Photosynthesis Part I
TA: Deepti
Lab 5: Photosynthesis Part I: Spinach Leaf Disc Assay
Spectrum of visible light: _____________ nm to ______________ nm
Part A: Preparation of equivalent light transmittance solutions at 50% transmittance
The final dye solution prepared will be for the whole class and shared.
Prepare dye solution using 150 ml of sodium bicarbonate solution in a large beaker and specific
dye (recommendations provided by TA- Instructions on the TA table)
Set the wavelength specified by TA for maximum transmittance for that specific dye solution
(Instructions for the suggested wavelengths on the TA table)
Recheck calibration of Spec 20 using the blank
Take the prepared dye solution and measure the transmittance
Now add more dye or bicarbonate to make the dye solution have a transmittance of 70%
To adjust transmittance add dye or sodium bicarbonate
To decrease transmittance you need to add more ___________
To increase the transmittance you need to add more ____________
The dye solution should have a transmittance of 50%.......This is your final dye solution.
Part B: Experimental Set-Up
Keep everything in the drawer and in the dark. Keep the lights on the table turned off.
Beaker 1: sodium bicarbonate (control/ comparison): 20 ml
Color Experiment: Use beakers with the same diameter
Beaker 2: blue dye solution: 20 ml
Beaker 3: green dye solution: 20 ml
Beaker 4: yellow dye solution: 20 ml
Beaker 5: red dye solution: 20 ml
Drug Experiment
Beaker 6: DCPIP solution: 10 ml bicarb + 1 drop/ ml DCPIP (10 drops)
Beaker 7: Diuron or DBMIB solution: 10 ml bicarb + 1 drop/ml Diuron (10 drops) or 1.5drop/ml
DBMIB (15 drops)
Beaker 8: Combination of DCPIP and drug used in beaker above (diuron or DBMIB):
10 ml bicarb + 1 drop/ ml DCPIP (10 drops) + diuron (10drops) or DBMIB (15 drops)
Beaker 9: sodium bicarbonate (control/ comparison): 20 ml (if possible). Put that in the dark.
Stir the solutions and keep in dark in the drawer.
Photosynthesis Part I
TA: Deepti
Part C: Spinach leaf discs:
You need 5 discs per beaker. A few may not be good to use……so cut 8 per beaker.
Cut out discs using the punch provided (Do not keep the spinach leaves outside…they wilt and
then cannot be cut)
Put the discs immediately into a Petri dish with bicarb.
When all the discs have been cut…..then transfer them all into a syringe with bicarb in it. Do not
damage the discs. Use a spatula provided to transfer the discs.
Now insert the plunger and point the syringe skyward and push out all the air.
Put your finger over the syringe tip and pull back on the plunger to create a vacuum within the
syringe. Pull back a little at a time so as not to damage the discs.
While still under vacuum remove your finger from the syringe tip. As the vacuum is released, the
leaf discs fill with the bicarb buffer.
Repeat until all the discs sink in the buffer.
Pour the discs into a Petri dish
Part D: Final Set-Up
Now transfer a fixed amount of discs in each beaker (5 or 6). You may need to tap the discs for
them to sink (surface tension of the water)
Turn on the light on the table
Immediately now put all the beakers under the light.
Note the time
Part E: Observing the photosynthetic efficiency in different colors of light and in the
presence of different drugs
Note the number of discs that rise to the surface.
Time Started:
Number of discs that rise to the surface
1 min
2 min
3 min
4 min
5 min
10 min
Bicarb buffer
Blue dye
Green dye
Yellow dye
Red dye
DCPIP
DCPIP +
PART E: Clean Up
Wash out all the beakers and wash the spinach discs down the drain
Dispose the herbicide treated solutions in marked containers in the hood
15
min
20 min