DLC onderzoek naar vreemde vette oliën
De Pharmacopoeia Europea geeft hiervoor een onderzoek dat berust op een hydrolyse van de oliën gevolgd door dunnelaagchromatografie met paraffine als stationaire fase en 90% azijnzuur als mobiele fase.2.4.21. FOREIGN OILS IN FATTY OILS BY THIN-LAYER CHROMATOGRAPHY
Examine by thin-layer chromatography ( 2.2.27) using kieselguhr G R as the coating substance.Impregnate a plate by placing it in a chromatographic tank containing the necessary quantity of a mixture of 10 volumes of liquid paraffin R and 90 volumes of light petroleum R so that the plate dips about 5 mm beneath the surface of the liquid. When the impregnation mixture has risen by at least 12 cm from the lower edge of the plate, remove the plate and allow the solvent to evaporate for 5 min.
Carry out the chromatography in the same direction as the impregnation.
Preparation of the mixture of fatty acids.
Heat 2 g of the oil with 30 ml of 0.5 M alcoholic potassium hydroxide under a reflux condenser for 45 min.
Add 50 ml of water R, allow to cool, transfer to a separating funnel and extract with three quantities, each of 50 ml, of ether R.
Discard the ether extracts, acidify the aqueous layer with hydrochloric acid R and extract with three quantities, each of 50 ml, of ether R.
Combine the ether extracts and wash with three quantities, each of 10 ml, of water R; discard the washings, dry the ether over anhydrous sodium sulphate R and filter.
Evaporate the ether on a water-bath. Use the residue to prepare the test solution. The fatty acids may also be obtained from the soap solution prepared during the determination of the unsaponifiable matter.Test solution.
Dissolve 40 mg of the mixture of fatty acids obtained from the substance to be examined in 4 ml of chloroform R. Reference solution.
Dissolve 40 mg of the mixture of fatty acids obtained from a mixture of 19 volumes of maize oil R and 1 volume of rapeseed oil R in 4 ml of chloroform R.Apply separately to the plate 3 ml of each solution.
Develop over a path of 8 cm using a mixture of 10 volumes of water R and 90 volumes of glacial acetic acid R.Dry the plate at 110 �C for 10 min. Allow to cool and, unless otherwise prescribed, place the plate in a chromatographic chamber, with a tightly fitting lid, that has previously been saturated with iodine vapour by placing iodine R in an evaporating dish at the bottom of the chamber. After some time brown or yellowish-brown spots become visible.
Remove the plate and allow to stand for a few minutes. When the brown background colour has disappeared, spray with starch solution R. Blue spots appear which may become brown on drying and again become blue after spraying with water R.The chromatogram obtained with the test solution always shows a spot with an R� of about 0.5 (oleic acid) and a spot with an R� of about 0.65 (linoleic acid) corresponding to the spots in the chromatogram obtained with the reference solution.
With some oils a spot with an R� of about 0.75 may be present (linolenic acid).
By comparison with the spot in the chromatogram obtained with the reference solution, verify the absence in the chromatogram obtained with the test solution of a spot with an R� of about 0.25 (erucic acid).
10 september 1999