Extraction and Functional Properties of Alfalfa Seed Protein for Human Food

The growing interest in sustainable protein sources has increased the demand for diverse and functional protein options. Alfalfa (Medicago sativa L.), a legume with a high protein content of 36.5% traditionally used as animal feed, alfalfa's potential as a protein source for food applications remains underexplored. This study optimizes alfalfa protein concentrate (APC) extraction from seeds using alkaline extraction process with varying buffers, NaCl concentrations, and pH levels (2, 10, and 12) across 15 treatments. Alfalfa Seeds, containing ~9.37% lipids, underwent hexane-based Soxhlet extraction, removing 9.25% of lipids prior to protein extraction. Antinutritional factors, a common challenge in legume proteins, were also evaluated before and after defatting. Defatted flour showed reduced phytic acid levels, though trypsin inhibitor concentration increased, and chymotrypsin inhibitor levels remained unchanged. Three extraction conditions (0% NaCl at pH 2, 10, and 12) yielded the highest protein recovery (19.39%, 34.91%, and 39.73%) and purity (60.36%, 58.32%, and 57.35%). Defatted flour-derived APC exhibited higher protein purity (69.76%, 67.84%, and 68.46%) than non-defatted samples. Total amino acid analysis revealed APC extracted at 0% NaCl, pH 10, had the highest amino acid content (71 g/100 g), and met FAO requirements for essential amino acids, suggesting as a complete protein. Protein solubility, a critical functional property, demonstrated promising results for APC (0%NaCl at pH10 and pH12), indicating enhanced bio-functional properties. Alfalfa seeds, particularly sprouts, contain L-canavanine, a non-protein amino acid potentially linked to autoimmune diseases like lupus. APC samples resulted below ( < 1%) in the previous report. Future studies will include in-vitro DIAAS (Digestible Indispensable Amino Acid Score) to assess the protein quality and the bioaccessibility of L-canavanine, followed by bioavailability studies to evaluate physiological effects. Comprehensive research into APC’s bio-functional and physiological properties will further solidify its potential as a sustainable protein source for food applications.