Concentration and Application Order Effects of Sodium Benzoate and Eugenol Mixtures on the Growth Inhibition of Saccharomyces Cerevisiae and Zygosaccharomyces Bailii

Growth of Saccharomyces cerevisiae and Zygosaccharomyces bailii cells was monitored in the presence of sodium benzoate and eugenol alone or combined. The two antimicrobials' concentration, addition order, and timing were varied to determine and quantify any additive inhibitory effect on the yeasts. The yeast growth was also followed in the presence of ethanol, which served as solubilizer, at pertinent concentrations. The growth patterns are depicted as adjusted optical density compared with time curves. They all had sigmoid shape, described mathematically by a shifted logistic model that had an almost perfect fit to the data. The model's 3 parameters accounted for the curve's asymptote, the location of its inflection point and slope, which are rough measures of the overall growth level and its degree of suppression, the time to reach the peak growth rate and its retardation, and the overall growth rate, respectively. Maximum growth inhibition was achieved when the sodium benzoate and eugenol were administered together or alone in full dose. When each was administered alone but in 2 half dose additions, their efficacy dropped. When they were used together but added sequentially with a 24 h pause, their administration order had a noticeable effect on the treatment's efficacy, which depended on their respective concentrations. These observations are presented in a slightly modified version of the "hurdle" ideogram. They suggest that sequencing the administration of antimicrobials can be a simple tool to probe their mode of activity and quantify their efficacy. Practical Application Reducing the amount of additives in foods is a goal pursued by many branches of the food industry. In microbial growth suppression, a promising way to accomplish such a reduction is through the administration of 2 or more antimicrobials, preferably natural, exploiting their synergism. To search for effective combinations, in respect to type and concentration, one needs an insight into their mode of activity. Sequencing their administration, as demonstrated with 2 antimicrobials and 2 common yeasts that are involved in beverage spoilage, has offered a simple way to probe into certain aspects of the effect of antimicrobial combinations on microorganisms. The presented algebraic growth model enables to quantify, separately, the growth overall suppression, its retardation, and lowering its peak rate. The proposed modified version of the "hurdles paradigm" helps to visualize the differences in the antimicrobials' general mode of activity and how it is affected by their sequencing.