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A bioactive <i>Pleurotus eryngii</i> aqueous extract powder (SPAE) was obtained by spray drying and its performance in terms of physicochemical properties, in vitro digestion, inflammatory factors, and modulation of the intestinal microbiota was explored. The results indicated that the SPAE exhibited a more uniform particle size distribution than <i>P. eryngii</i> polysaccharide (PEP). Meanwhile, a typical absorption peak observed at 843 cm⁻¹ in the SPAE FTIR spectra indicated the existence of α-glycosidic bonds. SPAE exhibited higher antioxidant abilities and superior resistance to digestion in vitro. In addition, SPAE supplementation to mice significantly reduced the release of factors that promote inflammation, enhanced the secretion of anti-inflammatory factors, and sustained maximum production of short-chain fatty acids (SCFAs). Additionally, it significantly enhanced the relative abundance of SCFAs-producing <i>Akkermansia</i> and reduced the abundance of <i>Ruminococcus</i> and <i>Clostridiides</i> in intestines of mice. These results show the potential of SPAE as a novel material with prebiotic effects for the food and pharmaceutical industries.