Abstract
A dichloromethane extraction system has been developed which is suitable for alfalfa carotenoids and is compatible with nonaqueous reverse phase liquid chromatographic (LC) analysis. Ground samples are extracted in the presence of antioxidant with dichloromethaneacetone (2 + 1). After an overnight soak, the mixture is saponified with 40% w/v potassium hydroxide in methanol and then is partitioned with 10% w/v aqueous sodium sulfate. An aliquot of the lower yellow organic phase is used for LC analysis after dilution (1 + 1) with acetonitrile. An isocratic 2 solvent system of ethyl acetateacetonitrile containing 0.1% n-decanol as mobile phase modifier is used for liquid chromatography with a high carbon loading non-endcapped ODS packing material. In comparisons with the existing AOAC method for carotenes and xanthophylls in dried plant materials and mixed feeds, the proposed method gave consistently higher results in a range of 7-28%, depending on the substrate. Possible causes for the differences were examined and include losses during partition and column chromatography with the AOAC method. Calculation methods and the choice of standard materials may also contribute to the difference in results.
