臺灣產稻米的機能性成分研究

Abstract

稻米中含有多種具有生物活性的植物化學成分，如生育醇 (tocopherols, Toc)、生育三烯醇 (tocotrienols, T3)、γ-谷維素 (γ-oryzanol) 以及多酚類成分 (polyphenolics)。這些植物化學成分分布在稻米的不同部位中，並各自有不同的生物活性。本研究的目的為 (1) 針對稻米中的維生素E異構物及γ-谷維素開發一套可以同步分析九種活性成分的方法，及 (2) 選定數種臺灣常見稻米進行生育醇、生育三烯醇、γ-谷維素及多酚類機能性成分含量分析。 本研究成功建立一套藉由正相高效能液相層析法 (NP-HPLC) 定量稻米中八種維生素E異構物及γ-谷維素的方法。使用Inertisil SIL 100A分離管柱，動相溶液由正己烷/異丙醇/乙酸乙酯/醋酸 (97.6:0.8:0.8:0.8, v/v/v/v) 組成，於25分鐘內完全分離九種活性成分。此方法具有良好的再現性及線性決定係數 (r2 > 0.99)，八種維生素E異構物的線性濃度範圍介於0.05-10 μg mL-1，而γ-谷維素則介於0.5-500 μg mL-1。此新建立之方法具有可靠的分離結果、分析時間短、及低成本等優點，因此可適用於大量稻米樣品的分析。 國產稻米不同部位中的生育醇、生育三醇及γ-谷維素的分析結果顯示，米糠具有最高濃度的生育醇 (42.5-123.5 mg kg-1)、生育三烯醇 (111.3-191.4 mg kg-1)及γ-谷維素(1728.4-3440.2 mg kg-1)。米糠中的總維生素E含量為糙米中的5.5倍，精白米中的20倍以上，而米糠中的γ-谷維素則是糙米中的5倍之多。稻米中的維生素E主要為γ-T3，共佔總維生素E含量約48.16%，而β-T3之含量為最低，僅佔0.20%。根據不同稻米品種的結果顯示，Japonica比Indica含有更高的生育醇、生育三烯醇及γ-谷維素含量，其中又以有色米種高於非有色米種。結果亦發現不同亞種稻米中維生素E組成有顯著差異，Indica具有較高的γ-T3含量比例，約佔總維生素E含量的54.70%，而Japonica則是具有較高的α-Toc含量比例，約佔總維生素E含量的30.79%。 多酚類分析結果顯示，米糠中的濃度最高，而稻殼中的種類最多。米糠中總酚含量 (TPC; 1.29-14.28 g of GAE kg-1)、總類黃酮含量 (TFC; 1.23-25.83 g of QE kg-1)、阿魏酸 (7.32-67.95 mg kg-1)及香豆酸 (4.64-39.73 mg kg-1)含量皆高於稻米其他部位。根據所分析的稻米品種中，總類黃酮含量皆高於總酚類含量。不同稻米品種結果顯示，Japonica比Indica含有較高濃度的總酚類及總類黃酮類，其中有色米品種之含量更顯著高於非有色米品種，這是因為有色米中含有大量的花青素。稻米中最主要的酚類化合物為阿魏酸及香豆酸，其他有兒茶素、香草酸及咖啡酸等。根據稻米不同部位分析結果發現，阿魏酸主要分布在米糠中，而香豆酸則主要分布在稻殼中。 總之，本研究成功建立一套快速、再現性好及低成本的稻米中九種機能性成分，如生育醇、生育三烯醇及γ-谷維素的分析方法，並依此方法完成二十種國產稻米中的這些機能性成分分析。結果發現，米糠中具有最高濃度的總維生素E、生育醇、生育三烯醇、γ-谷維素及多酚類化合物，同時亦發現Japonica稻米品種普遍上比Indica含有較高的機能性成分，而有色米品種中的這些成分含量普遍比非有色米高。

Whole rice contains several bioactive phytochemicals such as tocopherols (Toc), tocotrienols (T3), γ-oryzanol, and polyphenolic compounds. They have been reported to possess different biological activities and are distributed in different grain parts of rice. The objectives of this study were: (i) to develop an improved analytical method for simultaneous quantification of eight vitamin E analogs (α-, β-, γ- and δ-tocopherols and -tocotrienols) and γ-oryzanol in rice, and (ii) to examine the Toc, T3, γ-oryzanol, and polyphenolic contents in different grain parts of selected commercial rice varieties in Taiwan. A normal phase high performance liquid chromatography (NP-HPLC) was developed to analyze eight vitamin E isomers and γ-oryzanol in rice. A complete separation of all these nine compounds was achieved using an Inertisil SIL 100A column with the isocratic elution comprising of n-hexane/isopropanol/ethyl acetate/ acetic acid (97.6:0.8:0.8:0.8, v/v/v/v) as mobile phase within 25 min. A good reproducibility and a high coefficient of determination (r2 > 0.99) were obtained at concentrations ranging from 0.05-10 μg mL-1 for all vitamin E isomers and 0.5-500 μg mL-1 for γ-oryzanol, suggesting that the newly established method is accurate, fast and inexpensive for routine determinations of tocopherols, tocotrienols and γ-oryzanol in rice samples. Results on the quantification of Toc, T3, γ-oryzanol, polyphenolic contents in the different grain parts of the selected commercial rice varieties showed that rice bran contained the highest levels of phytochemicals, including Toc (42.5-123.5 mg kg-1), T3 (111.3-191.4 mg kg-1), γ-oryzanol (1728.4-3440.2 mg kg-1), total phenolic content (TPC; 1.29-14.28 g of GAE kg-1), total flavonoid content (TFC; 1.23-25.83 g of QE kg-1), ferulic acid (7.32-67.95 mg kg-1), and p-coumaric acid (4.64-39.73 mg kg-1). The level of vitamin E in rice bran was 5.5 times greater than in brown rice and over 20 times higher than in the polished rice, whereas the level of γ-oryzanol in rice bran was 5 times greater than in brown rice. Regardless of the grain parts, the Toc, T3 and γ-oryzanol contents present in the Japonica rice varieties was higher than in the Indica rice varieties, and contents of these compounds were found higher in the colored rice varieties than in the non-colored rice varieties. γ-T3 was the main vitamin E isomer (48.16%) found in all rice varieties, while β-T3 present only in trace amount (0.20%). The Indica rice varieties generally contained a higher proportion of γ-T3, which account for about 54.70% of the total vitamin E content, while the Japonica rice varieties had a higher proportion of α-Toc (about 30.79% of the total vitamin E). The polyphenolic content was present mainly in rice bran and rice husk. All analyzed rice varieties exhibited a higher level of TFC than the TPC. The Japonica rice varieties showed a significantly higher level of TPC and TFC than the Indica rice varieties. The major phenolic constituents present in the selected rice samples were ferulic acid, p-coumaric acid, catechin, vanillic acid and caffeic acid. Regardless of the grain parts, ferulic acid was the most abundant polyphenolic compounds in all rice samples and was mainly distributed in the rice bran, while p-coumaric acid was present mainly in the rice husk. The contents of individual polyphenolic compounds in the red and black rice varieties were higher than the other varieties, it was attributed to the high content of anthocyanin in the colored rice bran. In conclusion, the present study has established a rapid, accurate, reproducible and inexpensive NP-HPLC method for simultaneous determination of nine major bioactive compounds of rice, including four tocopherols (α-, β-, γ- and δ-T), four tocotrienols (α-, β-, γ- and δ-T3) and γ-oryzanol. Studies also showed that the rice bran present the highest content of total vitamin E, Toc, T3, γ-oryzanol, TPC, TFC, and polyphenolic compounds. The content of these phytochemicals in the Japonica rice varieties were generally higher than in the Indica rice varieties, while the colored rice varieties were higher than the non-colored rice varieties.