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The effect of homogenization, ultrasound and microwave extraction methods and conditions on fucoxanthin content, total phenolic content and antioxidant activity of extracts obtained from Phaeodactylum tricornutum were investigated in this study. The solvent/biomass ratio was the most effective parameter on fucoxanthin content, total phenolic content and antioxidant activity. The maximum fucoxanthin content (5.60±0.06 mg/g) and antioxidant activity (763.00±15.88 EC50 µg/mL extract) were obtained with the homogenization extraction method whose optimum conditions were 1.93% biomass/solvent ratio, ~5200 rpm homogenization rate and 14.2 min extraction time. Although the ultrasonic extraction method has reached the approximately same level of fucoxanthin content (5.24±0.07 mg/g)), TPC (67.68±1.58 mg gallic acid/L) and activity (619.90±17.16 EC50 µg/mL extract) at an amplitude of 55.72%, a higher biomass/solvent ratio (2.72%) and a longer extraction time (17.37 min) have been required. The lowest fucoxanthin content, total phenolic content and antioxidant activity were determined for the microwave extraction method.