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Abstract
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Mulberry juice is rich in bioactive moieties; however, its functional potential is often limited by low bioavailability and instability of phenolic compounds during processing and storage. This research addresses the challenge of improving bioactive components release and stability through the synergistic application of enzymatic hydrolysis and multifrequency ultrasound-assisted fermentation. The aim of this study was to develop a functionally enriched fermented mulberry juice with improved physicochemical, structural, antioxidant and sensory profiling. A combined treatment involving enzymatic hydrolysis, multifrequency ultrasound (20/28/40 kHz) and mono- and co-culture fermentation using Lactiplantibacillus plantarum, Lacticaseibacillus paracasei, and Lactobacillus helveticus was employed. Physicochemical attribites, antioxidant assay, structural characteristics (SEM, FTIR) and molecular interactions were systematically evaluated. The findings revealed a significant decrease in pH (from 4.01 to 3.43; p < 0.05) and a marked increase in total anthocyanin content (from 16.53 to 70.26 mg/L) and total sugars (from 2.66 to 24 g/L) following treatment. DPPH radical scavenging activity increased substantially in co-culture treatments, reaching up to ∼78-80%, presenting enhanced antioxidant activity (p < 0.05). Structural analysis exhibits increased porosity and molecular modifications, facilitating the release of bound phenolics. Molecular docking further confirmed moderate binding interactions (-1.83 kcal/mol) between peptidoglycan and cyanidin-3-rutinoside, revealing improved stability and potential bioavailability of anthocyanins. These outcomes demonstrate that the integration of enzymatic hydrolysis and multifrequency ultrasound creates a synergistic effect that boosts substrate accessibility, fermentation efficiency and bioactive compound release beyond conventional fermentation approaches. The study provides a novel strategy for improving the functional quality of f
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