Hydrogen peroxide h2o2

C. perfringens strains were incubated in six-well polystyrene plates at 37 °C for 48 h, under anaerobic conditions. After incubation, sessile cells were collected by removing the medium and washed with PBS one time, and the planktonic cells were obtained by centrifugation. The initial cell concentration of vegetative cells and spores were 10⁵ CFU/mL and 10⁴ CFU/mL, respectively. The cells were exposed to disinfectants for 10 min or to atmospheric conditions for 18 or 36 h. In the negative control, the disinfectants were not treated. Three types of disinfectants that are commonly used in the food processing industry were tested on TYJAM-D-66 in this study, at concentrations suggested by the manufacturers—0.01% sodium hypochlorite (Junsei Chemicals Co., Chuo-ku, Tokyo), 5% hydrogen peroxide (H₂O₂) (Daejung), and 80% ethyl alcohol (Duksan Pure Chemicals Co., Ltd., Ansan, Korea). After treatment, the vegetative cells and the germinated spores were determined by plating onto the BHI agar medium. The plates were incubated anaerobically at 37 °C for 24 h. After incubation, the colonies were counted to determine the surviving population size. The experiments for each strain were performed three times with duplicates.

Distilled water (H₂O), methanol (CH₃OH), acetone (C₃H₆O), ethyl alcohol (C₂H₅OH), and hydrogen peroxide (H₂O₂) were purchased from DaeJung Chemicals and Metals Co., Ltd. (Siheung, Korea). Lithium bromide (LiBr), sodium carbonate (Na₂CO₃), hexafluoro-iso-propanol (HFIP), horseradish peroxidase (HRP), and 4-aminobenzenethiol (4-ABT, C₆H₇NS) were purchased from Sigma-Aldrich (St. Louis, MO, USA). Gold nanoparticles with an average diameter of 30 nm were purchased from Nanopartz (Loveland, CO, USA). Bombyx mori cocoons were supported by the Rural Development Administration (Wanju, Korea).