Paeonia Radix Rubra (PRR) is a very common traditional Chinese medicine (TCM). The roots of Paeonia lactiflora and Paeonia anomala subsp. veitchii are used for the production of PRR. However, other species of section Paeonia in China are also used to produce PRR. The roots of section Paeonia from 20 populations of seven species and two subspecies in China were analyzed by high performance liquid chromatography quadrupole time-of-flight mass spectrometry (HPLC-Q-TOF- MS). A total of 21 metabolites were identified, including nine monoterpene glycosides, seven tannins, three phenols, one paeonol and one flavonoid. There were significant differences in the composition and content of metabolites among different populations. The relative contents of monoterpene glycosides and tannins were generally higher in most samples. Cluster analysis showed that 20 populations could be divided into four groups. Among them, the populations of P. lactiflora and Paeonia mairei were clustered together in one group. The content of paeoniflorin in P. lactiflora was high (>22.20 mg g-1, dry weight, the same below), and the content of lactiflorin in P. mairei (>17.50 mg g-1) was significantly higher than in other species (51.60 mg g-1) was significantly higher than in other species (<43.40 mg g-1), suggesting that it could be useful medicinal germplasm for the development of monoterpene glycosides. In addition, some populations of Paeonia emodi, Paeonia sterniana and P. mairei may also be used as potential germplasm resources for use in PRR. Genetic and environmental factors resulted in differences in the composition and content of metabolites in different populations of the same species. Therefore, it is necessary to carefully consider the origin of Paeonia germplasm when selecting medicinal materials to be used as resources for the production of PRR.