One of the fundamental carotenoid compounds available in microalga Chlorella pyrenoidosa is lutein, which is a commercially valuable antioxidant compound. The extraction of lutein from C. pyrenoidosa was studied in detail under various operating conditions. Acid pretreatment was done using 10% HCl. To analyze the effectiveness of ultrasound based extraction, experiments were carried out in presence and absence of ultrasonication. The results showed increased lutein yields in presence of ultrasonication. Scanning electron micrographs of pretreated and ultrasonicated algae cells showed increased surface area due to formation of porous surface. There were no significant contrasts in antioxidant activity of lutein between the samples obtained from ultrasound and acid treatment. Furthermore, to optimize the operating parameters like concentration of HCl, ultrasound exposure time and microalga biomass concentration, response surface methodology (RSM) design was applied. From the experimental results the optimum values derived are incubation time 41.34 min, HCl concentration 15.63 % and biomass concentration 309.21 g/L for a yield of 3.5 mg of lutein per gram of algae. Hence, ultrasound extraction with acid treatment is the most efficient technique for the extraction of microalgal lutein.