Fucoxanthin, a group of carotenoids abundantly present in the marine macro algae which exhibits diverse health benefits. Extraction of such potential biomolecules involves multiple steps in conventional extraction (CE) and also laborious and time-consuming. We attempted to extract high percentage fucoxanthin and phenolic compounds from macro alga, Padina tetrastromatica adopting ultrasound-assisted extraction (UAE). It was further optimized with solvent concentration; temperature and time as 80%, 50 degrees C and 30 min respectively using one factor at a time method (OFAT) followed by response surface methodology (RSM). It was observed that fucoxanthin yield of about 750 mu g g(-1) DW was obtained at the maximum by UAE than that of CE (430 mu g g(-1) DW). It was further purified and characterized using FTIR, HPLC and HRMS technique confirming the presence of purified fucoxanthin at the retention time of 3.14 min and molecular mass of 681.41 m/z. In addition, fucoxanthin exhibits a strong antioxidant activity by inhibiting DPPH and ABTS free radicals upon dose dependent manner. Interestingly, fucoxanthin showed protective effect against H2O2-induced intracellular ROS and cytotoxicity on fibroblast cells at the concentration of 120 mu M. Thusly, UAE could be implemented for the extraction at high percentage and eventually be utilized these biomolecules as nutraceuticals.