Bactericidal concentrations of methylglyoxal produced by heated cells of Streptococcus faecalis

Cells of Streptococcus faecalis that survived heating for 21 min at 60 degrees C were killed when resuspended at an initial cell density of about 1 X 10(8) viable units/mL and incubated at 33 degrees C for 24 h in a no-growth medium containing potassium phosphate buffer, pH 7.1, glucose, and casein hydrolysate. When such heated cells were resuspended at an initial cell density of about 1 X 10(7) viable units/mL or lower, subsequent cell death was reduced at least 10 000-fold. Unheated cells incubated under similar conditions at about 1 X 10(8) and 1 X 10(9) viable units/mL did not die. Cell death was due to a toxic compound synthesized by the heated cells, and supernatants from incubations showing a bactericidal effect contained a component, absent in nonlethal supernatants, that reacted with 2,4-dinitrophenylhydrazine. Thin-layer chromatography, mass spectrometer analysis, and the visible spectrum of the 2,4-dinitrophenylhydrazine derivative of the unknown and authentic methylglyoxal, and the positive response shown by the free unknown compound when used as a substrate for glyoxalase I, suggested that methylglyoxal was the bactericidal compound. Solutions of authentic methylglyoxal were bactericidal at concentrations above 0.2 mM and lethal supernatants contained about 1 mM methylglyoxal, whereas supernatants that were not lethal contained less than 0.02 mM methylglyoxal.