<p>The environmental crisis and global population growth have intensified efforts across multiple sectors to extend production chains by applying the potential of industrial biomasses, thereby enabling the use of “second-generation” food inputs rich in proteins and reducing the disposal of agro-industrial residues into the environment. In this context, the objective of this research was to evaluate the quality of industrial Brazil nut (<i>Bertholletia excelsa</i> HBK) biomass through protein isolation based on green chemistry principles. The methodology followed the standards of the Association of Official Analytical Chemists (AOAC) and other internationally recognized protocols. The results showed a yield of 43% for the Brazil nut protein isolate with a protein content of 88.14%. Its amino acid profile exhibited notably high levels of sulfur-containing amino acids (14.14%) and branched-chain amino acids (36.60%). FTIR analysis revealed bands characteristic of nitrogen-containing groups (N–H and C = O) such as type I, II, and III amines and amides. Thermogravimetric analysis confirmed thermal stability at processing-relevant temperatures. The techno-functional evaluation demonstrated high capacities for water absorption (195%), oil absorption (209.4%), foam formation (167.8%), and emulsifying activity (42.6%). These findings update research conducted more than two decades ago, demonstrating methodological improvements that increased yield and expanded knowledge of the nutritional and techno-functional potential of this biomass. The results support the extension of production chains within the framework of the circular bioeconomy of Amazonian agroextractivism, contributing simultaneously to environmental preservation and socioeconomic development.</p>