme to happen in a few seconds. To determine the rate of reaction one would measure the amount of substrate used or product generated over time. The rate of reaction which we determine can be varied by changing the reaction conditions. The reaction conditions we varied were time, temperature, substrate concentration, and enzyme concentration. To gauge the reaction progress, we used a substance called PNPP in which the absorbance changes as the product of the reaction is formed. I.Time CourseData of Absorbance for Each Time Assay with Calculation of MicromolesTubeAbsorbanceMicroMoles20.501111.340.798177.361.123249.581.384307.5101.604356.4121.86413.3The tube column has values for how many minutes the reaction was allowed to progress. The absorbance column was data taken directly from the spectrophotometer after using the blank (zero tube). The last column was calculated using a given formula with the extinction coefficient (18.8x10^3 liter mol^-1 cm^-1). 14-1-05Reaction Progression of Above Data Plotted on a GraphMicromoles vs. Timey = 66.439x - 355010015020025030035040045050024681012TimeMicro Moles of p-nitrophenolThis graph shows a plot of micromoles p-nitrophenol versus reaction time. The data points are the blue rectangulars and the blue line connecting them shows a slight trend. The red line is the actual trend line. The equation for that trend line is also given. A.) Is the time course linear throughout? If the time course is not linear, what are some factors that might contribute to the changed velocity at longer periods of time?B.) Is the 5-minute fixed-time assay valid for acid phosphatase? If not, how should it be changed?C.)Determine the slope of the graph.Our plot for the data collected yields a fairly straight line with a slight curve at the beginning and towards the end. The reason for the curve is that at around 9-10 minutes of reaction time, all of the substrate is being used up. This will cause a leveling out of the reaction progress, because after the substrate is consumed, the enzyme has no materials with which to make its products. The twelve minute assay we performed was a decent amount of time to get an accurate picture of how the reaction will plateau. If we had used a five minute assay, we would conclude from our data that this reaction would continue 24-1-05forever if we allowed. For better results we could use a twenty minute assay and take measurements from the plateau state of the graph. The initial velocity for this reaction was 35 micromoles p-nitrophenol/minute....
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This lab report was uploaded on 04/05/2008 for the course BIOL 4177 taught by Professor Pike during the Spring '08 term at East Tennessee State University.

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Wheat Germ Acid Phosphatase - 4-1-05The Affects of Varying...

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