## 3 Results and Discussion
### Analysis of Total Biomass and Lipid Content under Red Light Shift Treatment  
The biomass of _P. tricornutum_ was significantly greater with RS treatment by 2-fold and 1.4-fold compared to W and R conditions, respectively (Figure 3A). This correlates with increased cell density (Figure 2). In addition, our results confirm observations from another study on the impact of red light where biomass increased by 40% under enriched red light [40]. Similarly, another study reported better growth and higher fucoxanthin content in _P. tricornutum_ under red and blue light [30]. That study is similar to our treatment RS, where we have supplied a red wavelength for the first 7 days and then exposed the culture to the white light during the stationary phase. Furthermore, it has been reported that cultured _P. tricornutum_ under R contains a greater number of thylakoids, where light-dependent reactions occur, compared to _P. tricornutum_ cultured in W [41]. On a wider scale, the impact of  
Figure 2: Growth curve of _P. tricornutum_ in continuous white light (W), red light (R), and in red light shift (RS) (i.e., red light during the exponential phase and then white light during the stationary phase). Cells were grown on continuous shaking mode in 5 mL culture; each point represents the mean optical density at 680 nm of three independent experiments (\(\pm\)SD). red light on the ultrastructure of chloroplasts and creating superstacks of thylakoid membrane in diatoms suggest that R will affect photosynthesis, cell division, and consequently promote biomass accumulation [41]. Altogether, the data suggest that the use of red light and variations thereof improves the overall biomass of _P. tricornutum_ at the laboratory scale, offering a promising first stage, which should be pursued at a larger scale to be considered a useful strategy for industrial applications. Total lipids were also analyzed in the same conditions, using dry weight estimation. Nile red assay was used for rapid screening of lipid and confocal microscopy for visual analysis, as described previously [20]. _P. tricornutum_ produces lipids that are potentially significant for several industrial applications. It has been reported that the crude lipids extracted from _P. tricornutum_ are around 321.89 mg/g dry weight [42] or 34% of dry weight [43] and are comparable to the lipid extracted in W in our study, which is 0.4 g/g dry weight in control white light. Furthermore, the lipid dry weight in RS was significantly increased to 2.3-fold and 1.8-fold as compared to W and R, respectively (Figure 3B). It is interesting to note the 1.8 fold increase in lipid from R to RS, which suggests that shifting from red light to full spectrum during the stationary phase has an impact on lipid accumulation. It is known that the usual change in the lipid composition is influenced by the cellular C/N ratio, light intensity, nutrient depletion, and other possible inducible biotic/abiotic stress conditions [28; 29; 30]. An increase in C/N ratio, lipid, and carbohydrate in red light acclimated _P. tricornutum_ culture has been reported [11]. The same study reported an increase in the lipid content from red light to blue light shift, whereas the reverse shift, from blue to red light, promoted an increase in carbohydrates. In addition, lipid accumulation under red light has been observed in _Botryococcus_ sp. [7] and _Chlorella_ sp. [29], which can be linked with effective nitrogen consumption rate and a doubling of cell numbers in red light, which aids in increasing the total lipid content. In addition, high light intensity has been shown to affect lipid biosynthesis in _Nannocloropsis gaditana_, which has been correlated with the regulation of cytosolic fatty acid synthase of type 1 (_FAS1_) [44] and polyketide synthase (_PKS_) and the down-regulation of the chloroplast fatty acid synthase of type 2 (_FAS2_) [45]. One of the studies has reported a significant increase in lipids in _P.