# Study of Physicochemical and Gelatin Properties of Fish Gelatin from Different Sources
## 3 Results and Discussion
### Structural Detection of Self-Assembled and Enzymatic Gels  
Fourier transform infrared spectroscopy (FTIR) is a common technique for determining the secondary structure of proteins. FTIR spectra correlate with the vibrational state of chemical bonds in proteins [31]. As shown in Figure 4, the peaks and bands of amides A, I, II and III were similar between the FTIR spectra of the four gelatin samples, indicating that the four fish gelatin samples had similar secondary structures of proteins.  
We observed that the absorption peak of amide A that is attributed to the N-H stretching vibration was 3400-3440 cm\({}^{-1}\). When the N-H group was involved in hydrogen bonding, the absorption peak of amide A shifted to a lower position, with stronger hydrogen bonding causing a greater shift. We found that the position of the amide A absorption peak was essentially the same for all samples. However, as crosslinking by TG enzyme generated more amide bonds and increased covalent bonding energy in gelatin [32], we detected that the amide A absorption peak in enzymatic gels was mainly shifted to a lower position. We specifically observed a greater shift in the distance of the amide A peak between the self-assembled and enzymatic gels, suggesting changes in the physical and chemical properties of gelatin and enhancement of gel strength.  
\begin{table}
\begin{tabular}{c c c} \hline \hline
**Gelation** & **Water (\%)** & **Oil (\%)** \\ \hline M03 & 179 \(\pm\) 12 f & 157 \(\pm\) 5 f \\ M04 & 245 \(\pm\) 15 e & 122 \(\pm\) 4 g \\ M06 & 6 \(\pm\) 11 g & 155 \(\pm\) 6 f \\ M08 & 14 \(\pm\) 13 g & 253 \(\pm\) 3 c \\ M03+TG & 786 \(\pm\) 13 b & 247 \(\pm\) 4 d \\ M04+TG & 527 \(\pm\) 11 c & 286 \(\pm\) 5 b \\ M06+TG & 906 \(\pm\) 14 a & 293 \(\pm\) 3 a \\ M08+TG & 327 \(\pm\) 12 d & 180 \(\pm\) 6 e \\ \hline \hline \end{tabular} \(p\)-values \(<\) 0.05 were considered statistically significant. The result of abcd’s significant difference analysis on the data is obtained by SPSS software. As long as there are the same letters, there is no significant difference. The data has significant difference marked from a to d.  
\end{table}
Table 5: Water-holding properties of gelatin and its oil-holding properties.  
We tested the diffraction effect of gelatin by X-ray diffraction (XRD), as shown in Figure 5. We found that self-assembled gels exhibited two diffraction peaks, a small peak at 7\({}^{\circ}\) (20), the intensity of which reflected the triple helix content in gelatin, and a stronger broad peak at 20\({}^{\circ}\) (20) attributed to the large number of polar groups in the gelatin molecular chain, which will form intra- or intermolecular hydrogen bonds. The highly ordered arrangement of these hydrogen bonding networks can transform gelatin into an amorphous structure (non-crystalline structure) with a uniform polymeric network structure. We detected that the position of the diffraction peak at 20\({}^{\circ}\) did not change between gels, with the intensity of the amorphous characteristic peaks of these four gelatins being continuously weakened from high to low, thereby preventing the formation of an ordered microcrystalline structure. We further observed that after crosslinking by the TG enzyme, the position and intensity of the small peak at 7\({}^{\circ}\) (20) of the enzymatic gel did not differ from that of the self-assembled gel, indicating that the triple helix content remained unaltered. However, as crosslinking by the TG enzyme increased the hydrogen bonding between gelatin molecules, we noticed that the peak intensity at 20\({}^{\circ}\) in the enzymatic gel was decreased compared with that in the self-assembled gel, suggesting an increased structured order in the enzymatic gel.  
Figure 4: Infrared map of gelatin raw material and enzymatic gel (**A**) gelatin raw material self-gel; (**B**) TG enzyme cross-linked.  
Figure 5: X-ray diffraction (XRD) plots of self-gel and enzymatic gel (**A**) self-gel; (**B**) enzymatic gel.