Separation of alpha-tocopherol and its oxidation products by high performance liquid chromatography.

A very sensitive high performance liquid chromatographic (HPLC) method was developed for the separation of alpha-tocopherol (alpha-T) and its five oxidation products: alpha-tocopheryl quinone (TQ), dimer (D), dihydroxy dimer (DHD), trimer (T) and 9-methoxy-alpha-tocopherone commonly called alpha-tocopheroxide (TO). The separation was achieved on a normal-phase silica-based column (Ultrasphere-Si), using a mobile phase of hexane/chloroform/isopropanol (95:4.5:0.5, v/v/v) at a flow rate of 0.4 ml/min, and the eluants were monitored simultaneously at their maximum absorptions using a variable-wavelength UV detector. The minimum detection limit is 0.01 microgram for alpha-T, TQ and TO, 0.05 microgram for DHD and D, and 0.1 microgram for T/injection. This normal-phase method has the combined advantages of being very sensitive, fast and capable of separating all six compounds at the same time.