## 2 Results and Discussion
### In Silico Simulated Gastrointestinal (GI) Digestion of Bioactive Peptides  
Simulated GI digestion that mimics protein degradation in the stomach and small intestine is considered as a rapid and effective tool to determine the stability of peptides against GI proteases [30]. In this regard, in silico simulated GI digestion is an effective approach to investigate the bioactive potential of any peptide of interest prior to assessing their bioavailability and bioaccessibility in vivo. Thus, selected peptides were subjected to in silico proteolysis simulation with representative digestive enzymes; pepsin (EC 3.4.23.1), trypsin (EC 3.4.21.4) and chymotrypsin (EC 3.4.21.1) using \"BIOPEP Enzyme(s) Action\" program [19]. The current study predicted the potential of peptides derived from sea cucumber protein hydrolysates as the precursors of peptides with antioxidative or ACE inhibitory properties. Tables 6 and 7 represent the bioactive potential upon GI digestion of peptides derived from flower and internal organs, respectively. \begin{table}
\begin{tabular}{c c c c c c c} \hline \hline \multirow{2}{*}{**Sample**} & \multirow{2}{*}{**Peptide**} & \multirow{2}{*}{**Results of Enzyme Action**} & \multicolumn{2}{c}{**Location of Released Peptides**} & \multicolumn{2}{c}{**Active Fragment Sequence**} & \multirow{2}{*}{**Location**} & \multicolumn{2}{c}{**Bioactivity of Identified Peptide**} \\ \hline Flower & GPPCGPQWPLDF & GPPCGPQW\(\_\)PL\(\_\)—DF & (1–7), (8–9), (10–11) & PL & (8–9) & ACE inhibitor \\  & & & & DF & (10–11) & ACE inhibitor \\  & & (1–6), (7–11), & & No Antioxidant or \\  & GPPCPR\(\_\)GPTR\(\_\)–GPTR\(\_\)–G & (12–12), (13–13) & – & – & ACE inhibitory \\  & GPGGPGFM & – & – & – & – & – \\  & APDMAFPR & APDM\(\_\)–AF\(\_\)–PR & (1–4), (5–6), (7–8) & PR & (7–8) & ACE inhibitor \\  & & & AF & (5–6) & ACE inhibitor \\  & GGFFG & GGF\(\_\)PGG & (1–3), (4–8) & – & – & ACE inhibitory \\  & GPGM\(\_\)–M\(\_\)–GP & (1–4), (5–5), (6–7) & GP & (6–7) & ACE inhibitor \\  & GPPCAGSPL\(\_\)diagonal & GPPCASGPL\(\_\)diagonal & (1–9), (10–15) & – & – & ACE inhibitory \\  & & & & No Antioxidant or \\  & GPSCPPGP & – & – & – & – & – \\ \hline \hline \end{tabular}
\end{table} 
Table 6: Remaining bioactive properties after in silico simulated gastrointestinal digestion of peptides originated from flower protein hydrolysates. According to the predictions, none of the peptides showed antioxidative potential after the simulated digestion process. This may be due to the scarcity of information in databases related to the potent antioxidative active peptides. It has been suggested that bioinformatics strongly depend on the information included in the data base [1]. However, prediction of releasing ACE inhibitory peptides was found to be similar in all samples. Three peptides (GPPGPQWPLDF, APDMAFPR and GPGMMGP) from flower and internal organs were predicted to release potent ACE inhibitory fragments from their original sequences. Furthermore, frequency of releasing potent ACE inhibitory peptides from each identified peptide was varied as some peptides have more than one active fragment embedded in their sequences. For example, GPPGPQWPLDF sequence was predicted to release two active ACE inhibitory sequences, namely PL and DF, whereas APDMAFPR has the potential to release PR and AF following simulated GI digestion. The release of bioactive peptides was predicted after using the \"enzyme(s) action\" option of BIOPEF data base (Table 8).