透過您的圖書館登入
IP:54.162.130.75
  • 學位論文

在超臨界二氧化碳中利用脂肪酶對咖啡果皮中綠原酸酯化反應之研究

Direct lipase catalyzed chlorogenic acid from coffee pulp in supercritical carbon dioxide

指導教授 : 陳和賢 黃卓治

摘要


目前對於萃取天然抗氧化物質和生物轉化設計用在改善營養和健康食品的研究中,已受到日益關注。本研究利用一種普遍的熱帶農業廢棄物-咖啡殼,作為替代原料來生產具功能性的抗氧化物質。咖啡(Coffee arabica)殼中含有脂質和酚類化合物,故以超臨界二氧化碳(CO2) 分析其可溶性區分和不溶性區分的物質組成分。以GC-MS分析超臨界二氧化碳可溶性區分之組成分,含有咖啡因、C14、C16、C18和C20碳鏈脂肪酸、蠟酯和固醇等成分存在。以HPLC分析超臨界二氧化碳不溶性區分的組成分,證實咖啡殼中有高含量綠原酸。因此,第二部分,本研究旨在探討這一生物活性化合物 (綠原酸) 生物轉化反應。直接以固定化Candida antarctica脂肪酶B (Novozym 435) 在超臨界 CO2/三丁醇 (t-butanol) 中酯化綠原酸進行研究。使用田口方法評估反應溫度 (35-55°C),反應壓力 (150-250 bar) ,三丁醇用量 (2-10%,v/v) 和脂肪酶量 (10-30 mg/ml) 對酯化物濃度和整體轉換效率之影響。得到最佳反應條件為:壓力150 bar,溫度55°C, 三丁醇 10% (v/v),脂肪酶量20 mg/ml時,有最佳轉換率。另外,添加分子篩 (3 Å) 20 mg/ml,可減少因反應生成水所造成的抑制作用。在反應25小時的條件下,分別使用香葉醇(geraniol) 、戊醇 (pentanol) 和庚醇 (heptanol) 等醇類為脂肪酸碳鏈的提供者時,其最大的轉換率分別為77、82和85%。以HPLC分析證實,咖啡殼的水-甲醇萃取物在5-CGA選擇性酯化反應中,可達到轉換庚酯65%為最佳條件。

並列摘要


There has been a growing interest in the extraction and biotransformation of natural antioxidants for the design and improvement of nutraceuticals and foods beneficial to health. This research proposed the use of coffee pulp, a major tropical agroindustrial waste, as alternative bioresource for the production of functionalized antioxidants. Lipids and phenolic compounds contained in Coffea arabica pulp were analyzed as the SC CO2 soluble and non-soluble fractions, respectively. GC-MS analysis from SC CO2 soluble fractions showed the presence of caffeine, fatty acids of C14, C16, C18, and C20 carbon chains, wax esters, and sterols. HPLC results corresponding to SC CO2 non-soluble fractions confirmed a high content of chlorogenic acid in coffee pulp. Thus, the second part of this research aimed to study the biotransformation of this well known bioactive compound. The direct esterification of chlorogenic acid by immobilized Candida antarctica lipase B (Novozym 435) in supercritical CO2/t-butanol was studied. The Taguchi approach was applied to evaluate the effects of temperature (35-55 ◦C), pressure (150-250 bar), t-butanol (2-10 %, v/v), and the enzyme amount (10-30 mg/ml), on the ester concentration and overall conversion. Optimum reaction conditions were established at: 150 bar, 55 °C, 10 % t-butanol (v/v), 20 mg/ml of lipase. Addition of 20 mg/ml of molecular sieves (3Å) was also necessary to minimize the inhibiting effect of the increasing water concentration. Maximum conversions reached 77, 82 and 85 % in 25 h using geraniol, pentanol and heptanol as aliphatic chain donors, respectively. HPLC analysis demonstrated the selective esterification of chlorogenic acid from a coffee pulp aqueous-methanolic extract, which reached a conversion to heptyl ester of 65 % at the optimal condition.

參考文獻


van Dieren, S., C. S. P. M. Uiterwaal, Y. T. van der Schouw, D. L. van der A, J. M. A. Boer, A. Spijkerman, D. E. Grobbee, and J. W. J. Beulens (2009) Coffee and tea consumption and risk of type 2 diabetes. Diabetologia. 52(12): 2561–2569.
Adachi, S., and T. Kobayashi (2005) Synthesis of esters by immobilized-lipase-catalyzed condensation reaction of sugars and fatty acids in water-miscible organic solvent. J. Biosci. Bioeng. 99(2): 87-94.
Adams, M., and A. E. Ghaly (2007) Maximizing sustainability of the Costa Rican coffee industry. J. Cleaner Prod. 15(17): 1716-1729.
Andreasen, M. F., P. A. Kroon, G. Williamson, and M. T. Garcia Conesa (2001) Intestinal release and uptake of phenolic antioxidant diferulic acids. Free Radical Biol. Med. 31(3): 304-314.
Arlorio, M., J. D. Coisson, F. Travaglia, F. Varsaldi, G. Miglio, G. Lombardi, and A. Martelli (2005) Antioxidant and biological activity of phenolic pigments from Theobroma cacao hulls extracted with supercritical CO2. Food Res. Int. 38: 1009-1014.

延伸閱讀