The study aimed to characterize açaí seeds and explore their potential for producing mannooligosaccharides (MOS) through enzymatic hydrolysis. According to characterization tests by XRD, FTIR, and chemical analysis, acai seeds, a waste material from acai fruit processing, contain significant hemicellulose content, with a main content based on mannan. The study utilized Rohalase®GMP enzyme for hydrolysis and monitored reducing sugars and MOS production (8–10 g MOS/L) hydrolysate with varying lengths, including di-, tri-, tetra-, and pentasaccharide over time. Results showed a concentration plateau of reducing sugars at 13 h and increased MOS until hour nineteen. Mannobiose (M2) was the predominant MOS produced. Comparative analysis with prior research indicated that the açaí seed hydrolysate’s MOS content aligns with that from other sources. Importantly, the study achieved % hydrolysis and MOS yield of 10.79 ± 0.05% and 51.39 ± 0.11 g M2-M5/g mannan, respectively. Açaí seed is a promising source to produce enzymes, biofuels, or thermal energy, as well as highly valued chemical compounds in the industry. This work additionally demonstrates its potential as a sustainable source of valuable MOS, suggesting applications as prebiotics and functional food additives, with implications for various industries seeking eco-friendly alternatives.