A study of the mechanism of action of triclosan and PCMX
Triclosan and PCMX are two broad-spectrum antimicrobial agents of particular interest to
Reckitt Benckiser. Triclosan is one of the most widely used biocides today and is
incorporated into increasing numbers of household products such as toothpaste, mouth washes
and underann deodorants.
It is generally accepted that biocides have multiple target sites within the bacterial cell;
however, recent reports have identified mutations in the E. coli fabI gene as being linked to
decreased susceptibility to triclosan. The fabI gene encodes for enoyl reductase, an important
enzyme in fatty acid biosynthesis. Conclusions were drawn that triclosan had a single,
primary target in bacteria and the previously reported effects of triclosan on membrane
structure were secondary effects arising from specific inhibition of the fatty acid biosynthetic
In this study the mechanisms of action of both triclosan and PCMX were investigated against
the hygiene indicators E. coli NCTC 8196 and S. aureus NCTC 10788. Initially, interactions
of both biocides with the bacterial cell were investigated using adsorption isotherms.
Triclosan was found to possess an unusual Z-shaped isotherm, indicative of a concentration
dependant breakdown of cellular structure. Disruption of E. coli cells by sonication also
increased the cells sorptive capacity for triclosan, indicating that exposure to concentrations of
triclosan above 6ppm induced a structurally damaging event to the cell. PCMX was shown to
have a complex S-shaped isotherm, with 2 saturation plateuxs. The termination of the primary
uptake phase corresponded to the threshold concentration required for bactericidal activity
and leakage of cellular constituents, the secondary uptake was thought to represent
penetration to new sorptive sites within the cell itself.
The bacteriostatic and bactericidal properties of both biocides were probed by calculation of
MICs and MBCs and by production of time survivor curves. Both compounds were found to
have bacteriostatic or bactericidal activity against E. coli NCTC 8196 and S. aureus NCTC
10788 depending on concentration. Concentration exponents were detennined and were found
to be in the region of S for triclosan and 6 for PCMX, indicating that their activity is greatly
reduced on dilution.
Non-growing cells were used to investigate membrane damage caused by the two biocides.
The leakage of intracellular potassium and changes in the permeability of the membrane to
specific ions and protons were used as subtle indicators of membrane damage. PCMX was
shown to cause substantial membrane damage, indicated by rapid, gross potassium leakage
and by the rapid loss of absorbance of E. coli spheroplasts stabilised in salt solutions, when
exposed to bactericidal concentrations of the agent. The effect of triclosan on the membrane
appeared to be more subtle, with cells exhibiting a more concentration dependant loss of
potassium and spheroplast stability over time. Bactericidal concentrations of triclosan were
shown to induce the translocation of protons and the uncoupling of oxidative phosphorylation.
Both compounds were able to manifest significant damage to the bacterial cell under
conditions which limit the significance of the ENR system to bacterial survival. Whilst ENR
may be a target for the growth inhibitory properties of triclosan, this study indicates that the
disinfectant potential of the agent derives from additional target damage.
Combinations of triclosan and PCMX were also investigated for possible signs of synergy.
Whilst adsorption profiles for either compound in the presence of the other clearly indicated
changes in the sorptive capacity of the cell, the enhanced bacteriostatic and bactericidal
effects of combinations of the biocides were thought to have arisen from the non-linear
relationship between antibacterial activity and concentration, rather than 'true' synergy itself.