Y-Tocotrienol

Wehmeyer et al. (1969) published results on the content of B vitamins (thiamine, riboflavin, and nicotinic acid), vitamin C, and p-carotene and foimd that the morama bean is a good source of both B vitamins and vitamin C, but a poor source of p-carotene. Holse et al. (2010) investigated the content of the eight vitamin E isomers and found that the vitamin E composition in morama beans is dominated by y-tocopherol with 59-234 ng/g, followed by a- and p-tocopherols with 14- 8 gg/g and 1.1-3.3 ng/g, respectively. Eurthermore, traces of 8-tocopherol as well as p- and y-tocotrienols were present in some samples. The remaining two tocotrienols (a- and 8-) were not present in the beans. The presence of a-, p-, and y-tocopherols in the morama bean was also foimd by Mitei et al. (2009) who examined morama oil and by Dubois et al. (1995) who examined two samples of T.fassoglense. 

There are numerous papers that refer to the quantification of tocopherols and tocotrienols using HPLC methods. Normal-phase HPLC methods with a silica column as well as reversed-phase HPLC methods with a C18 column are commonly used. A silica normal-phase column is able to separate all eight tocopherols and tocotrienols in a typical chromatographic procedure. Because plant tissues possess most forms of tocopherol and tocotrienol, it is recommended that the normal-phase HPLC method be applied to food samples from plants. In the reversed-phase HPLC method, [3- and y-tocopherol and (3-and y-tocotrienol are not usually completely separated. This method can be used in animal tissues, which either lack or have reduced levels of [3- and y-tocopherol and (3- and y-tocotrienol. The resolution of the normal-phase HPLC method is higher than that of reversed-phase HPLC method however, the reversed-phase HPLC column is more long-lasting than the normal-phase HPLC column (see Critical Parameters and Troubleshooting). 

In normal-phase HPLC on a silica column, separation is based on the number and position of methyl substituents on the chromanol ring. In reversed-phase HPLC on a Cl8 column, separation is based on the structure of the side chain and the number of methyl substituents. It is therefore difficult to completely separate [3-and y-tocopherol and (3- and y-tocotrienols by reversed-phase HPLC, because both have the same side-chain structure and number of methyl substituents on the chromanol ring. Only six peaks are usually found in the reversed-phase HPLC method. Thus, reversed-phase HPLC is recommended for samples from animal tissues, which contain little or no P and y vitamers. 

In normal-phase HPLC on a silica column, the order of elution of tocopherols and tocotrienols is a-tocopherol, a-tocotrienol, [3-tocopherol, (3-tocotrienol, y-tocopherol, y-tocotrienol, 8-tocopherol, and 5-tocotrienol. Figure Dl.5.3 is an example of a normal-phase chromatograph of tocopherols and tocotrienols in rice bran oil. 

Fig. 13 HPLC of vitamin E. (A) Standards of vitamin E vitamers. Column, 5-p.m Supelcosil LC-Si (250 X 4.6-mm ID) mobile phase, isooctane/ethyl acetate (97.5 2.5), 1.6 ml/min fluorescence detection, excitation 290 nm, emission 330 nm. Peaks (1) a-tocopherol (2) a-tocotrienol (3) /3-tocopherol (4) y-tocopherol (5) /3-tocotrienol (6) y-tocotrienol (7) 5-tocopherol (8) 5-tocotrienol. (B) Saponified rice bran sample. Chromatographic conditions as in (A) except for mobile phase isooctane/ethyl acetate/2,2-dimethoxypropane (98.15 0.9 0.85 0.1). (From Ref. 228. AOCS Press.)...

The other five molecules which were identified using continuous-flow HPLC-1 H NMR spectroscopy at an observation frequency of 400 MHz were a-tocoenol with a mass of 428 (a double bond at Cl 1 ), a-tocotrienol with a mass of 424 (double bonds at C3, Cl and Cll ), (3-tocotrienol with a mass of 410 (double bonds at C3, Cl and Cl V, but with loss of a methyl on the phenyl ring), y-tocotrienol, also with a mass of 410 (like (3-tocotrienol but with loss of a different methyl group on the phenyl ring) and 8-tocotrienol with a mass of 396 (like (3-tocotrienol but with the loss of two methyl groups from the phenyl ring). 

Tocopherols and tocotrienols are precursors of vitamin E and are important antioxidants in oils. Their reactivity means that they are not stable to many oil processing procedures, including deodorization, which reduces levels by up to 15%. Levels of tocopherols in cocoa butter are usually about 100-300 mg/kg, with the y-isomer (IV) being the major component (about 90%), but they can be entirely absent (Lipp et al., 2001) (Figure 3.3). Tocotrienols have a similar structure with unsaturation of alternate bonds along the alkyl chain. Only y-tocotrienol is found in cocoa butter and this at low levels (< 5 mg/kg). Palm oil is notably high in tocopherols and tocotrienols, of which a-tocopherol and a-tocotrienol make up 20% to 30% each with most of the remainder as y-tocotrienol. 

Much less is known on the antioxidant activity of tocotrienols than tocopherols. Tocotrienols were shown to have similar reactivities to peroxyl radicals and antioxidant activities than tocopherols in solution and membranes (Yoshida et ah, 2003) also, in general, y-tocotrienol was a better antioxidant than a-tocotrienol, and tocotrienols were better than tocopherols in oil systems (Seppanen et ah, 2010). Recently, Muller et ah (2010) conducted a comparative study to investigate the four tocopherols, four tocotrienols, and a-tocopheryl acetate on their antioxidant activities in five different popular in vitro assays (FRAP, a-TEAC, DPPH, ORAC, and CL), which were adapted to nonpolar antioxidants. Most notably, they found that a-tocopheryl acetate, a popular ingredient in supplements, had no significant antioxidant activity in vitro. However, once ingested, tocol esters are hydrolyzed and antioxidant activities are retained. Overall, the eight tocols performed similarly in the five assays. The authors concluded that in vitro antioxidant assays performed in polar solvents are not a good way to predict in vivo antioxidant activity. 

Figure 4.1. Vitamin E vitamers, tocopherois and tocotrienois, and the synthetic water-soluble vitamin E analog, Trolox. Relative molecular masses (Mr) a-tocopherol, 430.7 (acetate 488.8, succinate 546.8) , 6-tocopherol, 419.7 y-tocopherol, 416.7 5-tocopherol, 402.7 a-tocotrienol, 424.7 , 6-tocotrienol, 410.7 y-tocotrienol, 410.7 5-tocotrienol, 396.7 and Trolox, 250.3.

From Table 8, it can be seen that ot-tocopherols and y-tocotrienols account for the major portion of the total tocopherols and tocotrienos present in palm oil. Gapor (9) confirms the presence of the above-listed tocopherols and tocotrienols by high-performance liquid chromatography (HPLC) and also indicated the probable presence of the esterified forms. 

The effect of tocotrienols on cancer progression was evaluated by Komi-yama and Yamaoka (177). The antitumor activity of tocotrienols was evaluated in terms of the increase in the lifespan of mice inoculated with tumor cells. a-Tocotrienols and y-tocotrienols were effective against the sarcoma cancer cell lines and Ehrlich carcinoma. When human lung carcinomas were challenged with these tocotrienols, a cytotoxic activity due to the tocotrienols was exhibited. Similarly. DMBA-treated rats responded with lower tumor numbers when their diets were supplemented with palm tocotrienols (178). Recently, a-carotene isolated from pahn oil has been shown to have antitumor activity against mouse lung cancer and against skin cancer (179). 

Rice bran oil and palm oil are the only readily available oils that contain significant levels (about 1000 ppm) tocotrienols (99). Tocotrienols belong to the Vitamin E family and have similar chemical stmcmres. According to Tomeo et al. (100), tocotrienols are powerful antioxidants. Commercially available RBO may contain 980 ppm y-tocotrienol (101). 

Palm oil is an exception among oil crops, because more than 85% of its vitamin E content consists of y-tocotrienol, a-tocotrienol and a-tocopherol (Tan, 1989). 

Barley grains contain all eight tocopherols and tocotrienols (Table 1.3), but there is a large variation in the total contents of tocopherols and tocotrienols. The major vitamers are a-tocotrienol (contributing to >50%), a-tocopherol, P-tocotrienol and y-tocotrienol, and they are generally correlated positively with each other (Peterson and Qureshi, 1993). The total vitamer content in whole-grain barley of 30 genotypes... 

Palm fatty acid distillate (PFAD) is a valuable by-product of palm oil processing, from which the production of tocopherols and tocotrienols is technically feasible (Ong and Choo, 1997). PFAD is wildly abundant, as palm oil is the second largest oil produced in the world. It contains 4000-8000 pg/g total vitamin E compounds, which is up to 10 times as much as crude palm oil contains (Tan, 1989 Ong and Choo, 1997). The tocopherol and tocotrienol profiles of PFAD and crude palm oil are similar, consisting of about 85% of tocotrienols, i.e., y-tocotrienol > a-tocotrienol > 8-tocotrienol (Tan, 1989). Because of cholesterol-lowering properties (Lane et al., 1999), the high level of tocotrienols makes PFAD an important source, because most other natural sources are devoid of tocotrienols. Fatty acids and esters, sterols and squalene are removed from PFAD as described above. Finally, the vitamin-E-rich product is purified and deodorized to yield a tocopherol/tocotrienol-rich product of 95-99% purity. 

In deep-fat frying, the temperature reaches >170°C, and tocopherols are unstable. The order of stability of different tocopherols remains similar to that at lower temperatures, i.e., a < y < 8 (Gordon and Kourimska, 1995 Lampi and Kamal-Eldin, 1998). However, an opposite order of stability has also been found. During deep-fat frying, the relative stabilities of natural tocopherols and tocotrienols in soybean oil were a-tocopherol > 8-tocopherol > P-tocopherol > y-tocopherol, in corn oil were a-tocopherol > y-tocopherol > 8-tocopherol > y-tocotrienol, and in palm oil were a-tocopherol > S-tocotrienol > a-tocotrienol > y-tocotrienol, respectively (Simonne and Eitenmiller, 1998). The authors assumed that tocotrienols were less stable than tocopherols, because they acted more effectively as antioxidants. 

Type of material a-Tocopherol a-Tocotrienol y-Tocotrienol 8-Tocotrienol Total (ppm)... 

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