## 2 Materials and Methods
### Antioxidant and Enzyme Activity Assessment in Prawn Hepatopancreas  
At the end of the feeding trial, a total of 100 prawns (20 per dietary treatment; 5 prawns per tank, samples analyzed individually) were randomly selected for hepatopancreas sampling to assess antioxidant and digestive enzyme activity, as well as lipid peroxidation levels. Prawns were euthanized by thermal shock (immersion in ice water for 5 min), and the body surface was disinfected using 70% (\(\
u/v\)) ethanol. The hepatopancreas was carefully dissected and immediately stored at \(-\)20 \({}^{\circ}\)C until further biochemical analysis. Tissue samples were homogenized in 50 mM Tris-HCl buffer (pH 7.4) using an electric homogenizer (IKA\({}^{\oplus}\) T10 basic; IKA\({}^{\oplus}\), Staufen, Germany). All procedures were performed on ice to maintain enzyme stability. The homogenates were centrifuged at 12,000\(\times\)\(g\) for \begin{table}
\begin{tabular}{l c c c c c} \hline  & \multicolumn{5}{c}{**Experimental Diets**} \\ \hline  & HFM 0\% & HFM 1.5\% & HFM 3.0\% & HFM 4.5\% & HFM 6.0\% \\ \hline C21:5 n-3 & 0.21 \(\pm\) 0.02 & 0.32 \(\pm\) 0.07 & 0.27 \(\pm\) 0.01 & 0.27 \(\pm\) 0.01 & 0.23 \(\pm\) 0.01 \\ C22:6 n-3 & 0.24 \(\pm\) 0.02 & 0.20 \(\pm\) 0.03 & 0.34 \(\pm\) 0.03 & 0.32 \(\pm\) 0.03 & 0.15 \(\pm\) 0.02 \\ C22:2 n-6 & 0.14 \(\pm\) 0.01 & 0.07 \(\pm\) 0.01 & 0.06 \(\pm\) 0.01 & 0.06 \(\pm\) 0.02 & 0.04 \(\pm\) 0.01 \\ C22:5 n-6 & 0.13 \(\pm\) 0.02 & 0.15 \(\pm\) 0.01 & 0.20 \(\pm\) 0.02 & 0.11 \(\pm\) 0.04 & 0.14 \(\pm\) 0.03 \\ C22:4 n-6 & 0.06 \(\pm\) 0.01 & 0.09 \(\pm\) 0.02 & 0.03 \(\pm\) 0.05 & 0.00 \(\pm\) 0.00 & 0.03 \(\pm\) 0.01 \\ SFA & 46.5 \(\pm\) 0.10 & 49.7 \(\pm\) 0.32 & 49.5 \(\pm\) 0.06 & 48.6 \(\pm\) 0.01 & 45.9 \(\pm\) 0.01 \\ MUFA & 47.9 \(\pm\) 0.12 & 45.0 \(\pm\) 0.12 & 45.4 \(\pm\) 0.23 & 47.0 \(\pm\) 0.18 & 48.0 \(\pm\) 0.01 \\ PUFA & 5.67 \(\pm\) 0.02 & 5.32 \(\pm\) 0.20 & 5.16 \(\pm\) 0.29 & 4.44 \(\pm\) 0.17 & 5.77 \(\pm\) 0.01 \\ PUFA n-6 & 4.54 \(\pm\) 0.04 & 4.04 \(\pm\) 0.09 & 3.86 \(\pm\) 0.21 & 3.24 \(\pm\) 0.17 & 4.63 \(\pm\) 0.02 \\ PUFA n-3 & 1.13 \(\pm\) 0.02 & 1.28 \(\pm\) 0.11 & 1.30 \(\pm\) 0.08 & 1.21 \(\pm\) 0.01 & 1.14 \(\pm\) 0.03 \\ \hline \end{tabular} HFM: Hydrolyzed feather meal; SFA: saturated fatty acids; MUFA: monounsaturated fatty acids; PUFA: polyunsaturatedarted fatty acids.  
\end{table} 
Table 3: _Cont._10 min at 4 \({}^{\circ}\)C in a refrigerated centrifuge (Sigma 3-16 KL; Sigma, St. Louis, MO, USA), and the resulting supernatant was collected for enzymatic assays. Total protein concentration in the homogenates was quantified using the Bradford method [28], employing bovine serum albumin as a standard and measuring absorbance at 595 nm. Protein concentration values were used to normalize the enzymatic activity measurements. Glutathione peroxidase (GPx) activity was evaluated based on the oxidation of NADPH during the reduction of cumene hydroperoxide, with absorbance read at 340 nm [29; 30]. Glutathione reductase (GR) activity was determined by quantifying NADPH oxidation during the conversion of glutathione disulfide to reduced glutathione, also measured at 340 nm [28; 31]. Reduced glutathione (GSH) levels were analyzed using the non-protein thiol detection method with Ellman reagent [31; 32]. Glutathione S-transferase (GST) activity was measured through the conjugation of GSH with 1-chloro-2/_A_-dinitrobenzene, monitored spectrophotometrically [33; 34]. Lipid peroxidation was determined by the quantification of malondialdehydedehyde (MDA) via its reaction with thiobarbituric acid reactive substances (TBARS) [35; 36]. The activities of digestive enzymes, such as maltase (U/mg protein), sucrase (U/mg protein), chymotrypsin (\(\mu\)mol/min/mg protein), and trypsin (nmol/min/mg protein), were also quantified to evaluate the digestive capacity of _M. rosenergii_ post-larvae. The activities of maltase and sucrase were measured following the protocol by [PERSON] et al. [37].