A Novel Method for the Reduction of Numbers of Listeria monocytogenes Cells by Freezing in Combination with an Essential Oil in Bacteriological Media

Abstract

The use of multiple freeze (−20°C)–thaw cycles in combination with isoeugenol and polysorbate 80 was investigated as a method for the reduction of numbers of Listeria monocytogenes cells in a bacteriological medium. Three freeze (1 h, −20°C)–thaw cycles in the presence of isoeugenol at concentrations of 0, 100, and 300 ppm resulted in average L. monocytogenes reductions of 0.69, 2.65, and 3.3 log10 MPN (most probable number) per ml, respectively. Increasing the number of freeze–thaw cycles further decreased cell numbers, with reductions of nearly 5 log10 MPN/ml being obtained with six freeze-thaw cycles. Freeze–thaw cycles were effective in reducing cell numbers at isoeugenol concentrations down to 25 ppm. Rapid freezing rates with liquid nitrogen were found to be less effective in reducing numbers of L. monocytogenes cells. Two rapid freeze–thaw cycles in the presence of 100 ppm isoeugenol and polysorbate 80 resulted in a reduction of 1.45 log10 MPN/ml. Two freezing (−20°C) cycles involving slow freezing and thawing rates with samples being held frozen for 6 h for each cycle resulted in reductions larger than those obtained with faster freezing rates. It was found that complete thawing in freeze-thaw cycles was not necessary to achieve bactericidal action. The application of multiple freeze-thaw cycles in combination with low concentrations of isoeugenol could effectively reduce numbers of L. monocytogenes cells in bacteriological media.