## 4 Conclusions  
In short, hydrolysis triggers the passage of _Anisakis_ secretory/secretory and somatic allergens to a more soluble or available state, initially increasing their allergenicity. After long silage periods, the lower molecular weight fragments of _Anisakis_ proteins become fully digested. However, some higher-molecular size fragments with potential allergenicity still appeared, but at very low concentrations. The silage duration should be adjusted to the \begin{table}
\begin{tabular}{l c c c c} \hline \hline  & **V3** & **F2** & **F5** & **Reference \({}^{*}\)** \\ \hline _Essential amino acids_ & & & & \\ Histidine & 3.3 & 3.3 & 3.2 & 2.1 \\ Threonine & 5.2 & 5.2 & 5.1 & 3.9 \\ Arginine & 6.4 & 6.8 & 6.9 & 1.3 \\ Valine & 6.1 & 5.5 & 5.2 & 3.6 \\ Methionine & 3.2 & 2.9 & 2.9 & 3.1 \\ Phenylalanine & 4.2 & 4.0 & 3.8 & 6.5 \\ Isoleucine & 5.3 & 4.4 & 4.3 & 2.5 \\ Leucine & 8.2 & 7.5 & 7.0 & 3.3 \\ Lysine & 9.2 & 8.3 & 8.0 & 5.7 \\ Tryptophan & 0.25 & \textless{}0.010 & \textless{}0.010 & 0.8 \\ _Non-essential amino acids_ & & & & \\ Aspartic & 9.7 & 9.7 & 9.7 & \\ Glutamic & 14.2 & 13.7 & 13.8 & \\ Serine & 5.0 & 5.4 & 5.3 & \\ Glycine & 6.3 & 5.4 & 7.2 & \\ Alanine & 6.6 & 6.5 & 6.6 & \\ Tyrosine & 1.1 & 3.8 & 3.9 & \\ Hydroxyproline & 0.9 & 1.1 & 1.5 & \\ Proline & 4.9 & 5.3 & 5.4 & \\ EAA (g/100 g silage DM) & 29.35 & 20.83 & 20.44 & \\ NEMA (g/100 g silage DM) & 27.96 & 22.77 & 23.36 & \\ TAA (g/100 g silage DM) & 57.31 & 43.60 & 43.80 & \\ FAA (g/100 g silage DM) & 48.88 & 10.75 & 12.29 & \\ \hline \hline \end{tabular} \({}^{*}\) Reference amino acid requirements of juvenile common carp (NRC,1993 [43]).  
\end{table} 
Table 4: Essential amino acids (EAAs), nonessential amino acids (NEAAs), total standard amino acids (AAs) and free amino acids (FAA) in the liquid fractions of the three silages obtained after 12 months of storage (V3) and 15 months of storage (F2 and F5). Single amino acids are expressed in g/100 g AA. minimal time necessary to minimize or eliminate allergens from _Anisakis_ while avoiding potential loss of sensitive amino acids. Although this product is not intended for human consumption, but rather for animal feed, smaller proteins or fragments can be expected to be the most easily absorbed in the digestive tract. Therefore, it is necessary to determine whether resistant and larger fragments with allergenic potential can pass through the intestinal tract of animals without being absorbed or incorporated into the edible fraction of animals. Thus, further work and additional sensitive proteomic techniques must be performed to identify the risks of potentially remaining IgE reactive proteins in foods obtained from animals fed with silages. This approach ensures that feeding this product to animals does not pose a potential risk to _Anisakis_ allergy sufferers. Conceptualization: C.B., B.I. and M.L. ; methodology: C.B., B.I. and M.L. ; formal analysis and investigation: C.B., B.I., M.L. and C.A. ; validation: C.B., M.L., B.I. and G.D.; visualization: C.B. and M.L. ; writing--original draft preparation: C.B. ; writing--review and editing: B.I., M.L. and G.D.; project administration: G.D.; funding acquisition: G.D.; resources: X.A. and G.D.; supervision: G.D. All authors have read and agreed to the published version of the manuscript. This work was supported by the Directorate of Fisheries and Aquaculture of the Basque Government (Department of Economic Development, Infrastructures and Environment-Vice Dept. of Agriculture, Fisheries and Food Policies) project ANIFEMP (grant number 00001-IRB2018-33). This study was conducted according to the guidelines of the Declaration of Helsinki. The collection of sera was approved by the Ethics Committee for Clinical Research (CEIC) of the Araba Integrated Health Organization (Vitoria, Spain), with internal Ref. PI2018118 (date of approval: 10 August 2018). All participants signed the corresponding informed consent. The anonymity of the participants was guaranteed throughout the study.