Background: Hypericum heterophyllum has been used as a traditional medicine and includes significant bioactive compounds. Objective: To quantitatively determine the phenolic and flavonoid contents of Hypericum hetero-phyllum flowers and evaluate the antioxidant activity. Also, to present the activity-compound relationship. Methods: Flowers of H. heterophyllum Vent., endemic species for Turkey were collected from Yozgat-Turkey. H. heterophyllum flowers were extracted with methanol, ethanol, acetone, and chlo-roform individually. LC-ESI-MS/MS analysis for phytochemicals was carried out for each extract. Total phenolic and flavonoid content were carried out for each extract. The antioxidant activity was performed using DPPH•, ABTS•+, hydroxyl radical, and superoxide radical scavenging assays. Results: Chlorogenic acid was found as a major compound in all extracts. Among the extracts, methanol extract included the most phenolic compound. The methanol extract and acetone extract consisted of the most flavonoids. Acetone and ethanol extracts revealed the most DPPH and ABTS activity respectively. Acetone extract exhibited the highest hydroxyl radical and superoxide radical scavenging effect Conclusion: Flowers of H. heterophyllum contain bioactive compounds and they revealed good antioxidant activity. Hence, H. heterophyllum flowers could be a promising agent for the pharmaceutical and food industries.