## 3 Results and Discussion
### Influence of Light on the H. pluvialis Cyst Cell Germination  
Light influences the speed of nitrate uptake, while nitrate plays an essential role in the growth and synthesis of astaxanthin [38] and cyst germination in _H. pluvialis_[25; 28; 29]. The time course of the nitrate concentrations of culture 5 to culture 8 is presented in Figure 14. As can be seen, all nitrate was nearly consumed 17 h after the start of germination in the cultures; an identical light intensity of 75 umol m\({}^{-2}\) s\({}^{-1}\) was applied to all cultures. The time course of the nitrate concentration for the third set of experiments (culture 9 to culture 12) is presented in Figure 15. In the illuminated phototrophic and mixotrophic (culture 11 and 12) nitrate is already consumed 17 h after the start of germination, while it is depleted after 25 h in heterotrophic conditions (culture 9 and 10). Similar tendencies were observed in the literature, where nitrate was consumed two days earlier in a _H. pluvialis_ cyst culture illuminated at 300 umol m\({}^{-2}\) s\({}^{-1}\) compared to 150 umol m\({}^{-2}\) s\({}^{-1}\)[25]. The faster nitrate consumption was accompanied by a trend toward the faster release of zoospores for the illuminated cultures, whereby for all cultivation conditions the maximum zoospore release was reached 49 h after the start of germination (Figure 8). As shown in Figure 16, a similar trend was also measured for the heterotrophic conditions of cultures 13 and 14, when compared to the experiments where mixotrophic and heterotrophic conditions were combined (cultures 15 and 16). These were cultivated at a light intensity of 75 umol m\({}^{-2}\) s\({}^{-1}\) until nitrate was consumed and further cultivated at heterotrophic conditions (without light). Figure 14: Time course of the nitrate concentration in the supernatant of the phototrophic culture 5 (phototrophic + CO\({}_{2}\) + 75 umol m\({}^{-2}\) s\({}^{-1}\)) and culture 6 (phototrophic + ambient air + 75 umol m\({}^{-2}\) s\({}^{-1}\)), and mixotrophic culture 7 (mixotrophic + 4 mM glucose + CO\({}_{2}\) + 75 umol m\({}^{-2}\) s\({}^{-1}\)) and culture 8 (mixotrophic + 4 mM glucose + ambient air + 75 umol m\({}^{-2}\) s\({}^{-1}\)). Figure 15: Time course of the nitrate concentration in the supernatant of the culture 9 (heterotrophic + \"no external carbon source\" + ambient air) and culture 10 (heterotrophic + 4 mM glucose + ambient air), phototrophic culture 11 (phototrophic + ambient air + 75 umol m\({}^{-2}\) s\({}^{-1}\)) and mixotrophic culture 12 (mixotrophic + 4 mM glucose + ambient air + 75 umol m\({}^{-2}\) s\({}^{-1}\)). In general, the rate of nitrate and nitrite uptake in microalgae depends on the nitrogen status of the cells, the presence of other nitrogen compounds, and light conditions [39; 40]. Nitrate enters the cell via specific transport systems, followed by a reduction to nitrite in the cytosol. Nitrite is then transported to the chloroplast where it is reduced to ammonium. Finally, ammonium is incorporated into L-glutamate through the glutamine/glutamate cycle [40] and allows the formation of amino acids and cell growth. In the absence of nitrate, supplemented carbon in the form of CO\({}_{2}\) or glucose is stored as starch, TAGs, or fatty acids. In conclusion, the nitrate uptake and formation of zoospores were accelerated in the illuminated phototrophic and mixotrophic conditions compared to the heterotrophic conditions. However, the illumination itself did not enhance the number of released zoospores, as more zoospores were released at heterotrophic conditions (culture 9 and 10) compared to the phototrophic and mixotrophic conditions (culture 11 and 12) (Figure 8).