Repurposing Success: Real-World Validation of a Novel Cephalosporin Decontamination Protocol Using Chlorine Dioxide...

Beta-lactam manufacturing facilities face strict regulatory challenges to repurposing for non-beta-lactam production due to the high risk of cross contamination. This study introduces a novel decontamination protocol for cephalosporin facilities, combining chlorine dioxide (ClO₂) treatment with disc diffusion testing to verify the complete degradation and inactivation of beta-lactam residues in a real-world setting.

Antimicrobial susceptibility discs (ASDs) contaminated with cephalosporins (Cefalexin, Cefaclor, Cefixime, Cefuroxime) were exposed to both aqueous (400 ppm for 24 h) and gaseous ClO₂ (5 mg/L for >5 h) in separate cycles, achieving a minimum cumulative exposure of 9600 ppm-hours in both. Post-treatment, replicates (4 = aqueous cycle, 5 = gaseous cycle) of each cephalosporin were analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS) and microbial challenge testing to assess degradation and biological inactivation, respectively.

For all, LC-MS/MS confirmed cephalosporin degradation below the reporting limit (<0.02 μg/disc), while microbial challenge testing demonstrated no significant biological activity with <1 mm bacterial inhibition. Microbial challenge and LC-MS/MS results showed significant agreement (Cohen's κ = 1.00) in both aqueous and gaseous ClO₂ cycles.

Gaseous ClO₂ application at 9600 ppm-hours was applied to a 450 m² cephalosporin facility. Analysis confirmed that cephalosporin residues were degraded to levels below the acceptable limits (<1 μg/disc) with microbial challenge testing supporting the success of the cycle, with no significant biological activity remaining in the discs placed throughout the facility.

This study presents the first successful protocol for cephalosporin facility decontamination using gaseous ClO₂ in a real-world setting.