2.2. Extraction and Chromatographic determination of glucose,
fructose, sucrose, myo-inositol, threhalose, FOS and RFOS.
For carbohydrates analyses, 2 g of frozen strawberry powder (wet) were
extracted with 5 ml of distilled water, homogenized and centrifuged at
10000 rpm for 30 min at 4 ºC, and determined by UPAEC-PAD with a
Carb2-250 (250 x 4mm) column, as previously described (Blanch et al.,
2015b). Samples were analysed on Bioscan module (817 IC Metrohm,
Herisau, Switzerland) equipped with a pulsed amperometric detector, (IC
Pump 812) and coupled degasser (IC-837). Isocratic elution was carried
out with 100 mM NaOH- 10 mM NaOAc, and the flow rate through the column
was 0.5 mL/min, leading to sampling times of 60 min. For FOS and RFOS
analyses, 3 g of frozen strawberry powder (wet) were homogenized in 4 ml
of ultra-pure water and the mixture was boiled under reflux in a water
bath for 15 min. After cooling, the samples were sonicated for 10 min at
40 ºC and the pH of each sample was adjusted to 7.5 with 10%
NH4OH. The samples were then centrifuged at 14700g for
15 min at 4 ºC and the supernatants were filtered thought a 0.45 µm pore
size membrane. The analyses of extracts and calibration standards were
performed using an Agilent 1200 liquid chromatography coupled to an
Agilent Triple Quadrupole MS detector G6410B (HPLC-QqQ-SIM, Agilent
Technologies, Palo Alto, CA, USA). The chromatographic separation was
achieved on an HYPERCARB (100 mm X 2.1μm X 5μm) column, with a mobile
phase consisting of water containing 5 mM ammonium formate (A) and
performing an elution using acetonitrile (B) in gradient as follows:
%B: 0 min, 5%; 5 min, 10%; 10 min, 10%; 20 min, 50%; 30 min, 5%;
40, min, 5% at 25 ºC with an injection volume of 5 μL.