Quantitative determination of tocopherols in single seeds of rapeseed (Brassica napus L.)

A high-performance liquid chromatography (HPLC) method was developed to det ermine tocopherols in single seeds of oilseed rape (Brassica napus L.). Sin gle seeds were extracted with isooctane, centrifuged, and the overlayer ext ract injected into the HPLC instrument without further purification. Separa tion was performed on a diol-column (250 mm x 3 mm I.D.) using a mobile pha se consisting of 94% isooctane and 6% tert-butyl methyl ether. Quantificati on of individual tocopherols was done by fluorescence detection and with be ta-tocopherol as internal standard. The standard error (SE) of the present method for total tocopherol content, as determined with 30 ground samples o f Samourai (mean = 304mg kg(-1) dry seed) and 20 ground samples of Lirabon (mean = 350mg kg(-1) dry seed), was 9.5mg tocopherol/kg dry seed. The SEs o f the method for the determination of alpha- and gamma-tocopherol percentag es were 1.2% and 1.3%, respectively. The low SE values indicate that the me thod can be effectively used to detect small differences for tocopherol con tent and composition in studies requiring the analysis of reduced amounts o f plant material. In particular, the comparison between the SE of the metho d and the SD of single seeds within individual plants of Samourai (36.8 mg kg(-1) dry seed) and Lirabon (37.6 mg kg(-1) dry seed) shows the potential of the present method for the detection of the tocopherol content within pl ant differences.