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Methods


HIP Extraction






Description:

About 0.5 g of frozen plant tissue is homogenized in 20 ml HIP solution - including a mixture of internal standards (standard amounts are described in the corresponding analytical methods) - with an Ultra Turrax (13000 min -1 ) under streaming argon for 45 s. The extract is shaken at 4 °C for 10 min and then centrifuged at 3200 x g at 4 °C for 10 min. To the clear upper phase, the potassium sulphate solution is added up to a volume of 32.5 ml. After vigorous shaking at 4 °C for 10 min, the extract is centrifuged at 3200 x g at 4 °C for 10 min. Subsequently, the upper hexane-rich layer is dried under streaming nitrogen. The remaining lipids are re-dissolved in 0.2 ml methanol and stored under argon atmosphere at -20°C until use.

HIP solution:
n-hexane/2-propanol (3:2, v/v) with 0.0025 % (w/v) 2-butyl-6-hydroxy toluene (SIGMA)
potassium sulphate solution:
6.7 % (w/v) K 2SO 4



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Oxylipin Profiling Database All rights reserved 2004 - 2005 A. Schomburg, Plant Biochemistry, University of Goettingen