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Commission Regulation (EEC) No 2568/91 of 11 July 1991 on the characteristics of olive oil and olive-residue oil and on the relevant methods of analysis
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This part specifies the preparation of the methyl esters of fatty acids. It includes methods for preparing fatty acid methyl esters from olive and olive-pomace oils.
The preparation of the fatty acid methyl esters from olive oils and olive-pomace oils are performed by transesterification with methanolic solution of potassium hydroxide at room temperature. The necessity of purification of the sample prior to the trans-esterification depends on the sample's free fatty acids content and the analytical parameter to be determined, it can be chosen according to the following table:
Category of oil | Method |
---|---|
Virgin olive oil with acidity ≤ 2,0 % | 1. Fatty acids 2. trans -Fatty acids 3. ΔECN42 (after purification with silica-gel SPE) |
Refined olive oil | |
Olive oil composed of refined olive oil and virgin olive oils | |
Refined olive pomace oil | |
Olive pomace oil | |
Virgin olive oil with acidity > 2,0 % Crude olive pomace oil | 1. Fatty acids (after purification with silica-gel SPE) 2. trans -Fatty acids (after purification with silica-gel SPE) 3. ΔECN42 (after purification with silica-gel SPE) |
Methyl esters are formed by trans-esterification with methanolic potassium hydroxide as an intermediate stage before saponification takes place.
When necessary, the samples will be purified by passing the oil through a silica gel solid-phase extraction cartridge. A silica gel cartridge (3.1.2.8) is placed in a vacuum elution apparatus and washed with 6 ml of hexane (3.1.2.2); washing is performed without vacuum. Then a solution of the oil (0,12 g approximately) in 0,5 ml of hexane (3.1.2.2) is loaded onto the column. The solution is pulled down and then eluted with 10 ml of hexane/diethyl ether (87:13 v/v) (3.1.2.6). The combined eluates are homogenised and divided in two similar volumes. An aliquot is evaporated to dryness in a rotary evaporator under reduced pressure at room temperature. The residue is dissolved in 1 ml of heptane and the solution is ready for fatty acid analysis by GC. The second aliquot is evaporated and the residue is dissolved in 1 ml of acetone for triglyceride analysis by HPLC, if necessary.
In a 5 ml screw-top test tube (3.1.3.1) weigh approximately 0,1 g of the oil sample. Add 2 ml of heptane (3.1.2.2), and shake. Add 0,2 ml of the methanolic potassium hydroxide solution (3.1.2.7), put on the cap fitted with a PTFE joint, tighten the cap, and shake vigorously for 30 seconds. Leave to stratify until the upper solution becomes clear. Decant the upper layer containing the methyl esters. The heptane solution is ready for injection into the gas chromatograph. It is advisable to keep the solution in the refrigerator until gas chromatographic analysis. Storage of the solution for more than 12 hours is not recommended.]
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