IntroductionTo investigate the mechanism of leaf color change at different stages inPopulus × euramericana‘Zhonghuahongye’ (‘Zhonghong’ poplar).MethodsLeaf color phenotypes were determined and a metabolomic analysis was performed on leaves at three stages (R1, R2 and R3).ResultsThea*, C*and chromatic light values of the leaves decreased by 108.91%, 52.08% and 113.34%, while the brightnessLvalues and chromaticb*values gradually increased by 36.01% and 13.94%, respectively. In the differential metabolite assay, 81 differentially expressed metabolites were detected in the R1 vs. R3 comparison, 45 were detected in the R1 vs. R2 comparison, and 75 were detected in the R2 vs. R3 comparison. Ten metabolites showed significant differences in all comparisons, which were mostly flavonoid metabolites. The metabolites that were upregulated in the three periods were cyanidin 3,5-O-diglucoside, delphinidin, and gallocatechin, with flavonoid metabolites accounting for the largest proportion and malvidin 3- O-galactoside as the primary downregulated metabolite. The color shift of red leaves from a bright purplish red to a brownish green was associated with the downregulation of malvidin 3-O-glucoside, cyanidin, naringenin, and dihydromyricetin.DiscussionHere, we analyzed the expression of flavonoid metabolites in the leaves of ‘Zhonghong’ poplar at three stages and identified key metabolites closely related to leaf color change, providing an important genetic basis for the genetic improvement of this cultivar.