This study was focused on the identification and quantification of betaxanthins using high-performance liquid chromatography with a diode array detector coupled to electrospray ionisation mass spectrometry (HPLC-DAD-ESI-MS) in leaves of various cultivars of Beta vulgaris (beet) and Gymnocalycium mihanovichii grafted cacti. In G. mihanovichii grafted cacti, four betaxanthins, namely histidine-Bx, histamine-Bx, serine-Bx, and proline-Bx, were tentatively identified in the yellow, orange, pink, and red varieties, with contents ranging from 0.09 to 1.55 mg/kg fresh weight (FW). Betaxanthins were not detected in the green cactus. Histidine-Bx was the prevailing betaxanthin compound in the majority of cultivars. Fifteen betaxanthins were successfully identified in the leaves of five B. vulgaris cultivars (cv.): Snow Ball, Boldor, Cylindra, Rhubarb, and Round Dark Red. Leaves of yellow beet (cv. Boldor) had the highest total betaxanthin content (20.4 mg/kg FW), while white beet (cv. Snow Ball) had the lowest one (3.43 mg/kg FW). The leaves of red cultivars had comparable betaxanthin contents, ranging from 13.4 to 18.8 mg/kg FW, similary to the yellow cultivar, indicating their potential as valuable sources of betaxanthins. There was no single dominant betaxanthin in B. vulgaris leaves. The leaves of B. vulgaris were found to be a richer source of betaxanthin than the grafted cactus G. mihanovichii, which opens up new prospects for research into the potential utilization of these compounds in the industry.