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The Amazonian <i>Pentaclethra macroloba</i> (Willd.) Kuntze nuts contain a lipidic fraction with health-promoting effects, but little is known about the bioactivity of other constituents. In this study, the lipidic fraction obtained using supercritical fluid extraction (SFE) with CO₂ was chemically characterized by using lipidomics techniques. The SFE-CO₂ residue, named as pracaxi cake, was re-extracted by pressurized liquid extraction following a biorefinery approach. Using a response surface methodology and based on the extraction yield and different <i>in vitro</i> assays, two optimum conditions were obtained: 80% and 12.5% of ethanol at 180 °C. Under these conditions, extraction yield and different <i>in vitro</i> measurements related to neuroprotection were assessed. Chemical characterization of these extracts suggested the presence of triterpenoid saponins and spermidine phenolamides, which were not previously reported in pracaxi nuts. These results suggest that pracaxi oil extraction by-products are a valuable source of bioactive compounds with neuroprotective potential.