Prebiotic Potential of Oligosaccharides and Polysaccharides Extracted from Leucaena leucocephala Seeds

Background/Objectives: Plant-derived soluble fibers are being explored as sustainable prebiotic ingredients; however, tropical legumes such as Leucaena leucocephala remain understudied. This study evaluated soluble fibers from L. leucocephala seeds after simulated gastrointestinal digestion, focusing on rheological properties, microbial selectivity, metabolite production, and intestinal safety. Methods: The anatomical parts of the seed underwent INFOGEST 2.0 digestion. Soluble fibers were characterized by GC-MS monosaccharide profiling, viscosity, and SEM/EDS analyses, and were used as substrates for both probiotic and pathogenic bacteria. Fermentation supernatants were analyzed for short-chain fatty acids and lactate, and cytotoxicity was assessed using Caco-2 cells. Results: Endosperm polysaccharides exhibited high apparent viscosity (>300 cP) and pseudoplastic behavior. Monosaccharide profiles revealed the presence of galacto-oligosaccharides and arabinoxylo-oligosaccharides in the oligosaccharide fraction, and galactomannans, xylans, and arabinoxylans in the polysaccharide fraction. Polysaccharides selectively promoted the growth of Lacticaseibacillus rhamnosus GG and Bifidobacterium spp., comparable to or exceeding that of fructo-oligosaccharides (p < 0.05), while limiting pathogenic bacteria. Fermentation produced acetate and lactate concentrations of >4500 ppm and >1000 ppm, respectively. Caco-2 viability remained >90% across all treatments. Conclusions: Compartment-resolved analysis identified the endosperm as the principal source of digestion-resistant viscous fiber, selectively fermented by probiotic bacteria at levels matching or exceeding fructo-oligosaccharides. These findings position L. leucocephala endosperm fiber as a candidate prebiotic substrate, warranting further preclinical evaluation.