Abstract

Diatom Thalassiosira weissflogii fluorescence
1000X O.I. Red color is the natural chlorophyll
and the blue color is the  membrane stain.

 Diatoms are very abundant in the ocean.  Diatoms fix carbon dioxide and incorporate it into their biomass. These organisms provide approximately 40% of the primary production of the earth.   Studying diatoms and their physiological response to various concentrations of carbon dioxide enables investigators to analyze carbon cycles that are involved in the global carbon cycle.  Studying diatoms and their responses to CO2 levels is important in understanding future climate. 

This investigation studied the effects of carbon dioxide concentrations on cell count, fluorescence, pigmentation, and activity of carbonic anhydrase (CA)  in diatoms.

The carbon dioxide levels used were 100 ppm, 350 ppm, and 750 ppm. These concentrations are physiological values found in nature.   These values represent an algal bloom, ocean equilibrium, and upwelling conditions, respectively.  How these cells utilize carbon at these levels is the focus of this study.  The level of 750 ppm is also the level of CO2 projected within the next 100 years.

The enzyme ribulose 1, 5-bisphosphate carboxylase-oxygenase (RubisCO) requires higher concentrations of carbon dioxide than found in the surrounding waters. This enzyme fixes carbon and limits the rate of photosynthesis. Many algae  are known to use a carbon concentrating mechanism (CCM) so that they can actively transport inorganic carbon for utilization when carbon dioxide quantities are limited. The CCM contains CA,  an enzyme  that catalyzes the reversible reaction:

It has been shown in earlier studies that the quantity of  CA in the cell is regulated by carbon dioxide. Its actual role is not well understood. It might dehydrate the bicarbonate ion to carbon dioxide in the chloroplast and then make it available for fixation by RubisCO. CA could be involved in the active uptake of bicarbonate from outside of the cell.  There may be other and varied roles in which it is involved, so continued research is needed.

 
 
 

Results and Conclusions


Data Table1.   Thalassiosira weissflogii Cell Density/Time

The cell density for the 750 ppm is erroneous due to mechanical problems.
 
Day
100 ppm CO2
350 ppm CO2
750 ppm CO2
1
 774
1022
910
3
25658
 31863
31515
6
299685
402429
295802

 
 

Graph 1.  Thalassiosira weissflogii Cell Density versus Time





Table 2.  Average Pigments Per Cell on Day 3 (x1000)
 
Pigment
100 ppm CO2
350 ppm CO2
750 ppm CO2
Chlorophyll a
4.6
6.6
6.4
Chlorophyll c
0.46
0.55
0.61
Fucoxanthin
1.5
2.0
2.1

 

All populations of the diatom Thalassiosira weissflogii grew during the six days of incubation. The population in the 350 ppm CO2 grew at the fastest rate.  The population in the 100 ppm CO2 and the 750 ppm CO2 grew at approximately the same rate.  Due to a mechanical problem, the 750 ppm CO2 did not aerate the culture properly; therefore, the values for the 750 ppm CO2 are questionable and should be repeated.

In the CA assays, the greatest amount of CA was detected in the 100 ppm CO2 environment.  This is to be expected because as CO2 levels decrease, the concentration of CA increases to catalyze the bicarbonate to carbon dioxide reaction.

In the pigment extraction,  chlorophyll a, chlorophyll c, and fucoxanthin, a carotenoid were measured.  All three pigments increased in concentration during the six days, due to populations increases.  Chlorophyll a and fucoxanthin are more abundant in the cells than chlorophyll c.  Chlorophyll a and fucoxanthin were easily separated using thin layer chromatography; chlorophyll c, because of its low concentration, was not detected using chromatography.

Day three was used to compare the amount of pigmentation per cell count.  Day one was not chosen because of the small values for both pigments and cell numbers which introduced large errors to the quotas.  Day six was not used due to the mechanical problems in the cultures which occurred over the weekend.  All cells contained relatively the same amounts of the three pigments--chlorophyll a, chlorophyll c, and fucoxanthin.  However, the total amounts of pigment per cell depended upon the culture conditions.  Cells cultured at 100 ppm have less pigment per cell than the other two concentrations.  This suggests that limiting the amount of carbon dioxide influences the amount of pigmentation.  This is a surprising result, since all of the cells showed similar growth rates.  The reason for this is unknown and more research in this area is needed.

Inquiry and Extensions
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b
The Woodrow Wilson National Fellowship Foundation
CN 5281, Princeton NJ 08543-5281 - Tel:(609)452-7007 - Fax:(609)452-0066
Technical contact: lpt@woodrow.org