Folate concentrations were determined by a previously
described microbiological assay using Lb. rhamnosus NCIMB 10463
as the indicator organism (Lai~no et al., 2012). Briefly, diluted samples
or different concentrations of HPLC-grade folic acid (Fluka
BioChemica, SigmaeAldrich, Switzerland) were placed with the
indicator strain and incubated statically during 48 h at 37 C in 96-
well sterile microplates containing the folate-free medium (Difco,
USA). The optical density was read at 580 nm (OD580) using a
microplate reader (VERSAmax tuneable microplate reader, Molecular
Devices, USA). The folate concentration of the samples was
determined by comparing the OD with those obtained with the
standard curve prepared using commercial folic acid.