Abstract: To clarify the metabolites that affect the appearance and sensory quality of flue-cured tobacco from Guizhou Province, a total of 44 tobacco leaf samples with distinct appearance characteristics, harvested in 2021 from Guizhou, were prepared on the principle of unique factor differences. The appearance and sensory quality of these samples were evaluated, and their routine chemical components and metabolites were determined. The correlations between the metabolites and the appearance and sensory quality were investigated through correlation analysis, principal component analysis and cluster analysis. The results showed that: 1) Three appearance quality indicators, i.e., leaf structure, body and oil content, were closely correlated with the sensory quality of the samples. 2) L-lysine, D-glucose, myristic acid, melezitose, lactic acid, trehalose, and sucrose were closely correlated with the sensory quality of the samples. Specifically, trehalose and sucrose were closely correlated with both leaf structure and body. 3) The results of both correlation analysis and metabolite differential analysis indicated that seven metabolites, including L-lysine, D-glucose, myristic acid, melezitose, lactic acid, trehalose, and sucrose, might be the key metabolites affecting the sensory and appearance quality of flue-cured tobacco leaves. The above findings provide a reference for industrial enterprises aiming to efficiently utilize tobacco leaves from Guizhou Province.