Reuterin concentrationwas quantified by a colorimetric method adapted. Briefly, serial dilutions of purified
reuterin in sterile distilled water were prepared and colorimetric reaction after the addition of tryptophan solution (0.01 M solution in 0.1 N HCl) and concentrated HCl (32%, 10 N) were carried out. Solutions were incubated at 60 C for 5 min in a Thermomixer compact (Eppendorf, Hamburg, Germany) to develop colour. The absorbance was measured at 490 nm with a Beckman DU-650 spectrophotometer (Beckman Coulter, Inc., Palo Alto, CA, USA). A standard curve of acrolein in water (0e4 mM) was also prepared plotting acrolein concentrations against the average of three absorbance measurements for each concentration. Samples with reuterin were compared against the acrolein standard to calculate molar concentration of reuterin stock solution. Experiments were carried out in triplicate.