## 3 Results and Discussion
### Antioxidant Activity in Frozen and Freeze-Dried Materials  
Drying by the use of elevated temperature is a process that may imply thermal degradation and reduce the antioxidant activity in the raw materials. Freeze-drying is one of the most gentle drying processes, applied to increase the shelf-life and retain the nutrients of raw materials [13; 50]. It has been previously shown that the freeze-drying process was the only treatment which did not reduce the antioxidant activity compared to normal drying and microwave-vacuum drying in strawberry [60]. Another study found that the freeze-drying process yielded the best quality of dried sea cucumber (_Stichopus japonicus_) [53]. Despite obtaining a good quality of food products using the freeze-drying process, the drying process has its drawbacks. The freeze-drying process is time consuming and costly, and hence, it is not always relevant in the industry [53]. Furthermore, the freeze-dried products could be more susceptible to lipid oxidation due to their porosity and exposure to oxygen in the food product after the removal of water [61].  
The capability of a substance to decrease ferric (Fe\({}^{3+}\)) to ferrous (Fe\({}^{2+}\)) iron is evaluated by the ferric reducing antioxidant power (FRAP) assay [28]. The FRAP activity in both frozen and freeze-dried samples ranged between 1 and 3 mmol TE/100 g dry weight (Table 3). When calculating the FRAP activity of frozen samples based on dry weight, they showed significantly higher (\(p<0.05\)) FRAP activity compared to freeze-dried samples. According to a previous study by [PERSON] et al. [62] on the antioxidant activity of dried sea cucumber, _Holothuria edulis_ resulted in the highest FRAP activity, followed by _Pearsonothuria graeffei_ and _Stichopus hermanni_ (48.5, 28.3, and 12.0 umol Fe(II), respectively). The result in this study is lower compared to previous results. The reason for this difference could be due to the extraction and drying technique, resulting in a higher concentration [62], compared to the extract obtained in this study. However, the FRAP activity in both frozen and freeze-dried samples in this study exhibited similar FRAP activity compared to the protein hydrolysate of orange-footed sea cucumber (_Cucumania fondosa_) [13], protein hydrolysate of fried herring (_Clupea harengus_) [63], and saithe (_Pollachius virens_) [64].  
Figure 5: True retention (%) of essential amino acids (EAAs) after freeze-drying of frozen red sea cucumber (_Parastichopus tremulus_). The results are presented as average \(\pm\) standard error, \(n=10\).  
The ABTS assay evaluates a substance's ability to reduce reactive oxygen species by electron transfer [29; 31]. The ABTS activity ranged between 0.1 and 0.3 mmol propyl gallate equivalents (PGE)/g (Table 3). The freeze-dried samples exhibited the highest ABTS activity. A previous study found freeze-dried and hydrolysates of _C. frondosa_ ranging between 2 and 13 umol PGE/g dry weight in a sea cucumber [13].  
The ORAC assay measures the ability of bioactive peptides to neutralize reactive oxygen species by hydrogen transfer [32]. The antioxidant activity ranged between 130 and 230 mmol TE/g (Table 3), where freeze-dried samples exhibited the highest ORAC activity. According to a study by [PERSON] et al. [65] on a peptide fraction of _P. tremulus_, the ORAC activity was found to be 0.4 mmol TE/ug protein. The ORAC activity found in protein hydrolysate of saithe (_Pollachius virens_) ranged from 331 to 558 umol TE/g [64]. The ORAC activity is higher compared to previous results. Regardless, the ORAC activity is lower compared to a previous study by [PERSON] et al. [13] on freeze-dried and hydrolysates of _C. frondosa_.