Objective To investigate the biological characteristics and in vitro probiotic functions of the food-derived Pediococcus acidilactici N8, and to screen effective protectants for the preparation of high-viability (>10¹² CFU/g) spray-dried probiotic preparations.Methods A lactic acid bacterium (strain N8) isolated from cheese was identified through morphological analysis and 16S rDNA gene sequencing. Its probiotic potential was determined through tests on growth characteristics, safety, tolerance, antibacterial activity, adhesion ability, and antioxidant capacity. Spray drying was employed to prepare probiotic formulations, and high-viability spray-drying protectants were screened.Results Based on morphological characteristics and 16S rDNA gene sequence analysis, strain N8 was identified as P. acidilactici. It exhibited a rapid S-shaped growth curve and probiotic traits including non-hemolytic activity and sensitivity to multiple antibiotics. Its supernatant effectively inhibited the growth of various pathogenic bacteria. The strain showed strong stability under heat, varying pH levels, bile salts, and simulated gastrointestinal fluid conditions, and possessed strong adhesion ability. Both the cell-free supernatant and intact cell suspension of P. acidilactici N8 demonstrated strong scavenging abilities against DPPH, ABTS, and hydroxyl radicals. When buffalo milk, Cordyceps militaris polysaccharides, and sodium glutamate were used as protectants, the viable count of the probiotic preparation reached 10¹² CFU/g.Conclusion P. acidilactici N8 is a food-derived lactic acid bacterium with good probiotic potential. Using buffalo milk, C. militaris polysaccharides, and sodium glutamate as spray-drying protectants can yield high-viability probiotic preparations.