Abstract Nannochloropsis oculata is naturally rich in eicosapentaenoic acid (EPA). To turn this microalga into an economically viable source for commercial applications, extraction efficiency must be achieved. Pursuing this goal, emerging technologies such as high hydrostatic pressure (HHP) and moderate electric fields (MEF) were tested, aiming to increase EPA accessibility and subsequent extraction yields. The innovative approach used in this study combined these technologies and associated tailored, less hazardous different solvent mixtures (SM) with distinct polarity indexes. Although the classical Folch SM with chloroform: methanol (PI 4.4) provided the highest yield concerning total lipids (166.4 mglipid/gbiomass), diethyl ether: ethanol (PI 3.6) presented statistically higher values in terms of EPA per biomass, corresponding to 1.3-fold increase. When SM were used in HHP and MEF, neither technology independently improved EPA extraction yields, although the sequential combination of technologies did result in 62% increment in EPA extraction. Overall, the SM and extraction methodologies tested (HHP—200 MPa, 21 °C, 15 min, followed by MEF processing at 40 °C, 15 min) enabled increased EPA extraction yields from wet N. oculata biomass. These findings are of high relevance for the food and pharmaceutical industries, providing viable alternatives to the “classical” extraction methodologies and solvents, with increased yields and lower environmental impact. Key points • Et2O: EtOH is a less toxic and more efficient alternative to Folch solvent mixture • HHP or MEF per se was not able to significantly increase EPA extraction yield • Combinations of HHP and MEF technologies increased both lipids and EPA yields Graphical abstract