2.5 Extraction and derivatization of lipids
1 mL of cultivation broth was sampled into 4 mL glass vials and
centrifuged at 3,500 g for 5 min at 20°C. The supernatant was removed,
and 1 mL of 1 M HCl in anhydrous methanol was added to the pellet.
Samples were vortexed for 20 s and incubated at 70°C for 2 h for
methanolysis reaction to proceed. Samples were cooled down to room
temperature, 1 mL of 1 M NaOH in methanol, 500 µL of saturated NaCl
solution in water, 1 mL of hexane, and 10 µL of internal standard (2 g/L
of methyl nonadecanoate (19:Me) in hexane) were added sequentially.
Vials were vortexed for 10 s, centrifuged at 3,500 g for 5 min at 20°C,
and the upper organic layer was taken for analysis.