Determination of the Effects of Nitrogen and Sulfur Starvationon Chlorophyll Content, Cell Density and Lipid Content in Chlamydomonas reinhardtii
Authors: Franco Yahaira , Lopez Ramiro , Ynalvez Ruby A. 
Published: April 2020

The severe environmental impacts of the use of fossil fuels has led to the development of biofuels as an alternative source of renewable green energy. Currently, algae is being studied as a source for biofuel because it does not compete for arable land and has a fast generation time. The microalgae, Chlamydomonas reinhardtii has potential for biofuel production due to its high lipid body synthesis when subjected to nutrient deprivation. The lipid bodies, predominantly triacylglycerides (TAGs) can be converted into biodiesel which could then be used as a form of biofuel. There is still a lack of research comparing the effects of nutrient starvation in strains of C. reinhardtii. The objectives of this study are: to determine the optimum time at which C. reinhardtii produces the highest amount of lipids under nutrient starvation and to compare the effects of nitrogen, sulfur and both nitrogen sulfur starvation on cell density, chlorophyll content and lipid content of strains cc125. We hypothesize that the nitrogen starvation treatment of C. reinhardtii strain cc125 will produce higher lipid synthesis and the sulfur starvation treatment will produce the highest biomass synthesis.

C. reinhardtii strains cc125 were grown to early logarithmic phase in TAP medium liquid culture under constant light on a rotary shaker. Once logarithmic phase was reached, cells were subjected to nitrogen (TAP‐N), sulfur (TAP‐S) and combined nitrogen and sulfur (TAP‐NS) deprivation over a 6 day period to promote lipid accumulation. Samples for cell density, chlorophyll content, and confocal imaging were collected for each strain and nutrient treatment every 2 days. The change in chlorophyll a, chlorophyll b, total chlorophyll and cell density at day 2, 4 and 6 from day 0 were determined. Results showed that changes in chlorophyll a, chlorophyll b and total chlorophyll among all treatments were statistically not significantly different (p>0.05). Chlorophyll a (3.02 ± 2.31), chlorophyll b (1.68 ± 0.85) and total chlorophyll (4.66 ± 2.98) were all highest in treatment TAP‐N and was lowest in TAP‐NS, (chlorophyll a, 0.34 ± 2.30; chlorophyll b, 1.12 ± 0.86; total chlorophyll, 0.6 ± 2.99). There was no statistical difference in cell density among treatments (p>0.05). There was highest increase in cell density in TAP‐S treatment while highest decrease in cell density was observed in TAP‐NS treatment. In addition, there was no statistical difference among days for all treatments in chlorophyll a, chlorophyll b, total chlorophyll and cell density. Results for biomass determination, lipid extractions and quantification; and the qualitative detection of lipid bodies by confocal microscopy will be reported. C. reinhardtii strain sta6 will also be studied. Determining which strain of C. reinhardtii and at which condition will produce high TAGs is essential for engineering and mass producing algae as an established source of biofuel production. This can lead to the adaptation of a viable fuel production system that would resolve the ever growing demand for environmentally sustainable energy resources.

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