The bactericidal effect of an ionizer under low concentration of ozone

Abstract

Background: Several mechanisms have been suggested for the bactericidal action of ionizers including electrical phenomena, effects of negative and positive ions and electrostatic repulsion. Negative and positive ions have indeed been shown to have bactericidal effects. In addition, since ozone is generated along with ions, these may contribute to the bacterial killing. In this study, we used a newly developed ionizer, which generates a relatively low concentration of ozone, to determine whether its effect on bacterial cells were due to ions or ozone, and, if ions, how the ions exerted their effects.

Results: The effect of ions on bacterial killing was compared with that of the ozone produced using an ion trap to remove the ions. The ionizer had the ability to kill the bacteria, and ion capture dramatically reduced its bactericidal effect, indicating that the ozone generated had little or no bactericidal effect under these conditions, and the ions produced were responsible for almost all the bacterial killing. Operation of the ionizer increased the level of 8-oxo-dG, a marker of oxidative DNA damage, and decreased aconitase activity, which is known to be sensitive to ROS. The ionizer further affected the adenylate energy charge of bacterial cells. Removal of the ions with the ion trap greatly reduced all these effects.

Conclusion: These results indicate that negative and positive ions generated by the ionizer are responsible for inducing oxidative stress and so reducing bacterial survival.

Keywords: Bactericidal effect; Ionizer; Low concentration of ozone; Negative and positive ions; ROS.

Fig. 1

The experimental set-up and the ionizer. a The configuration and size of the test chamber and the locations of the ionizer and fan. The ionizer is located 30 cm from the bottom of the box, and the fan is located 7 cm from the top. b The ionizer has three components: high voltage generator (power pack), ground electrode and discharge electrode. It is characterized by a specific frequency and an alternate D.C. pulse-high positive/negative voltage. The input voltage of the power pack is 12 V in D.C., and the output voltage is peak-to-peak ± 2.8 kV

Fig. 2

Survival after exposure to negative and positive ions. Each bacterial species was filtered and exposed to ion-with-fan (IZ + F) and fan only (F-mode) conditions. The percent survival shown are the means of 5 replicates in each experiment. a E. coli, b E. faecalis, c B. subtilis, d S. aureus

Fig. 3

Effect of ion capture. a S. aureus was exposed to negative and positive ions with and without ion capture. The percent survival shown are the means of 5 replicates in each experiment. b The concentrations of ions measured with an ion meter during exposure to one ionizer with and without ion capture. c The concentrations of ozone measured with an ozone meter during exposure to one ionizer with and without ion capture. d The concentration of ozone was assessed from the discoloring of a solution of blue potassium indigotrisulfonate. Statistically significant differences are shown, ***p < 0.001

Fig. 4

Generation of ROS examined with a fluorescence microscope. S. aureus suspensions were spread on glass slides after incubation with DCFH-DA for 1 h, and exposed to the F-mode, IZ + F-mode and IZ + F + C-mode for 120mins; they were then examined with a fluorescence microscope (1,000X, confocal microscope, Olympus FV1200)

Fig. 5

Generation of ROS by the ionizer and their effects on S. aureus. a S. aureus suspensions were incubated with DCFH-DA for 1 h, washed with PBS, and exposed to the three modes as above and DCF fluorescence intensity was measured. b Formation of 8-hydroxydeoxyguanosine (8-OHdG) in S. aureus after exposure to the three modes. c The effect of ion exposure on aconitase activity. Bacteria were exposed to the three modes for 120 min. d Resulting levels of adenylate energy charge [(ATP) + 0.5(ADP)] / [(ATP) + (ADP) + (AMP)]. Each experiment was repeated at least 5 times. Statistically significant differences are shown. *p < 0.05, **p < 0.01, ***p < 0.001