Garlic Extract Induces Dissociation of Hydroxyapatite-Bound S. mutans

OBJECTIVES: Garlic extract, GE, has been reported to inhibit S. mutans, SM, growth. Our goal is to characterize the effects of GE on SM viability, on SM reversible binding, and on biofilm formation on hydroxyapatite, HA. We are interested in electrostatic interactions. HYPOTHESES: 1. GE inhibition of SM growth is reversible, and removing GE will remove inhibition; 2. An electrically neutral GE constituent, likely allicin, binds SM and HA, reducing negative surface charge and increasing SM to HA binding. METHODS: SM is suspended in neutral phosphate buffer for planktonic states, incubated with HA and washed to remove free SM for single-layer HA-SM, or grown for 48 hours in a medium containing 20 μm HA beads for biofilms. GE is from white garlic. Excess GE is removed by washing. Viable SM is determined from SYTO9/propidium iodide fluorescence ratios. Zeta potentials, ζ, are calculated from electrophoretic mobilities using the Smoluchowski equation, and are taken as an estimate of surface charge. Fluorescence micrographs are obtained after staining with SYTO-9. RESULTS: Planktonic SM exposed to GE is 43% dead. Washing had no significant effect. Growth rates after treatment confirm that GE irreversibly kills ~40% of the SM. In the absence of GE, values of ζ for HA, SM and single-layer SM on HA are -2.61, -0.51 and -1.10 mV, respectively. GE does not changes ζ for SM, but changes both HA and HA-SM to the same value, -1.55 mV, consistent with SM not binding. Fluorescence micrographs HA-SM and of biofilms grown on HA show that GE causes the SM is dissociate from HA. CONCLUSIONS: the hypotheses are wrong. GE inhibition of planktonic SM growth is irreversible. GE exposure causes bound SM to dissociate from HA-SM.