(602d) Effect of Nitrogen Starvation in Chlorophyll and Carotenoids Production By Chlorella vulgaris Under Photoeterotrophic Culture

Chlorella vulgaris is a green microalga that has been extensively studied for various purposes, such as the production of biomass as a source of valuable chemicals or healthy foods, and wastewater treatment and aquaculture feed. This microalga has one of the highest chlorophyll content found in nature. The content of pigment and other compounds in C. vulgaris, like other microalgae, can be improved by nutritional stress conditions as nitrogen and phosphorus limitation and iron and magnesium deficiency. After carbon, oxygen and hydrogen, nitrogen is the most abundant element in algae without mineralized walls. Therefore, as cells grow and divide they require a supply of nitrogen. If the supply of nitrogen is limited in proportion to other elements, photosynthesis may continue but the resultant compounds will include a smaller proportion of nitrogen-rich components and more energy-rich components such as lipids. In this study the effect of nitrogen starvation on growth and pigment production by Chlorella vulgaris under photoheterotrophic culture was evaluated.

The freshwater microalga Chlorella vulgaris was used. Nitrogen starvation consisted in remove the nitrogen sources of the culture media (g L^-1: NaNO₃ 1.0, K₂HPO₄ 0.25, MgSO₄ 0.5, NH₄Cl 0.05, CaCl₂ 0.058, FeCl₃ 0.003). Three conditions were evaluated: without NaNO₃, without NH₄Cl and without either nitrogen sources. Experiments were carried out in 250 mL Erlenmeyer flask containing 150 mL of media under photoheterotrophic conditions using glucose as sole carbon source (2 g L^-1) and white cool fluorescent light as energy source (45 μmol photons m⁻² s⁻¹) at 27±2 °C for 21 days. The final cell concentration was reported as dry weight (g L^-1). Pigments were extracted using dimethylsulphoxide (DMSO) and OD was measured at 649, 665 and 480 nm to calculated pigment content.

In photosynthetic microorganism as long as nitrogen was available, the rapid chlorophyll accumulation and cell division was observed, however, when the nitrogen was consumed, cell division stopped. Biomass yield was the same when NaNO₃ (1.42 g L^-1) and NH₄Cl (1.51 g L^-1) were deprived, however when nitrogen sources are limited, the biomass decreased in 48.8 and 45.8% respectively in comparison with the complete nitrogen stress. The consumption of nitrogen by the cells resulted in a fast decrease in the level of chlorophyll in the cultures, suggesting that cells metabolize chlorophyll during periods of nitrogen stress. There was also an increase in the ratio of carotenoid to chlorophyll. This behavior was observed in cultures without NaNO₃ (0.51) and NH₄Cl (0.48) where carotenoid/chlorophyll ratio was higher in comparison with control (0.09). According to the results, nitrogen deprivation represents a feasible nutritional stress to increase carotenoids production without affecting biomass yield.