In a first stage, the extraction experiments were performed
using different conditions of methanol concentration (60–100%),
solvent/solid ratio (10–30 ml/g), and extraction time (30–
90 min). The conditions used in each experiment were settled
according to the 2
3
full factorial design presented inTable 1. Subsequently, additional extraction experiments were performed by
varying the methanol concentration between 20% and 80%, and
the solvent/solid ratio between 10 and 40 ml/g SCG, during
90 min (Table 2). For comparison, assays using only distilled water
as extraction solvent were also performed.
For the reactions, the extraction solvent and the SCG were
poured into 100-ml Erlenmeyer flasks, which were duly covered
and maintained during the desired time in a water-bath with magnetic agitation at 60–65C. Subsequently, the total content of each
flask was centrifuged (2500g,4C, 20 min) and the supernatant
(SCG extract) was filtered through 0.22lm filters and stored at
20C in darkness until analyses. The volume of extract recovered
after each extraction was quantified and used for calculations