Abstract: In this study, AMF was fractionated into various fractions with different melting points using a multi-stage gradient dry fractionation approach. The physicochemical properties of the AMF fractionates, including composition, solid fat content, slip melting point, thermal properties, and microstructure, were determined to investigate the mechanism by which the fractionation process affects these properties. Two typical gradient fractionation strategies were adopted to compare the properties of the final products. The results indicated that as the fractionation temperature increased, the long-chain saturated fatty acids in the solid fractions gradually increased, while the unsaturated fatty acids and short- to medium-chain saturated fatty acids gradually decreased. Regarding thermal properties, the fractionation temperature significantly impacted the thermal behavior of the fractionates. Both the crystallization and melting temperatures of all fractions increased significantly with rising fractionation temperature. In terms of crystal microstructure, the fractions were dominated by the β′ form, with crystals packed in a double chain length, representing the common stable crystal structure of milk fat products. At the same fractionation temperature, the saturated fatty acid content of H-S35 reached 72.73%, which was 4.35 percentage points higher than that of C-S35. Furthermore, the slip melting point of H-S35 was 41.1℃, which was 1.9℃ higher than that of C-S35. The solid fractionates obtained by fractionation under progressively increasing temperatures had significantly higher saturated fatty acid contents and slip melting points than those prepared under progressively decreasing temperatures. These results verify that temperature-rising fractionation is more conducive to enriching high-melting-point components in solid fat fractions. This study provides a theoretical basis and technical support for the deep processing and high-value product development of AMF, promoting its industrial application in bakery fats and other food fields.