Phenolic Acid Composition of Wheat Bran - American Chemical Society

Chapter 2

Downloaded by UNIV OF MICHIGAN ANN ARBOR on February 18, 2015 | http://pubs.acs.org Publication Date: July 21, 2005 | doi: 10.1021/bk-2005-0909.ch002

Phenolic Acid Composition of Wheat Bran Kequan Zhou, John W. Parry, and Liangli (Lucy) Yu Department of Nutrition and Food Science, 0112 Skinner Building, University of Maryland, College Park, MD 20742

Bran samples of wheat grown in Colorado were extracted with aqueous acetone followed by alkaline hydrolysis. After neutralization with HC1, the total free phenolic acids were extracted by ethyl acetate/ethyl ether (1:1, v/v), and analyzed by a reverse phase HPLC and LC-MS. The major phenolic acids detected in the wheat bran extracts were ferulic, syringic, p-coumaric, p-hydroxybenzoic, and vanillic acids. The results also showed that both wheat variety and the growing condition may influence the phenolic acid composition in wheat bran.

Reactive oxygen species, such as hydroxyl radicals and peroxyl radicals, may attack DNA, bioactive proteins, membrane lipids, and carbohydrates and cause damage, which may result in cell injury and death, and consequently the development of several aging-related chronic diseases including cancer and heart disease (/). Antioxidants may prevent these physiologically important molecules from oxidative damages, and reduce the risk of aging-related diseases (2,3). In addition to their health benefits, antioxidants may also suppress lipid oxidation in food products and improve food quality, stability and safety. Natural antioxidants are in high demand because of consumer preference and the potential safety concerns of synthetic antioxidants such as butylated hydroxytoluene (BHT), which is currently used as an antioxidative food additive.

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© 2005 American Chemical Society In Phenolic Compounds in Foods and Natural Health Products; Shahidi, F., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 2005.


11 There has been a continuous effort to develop novel edible natural antioxidants for disease prevention and health promotion, as well as for food preservation. Previous research has shown that wheat and wheat based food products have significant antioxidative activities and may serve as dietary sources of natural antioxidants (2-/0). Wheat antioxidants were shown to directly react with and quench free radicals, chelate transition metals, suppress lipid peroxidation in fish and soy oils, and prevent liposome peroxidation (3,4,6-8). Phenolics were present in several varieties of wheat and are believed to contribute to the overall antioxidant properties of wheat. A group of phenolic acids have been detected in wheat extracts. In 1982, Sosulski and others (77) reported that ferulic, syringic, and vanillic acids were the most prevalent phenolic acids present in the flour of Neepawa wheat, with ferulic acid accounting for 89.1% of the total. Later in 1992, Onyeneho and Hettiarachchy (7) detected ferulic, vanillic, p-coumaric, caffeic, chlorogenic, gentisic, syringic and p-hydroxybenzoic acids in bran extracts of durum wheat (Triticum durum). Among these phenolic acids, ferulic, vanillic, and p-coumaric acids were present in the greatest amounts. In addition, Hatcher and Kruger (72) detected six phenolic acids in extracts of five varieties of Canadian wheat. These included sinapic, ferulic, and vanillic acids, along with minor amounts of coumaric, caffeic and syringic acids. Previous studies have also shown that antioxidative properties of wheat might vary among wheat cultivars, and may be significantly altered by growing conditions and the possible interactions between genotype and growing condition (3,4,6). It is of interest whether growing condition and wheat cultivar may influence the contents of phenolic acids in wheat. Therefore, the present study was conducted to examine and compare the phenolic acid composition of bran extracts from Lakin, Venago, and Enhancer wheat grown at Burlington and Walsh in Colorado

Materials and Methods

Materials. Bran samples of Lakin, Venago, and Enhancer wheat from Burlington and Walsh in Colorado were provided by Dr. Scott Haley in the Department of Soil and Crop Science at Colorado State University, Fort Collins, Colorado. All other chemicals and solvents were of the highest commercial grade and used without further purification.

In Phenolic Compounds in Foods and Natural Health Products; Shahidi, F., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 2005.

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Extraction and testing sample preparation 4 grams of each bran sample was ground and extracted for 15 hours with 40 mL of 50% acetone under nitrogen at ambient temperature. After removing acetone using a rotary evaporator at 35°C, the extracts were hydrolyzed with 4N NaOH for 4 hours at 55°C under nitrogen, acidified using 6N HC1, and extracted with ethyl ether/ethyl acetate (1:1, v/v) according to the procedure described previously (13). The ethyl ether/ethyl acetate was evaporated at 25°C using a nitrogen evaporator, and the solid residue was re-dissolved in methanol, filtered through a 0.45 μιη membrane filter, and kept in dark under nitrogen until HPLC analysis.

HPLC analysis Phenolic acid composition in the methanol solution was analyzed by HPLC using a Phenomenex CI8 column (250 mm χ 4.6 mm). Phenolic acids were separated using a linear gradient elution program with a mobile phase containing solvent A (acetic acid/H 0, 2:98, v/v) and solvent Β (acetic acid/acetonitrile/ H 0, 2:30:68, v/v/v). The solvent gradient was programmed from 10 to 100% Β in 42 min with a flow rate of 1.5 mL/min (14). Identification of phenolic acids was accomplished by HPLC-MS and comparing the retention time of peaks in wheat samples to that of the standard compounds. 2

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Statistic analysis Data were reported as mean ± SD for triplicate determinations. Analysis of variance and least significant difference tests (SPSS for Windows, Version Rel. 10.0.5., 1999, SPSS Inc., Chicago, IL) were conducted to identify differences among means. Statistical significance was declared at PO.05.

Results and Discussion Phenolic acid composition in the hydrolysate of bran extracts was examined. Five phenolic acids, including p-hydroxy benzoic, coumaric, ferulic, syringic, and vanillic acids, were detected in both hard winter red and white varieties grown in Colorado (Figure 1A-B). Among these phenolic acids, ferulic acid had the greatest concentration in all three tested hard winter wheat varieties, and followed by syringic acid (Figure IB), regardless of wheat genotype. This is in agreement to the observation of Sosulski and others (//) and Onyeneho and


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Hettiarachchy (7) that ferulic acid was the predominant phenolic acid in wheat samples. Ferulic acid accounted 65.3-74.4% and 65.4-73.8% of total phenolic acids on per weight basis in hard winter red and white wheat bran samples, respectively.

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Figure 1. HPLC chromatograms of standard phenolic acids and the hydrolysate of bran extract. A: represents the HPLC of the standard phenolic acids, while B: is the HPLC result of the hydrolate of bran extract Gallic, protocatechuic, p-OH benzoic, chlorogenic, vanillic, caffeic, syringic, coumaric, andferulic standfor gallic, protocatechuic, p-OH benzoic, chlorogenic, vanillic, caffeic, syringic, coumaric andferulic acids, respectively.


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Figure 2. Comparison of white and red wheat for their phenolic acid contents. A: represents the phenolic acid composition ofLakin and Enhancer wheat grown at Burlington, while Β represents that ofLakin and Enhancer wheat grown at Walsh Colorado, respectively. Lakin is hard white winter wheat, and Enhancer is hard red winter wheat. The clear column stands for Lakin extract, while the solid column represents Enhancer extract, respectively. Vertical bar on the top of each column is the standard deviation (n = 3).



15 Lakin wheat, a hard white winter variety, was compared with Enhancer wheat, a hard red winter variety, for their phenolic acid compositions using bran samples from wheat grown at Burlington and Walsh testing locations in Colorado. Significant differences were observed in all of their phenolic acid compositions, except for the vanillic acid content in the two wheat brans from the Burlington location and the coumaric acid concentration in that from the Walsh location, suggesting that white and red wheat may differ in phenolic acid composition. Bran extracts prepared from Enhancer wheat grown at both locations had greater concentrations of total phenolics, ferulic, and syringic acids than that in Lakin bran extracts (Figure 2) under the experimental conditions. More varieties of both white and red wheat grown at different locations are required to compare and make a conclusion how red and white wheat differ in their phenolic acid composition in general. It is interesting to compare the two same colored wheat varieties for their phenolic acid compositions. Both Enhancer and Venago are hard red winter wheat. Bran extracts were prepared from Enhancer and Venago bran from Burlington and Walsh, and analyzed for their phenolic acid compositions. The results showed that the two red wheat extracts differed in their contents of total phenolic, ferulic, syringic, and vanillic acid contents (Figure 3). Enhancer bran had greater total and individual phenolic acid contents than Venago bran, regardless of growing location, suggesting the dependence of phenolic acid composition on wheat genotype. In addition, bran extracts of both varieties from Burlington exhibited greater total and individual phenolic acid contents than those grown at Walsh, indicating the potential effect of growing conditions on the phenolic acid content of wheat. These are in agreement with previous observations that both genotype and growing conditions may influence the antioxidant properties and total phenolic contents of wheat (3,4 6 - 8). An additional experiment was conducted to further evaluate the effect of growing condition on phenolic acid composition of wheat using Lakin (white wheat) and Enhancer (red wheat) bran samples. Figure 4A represents the phenolic acid composition of Lakin bran, and Figure 4B represents that of Enhancer bran from Burlington and Walsh, respectively. Lakin bran from Burlington exhibited greater total and all five individual phenolic acid contents than that from Walsh (Figure 4A). Enhancer bran from Burlington also had greater total, ferulic, syringic acid contents than that from Walsh, but the bran samples from the two locations had no difference in their coumaric and phydroxy benzoic acid contents (Figure 4B). These results suggest that growing condition may alter the total and individual phenolic acid contents in wheat, and the interaction between genotype and environment may also play a role in phenolic acid composition of wheat.


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Ferulic

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Figure 3. Phenolic acid compositions of two red wheat bran extracts. A: represents the phenolic acid composition ofEnhancer and Venago wheat grown at Burlington, while Β represents that ofEnhancer and Venago wheat grown at Walsh Colorado, respectively. Both Enhancer and Venago are hard red winter wheat. The clear column stands for Enhancer extract, while the solid column represents Venago extract, respectively. Vertical bar on the top of each column is the standard deviation (n = 3).


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Figure 4. Effect ofgrowing condition on phenolic acid composition of wheat. A: represents the phenolic acid composition of Lakin wheat grown at Burlington and Walsh, while Β represents that of Enhancer wheat grown at Burlington and Walsh, Colorado. Lakin is hard white winter wheat, and Enhancer is hard red winter wheat. The clear column stands for wheat grown at Burlington, while the solid column represents thatfromWalsh, Colorado, respectively. Vertical bar on the top of each column is the standard deviation (n = 3).


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Acknowledgement The author would like to thank Dr. Scott Haley in the Department of Soil and Crop Science, Colorado State University for providing wheat bran samples.


References 1.

Halliwell, B.; Gutteridge, J. M . C.; Cross, C. E. J. Lab. Clin. Med. 1992, 119, 598-620. 2. Baublis, A. J.; Clydesdale, Ε. M . ; Decker, E. A. Cereal Foods World. 2000, 45, 71-74. 3. Yu, L.; Haley, S.; Perret, J.; Harris, M . ; Wilson, J.; Qian, M . J. Agric. Food Chem. 2002a, 50, 1619-1624. 4. Yu, L.; Haley, S.; Perret, J.; Harris, M . Food Chem. 2002b, 78, 457-461. 5. Yu, L.; Davy, B.; Wilson, J.; Melby, C. L. J. Food Sci. 2002c, 67, 26002603. 6. Yu, L.; Perret, J.; Harris, M.; Wilson, J.; Haley, S. J. Agric. Food Chem. 2003, 51, 1566-1570. 7. Onyeneho, S. N . ; Hettiarachchy, N . S. J. Agric. Food Chem. 1992, 40, 1496-1500. 8. Zielinski, H.; Kozlowska, H. J. Agric. Food Chem. 2000, 48, 2008-2016. 9. Miller, H. E.; Rigelhof, F.; Marquart, L.; Prakash, Α.; Kanter, M . J. American College ofNutrition. 2000, 19, 312S-319S. 10. Abdel-Aal, E.; Hucl, P. J. Agric. Food Chem. 2003, 51, 2174-2180. 11. Sosulski, F.; Krygier, K.; Hogge, L. J. Agric. Food Chem. 1982, 30, 337340. 12. Hatcher, D. W.; Kruger, J. E. Cereal Chem. 1997, 74, 337-343. 13. Krygier, K.; Sosulski, F.; Hogge, L. J. Agric. Food Chem. 1982, 30, 330334. 14. Tüzen, M.; Özdemir, M . Turk. J. Chem. 2003, 27, 49-54.


Phenolic Acid Composition of Wheat Bran - American Chemical Society

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