Tocotrienols analytical methods

Determination of four tocopherols and four tocotrienols in vegetable oils and fats by the official American Oil Chemists Society method is based on separation by normal-phase HPLC and fluorescence detection (AOCS, 1990). Oil samples are dissolved in hexane, whereas margarines and other fats containing vitamer esters need a cold saponification step to liberate the vitamers. The American Association of Cereal Chemists has a method to analyze vitamin E in various foods. This method (AACC, 1997) is applicable to a vitamin E range of 1 x 10" - 100%, and it includes hot saponification and separation by reversed-phase HPLC. Results are calculated as a-tocopherol acetate. The Royal Society of Chemistry has approved a method to analyze vitamin E in animal feedstuffs by normal-phase HPLC after the vitamers have been liberated by hot saponification (Analytical Methods Committee, 1990). 

Vitamins The data for vitamins come mainly from tables. The analytical methods were rarely indicated in the sources. Vitamin A vitamin A activity expressed in international units (1 ID = 0.3 pg of retinol) Vitamin D vitamin D2 in plant products and D3 in animal products. Vitamin D activity is expressed in international units (1 ID = 0.025 pg of cholecalciferol) Vitamin E the vitamin activity of tocopherols and tocotrienols is expressed as alpha-tocopherol. 

Currently, high-performance liquid chromatography (HPLC) methods have been widely used in the analysis of tocopherols and tocotrienols in food and nutrition areas. Each form of tocopherol and tocotrienol can be separated and quantified individually using HPLC with either a UV or fluorescence detector. The interferences are largely reduced after separation by HPLC. Therefore, the sensitivity and specificity of HPLC methods are much higher than those obtained with the colorimetric, polarimetric, and GC methods. Also, sample preparation in the HPLC methods is simpler and more efficiently duplicated than in the older methods. Many HPLC methods for the quantification of tocopherols and tocotrienols in various foods and biological samples have been reported. Method number 992.03 of the AOAC International Official Methods of Analysis provides an HPLC method to determine vitamin E in milk-based infant formula. It could probably be said that HPLC methods have become dominant in the analysis of tocopherols and tocotrienols. Therefore, the analytical protocols for tocopherols and tocotrienols in this unit are focused on HPLC methods. Normal and reversed-phase HPLC methods are discussed in the separation and quantification of tocopherols and tocotrienols (see Basic Protocol). Sample... 

The decreasing importance of GC as an analytical technique for vitamin E becomes evident from the number of systems/applications inventoried in the first and second editions of this book. In the 1985 edition, 64 systems, 61 of which used packed columns, were listed. However, the number of papers that appeared between 1985 and 1989, included in the second edition was only five. A milestone in the history of GC of vitamin E was the first and, to the best of our knowledge so far only separation, in 1967, of p- and y-tocopherols as their quinone derivatives on a packed column, using a special binary liquid phase. The separation of TMS ethers of both isomers was achieved in 1975 on a 32-m open tubular column coated with a polar PZ-176 liquid phase and, later on a 30-m DB-5 capillary column. All tocopherols and tocotrienols were successfully resolved in as little as 15 min by capillary GC on a 20 m OV-17 column. GC was also the first method to distinguish (partially) between stereoisomers (see VIA). 

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