A total of 250 μL of
each sample extract and 2.5 mL of the described reagent were incubated
at 90 °C for 15 min. The blank consisted of 2.5 mL of the reagent
and 250 μL of methanol:acetic acid (99.5:0.5). The absorbance
was measured at 540 nm using a model U1100 UV/Visible spectrophotometer
(Hitachi, Japan). The results were expressed as mg of
quebracho tannin/g DW.