In order to disclose the phylogeny and genetic diversity of wild wolfberry germplasm resources in Ningxia and Shaanxi Province, as well as the mechanism on taste of mature fresh fruits (i.e., bitter and sweet), the substances of wild bitter Lycium, several methods including systematic investigation, SLAF sequencing technology, ultra-high performance liquid chromatography quadrupole time-of-flight high-resolution mass spectrometry (uplc-q / TOF-MS) and ultraviolet wavelength identification were used in this study. The results showed that from Qishan County and Fufeng County in Shaanxi Province to the mountainous area in Southern Ningxia, the Yellow River Ningxia Basin and the eastern foot of Helan, the mature fresh fruits showed a general trend from bitter to half bitter, bitter after sweet (or hemp bitter after sweet), sweet, slightly bitter after sweet and whole bitter then. The phylogenetic tree analysis revealed five groups, consisting of Group I, group II, group III and group IV that were different types of wild bitter wolfberry. Group V was a large number of wild germplasm with a mixture of germplasm showing bitter to sweet taste. There is a genetic diversity in the wild bitter wolfberry ceollection. By analyzing the liquid total ion flow diagram from the four samples (Ningxia sweet wolfberry cultivar ‘Ningqi 7’, ‘Haiyuan wild bitter wolfberry’, ‘Xiji wild bitter wolfberry’ and ‘Shaanxi Qishan wild bitter wolfberry’), we observed an absent signal in the 30-50min region in ‘Ningqi 7’ if compared to other samples. These chemical components mainly were steroidal alkaloids, namely Solanine, and 5,6-dihydrosolanine, which might be causal agents resulting in bitter taste in wild bitter wolfberry. Collectively, these results of this study provided a theoretical basis for in-depth study and utilization of wild bitter wolfberry germplasm and new cultivar breeding for medicine.