Introduction: Eclipta alba Linn. (Asteraceae) is an important ingredient of several Ayurvedic formulations. The monograph on different parts of plant like flowers, leaves, roots are listed in Ayurvedic pharmacopeia of India. The plant is reported to be effective for broad range maladies like inflammation, reproductive problems of females etc. It is also used as a hepatoprotectant, analgesic, antibacterial and antidiabetic agent. Wedelolactone is used as a bioactive marker to establish the quality of the crude drug and its formulations. In the present study, wedelolactone-based standardization of Eclipta alba and its quantitation from marketed herbal and Ayurvedic formulations has been documented using RP-HPLC. Methods: In the current work, an isocratic method has been developed and validated to quantitate wedelolactone from whole plant of Eclipta alba. This method is validated as per ICH guidelines and is used to quantitate the content of wedelolactone in polyherbal formulations like Liv52, Geriforte, Mahabhringaraj oil etc. Results: The LOD is found to be 0.5 μg/mL and the LOQ is 1 μg/mL. The linearity range of response is from 5 μg/mL to 100 μg/mL. Conclusion: The validated method is found to be simple, sensitive, accurate, rugged and reproducible. This developed method can be recommended for quality assurance and marker-based standardization of polyherbal formulations containing whole plant of Eclipta alba.