Aglycone-type soy isoflavones, recognized for their bioactive phytoestrogen properties, face industrial limitations due to their low natural abundance and inefficient conversion. This study optimized a multi-enzyme synergistic catalysis system using soybean sprout powder, achieving high conversion rates and purity through response surface methodology. The optimal enzyme system comprised β-glucosidase (25 U/mL), cellulase (200 U/mL), hemicellulase (400 U/mL), and β-galactosidase (900 U/mL) at pH 5.0, 50 °C, and 3.2 h. This system yielded an aglycone conversion rate of 92% and glycoside hydrolysis rate of 97%, outperforming single-enzyme approaches. Upon post-purification with AB-8 macroporous resin, the product reached a purity of 58.1 ± 0.54% and exhibited strong antioxidant activity, with DPPH and ABTS radical scavenging rates of 81.01 ± 0.78% and 71.37 ± 1.01%, respectively. In a zebrafish central nervous system injury model induced by mycophenolate mofetil, the 500 μg/mL sample group significantly reduced neural apoptosis fluorescence intensity compared to controls (p < 0.05), achieving a neuroprotective rate of 76.58%, which was similar to the effect of L-reducing glutathione. This study offers an efficient, cost-effective enzymatic strategy for producing aglycone soy isoflavones, highlighting their potential in functional foods and neuroprotective applications.