*3.8. Determination of Pigments*

For chlorophyll a, chlorophyll b and total carotenoid quantification, an acetone 90% extract was done with 5 mg of freeze-dried biomass in 1 mL of solvent. The extract was sonicated for 3 min and remained 24 h in darkness at 4 ◦C. Chlorophyll a was determined according to Equation (3) [89], chlorophyll b was determined according to Equation 4 [90] and total carotenoids were determined according to Equation (5) [91].

$$\left[ \left( 11.8668 \times \text{A}\_{664} \right) - \left( -1.7858 \times \text{A}\_{647} \right) \right] \tag{3}$$

$$\left[\left(18.9775 \times \text{A}\_{647}\right) - \left(-4.8950 \times \text{A}\_{664}\right)\right] \tag{4}$$

$$\left[\left(\mathbf{A}\_{480} \times \mathbf{4}.0\right)\right] \tag{5}$$

where A664, A647 and A480 are the measured absorbance at 667, 647 and 480 nm, respectively.

### *3.9. Lipopolysaccharides (LPS) Contamination Assay*

The presence of lipopolysaccharides (LPS) in the EPS fractions isolated from *T. suecica* was evaluated using the *Limulus* amebocyte lysate (LAL) assay kit (Endosafe®-PTS, Charles River Laboratories, Charleston, SC, USA). In brief, 25 μL of the EPS solution (concentration 50 μg mL−1) in distilled water was loaded into each of the four channels of the cartridge. The reader automatically mixed the sample with the LAL reagen<sup>t</sup> in two channels. Additionally, the LAL reagen<sup>t</sup> was mixed with the positive control in the other two channels. These samples were used as the control. Afterward, all samples were incubated and combined with the chromogenic substrate. After mixing, the optical density of the four channels was measured and compared with an internal standard curve. The amount of endotoxin in the sample was expressed as endotoxin units (EU) mL−1.
