Dietary supplements high in isolated isoflavones are commercially available for human consumption primarily to alleviate menopausal symptoms in women. The isoflavone composition, quantity and importantly their estrogenic potency are poorly standardised and can vary considerably between different products. The aim of this study was to analyse the isoflavone composition of 11 dietary supplements based on soy or red clover using the HPLC/MS/MS technique. Furthermore, we investigated the transactivational potential of the supplements on the estrogen receptors (ER), ERα and ERβ, performing luciferase reporter gene assays. As expected, we found that the isoflavone composition varies between different products. The measured total isoflavone contents in various supplements were mostly comparable to those claimed by the manufacturers in their product information. However expressing the isoflavone content as isoflavone aglycone equivalents, soy-based supplements had a clearly lower quantity compared to the manufacturer information. All supplements transactivated more or less ERα and ERβ with a preference for ERβ. The transactivational efficiency exceeded partly the maximal 17β-estradiol induced ER activation. While the different soy-based supplements revealed similar transactivation potential to both ERs, red clover-based supplements differed considerably. We conclude that different commercial dietary supplements based on soy or red clover vary in their isoflavone composition and quantity. They are estrogenically active, although especially the red clover-based supplements show considerable differences in their estrogenic potential to ERα and ERβ. Thus, different isoflavone-rich products cannot be necessarily compared regarding possible biological effects.