In order to study the effect of extraction pH on gelling properties and characteristic peptide identification of pigskin gelatin, the molecular mass distribution and gelling properties of pigskin gelatin extracted under different pH conditions were studied by dodecyl sodium sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), a texture analyzer and a rheometer. Moreover, the characteristic peptides were identified by high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). The results showed that with a decrease in pH, the relative molecular mass of pigskin gelatin initially increased and then decreased, the gel intensity gradually declined, and the gel melting temperature and gelling temperature initial increased and then decreased. Gelatin extracted at pH 1 had the worst gelling properties. HPLC-MS/MS analysis showed that pH significantly affected the traceability of gelatin. In this study, 62, 71, 79 and 76 characteristic peptides were detected from pigskin gelatins extracted at pH 1, 3, 5 and 7, respectively. Among them, 37 characteristic peptides were common to all pigskin gelatins. Compared with our previous research, 17 characteristic peptides were found to be common to pigskin gelatins extracted under different conditions. These stable common characteristic peptides could be used as an important basis for pigskin gelatin traceability with high accuracy.

In this study, by using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and double-sandwich enzyme-linked immunosorbent assay (ELISA), Juglans sigillata kernels from seven different production areas in Yunnan province were screened for their Jug r 1 contents. The preparation of Jug r 1 through ammonium sulfate precipitation, gel filtration chromatography (GFC), and liquid chromatography-tandem mass spectrometry (LC-MS/MS) was optimized and the product was identified. Its structure was characterized by circular dichroism (CD) spectroscopy. Finally, Jug r 1 content in walnut protein was measured by ELISA. The results showed that walnuts from Baoshan, which had the highest contents of total protein and Jug r 1, were used for subsequent experiments. The optimal concentration of ammonium sulfate saturation was in the range from 40% to 80%, and the optimal conditions for GFC were sample concentration of 30 mg/mL, loading volume of 4 mL, and elution flow rate of 1 mL/min. Under these conditions, the recovery of Jug r 1 was 16.58%. Mass spectrometry analysis showed that this protein had the typical characterisitcs of Jug r 1. Circular dichroism spectroscopy indicated that the secondary structure of Jug r 1 was dominated by α-helix and consisted of multiple conformations. After separation and purification by a two-step process, Jug r 1 with purity more than 96% was obtained. This study can provide a scientific basis for further research on Jug r 1 and also provide a reference for the separation and purification of allergic proteins from other nuts.