Objective · Both Lycium barbarum L. (LB) and Lycium ruthenicum Murr. (LR) belong to the Lycium genus of Solanaceae family but their fruits have significant phenotypic differences in terms of color and shapes. This study is aimed to investigate the inter-species difference of their fruit polyphenol composition. Methods · The polyphenol composition of fresh and dried fruits of two Lycium species were comprehensively analyzed using ultra-high performance liquid chromatography-electrospray ionization-quadrupole-time of flight mass spectrometry (UHPLC-ESIQ-TOF-MS). Results · 35 polyphenolic compounds were detected and quantified in the fresh and dried Lycium fruits including phenolic acids, anthocyanins and phenolamides (i.e., the amide-adducts of polyphenolic acids and polyamines). For both species, the dried fruits had more polyphenolic compounds than their fresh fruits with significant inter-species difference; for both fresh and dried fruits, LR had more polyphenolic compounds than LB. Conclusion · There are significant inter-species differences between these two Lycium in both fresh and dried fruits. These results offer essential information for further understanding of the biological functions of these two Lycium fruits as phytomedicines and functional food.