Objective: The differences in the chemical composition of Morus nigra (M. nigra) extracts from four different extraction methods, ultrasound-Assisted extraction with pure water (WU), pure water decoction extraction (WD), ultrasonic-Assisted extraction with formic acid water (FAU), and pure water heat reflux extraction (WHR), were identified using ultra-high performance liquid chromatography-quadrupole time-of-flight tandem mass spectrometry (UPLC-Q-TOF-MS/MS) technology. Method: A Waters ACQUITY BEH C18 (1.7 μm, 2.1 mm × 100 mm) was used, with a column temperature of 45°C, mobile phase of methanol and 0.1% formic acid aqueous solution, and gradient elution with a flow rate of 0.4 mL/min. Detection was performed in positive and negative ion modes, and compounds were identified using Progenesis QI software and mass spectrometry data reported according to the literature and laboratory self-built databases of the Mulberry genus. Multivariate statistical techniques, such as principal component analysis and orthogonal partial least squares-discriminant analysis, were applied to differential cluster metabolic profiles and chemical components and to screen the differential chemical components of M. nigra leaves. Results: There were significant differences in the chemical composition between WD and the other extraction methods of M. nigra leaves. A total of 13 differential metabolites (4 flavonoids, 3 organic acids, 3 phenylpropanoids, 2 alkaloids, and 1 trisaccharide) were identified. Conclusion: The multivariate statistical analysis and UPLC-Q-TOF-MS/MS method established in this study identified the differential chemical constituents of Xinjiang M. nigra leaves using different extraction methods, which provides a basis for the quality control of M. nigra leaves, and provides basic data for revealing the influence of extraction methods on the synthesis and accumulation of M. nigra leaf metabolites, which has certain reference significance. © 2023 the author(s), published by De Gruyter.