## 2 Materials and Methods
### Experimental Design  
To carry out the experiment, four feeds (40% crude protein, 8% ether extract) were produced at the Aquatic Organism Nutrition Laboratory (LANOA) at FURG: control = control feed produced in the laboratory without the addition of macroalgae; T5 = 5% inclusion of macroalgae in the feed; T10 = 10% inclusion of macroalgae; T15 = 15% inclusion of macroalgae; all experimental diets were tested in quadruplicate (Table 2). To make the diets, the dry ingredients were mixed, from the smallest to the largest, finishing with the liquid ingredients. The mixture was then pelletized in a meat grinder (Metalurgica 9000, PC- 22, Sao Paulo, Brazil) and the feed was dried in an oven at 60 \({}^{\circ}\)C for 24 h. The pellets were broken down to the desired diameter.  
The experiment was carried out in tanks with recirculating water that contained a macrocosm of 800 L of useful volume, with the presence of artificial substrate (0.60 m\({}^{2}\)), and, using a submerged pump (SPA 4000 L/h, BOYU(c), Chaozhou, China), the water was recirculated to 16 units with 50 L of useful volume at a speed of 2.60 L min\({}^{-1}\). The recirculation system was operated at a temperature of 26.20 \({}^{\circ}\)C, dissolved oxygen at 6.95 mg L\({}^{-1}\), pH 7.92, salinity at 14.82 ppt, and alkalinity at 150 mg CaCO\({}_{3}\) L\({}^{-1}\). Nutrients were maintained at 0.29, 0.18, 12.00, and 0.42 mg L\({}^{-1}\), for total ammoniacal nitrogen, nitrite,  
\begin{table}
\begin{tabular}{c c c c c} \hline \hline  & **Protein (\%)** & **Lipid (\%)** & **Fiber (\%)** & **Ash (\%)** \\ \hline _Ulva meal_ & 19.31 \(\pm\) 0.30 & 0.46 \(\pm\) 0.04 & 10.34 \(\pm\) 0.72 & 29.05 \(\pm\) 0.63 \\ \hline \hline \end{tabular}
\end{table}
Table 1: Proximal composition of the macroalgae _U. lactuca_ (% of dry matter) cultivated in an integrated biofloc system.  
n nitrate, and phosphate, respectively. The tanks were siphoned three times a week to remove organic matter from the bottom.  
Ten fish were used per tank with an initial weight of 0.94 \(\pm\) 0.01 g. The fish were fed at a fixed rate of 6% of the total biomass per day, divided into three feedings during the day (8:00 a.m., 12:00 p.m., and 5:00 p.m.) [29]. Initial and final weightings were performed, and the feed was adjusted. At the end of the experiment, the fish were anesthetized with benzocaine [30] and weighed to determine performance variables. Nine samples for treatment (three fish per tank) for proximal composition and nine samples for treatment for hematology were collected at the end of the experiment after the animals were anesthetized and euthanized with benzocaine at a concentration of 400 ppm [30].  
After collecting samples and weighing the animals, three fish were selected from each tank of the control, T5, T10, and T15 treatments to carry out a salinity stress test. The fish were transferred and maintained for 24 h in new experimental units composed of polyethylene tanks with a useful volume of 30 L, and the salinity was reduced from 15 to 0 ppt without acclimatization. As a control for saline stress, three fish were collected from the control treatment and kept at the salinity of the experiment (15 ppt). Fifteen experimental units and five treatments were used for saline stress: Sal15 (control without algae at a salinity of 15 ppt), Sal0C0 (control without algae at a salinity of 0 ppt), Sal0T5 (diet with 5% macroalgae at a salinity of 0 ppt), and Sal0T15 (diet with 15% macroalgae at a salinity of 0 ppt). After 24 h of exposure, three fish from each tank (nine from each treatment group) were euthanized by freezing and dissected. The gills, liver, and muscle of each fish were collected and preserved at \(-\)80 \({}^{\circ}\)C for future biochemical analysis.