L. reuteri biofilms produce reuterin. L. reuteri biofilms were cultured in MRS for 48 hours at 37°C in ambient atmosphere in multiwell plates. In order to measure reuterin production, biofilms were incubated in the presence of glycerol in anaerobic conditions. Reuterin concentrations were determined using a colorimetric assay and were normalized with respect to viable colony counts prior to the addition of glycerol. All L. reuteri biofilms produced detectable amounts of reuterin, although inter-strain differences were observed. ATCC PTA 6475 and ATCC 5289 produced less reuterin than ATCC 55730 and CF48-3A (ANOVA, p < 0.05).