Molecular Soil Biology 2026, Vol.17, No.1, 1-11 http://bioscipublisher.com/index.php/msb 6 Figure 6 Changes in lipid content of microalgae SA-2 Note: (a) Standard curve of lipid content; (b)treatment groups with different concentrations of EDTA-Ca; (c) treatment groups with different concentrations of EDTA-Fe. Different lowercase letters indicate a significant difference (P<0.05) 2.6 Effects of EDTA-Fe and EDTA-Ca treatments on microalgal pigment content Pigment content of microalgae under different concentrations of EDTA-Ca and EDTA-Fe was measured at various wavelengths. Total chlorophyll content reached its maximum under 0.3 mg/L EDTA-Ca, at 12.24 mg/L (Figure 7a), and under 0.2 mg/L EDTA-Fe, at 10.19 mg/L (Figure 7b). Chlorophyll a content peaked at 10.60 mg/L with 0.3 mg/L EDTA-Ca (Figure 7c) and at 16.34 mg/L with 0.2 mg/L EDTA-Fe (Figure 7d). Chlorophyll b content was highest at 2.74 mg/L under 0.3 mg/L EDTA-Ca (Figure 7e) and 13.32 mg/L under 0.2 mg/L EDTA-Fe (Figure 7f). Carotenoid content reached a maximum of 2.74 mg/L under 0.3 mg/L EDTA-Ca (Figure 7g) and 4.47 mg/L under 0.2 mg/L EDTA-Fe (Figure 7h).These results indicate that EDTA-Ca at concentrations below 0.3 mg/L and EDTA-Fe at concentrations below 0.2 mg/L promoted the synthesis of various pigments in SA-2, whereas higher concentrations of EDTA-Ca (>0.3 mg/L) and EDTA-Fe (>0.2 mg/L) inhibited pigment synthesis. The promotive effect of low-concentration EDTA-Fe (<0.2 mg/L) was more pronounced than that of low-concentration EDTA-Ca (<0.3 mg/L), while the inhibitory effect of high-concentration EDTA-Fe (>0.2 mg/L) was stronger than that of high-concentration EDTA-Ca (>0.3 mg/L). This difference may be attributed to the role of iron in multiple aspects of microalgal physiology, including photosynthetic electron transport, chloroplast development, key enzyme activities, and photoprotective mechanisms, suggesting that iron is more critical than calcium for chlorophyll synthesis in microalgae. 2.7 Effects of EDTA-Fe and EDTA-Ca treatments on microalgal carbohydrate content Changes in microalgal carbohydrate content on day 12 were measured under different concentrations of EDTA-Ca and EDTA-Fe (Figure 8). Carbohydrate content reached its maximum at 58.13 mg/L under 0.3 mg/L EDTA-Ca. For EDTA-Fe, the highest carbohydrate content of 80.04 mg/L was observed at 0.2 mg/L. These results indicate that EDTA-Ca at concentrations below 0.3 mg/L and EDTA-Fe at concentrations below 0.2 mg/L promoted carbohydrate synthesis in SA-2. Conversely, EDTA-Ca concentrations above 0.3 mg/L and EDTA-Fe concentrations above 0.2 mg/L inhibited carbohydrate accumulation in the microalgae. 3 Discussion In this study, EDTA-Fe and EDTA-Ca exhibited differential effects on microalgal biomass accumulation. The results indicate that supplementation with appropriate concentrations of EDTA-Fe and EDTA-Ca generally enhanced biomass content, whereas excessive concentrations of either EDTA-Fe or EDTA-Ca exerted inhibitory effects on SA-2. This observation is closely related to the distinct roles of the two elements in microalgal metabolism. Iron ions function as cofactors in enzymatic reactions, particularly those involving iron-containing proteins, and participate in the photosynthetic electron transport chain. Microalgae are rich in iron-binding proteins such as iron–sulfur (Fe-S) proteins, ribonucleotide reductase (RNR), and hemoproteins, with iron serving as a cofactor in DNA replication, DNA repair, cell cycle progression, metabolic catalysis, and iron homeostasis (Zhang, 2014). Iron is also a core element for photosynthetic electron transport, chlorophyll synthesis, and multiple enzymatic reactions, with its sufficient supply directly determining photosynthetic efficiency and organic carbon fixation rates.In contrast, calcium primarily contributes to cellular structure, signal transduction, and physiological regulation rather than directly participating in photosynthesis. Unlike nitrogen or phosphorus, it
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