Kromasil 100 5c18

For the separation of TAGs, Shimadzu LC20 chromatographic system with refractive index detector (RID 10A) and an Agilent 1200 Infinity chromatograph system with diode array and MS detectors were used. Chromatograms were recorded using mobile phases of the systems “acetonitrile-propan-2-ol” and “acetone – acetonitrile” for refractive index (RI) and DAD detector, respectively. The speed of the mobile phase was 0.8 mL/min; chromatographic columns of 250 × 4.6 mm Kromasil 100-5C18 (for HPLC with spectrophotometric detection) and 150 × 2.1 mm Kromasil 100–5C18 (for MS detection) were used at a thermostat temperature of 30°C.
Mass spectrometric detection (6130 Quadrupole MS, Agilent) was carried out in the atmospheric pressure chemical ionization mode under standard conditions at a fragmentor voltage of 150 V; signals were recorded for positively charged ions. The Kromasil 110-3.5C18 2.1 × 150 mm column was used, the mobile phase speed was 0.1–0.2 mL/min, and the eluent system was acetonitrile– propanol-2 with additions of ammonium formate (HCOONH₄) 0.2 mM. All the chromatograms were performed in isocratic mode. The MagicPlot Student software was used for the resolution of “problem” (with a low value of R_S) TAG.

The HPLC analysis was performed on a Dionex UltiMate 3000 LC series–diode array detector (DAD) system with a quaternary pump and an autosampler that could thermostat samples. Separation was achieved on Kromasil 100-5C18 (4.6 mm × 250 mm, 5 µm, Agilent Technologies, Wilmington, DE, USA). The detection wavelength was set at 235 nm for HPLC fingerprinting, and 220 nm and 280 nm for HPLC quantification. The injection volume was 20 µL.
The chromatographic separation was performed using acetonitrile (solvent A) and water (solvent B) as mobile phase at a flow rate of 1 mL·min⁻¹. The gradient program was set as follows: 0–35 min: 5% A–65% A; 35–40 min: isocratic 65% A; 40–45 min: 65% A–5% A; 45–50 min: 5% A–80% A.