To explore the effect of electron beam irradiation (EBI) pretreatment on the content of active substances in walnut green husk (WGH) and its bacteriostatic activity of four bacteria, WGH was pretreated by EBI, with ultrasonic assisted extraction for polyphenol, flavonoid, triterpene active substances. Scanning electron microscopy (SEM) and X-ray diffraction were used to observe the effect of EBI on the structure and cell wall of WGH powder. UHPLC-MS/MS was used to analyze the composition of active substances polyphenol, flavonoid and triterpene in WGH. The antimicrobial effects of WGH extracts on Escherichia coli, Staphylococcus aureus, Bacillus cereus and Salmonella were analyzed by the diameters of inhibition zones (DIZ), the minimum inhibitory concentration (MIC) and the growth curve. The results showed that the irradiation dose of 30 kGy electron beam could effectively destroy the tissue and cell structure of WGH, and the extraction contents of polyphenols, flavonoids and triterpenoids were 20.89 mg GAE/g, 38.01 mg RE/g and 14.73 mg OA/g, respectively, which increased by 14.78%, 43.0% and 13.69% compared with the non-irradiated samples, respectively. After irradiation pretreatment, active substances in WGH including 16 polyphenols, 24 flavonoids and 14 triterpenoids. Additionally, the bacteriostasis test showed that the extracts of WGH had better bacteriostasis after irradiation, and had the strongest inhibitory effect on B. cereus, followed by S. aureus, E. coli, and Salmonella, with MIC values of 0.008, 0.008, 0.016 and 0.032 g/mL, respectively. The WGH extracts changed the growth curves of four bacteria significantly and inhibited their growth and proliferation. Therefore, EBI pretreatment can significantly improve the extraction contents and antibacterial activity of active substances in WGH, which can provide a new idea for the extraction of biologically active ingredients in agricultural by-products and the development of natural antibacterial products.
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