Abstract: To clarify the impact and contribution of sugar ester on the quality of flue-cured tobacco leaves, near-isogenic lines with the same genetic background but significantly different sugar ester contents were used for systematic comparative analysis. Appearance quality, conventional chemical composition, sensory quality, and untargeted metabolome were evaluated. The results revealed significant differences in both the appearance and intrinsic quality of flue-cured tobacco leaves with different sugar ester contents. Increasing the sugar ester contents can effectively enhance the sensory quality of the tobacco leaves, improve the remaining flavor, reduce irritation and offensive taste, but the increase of Ⅲ-Ⅴ sugar ester contents may have a certain negative impact on the appearance quality of tobacco leaves. The untargeted metabolome of tobacco materials with varying sugar ester contents exhibited clear and regular differences. As sugar ester content changed, primary metabolic pathways directly related to sugar ester synthesis, as well as secondary metabolic pathways such as flavonoids and alkaloids, showed significant coordinated changes. Compared with medium and low sugar ester materials, the aroma material basis of high sugar ester materials is richer, which may be an important reason for the best sensory quality performance. Therefore, sugar ester can be used as a key regulatory target for improving the sensory quality of tobacco leaves.