This study employed gas chromatography-mass spectrometry with olfactory (GC-MS-O) and multi-omics methods to investigate the changes in volatile flavor compounds during the freezing process of Pacific chub mackerel (Scomber japonicus) from Japan and China, and Spanish mackerel (Scomberomorus niphonius). A total of 18 volatile flavor compounds were identified, and significant differences in volatile flavor components were observed among samples frozen for 1 week, 1 year, and 2 years. The results of the Partial least squares regression (PLSR) indicated that the fishy odor was correlated with independent variables such as fatty acids (FA 22:4, FA 28:6, FA 24:4), differentially expressed genes (Gene.2425 (NDUFA5), Gene.38 (GPX1), and Gene.2844 (DAD1)). Classification and regression tree (CART) analysis revealed that the peak area values of fatty acids (FA 22:5, FA 20:4) and fatty acid esters of hydroxy fatty acids (FAHFA 18:0/22:3) were the main differentiating factors for fishy odor perception.