The rotifer Brachionus plicatilis is the first live feed in larviculture of marine fish species. Rotifer diets differ in their biochemical composition, physical properties, and production technology while feeding protocols largely vary among facilities. The objective of the present study was to determine the effects of two different forms of Nannochloropsis oculata and commonly used commercial diets on growth performance and biochemical composition of rotifers produced under commercial conditions. Rotifers were fed one of five different types of feed: Algome (R) (dried Schizochytrium sp.), Protein Plus (R) (PP), Inactive Baker's Yeast (R) (INBY), spray-dried Nannochloropsis oculata (SDN), or freshly cultured Nannochloropsis oculata (FN). Rotifers fed SDN diet resulted in significantly higher rotifer biomass during 16days of semi-continuous culture, with an increasing biomass trend that lasted 11days, high egg production, and egg-carrying female numbers, whereas rotifers fed PP showed highest Sigma n-3, arachidonic acid, eicosapentaenoic acid, and docosahexaenoic acid contents. Amino acid profiles of rotifers were enhanced by utilization of both INBY and SDN diets. Overall, the results indicated that SDN is optimal for long-term biomass production of rotifers. However, their nutritional profile needs to be enriched by feeding PP (EFA source) and INBY (EAA source) once desired biomass production is obtained.