Introduction: Arenaria kansuensis Maxim. (AKM) is one of the most valued medicinal and edible herbs widely used in Qinghai-Tibet Plateau and there is also a large number of AKM bioactive constituents for health benefits of human beings. However, few works have referred to phytochemical content, fingerprint analysis and quality control of AKM. Therefore, the establishment of validated analytical methods is urgently needed for fingerprint comparison and quantitative analysis of AKM multicomponent. Objectives: To determine quantitatively and compare the phytochemical constituents of AKM located at different areas. Methodology: The chemical constituents in AKM samples were separated, identified, and quantified by high-performance liquid chromatography (HPLC) with a diode array detector. The discrimination and separation models for the chemical constituents were developed by chemometric analysis. Results: The flavones and β-carboline alkaloids were rich in AKM herbs, and the overall pattern of phytochemical profiles was the same, while the significant differences were detected in the total flavonoids, total β-carboline alkaloids and individual contents, especially the predominant compounds such as tricin and arenarine B. This demonstrated that ecogeographical origin gave an important impact on phytochemical compositions which could be considered as reliable parameters for classifying the AKM resources. Moreover, the contents of AKM constituents were higher in July and/or August than other months of the year, and there were no significant differences in the main phytochemical contents between cultivated and wild AKM herbs. Conclusion: This study could provide credible data and method for geographical origin trace, comprehensive evaluation and further utilization of AKM resources. © 2021 John Wiley & Sons, Ltd.