Investigating the factors that affect the rate of photosynthesis of submerged cotyledon (seed leaf) discs

Introduction:

Photosynthesis is the trapping of carbon dioxide and its subsequent reduction to carbohydrate, using hydrogen from water. Hexose sugars and starch are commonly formed so the following equation is often used:

light energy

6CO2 + 6H2O C6H12O6 + 6O2

carbon dioxide water chlorophyll carbohydrate oxygen

Photosynthesis consists of two stages: These are the light-dependent reactions, for which light energy is necessary, and the light independent reactions

series of light-dependent reactions that are temperature-independent and a series of temperature-dependent reactions that are light-independent. The rate of the first series, called the light reaction, can be increased by increasing light intensity (within certain limits) but not by increasing temperature. In the second series, called the dark reaction, the rate can be increased by increasing temperature (within certain limits) but not by increasing light intensity

Photosynthesis involves the production of oxygen, and the uptake of carbon dioxide. These can be used as a measure of the rate of photosynthesis.

This experiment will be an investigation into the photosynthetic rate, the rate at which plants produce all their own organic substances (food) using only light energy and simple inorganic substances. The rate of photosynthesis is dependent on environmental factors, particularly the amount of light available, the level of carbon dioxide and the temperature. The effect of these factors can be tested experimentally by altering one of these factors while holding others constant.

We know that light intensity, carbon dioxide concentration and temperature are three factors that can determine the rate of photosynthesis. If the level of one of these factors is changed, rate of photosynthesis changes. Normally, only changes to one of the factors will affect the rate of photosynthesis in a plant at a time. This is the factor that is nearest to its minimum, the limiting factor. Changing the limiting factor increases or decreases the rate, but changes to the other factors have no effect. This is because photosynthesis is a complex is a complex involving many steps. The overall rate of photosynthesis in a plant is determined by the rate of whichever step is proceeding most slowly at a particular time. This is called the rate-limiting step. The three limiting factors affect different rate-limiting steps.

Carbon dioxide concentration:

At low and medium CO2 concentrations, the rate-limiting step in the Calvin cycle is the point where CO2 is fixed to produce glycerate 3-phosphate. Ribulose bisphosphate (RuBP) and NADPH accumulate. At high CO2 concentrations some other factor is limiting.

Light intensity:

At low light intensities, there is a shortage of the products of the light-dependant reactions - NADPH and ATP. The point where glycerate 3-phosphate is reduced is the rate-limiting step in the Calvin cycle. At high light intensities some other factor is limiting.

Temperature:

At low temperatures, all of the enzymes that catalyse the reactions of the Calvin cycle work slowly. NADPH accumulates. At intermediate temperatures, some other factor is limiting. At high temperatures, RuBP carboxylase does not work effectively, so the rate-limiting step in the Calvin cycle is the point where CO2 is fixed. NADPH accumulates.

Research question: How does altering the carbon dioxide concentration affect the rate of photosynthesis?

Prediction:

I predict that the rate of photosynthesis is positively related with CO2 concentration. I predict that increasing the CO2 concentration will increase the rate of photosynthesis proportionally to it. Applied to my experiment, that means the higher the sodium hydrogen carbonate solution concentration (which provides the CO2 for the cress disks in order to photosynthesize), the faster oxygen is produced as a waste product and the faster the cress discs rise to the top of the beaker.

Variables

The independent variable:

Carbon dioxide concentration: Sodium hydrogen carbonate solution was used to provide the CO2 for the cress disks in order to photosynthesize. In the experiment different NaHCO3 concentrations were made up using pure NaHCO3 solution and distilled water, always making up 60cm3 of liquid.

Controlled Variables:

Light intensity: A bench lamp was used as the light source in the experiment. It was always constantly placed 5cm above the liquid surface so that the same amount of light falls on the beaker with the cress disks inside the hydrogen carbonate solution for every experiment.

Temperature: The temperature should stay constant for all of the carried out experiments, room temperature of about 20°C being the case.

Dependent variable:

Amount of oxygen as a waste product of photosynthesis produced:

In the experiment we didn't measure the amount of oxygen produced directly. Instead a stop watch was used to monitor how long it took for the individual cress disks to rise to the top of the beaker. Being the dependent variable, the time it took for cress disks to rise at different NaHCO3 concentrations, was the recorded data. In the experiment the NaHCO3 solution enters the air spaces of the cress disks. As it provides the cress disks with the CO2, say the higher the NaHCO3 concentration meant the more oxygen is produced as a waste product of photosynthesis. The quicker the cress disks rose to the top of the beaker means that more oxygen is produced during photosynthesis.

Apparatus

1 Plastic straw

3 Small Beakers (100cm")

2 Plastic Syringes (20cm")

1 Plastic Syringe (10cm")

1 stopwatch

1 Thermometer

1 cress plant

Method

1. Use a plastic straw to punch out 5 discs from the cotyledons of the cress plants provided. Do this by placing a finger underneath the cotyledon to support it. Keep the discs in the straw for the moment.

2. The plunger from a 10cm3 plastic syringe was removed. Then I placed a finger over the nozzle and approximately half-filled the syringe with sodium hydrogen carbonate solution. Gently blow the discs from the straw into the syringe. I replace the plunger, inverted the syringe and then pushed the plunger up far enough to expel the air from the syringe.

3. Place a finger over the nozzle of the syringe and gently pull out the plunger a short distance (past a 3cm3 distance on the syringe barrel). This procedure was done in order to pull the air out of the air spaces of the cress discs and replace it with the surrounding solution. I had to hold the plunger at this position for a few seconds to make sure the air really gets pulled out the discs air spaces, and then removed my finger from the nozzle.

4. Repeat this procedure twice more. Tap the syringe barrel between each evacuation. At this stage the discs should sink. If this does not happen repeat the evacuation process.

5. Use pure sodium hydrogen carbonate solution and distilled water in order to always make up 60cm3 liquid of different NaHCO3 concentrations Using two 20cm3 plastic syringes for each the pure sodium hydrogen carbonate solution and the distilled water, the needed quantities are given into a beaker to make up the wanted NaHCO3 concentrations.

6. Then the syringe plunger was removed and the contents are tipped into the beaker.

7. Check that the discs have sunk to the bottom and if not remove and discard those ones. A minimum of 4 sunken discs is required in order to have sufficient data for the experiment.

8. The beaker with the contents is placed immediately under a bench lamp. This has to stand not more than 5 cm above the solutions surface but so that it still allows one to see the discs.

9. The bench lamp is turned on and the stopwatch is started. The discs are observed until they have all risen to the surface. The time in seconds it takes for each disc to rise is recorded.

10. This procedure is repeated using different concentrations of sodium hydrogen carbonate solution.

Results

The following tables show the amount of time (in seconds) it took for the discs under different NaHCO3 concentrations to rise to the water surface. Every experiment was done twice in order to account for reliability of the first results.

Time taken in seconds for discs to rise in 10 cm" of NaHCO3

10 cm" pure NaHCO3 solution + 50 cm" H2O 1st reading per seconds 2nd in seconds Average in

seconds

disc 1 524 502 513.0

disc 2 592 575 583.5

disc 3 642 618 630.0

disc 4 680 658 668.5

disc 5 767 698 732.5

Time taken in seconds for discs to rise in 20 cm" of NaHCO3

20 cm" pure NaHCO3 solution + 40 cm" H2O 1st

experiment in seconds 2nd in seconds Average in

seconds .

disc 1 387 362 374.5

disc 2 428 401 414.5

disc 3 470 452 461.0

disc 4 502 486 494.0

disc 5 544 587 565.5

Time taken in seconds for discs to rise in 30 cm" of NaHCO3

30 cm" pure NaHCO3 solution + 30 cm" H2O 1st experiment in seconds 2nd in seconds Average in

seconds

disc 1 265 282 273.5

disc 2 279 336 307.5

disc 3 360 398 379.0

disc 4 446 458 452.0

disc 5 507 532 519.5

Time taken in seconds for discs to rise in 40 cm" of NaHCO3

40 cm" pure NaHCO3 solution + 20 cm" H2O 1st experiment in seconds 2nd in seconds . Average in

seconds

disc 1 313 301 307.0

disc 2 354 360 357.0

disc 3 378 401 389.0

disc 4 428 476 452.0

disc 5 487 498 492.5

Time taken in seconds for discs to rise in 50 cm" of NaHCO3

50 cm" pure NaHCO3 solution + 10 cm" H2O 1st experiment in seconds 2nd in seconds Average in

seconds

disc 1 263 282 272.5

disc 2 287 299 293.0

disc 3 343 318 330.5

disc 4 398 419 408.5

disc 5 456 443 449.5

Time taken in seconds for discs to rise in 60 cm" of NaHCO3

60 cm" pure NaHCO3 solution 1st experiment in seconds 2nd in seconds Average in

seconds

disc 1 259 245 252.0

disc 2 265 271 268.0

disc 3 303 288 295.5

disc 4 313 302 295.5

disc 5 319 311 315.0

Table showing 1/time for the different NaHCO3 solutions (CO2 concentration)

1st run

Concentration of CO2 (NaHCO3 solution in cm3) 1 / time it takes for disc 1 to rise to surface in seconds 1 / time it takes for disc 2 to rise to surface in seconds 1 / time it takes for disc 3 to rise to surface in seconds 1 / time it takes for disc 4 to rise to surface in seconds 1 / time it takes for disc 5 to rise to surface in seconds Average of 1 / time for the 5 disks

0 0 0 0 0 0 0

10 0.0019084 0.00168919 0.00155763 0.00147059 0.00130378 0.00158592

20 0.00258398 0.00233645 0.00212766 0.00199203 0.00183824 0.00217567

30 0.00377358 0.00358423 0.00277778 0.00224215 0.00197239 0.00287003

40 0.00319489 0.00282486 0.0026455 0.00233645 0.00205339 0.00261102

50 0.00380228 0.00348432 0.00291545 0.00251256 0.00219298 0.00298152

60 0.003861 0.00377358 0.00330033 0.00319489 0.0031348 0.00345292

Table showing 1/time for the different NaHCO3 solutions (CO2 concentration)

2nd run

Concentration of CO2 (NaHCO3 solution in cm3) 1 / time it takes for disc 1 to rise to surface in seconds 1 / time it takes for disc 2 to rise to surface in seconds 1 / time it takes for disc 3 to rise to surface in seconds 1 / time it takes for disc 4 to rise to surface in seconds 1 / time it takes for disc 5 to rise to surface in seconds Average of 1 / time for the 5 disks

0 0 0 0 0 0 0

10 0.00199203 0.00173913 0.00161812 0.00151976 0.00143266 0.00166034

20 0.00276243 0.00249377 0.00221239 0.00205761 0.00170358 0.00224596

30 0.0035461 0.00297619 0.00251256 0.00218341 0.0018797 0.00261959

40 0.00332226 0.00277778 0.00249377 0.00210084 0.00200803 0.00254053

50 0.0035461 0.00334448 0.00314465 0.00238663 0.00225734 0.00293584

60 0.00408163 0.00369004 0.00347222 0.00331126 0.00321543 0.00355412

Table showing 1/time for the different NaHCO3 solutions (CO2 concentration)

Average

Concentration of CO2 (NaHCO3 solution in cm3) 1 / time it takes for disc 1 to rise to surface in seconds 1 / time it takes for disc 2 to rise to surface in seconds 1 / time it takes for disc 3 to rise to surface in seconds 1 / time it takes for disc 4 to rise to surface in seconds 1 / time it takes for disc 5 to rise to surface in seconds Average of 1 / time for the 5 disks

0 0 0 0 0 0 0

10 0.00194932 0.0017138 0.0015873 0.00149589 0.00136519 0.0016223

20 0.00267023 0.00241255 0.0021692 0.00202429 0.00176835 0.00220892

30 0.00365631 0.00325203 0.00263852 0.00221239 0.00192493 0.00273684

40 0.00325733 0.00280112 0.00257069 0.00221239 0.00203046 0.0025744

50 0.00366972 0.00341297 0.00302572 0.00244798 0.00222469 0.00295622

60 0.00396825 0.00373134 0.00338409 0.00338409 0.0031746 0.00352848

Comments on results

As one can see from the tables, in general the time it takes for the cress discs to rise to the surface of the beaker decreases as the concentration of NaHCO3 increases. In average, when 10cm3 of NaHCO3 where used, disc 1 rose after 513.0 seconds, disc 5 after 732.5 seconds. When 60cm" of NaHCO3 where it took only 252.0 seconds for disc 1, and 315.0 seconds for disc 5 to rise. The basic pattern that as NaHCO3 concentration increases the time for the discs to come to the top decreases can be seen.

This would mean that more oxygen is produced by the leaf discs as NaHCO3 concentration increases, and as a result the time to rise to the top decreases. As oxygen production can be used as a measure of photosynthesis, in taking the time of how long it takes for the discs to come to the surface, we get values that are proportional to the oxygen production. One can therefore calculate the rate of photosynthesis by dividing 1 over the average time it took for the discs to rise to the top. From the results table above the general pattern that the rate of photosynthesis increases as the concentration of CO2 (NaHCO3) is raised.

At a CO2 concentration when 10cm3 NaHCO3 are used the rate of photosynthesis is 0.0016223. At a CO2 concentration when 60cm3 NaHCO3 are used the rate of photosynthesis is 0.00352848.

However the table shows that at a CO2 concentration when 40cm3 NaHCO3 were used the average rate is 0.0025744 which is lower than at 30cm3 NaHCO3 which gave a rate of 0.00273684. This explains the little dink in the graph at the CO2 concentration of 40cm3 NaHCO3 before the line continues to go up again from a CO2 concentration of 50cm3 NaHCO3.

The graph very well displays that the rate of photosynthesis increases fairly quickly as the CO2 concentration is increased and that the line is starting to level off at higher CO2 concentrations.

Analysis

My results clearly show that the average time it takes for the cress discs to rise decreases as the concentration of NaHCO3 increases. This is because the higher the CO2 concentration the higher the rate of photosynthesis. As a result of the photosynthetic rate increasing, the production of oxygen as a waste product of photosynthesis increases as well. As the cress discs produce oxygen quicker at higher CO2 concentrations that means the oxygen will make the discs rise quicker. With more oxygen produced the buoyancy of the cress discs increases and this leads to the discs floating to the top of the beaker. With the discs rising faster and indicating that the amount of oxygen produced increases with higher CO2 concentration, will mean that the rate of photosynthesis increases as well.

My results support this statement, too. My graph shows that at low to fairly high CO2 concentrations the rate of photosynthesis is positively correlated with CO2 concentration. This implies for my graph except for the CO2 concentration of 40cm3 which is therefore clearly an anomalous result. The graph also shows that at high CO2 concentrations the rate of photosynthesis is slowing down and moving towards a plateau. This is because at high CO2 concentrations there is some other factor limiting the rate of photosynthesis.

Conclusion

In general my results support my hypothesis that the rate of photosynthesis is positively related with CO2 concentration. My results from experiment have shown that at higher CO2 concentrations more oxygen is produced by the cress discs. This oxygen will make them rise to the surface more quickly as the leaf discs buoyancy increases.

My results table and graph show that at constant light intensities and temperature, the rate of photosynthesis initially increases with an increasing concentration of carbon dioxide, but is starting to reach a plateau at higher concentrations. At low concentrations of carbon dioxide, the supply of carbon dioxide is the rate-limiting factor. At higher concentrations of carbon dioxide, other factors such as light intensity and temperature are rate limiting.

The rate of photosynthesis is determined by the rate-limiting step which is the step that is proceeding most slowly at a time. At low to medium CO2 concentrations, the rate-limiting step in the Calvin cycle is the point where CO2 is fixed to produce glycerate 3-phosphate. RuBP and NADPH accumulate. The plateau on my graph however shows that at higher CO2 concentrations some other factor is limiting, meaning either light intensity or temperature are too low for the rate of photosynthesis to increase further.

To the extent that the rate of photosynthesis increases as the rate of CO2 concentration increases, my prediction overlaps with my results. However my results have also shown me the fact that this relationship doesn't continue like that forever. Having done this experiment, has shown me that the rate of photosynthesis increases with increasing CO2 but is limited by the factor which is nearest to its lowest value.

Accuracy of observations

In general, the accuracy of the equipment is very good, however, for each of them there is some element of inaccuracy in terms of the readings to be made - this also includes the human element in making the reading. For the most accurate results, the reading has to be made with the scale being on eye level.

The 20cm" plastic syringes have an accuracy of + 0.5cm3; the beakers as well show an accuracy of + 0.5cm3. The 10cm3 syringes however are accurate to + 0.25cm3

Thus for all the solutions we have to assume that the maximum error of the readings made could be + 1.25cm", which is very important.

Improvements to method:

Even though, the method could be improved still. One issue for instance that could be used to slightly improve the method and thus the accuracy of the results would be to use a water bath in order to make sure that the experiment is conducted under generally stable conditions as for this will ensure that the temperature stays the same throughout the whole experiment. This would be of great importance for conducting an experiment which implies of temperature being one of the limiting factors. Using a water bath one could also set up the experiment in a way that the oxygen produced as waste product of photosynthesis could be directly collected under water.

Evaluation and anomalous results

In general the method wasn't changed much to the preliminary work. However, in my preliminary work I first used a total amount of 100cm3 NaHCO3 solution to make up the different NaHCO3 concentrations. This showed that when low NaHCO3 concentrations (little pure NaHCO3 being used) were used, it took more than 10minutes for the cress discs to rise to the surface of the beaker. This is simply too long and wouldn't have given me enough time to do sufficient repeats of the experiment. That's why I decided to reduce the total amount of NaHCO3 solution used to 60cm3.

I think that my results have shown that a general pattern can be seen. The collected data, illustrated in tables and diagrams backs up my prediction. However the reliability is not too strong, as significant differences can be seen in the time taken for the discs to rise, especially at a NaHCO3 concentration of 40cm3.

Throughout the experiment the same plastic straw was used to cut discs out from cress leaves. The plastic straw had a diameter of about 4mm, meaning that all cress discs had the same diameter throughout the investigation. However other variations in the size of the cress discs could have occurred. For example the thickness of the cress leaves might have varied, resulting in thicker and thinner and cress discs between experiments. These structural differences might have accounted for the anomalous results but also for the significant differences in the time it took for the discs to come up to the surface, between run 1 and 2 of the same concentration.

A factor that wasn't particularly controlled at all was temperature. The light source of the investigation was a simple bench lamp. As I have observed the light bulb got really hot after a few minutes of usage. This would mean that at the beginning, namely the very first concentration of the first experiment wouldn't have been affected by the light bulb. However by the time the next concentration was used the light bulb was already hot. This would have resulted in the heat that given off by the light bulb to increase the temperature of the NaHCO3 solution the seed discs were floating in. Even further as the time it took for discs to rise varied with different NaHCO3 concentrations means that the time the NaHCO3 solution exposed to the heat given off by the light bulb varied, too. This means the temperature of the different NaHCO3 solutions must have varied as well. As temperature is one of the limiting factors of photosynthesis this could have had significant effects on the experiment. Temperature being higher at some NaHCO3 concentrations means that the reaction of photosynthesis must have taken place faster, resulting in a faster production of oxygen, meaning that the cress discs rose to the surface quicker.

In addition there are some anomalous results found in the graph. However, the graph shows a trough at a CO2 concentration of 40cm" NaHCO".

This should not be the case; the line should go up further and then level off properly. My graph however doesn't show a clear plateau which should be seen when high CO2 concentrations have been reached as slowly no more oxygen can be produced in the same time. Optimally at low to fairly high CO2 concentrations the graph should show that the rate of photosynthesis is directly proportional to CO2 concentration.

These factors could be down to the fact mentioned earlier that the cress discs might have been of different structures, e.g. thicker and therefore affected the experiment.

However I think that one also has to consider the fact that the experiment involved living organisms. Just like human beings, plants don't always act in an expected way. This is what essentially makes biology interesting in that the expected is not always happening.

In doing this experiment we were measuring the time it took for cress discs to rise to the surface of a beaker at different NaHCO3 concentrations. In doing so we were effectively trying to collect data, namely time, which is proportional to the production of oxygen of the seed discs in order to get information about the rate of photosynthesis. The seed discs producing more oxygen meant that they would rise faster. Oxygen being a by-product of photosynthesis can be used to get a picture of the rate of photosynthesis of a plant. So ideally an experiment carried out measure the amount of this oxygen production would be better designed to get an indication of the rate of photosynthesis of a plant. The oxygen collection would take place in water; a water bath could be used for example.