## 3 Discussion
### Scavenging of Reactive Oxygen Species under Blue Light Irradiation  
In general, organisms synthesize 5-ALA in one of the two ways, i.e., the C4 pathway and the C5 pathway. The C4 pathway mainly exists in animals and fungi, while the C5 pathway is utilized by plants, algae, and bacteria [60]. The 5-ALA is the precursor of chlorophyll, heme, and vitamin B12 biosynthesis pathways, with chlorophyll and heme synthesis pathways sharing several intermediate products. Our results showed that expression levels were decreased in genes (i.e., _hemL_, _hemB_, _hemC_, _hemD_, and _hemF_) involved in the sharedportions between chlorophyll and heme synthesis pathways, whereas the expression of _ho_ was significantly increased under blue light conditions (Figure 8). As the cofactor of catalase and many types of peroxididases, the increase in heme content removes excessive ROS produced in cells. Although the oxidation effect of the free heme damages cells under oxidative stress, cells develop a variety of protection mechanisms [61]. For example, studies have shown that heme oxygenase is involved in the antioxidant mechanism by catalyzing the degradation of heme to produce biliverdin, which shows antioxidant effect. The other two end products derived from biliverdin, i.e., CO and Fe/FTH, have also shown cytoprotective effect [62]. Compared with white light, the expression of heme oxygenase in blue light treatment group was significantly increased, leading to the increase in heme degradation products and the production of antioxidants. However, the expression of _ho_ was decreased significantly on day 7 after the treatment with SA (Figure 8). As an important signal molecule, SA regulates a variety of oxidative stress responses to improve the efficiency of antioxidant system in plants. In _H. pluvialis_, a large amount of astaxanthin is generated to reduce the damage caused by the detrimental environment, complementing, or ultimately replacing, the antioxidant system of the microalgae. Therefore, _H. pluvialis_ protects itself from the adverse effects of stress by accumulating more astaxanthin [29]. Our previous studies demonstrated that the addition of SA increased the astaxanthin content in _H. pluvialis_[4], suggesting that astaxanthin complements the antioxidant effect of heme degradation products and reduces the expression of _ho_. Similarly, in the blue light treatment groups, the expression of genes involved in astaxanthin synthesis (i.e., _psy, pds, lcyb,_ and _crtr-b_) was also significantly down-regulated (Figure 6), whereas the expression of heme oxygenase was significantly up-regulated (Figure 8), suggesting that heme degradation products help maintain the intracellular homeostasis in _H. pluvialis_. Previous studies showed that scavenging ROS was essential for the survival of _H. pluvialis_ under high light intensity [63]. Our results showed that the expression levels of most genes related to ROS in _H. pluvialis_ treated with blue light were significantly higher than those in white light, indicating that the damage of blue light to _H. pluvialis_ was higher than that of white light (Figure 9). Previous studies showed that APX, CAT, GRX, GPX, and PRDX removed hydrogen peroxide [64], while SOD scavenged superoxide to form hydrogen peroxide [65]. Studies have shown that the antioxidant systems in different species of plants respond differently to the treatment with SA. For example, the activities of ascorbic APX and SOD were enhanced, and the activities of CAT were reduced in corn, whereas the activities of antioxidant enzymes CAT, peroxidase (POX), SOD, and glutathione reductase were decreased in rice [66]. Moreover, the level of H\({}_{2}\)O\({}_{2}\) and the activities of antioxidant enzymes CAT, POX, and SOD were enhanced [66]. Our results showed that the treatment with SA led to the significant down-regulation of many genes involved in the ROS clearance pathway (Figure 9), suggesting the overall adjustment of resistance to stress in _H. pluvialis_.