## 3 Results  
### Amino Acid Composition  
The amino acid contents in protein isolates and hydrolyses, as well as raw materials, are presented in Table 2. There was little variation in the amino acid contents within isolates/hydrolysates produced using the same type of fractionation method (pH shift or enzymatic hydrolysis) with few statistically significant differences. In the case of roach isolates produced using acidic and alkaline pH shift, the alkali-processed isolate had a statistically significantly higher (\(p<\) 0.05) content of tryptophan (13.5 mg/g protein) and lower content of arginine (55.0 mg/g protein) compared with acidic isolate (11.7 and 57.7 mg/g protein, respectively). However, in the case of B. herring, the alkaline isolate had a statistically significantly higher content of arginine (63.8 mg/g protein) and lower content of glutamine + glutamic acid (125.5 mg/g protein) than the acid-extracted isolate (59.7 and 131.5 mg/g protein, respectively). In the case of enzymatically produced hydrolyses there were only minor differences in the amino acid composition depending on the enzyme used. However, the use of different types of extraction methods (hydrolysis vs. pH shift) was reflected in the amino acid composition. Most notable differences were observed in the contents of asparagine + aspartic acid, glycine, hydroxyproline, tyrosine, isoleucine, phenylalanine, tryptophan, and valine. Protein isolates produced using pH shift had a significantly lower content of glycine (28.5-31.0 mg/g protein) compared with that of the fish raw material (38.0-42.4 mg/g protein), or enzymatically produced hydrolyses (44.8-51.9 mg/g protein). This finding is in line with studies by [PERSON] and [PERSON] [19] and [PERSON] and [PERSON] [14], where protein isolates produced by pH-shift processes from herring had lower glycine contents compared with the level in herring itself. The authors of the latter study hypothesized that this could be either due to loss of protein(s) rich in glycine or the fact that glycine is one of the most abundant free amino acids in herring, and is likely discarded with the water phase during the second separation of the process. In the present study, enzymatically produced hydrolyses showed even higher contents of glycine compared with the levels in raw fish, and since these hydrolyses are collected with the water phase, it can be expected that the loss of free amino acids occurs to a lesser extent during the enzymatic fractionation. \begin{table}
\begin{tabular}{l c c c c c c c c c c c c c c c c} \hline \hline  & \multicolumn{6}{c}{**Rach**} & \multicolumn{6}{c}{**Rach**} & \multicolumn{6}{c}{**Racing**} \\ \cline{3-14}  & **Rare Material** & **Audio-pfil** & **Audio-pfil** & **Audio-pfil** & **Audio-  
The content of hydroxyproline was expectedly lower in pH-shift-processed isolates (1.3 mg/g protein in both acid and alkali-extracted isolates of roach, and 2.0 and 2.1 mg/g protein in acid and alkali-extracted isolates of B. herring, respectively) compared with contents in the raw materials (5.3 and 6.7 mg/g protein in roach and B. herring, respectively) and in the enzymatically produced hydrolysates (10.6-11.9 and 8.7-9.1 mg/g protein, in the case of roach and B. herring hydrolysates, respectively). Hydroxproline is almost exclusively found in collagen, which is efficiently removed in the precipitate of the first centrifugation of the pH shift [42]. On the other hand, higher hydroxyproline levels in the hydrolysates indicate that the use of proteases, or heating applied during hydrolysis, resulted in the extraction of collagen. Collagen is also rich in glycine and proline [42] and hence the removal of collagen may also have contributed to lower glycine levels in protein isolates produced by pH shift.