Jiang-Flavor Daqu (JFDQ) is a grain-type fermented starter for brewing Chinese liquor. Peptides, the metabolites of proteins in JFDQ, are important for the quality and flavor of JFDQ or even the liquor. The peptide variations in the progress of JFDQ preparation were investigated using RPLC-MS/MS. The JFDQ after high-temperature fermenting (HTF_SU) and after ripening (M_SU), as well as the mature JFDQ prepared in spring (M_SP) and in summer (M_SU), were compared respectively. These two groups were investigated from peptides, precursor proteins, abundance, interactions, and potential antimicrobial peptides (pAMPs). A total of 177, 158, and 262 peptides from HTF_SU, M_SP, and M_SU were identified, respectively. Significant differences (P < 0.01) in the abundance of shared peptides were found in different fermentation stage group (HTF_M), and stronger positive correlations were observed in different preparation season group (MSP_MSU). The interactions of the shared peptides in HTF_M and in MSP_MSU were investigated respectively. In addition, 8 pAMPs in HTF_SU, 5 in M_SP, and 22 in M_SU were predicted using CAMPR3, and their core functional regions were analyzed. This systematic study demonstrated the influences of fermentation stage and preparation season on the peptide profiles in JFDQ, which would provide theoretical guidance and be helpful for JFDQ production. SignificancePeptidomics analysis showed that the peptide profiles of JFDQ varied in different fermentation stages and different preparation seasons, which mainly resulted from the peptides with high abundance, high interaction degrees, and potential antimicrobial activity, as well as the important precursor proteins such as glutens. This systematic study would benefit for the insufficiency of peptide research of JFDQ till now, and provide theoretical guidance for JFDQ production.