## 3 Results and Discussion  
### The Hydrolysates (PH and AH) Characterization  
In this study, we used by-products of silver carp to produce hydrolyses. The yield of the hydrolyses prepared by PH was about 42.1% and the yield of the hydrolyses prepared by Alcalase hydrolysis was about 43.6%. Proximate composition of the defatted by-products and the hydrolyses are shown in Table 1.  
DH values and zeta potential values of PH and AH are shown in Table 2. Zeta potential value is an indicator of the surface charge of the hydrolysate in solution, which can reflect the stability and possible binding ability of the hydrolysate to ice crystal and/or protein via hydrogen bond and electrostatic interaction [20]. Our previous studies have revealed that both Protamex and Alcalase are efficient enzyme choices for preparing hydrolyses from silver carp processing by-products. Although enzyme type employed could determine the hydrolysate structural properties [4], the differences in DH values and zeta potential values of the hydrolyses prepared by hydrolyzation of silver carp by-products for 30 min by both enzymes were not significant in the present study (\(P>0.05\)). Calibration curve of five standard substances on the TSK-gel G3000 PWXL column (Tosoh Corporation, Yamaguchi, Japan, 2017) is shown in Figure 1a. Molecular weight distribution results (shown in Figure 1b) indicated that PH and AH demonstrated a wide variation in molecular weight (Mw) of \(\geq\)7500, 2027, about 1420, 286-780 and \(\leq\)138 Da, respectively. Relative proportions (%) of \begin{table}
\begin{tabular}{c c c c} \hline \hline \multirow{2}{*}{**Sample**} & **Crude Protein** & **Ash** & **Moisture** \\  & **(g/100 g)** & **(g/100 g)** & **(g/100 g)** \\ \hline Defatted by-products & 89.7 \(\pm\) 2.52 & 9.75 \(\pm\) 0.88 & 3.72 \(\pm\) 0.61 \\ PH & 85.2 \(\pm\) 3.11 & 9.74 \(\pm\) 0.12 & 3.52 \(\pm\) 0.14 \\ AH & 84.3 \(\pm\) 3.25 & 9.79 \(\pm\) 0.45 & 3.66 \(\pm\) 0.10 \\ \hline \hline \end{tabular}  
* PH represents the hydrolysate prepared by Protamex hydrolysis; AH represents the hydrolysate prepared by Alcalase hydrolysis; Mean values (\(\pm\)SD) of at least three measurements are shown.  
\end{table} 
Table 1: Proximate composition of the defatted by-products and hydrolyses. \begin{table} \begin{tabular}{c c c c c c c c} \hline \hline \multirow{2}{*}{**Sample**} & \multirow{2}{*}{**DH (\%)**} & \multicolumn{2}{c}{\(\zeta\) **Potential**} & \multicolumn{4}{c}{**Relative Proportion (\%) of Each Molecular Weight**} \\  & & **(mV)** & \(\geq\)**7500 Da** & **2027 Da** & **-1420 Da** & **286–780 Da** & \(\leq\)**138 Da** \\ \hline PH & 13.2 (\(\pm\)1.5) a  & -23.8 (\(\pm\)2.4) a  & 7.7 (\(\pm\)1.6) a  & 32.6 (\(\pm\)2.8) a  & 37.4 (\(\pm\)3.1) b  & 16.8 (\(\pm\)2.4) b  & 4.7 (\(\pm\)1.2) a  \\ \hline AH & 12.9 (\(\pm\)1.1) a  & -23.1 (\(\pm\)2.8) a  & 5.1 (\(\pm\)0.9) a  & - & 50.6 (\(\pm\)5.2) a  & 39.1 (\(\pm\)2.6) a  & 3.5 (\(\pm\)0.9) a  \\ \hline \hline \end{tabular}   
* Calibration curve of five standard substances on the TSK-gel G3000 PWXL column (Tosoh Corporation, Yamaguchi, Japan, 2017) is shown in Figure 1a. Molecular weight distribution results (shown in Figure 1b) indicated that PH and AH demonstrated a wide variation in molecular weight (Mw) of \(\geq\)7500, 2027, about 1420, 286–780 and \(\leq\)138 Da, respectively. Relative proportions (%) of each molecular weight are presented in Table 1. There were a small number of larger molecules (Mw \(\geq\) 7500 Da) in PH and AH, about 7.7 \(\pm\) 1.6 and 5.1 \(\pm\) 0.9%, respectively. Larger molecules and their proportion in the hydrolysates are crucial factors that affect gelling properties [4]. PH contained peptides at Mw of 2027 Da, which was absent in AH, at a relative proportion that reached 32.6 \(\pm\) 2.8%. Compared to PH, AH exhibited higher proportions of peptides at Mw of about 1420 and 286-780 Da (\(P\) < 0.05), which comprised of 50.6 \(\pm\) 5.2% and 39.1 \(\pm\) 2.6% of the hydrolysates, respectively.