**1. Introduction**

Grouper (*Epinephelus coioides*), which belongs to the order Perciformes and the family Serranidae, is a warm-water, offshore demersal fish that is referred to as "marine chicken" because of its considerable similarity in taste to chicken meat. With the development of artificial breeding and breeding technology, grouper has become an important economic fish along the coast of China [1]. Furthermore, the living standards of residents have been significantly enhanced as the economy has developed. Due to the quickening pace of life of the younger generation, fresh fish fillets are becoming the main marketing form of fish products [2,3]. However, grouper can easily decompose, because of its abundant nutrition, high water content, and excellent protease activity. Protein is one of the most important nutrients in aquatic products. During cold storage, the changing forms of protein mainly include protein oxidation and degradation. Carbonyl compounds formed by protein oxidation can change the cell structure of the myofibrillar protein, affecting the hydrophobicity index of fish protein [4,5]. Soon after fish die, the protein initially breaks down into many intermediates with the hydrolysis of endogenous

protease [6,7]. In the later stages, the enzymes produced by microbial reproduction lead to protein degradation, resulting in spoilage of the fish. Therefore, the change of biochemical characteristics of proteins in aquatic products is an important reason for the deterioration of such products' quality during cold storage [8]. Refrigerated fish fillets need to be packaged to ensure their freshness and to meet the needs of different customers. Presently, the main packaging methods of grouper are air packaging (AP), vacuum packaging (VP), and modified atmosphere packaging (MAP). AP and MAP have been considered to be effective ways of preserving food, due to the excellent isolation of oxygen and food [9,10]. MAP, due to the bacteriostatic effect of CO2 inducing anaerobic conditions into the packaging environment, can effectively reduce apparent changes of aquatic products [11]. In our previous study, it was found that a high concentration of CO2 in MAP (30% N2 and 70% CO2) could prolong the shelf life of cold-stored grouper. Currently, research on the effects of VP and MAP on the quality of aquatic products mainly focuses on the quality attributes, shelf-life assessment, changes of proteins in aquatic products during storage, and the denaturation of proteins [12–14]. Also, trichloroacetic acid (TCA)-soluble peptides, the myofibril fragmentation index (MFI), free amino acids (FAAs), SDS-PAGE, and FTIR of myofibrils are usually evaluated during storage.

However, the relationship between protein oxidation and degradation is controversial. On the one hand, the hydrophobicity caused by protein oxidation increases protease recognition and the subsequent oxidation of protein degradation [15]. On the other hand, the polymer caused by protein oxidation between protein crosslinking could affect the further degradation of proteins, and is a poor substrate of protease [16]—something which has not been studied in research on grouper. It is not clear if MAP or VP can isolate oxygen as an effective method of food preservation and slow down the progress of protein oxidation in grouper. Therefore, the purpose of this study is to investigate the relationship between protein oxidation and degradation, and to compare the preservation effects of protein oxidation and degradation among AP, MAP, and VP refrigerated storage on grouper fillets.
