Abstract: In order to explore the changes in nutritional ingredients of honey during the natural ripening process, the contents of moisture, monosaccharide, disaccharide, 9 kinds of oligosaccharides and 18 kinds of polyphenols in acacia honey were determined by the Abbe refractometer, high performance liquid chromatography(HPLC), gas chromatography-mass spectrometry(GC-MS) and high performance liquid chromatography-tandem mass spectrometry(HPLC-MS/MS), respectively, combined with hierarchical cluster analysis(HCA) and principal component analysis(PCA) to distinguish acacia honey during different ripening days. The results showed that during the ripening process of acacia honey, the contents of moisture, oligosaccharides and polyphenols changed significantly. Among them, the contents of moisture, kestose, and melezitose decreased by 5.77, 0.09, and 3.47 g/100 g, respectively, while maltulose, turanose, erlose, and isomaltose increased 0.85, 1.65, 2.43, and 0.25 g/100 g, respectively. For the polyphenols, the contents of rutin, kaempferol, protocatechuic acid, and p-hydroxybenzoic acid generally increased; gallic acid, morin, ferulic acid, galangin, caffeic acid phenethyl ester, caffeic acid, and chrysin increased first and then decreased; benzoic acid and p-coumaric acid decreased first and then increased. In addition, the contents of melezitose under different ripening days of acacia honey were reduced significantly(P<0.05), while p-hydroxybenzoic acid and caffeic acid increased significantly(P<0.05), which could be used as potential characteristic indexes for judging the ripeness of acacia honey. The chemometrics analysis results showed that oligosaccharides and polyphenols could obviously distinguish acacia honey during the different ripening days.