## 4 Discussion  
### Optimization Effect of Electrical Stimulation on N. closterium f. minutissima Under Laboratory  
Previous research has primarily focused on the effects of electrical stimulation on freshwater microalgae and species from the _Chlorophyta_ phylum. This study confirms that _Nitzschia closterium f. minutissima_, a marine diatom from the _Bacilliariophyta_, can also experience growth promotion and nutrient accumulation through electrical stimulation. Consistent with previous findings on freshwater microalgae and green algae, electrical stimulation also optimizes the cultivation of _N. closterium f. minutissima_ in several aspects, including increased cell density, enhanced biomass dry weight, and accelerated nutrient accumulation. Given the diversity and nutritional richness of microalgae, future research should explore whether electrical stimulation promotes growth across all phyla and how different electrical stimulation conditions can regulate the accumulation of specific target products.  
To comprehensively evaluate the optimal voltage for cultivation optimization, Z-score standardization was applied to data across seven dimensions, and a radar chart was generated, with four dimensions representing nutrient levels in the culture. The results indicated that 3 V was the optimal voltage for promoting the growth and nutrient accumulation of _Nitzschia closterium f. minutissima_ under short-term direct current stimulation, as evidenced by a shortened lag phase and a 67.66% increase in lipid content compared to the control group, reaching 94.95 \(\pm\) 5.66 \(\upmu\)g/mL. Polysaccharide and protein contents were also the highest among the short-term electrical stimulation groups (Figure 11a). For continuous direct current stimulation, 0.5 V was the optimal voltage, with a 39.47% increase in biomass dry weight compared to the control group, reaching 0.35 \(\pm\) 0.02 \(\upmu\)g/mL, while polysaccharide, protein, and lipid contents were the highest among the continuous stimulation groups (Figure 11b). By overlaying radar charts for the control group (CK), D-3 V, and C-0.5 V, the area sizes follow the order D-3 V \(>\) CK \(>\) C-0.5 V (Figure 11c), indicating that 3 V short-term stimulation was the optimal voltage in this study.  
It is noteworthy that although 3 V was the optimal voltage according to the comprehensive evaluation, it was not the best for maximizing cell density or chlorophyll a accumulation, and its impact on biomass dry weight was limited. This suggests that different metabolic pathways respond uniquely to electrical stimulation. Similar findings were reported by [PERSON] et al. in their study on the electrical stimulation of _Scenedesmus obliquus_, where 1 A/m\({}^{2}\) stimulation resulted in the most significant increases in cell density, biomass, and chlorophyll a content, while 2 A/m\({}^{2}\) stimulation yielded the highest accumulation of polysaccharides [13; 14]. This indicates that the optimal stimulation conditions may vary depending on the target product. Additionally, while different phyla of microalgae exhibit varying response patterns to electrical stimulation, they also share certain commonalities, which warrants further exploration through studies across diverse microalgal phyla.  
In this study, the most notable results of electrical stimulation on _N. closterium_ f. _minutissima_ include a 13.14% increase in microalgal cell density and a 67.66% increase in lipid content with short-term DC stimulation. However, compared to previous studies, there is still considerable room for improvement. For instance, in [PERSON] et al.'s study on _Haematococcus pluvialis_, cell density increased by approximately 20% [15], and in [PERSON] et al.'s research on _Chorella sp._ KR-1, triglyceride yield increased by a factor of 2.1 compared to the untreated control [11]. Nonetheless, we believe that through further optimization of the electrical stimulation method and conditions, the overall effectiveness of electrical stimulation on _N. closterium_ f. _minutissima_ can be further enhanced.