*4.6. Determination of Lipid Concentration*

The lipid concentration was measured by chloroform–methanol 2:1 (v/v) according to the method by Folch et al. [49]. For each sample, a mixed solution of chloroform–methanol (2:1) was added to the dried fine seed samples in a vented conical Erlenmeyer flask, and heated at 65°C for 30 min and boiled for 1 hour. After extraction, the samples were cooled and filtered in an eggplant-type flask using a glass filter with additional mixed solution of chloroform-methanol. The chloroform–methanol solution was evaporated from the sample and were petroleum ether and sodium sulfate were added, the solution was shaken. The samples were centrifuged at 3000 rpm, and the supernatant was transferred to a weighing glass tube and dried at 105°C. After evaporation of the ether, the glass tube was weighed and the lipid concentration calculated.
