Apparatus: ... Materials and Methods P1000 Gilson and blue gilson tips 3 disposable cuvettes Spectrophotometer parafilm stop watch tissue Chemicals: These chemicals are toxic, gloves and eye protection must be worn. All spills must be reported to the lecturer or a demonstrator and cleaned up immediately. Students must wash their hands before the laboratory is left. 50 mM sodium phosphate buffer, pH 7.5 (20ml) Distilled water 2 mM 2-nitrophenyl-ß-D-galactopyranoside (ONP-Gal) (5ml) 2 mM 2-nitrophenyl-6-glu

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Author:Steven S. Zumdahl, Susan A. Zumdahl, Donald J. DeCoste
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Chapter1: Chemical Foundations
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Problem 1RQ: Define and explain the differences between the following terms. a. law and theory b. theory and...
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Plot a Graph and Determine the Rate:

Make a graphical plot of reaction time (× axis) and average (y axis).

Determine the initial velocity (rate of the reaction by extending the initial straight-line portion of your reaction curve and calculating the gradient of the straight line (difference in y/ difference in ×).

  1.  What is the reaction rate based on your plotted values
Apparatus:
●●●●
●●●
P1000 Gilson and blue gilson tips
3 disposable cuvettes
Spectrophotometer
parafilm
stop watch
tissue
Materials and Methods
Chemicals:
These chemicals are toxic, gloves and eye protection must be worn. All spills must be reported
to the lecturer or a demonstrator and cleaned up immediately. Students must wash their hands
before the laboratory is left.
50 mM sodium phosphate buffer, pH 7.5 (20ml)
Distilled water
2 mM 2-nitrophenyl-ß-D-galactopyranoside (ONP-Gal) (5ml)
2 mM 2-nitrophenyl-ß-glucopyranoside (ONP-Glu) (5ml)
2 units.ml-¹ B-galactosidase (from E. coli) (10ml)
Method:
1. Check that the spectrophotometer is on and set at 440 nm. If it is not set up correctly ASK for
help.
3. In the second cuvette add:
beslist elven
2. Fill the first cuvette with 3 ml distilled water and zero the spectrophotometer by placing the
cuvette in the machine in the correct orientation and pressing the BLUE button. If you follow the
instructions below you will not need to repeat this.
1 ml 50 mM sodium phosphate buffer, pH 7.5
1 ml Distilled water
0.5 ml 2-nitrophenyl-ß-D-galactopyranoside (ONP-Gal)
<pause here before moving to next step>
add 0.5ml ß-galactosidase to the cuvette WHEN YOU ARE READY TO START
THE CLOCK
Mix by covering the top of the cuvette with parafilm, ensure the outside of the cuvette is dry and
place in the correct orientation (bevelled sides to the left and right) in the spectrophotometer.
Press the GREEN button to start measuring the absorbance.
4. Take your first absorbance reading at 30 seconds and continue taking readings every 30
seconds for 5 minutes, record the reading in the form of a table (for table, see next page).
5. REPEAT the experiment starting at step 3.
6. Calculate the average from the 2 experiments for each time point. Plot your data on a graph
and calculate the rate (absorbance units minute).
Rate of the reaction based on your plotted values:
Transcribed Image Text:Apparatus: ●●●● ●●● P1000 Gilson and blue gilson tips 3 disposable cuvettes Spectrophotometer parafilm stop watch tissue Materials and Methods Chemicals: These chemicals are toxic, gloves and eye protection must be worn. All spills must be reported to the lecturer or a demonstrator and cleaned up immediately. Students must wash their hands before the laboratory is left. 50 mM sodium phosphate buffer, pH 7.5 (20ml) Distilled water 2 mM 2-nitrophenyl-ß-D-galactopyranoside (ONP-Gal) (5ml) 2 mM 2-nitrophenyl-ß-glucopyranoside (ONP-Glu) (5ml) 2 units.ml-¹ B-galactosidase (from E. coli) (10ml) Method: 1. Check that the spectrophotometer is on and set at 440 nm. If it is not set up correctly ASK for help. 3. In the second cuvette add: beslist elven 2. Fill the first cuvette with 3 ml distilled water and zero the spectrophotometer by placing the cuvette in the machine in the correct orientation and pressing the BLUE button. If you follow the instructions below you will not need to repeat this. 1 ml 50 mM sodium phosphate buffer, pH 7.5 1 ml Distilled water 0.5 ml 2-nitrophenyl-ß-D-galactopyranoside (ONP-Gal) <pause here before moving to next step> add 0.5ml ß-galactosidase to the cuvette WHEN YOU ARE READY TO START THE CLOCK Mix by covering the top of the cuvette with parafilm, ensure the outside of the cuvette is dry and place in the correct orientation (bevelled sides to the left and right) in the spectrophotometer. Press the GREEN button to start measuring the absorbance. 4. Take your first absorbance reading at 30 seconds and continue taking readings every 30 seconds for 5 minutes, record the reading in the form of a table (for table, see next page). 5. REPEAT the experiment starting at step 3. 6. Calculate the average from the 2 experiments for each time point. Plot your data on a graph and calculate the rate (absorbance units minute). Rate of the reaction based on your plotted values:
Time (Minutes)
0.5
1.0
1.5
200
205
3.0
3.5
4.0
4.5
6
5.0
Average
0.1045
001745
0.2495
0.3255
0.4005
0.4695
0.54
0.6035
0.6595
0.712
Transcribed Image Text:Time (Minutes) 0.5 1.0 1.5 200 205 3.0 3.5 4.0 4.5 6 5.0 Average 0.1045 001745 0.2495 0.3255 0.4005 0.4695 0.54 0.6035 0.6595 0.712
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