*3.2. Lipids Analysis*

For fatty acids (FAs) analysis, lyophilized algal cells were directly transesterificated with 1% (*v/v*) sulfuric acid in methanol and methylbenzene at 50 ◦C overnight, and analyzed using gas chromatography–mass spectrometry (GC–MS) equipped with a DB-WAX capillary column (30 m × 0.25 mm × 0.25 μm) (Agilent, Palo Alto, CA, USA). Helium was used as the carrier gas with the flow rate of 1.2 mL/min. Samples were injected in split mode (5:1 split ratio) at an oven temperature of 45 ◦C with an injection volume of 1 μL. The oven temperature was raised from to 45 ◦C 150 ◦C at 15 ◦C/min, then to 240 ◦C at 6 ◦C/min, and held at 240 ◦C for 8 min. Total lipids were quantified as the fatty acids contained in the total lipids, namely total fatty acids (TFA). EPA peaks were identified by the MS spectrum matching from database. Heptadecanoic acid (C17:0, Sigma-Aldrich, St. Louis, MO, USA) was used as the internal standard.

#### *3.3. Polysaccharides Content Analysis*

Microalgal cells were collected by centrifugation. Cells were homogenized by a tissue lyser at 30 Hz for 1 min. After homogenization, intracellular soluble polysaccharides were extracted in distilled water. Four times volume of ethanol was added for polysaccharides precipitation overnight. Then, the precipitates were collected and redissolved in distilled water for soluble polysaccharides analysis. The glucose was used as standard to draw a standard curve. One milliliter of 6% phenol and 5 mL of concentrated sulfuric acid were added to each tube. After mixing and reaction for 10 min at room temperature, samples were cooled to room temperature and measure the absorbance at 490 nm wavelength.

#### *3.4. Pigments in Photosynthesis Complexes*

Pigments were extracted from fresh algal cells using methanol. By measuring the absorbance, respectively, at 664 nm and 630 nm with a spectrophotometer (NanoDrop Technologies, Wilmington, Delaware, USA), chlorophyll concentration was calculated with the following equation [47]:

$$\text{Chl } a = 13.2654 \times \text{A664} - 2.6839 \times \text{A630} \tag{1}$$

$$\text{Chl } \mathcal{c} = 28.8191 \times \text{A630} - 6.0138 \times \text{A664} \tag{2}$$

Identification and quantification of fucoxanthin were performed on ACQUITY Ultra Performance LC H-Class coupled in-line to a Xevo TQ-XS triple quadrupole mass spectrometer with a ESI probe (Waters, Milford, Massachusetts, USA). Samples were injected on a ACQUITY UPLC HSS T3 column (2.1 mm × 100 mm, 1.8 μm, Waters), and the eluents were acetonitrile (A), methanol (B) and demineralized water contained 0.1% (*v/v*) formic acid (C). The gradient was initiated from 58% A, 27% B, and 15% C, then transiently to 92%A and 8% C at 4 min, and finally reverted to its initial composition at 8 min followed by an equilibration phase of 2 min. The flow rate was at 350 μL/min. Nitrogen was used as desolvation gas (800 L/h at 400 ◦C). The spray capillary voltage was 3 kV for the positive ion mode. Multiple reaction monitoring (MRM) scanning mode was used for mass spectrometry detection. The standards of fucoxanthin were purchased from Sigma (St. Louis, MO, USA).
