Development of an Experimental Method to Evaluate Inhibitory Activities of Gaseous Antimicrobial Agents

Introduction: There is a growing interest in using gaseous antimicrobial agents to inhibit the growth of hazardous microorganisms in foods. Compared to antimicrobial agents in liquid phase, antimicrobial agents in vapor phase are more effective, have fewer residues in foods, and change the sensory properties of foods less. However, there is no standard method to evaluate the lethal activities of gaseous antimicrobial agents.

Purpose: This study was done to develop a method to determine the minimal inhibitory concentrations (MIC) and minimal lethal concentrations (MLC) of gaseous antimicrobial agents.

Methods: Experimental apparatus with upper and lower polycarbonate chambers was constructed. Nutrient agar containing 0.025% bromocresol purple and 1% glucose was poured into wells in the upper chamber and Escherichia coli O157:H7 (5 log CFU/well) was inoculated. Sterile filter discs were placed on the wells in the lower chamber and essential oils (EO) serially diluted were deposited. The chambers were sealed and incubated at 30°C for 48h. After incubation, the lowest concentration of EO which did not change the color of agar into purple was determined. The agar whose color was not changed was streaked on nutrient agar to determine the MLC.

Results: MIC values of 30 essential oils in vapor phase against E. coli O157:H7 were determined.  Vapor of cinnamon bark oil showed the lowest MIC (0.0391 µl/ml). The order of MICs of EOs in vapor phase was cinnamon bark < thyme-thymol < peppermint = oregano < thyme-linalool = clove = spearmint < tea tree = thyme Spanish = clary sage. The order of MLCs was cinnamon bark < thyme-thymol < oregano < peppermint = thyme-linalool < clove = spearmint = tea tree = thyme Spanish < clary sage.

Significance: An experimental apparatus and standardized method to determine MIC and MLC of gaseous antimicrobial agents were developed.