本實驗的主要目的，是以離心分配層析來製備分離、純化桑葉中 五種具有療效的成分，包括Chlorogenic acid、Isoquercitrin、Astragalin、
Quercetin 3-(6-malonylglucoside)、Kaempferol 3-(6-malonylglucoside) 等，經由偵測ESI-MS 已可初步確認這些成分結構。這些成分的功效 包括：Chlorogenic acid 可以減緩釋放葡萄糖進入血液中； Isoquercitrin 能抑制肥胖細胞的去顆粒化；Astragalin 可有效減少發炎反應；Q3MG 和K3MG 具有預防罹患動脈粥樣硬化之相關病症。
由於桑葉中的成分相當複雜，所以藉由EA 和 H2O 初步的批式萃 取除去極性較大的化合物，使欲分離的目標物單純化，因此更容易嘗
試出最佳化的溶劑系統。本實驗使用兩組 CPC 的溶劑系統，分別是
(MTBE/acetone/H2O = 6：4：10) 和 (MTBE/acetone/H2O = 6：4：
10)額外加入溶劑系統總體積 0.6% (v/v)的 formic acid，都是以 descending mode 進行操作，靜相滯留率均可以高達 85%以上，實驗
結果可成功分離出五種有效成分，且純度在 90%以上，本實驗雖然採
取相對小量的製備，但 CPC 具有在實驗條件不變的情況下容易被放
大之特性。最後再藉由NMR 進一步做結構鑑定。
在本實驗中這五種目標物的純度仍無法到達99%以上，若要提高 純度，未來還可以將第一次分管收集回收的樣品，再嘗試尋找另一組 合適的溶劑系統，然後藉由 HPLC 的分析，並計算出 KD值，若欲分 離的目標物在選用的CPC 溶劑系統中有好的分佈(KD值介於0.2~5 之 間)，就可再進行一次 CPC 的製備分離，經由分管收集然後進行 HPLC 的分析並計算純度，希望可達到純度提高的效果。
參考文獻
[1] Z. S. Jia, M. C. Tang, J. M. Wu, "The determination of flavonoid
contents in mulberry and their scavenging effects on superoxide radicals". Food Chem. 64 (1995) 555–559.
[2] K. Doi, T. Kojima, M. Makino, Y. Kimura, Y. Fujimoto, "Studies on the constituents of the leaves of M. alba L.". Chem. Pharm. Bull. 49 (2001) 151–153.
[3] T. Katsube, Y. Tsurunaga, M. Sugiyama, T. Furuno, Y. Yamasaki,
"Effect of air-drying temperature on antioxidant capacity and stability of polyphenolic compounds in mulberry (Morus alba L.) leaves".
Food Chem. 113 (2009) 964–969.
[4]T. Matsuoka, T. Kimura, N. Muraoka, "Research of the available constituents from mulberry tree". Tohoku Agricultural Research 47 (1994) 361–362.
[5] A. Onogi, K. Osawa, H. Yasuda, A. Sakai, H. Morita, H. Itokawa, "
Flavonol glycosides from the leaves of M. alba L.". Shoyakugaku Zasshi 47 (1993) 423–425.
[6] B. Enkhmaa, K. Shiwaku, T. Katsube, K. Kitajima, E. Anuurad, M.
Yamasaki, Y. Yamane, "Mulberry (Morus alba L.) Leaves and Their Major Flavonol Quercetin 3-(6-Malonylglucoside) Attenuate Atherosclerotic Lesion Development in LDL Receptor–Deficient Mice". Shimane Institute for Industrial Technology, Matsue City, Shimane 690-0816, Japan.
[7] W. Zheng, M. N. Clifford, "Profiling the chlorogenic acids of sweet potato (Ipomoea batatas) from China". Food Chem. 106 (2008) 147–152.
[8] K. L. Johnston, M. N. Clifford, L. M. Morgan, "Coffee acutely modifies gastrointestinal hormone secretion and glucose tolerance in humans: Glycemic effects of chlorogenic acid and caffeine". Am. J.
Clinical Nutrition 78 (2003) 728–733.
[9] K. L. Johnston, M. N. Clifford, L. M. Morgan, "Coffee acutely modifies gastrointestinal hormone secretion and glucose tolerance in humans: Glycemic effects of chlorogenic acid and caffeine". Am. J.
Clinical Nutrition 78 (2003) 728–733.
[10] T. Koda, Y. Kuroda, H. Imai, "Protective effect of rutin against spatial memory impairment induced by trimethyltin in rats".
Nutrition Research 28 (2008) 629–634.
[11] C. L. Casa, I. Villegas, C. A. Lastra, V. Motilva, M. Calero,
"Evidence for protective and antioxidant properties of rutin, a natural flavone, against ethanol induced gastric lesions". J.
Ethnopharmacol. 71 (2000) 45-53.
[12] N. Kamalakkannan, P. Prince, "Rutin improves the antioxidant status in streptozotocin-induced diabetic rat tissues". Mol. Cell Biochem 293 (2006) 211-9.
[13] C. G. Silva, R. J. Raulino, D. M. Cerqueira, S. C. Mannarino, M. D.
Pereira, A. D. Panek, J. F. M.Silva, F. S. Menezes, E. C. A.
Eleutherio, "In vitro and in vivo determination of antioxidant activity and mode of action of isoquercitrin and Hyptis fasciculata".
Phytomedicine 16 (2009) 761–767.
[14] W.H. Mager, J. Winderickx, "Yeast as a model for medical and medicinal research". Trends Pharmacol. Sci. 26 2005 265–273.
[15] M. Kotani, M. Matsumoto, A. Fujita, S. Higa, W. Wang, M.
Suemura, T. Kishimoto, T. Tanaka, "Persimmon leaf extract and astragalin inhibit development of dermatitis and IgE elevation in NC/Nga mice". J. Allergy Clin. Immunol. July (2000).
[16] K. Ishiguro, H. Oku, "Antipruritic effect of flavonol and 1,4-naphthoquinone derivatives from Impatiens balsamina L.".
Phytother Res 11 (1997) 343-7.
[17] L. Wang, Y. C. Tu, T. W. Lian, J. T. Hung, J. H. Yen, M. J. Wu,.
"Distinctive antioxidant and anti-inflammatory effects of flavonols".
J. Agric and Food Chem. 54 (2006) 9798-9804.
[18] F. Li, F. Wang, F. Yu, Y. Fang, Z. Xin, F. Yang, J. Xu, L. Zhao, Q.
Hu, "In vitro antioxidant and anticancer activities of ethanolic extract of selenium-enriched green tea". Food Chem. 111 (2008) 165-170.
[19] S. Komatsu, N. Ahsan, "Soybean proteomics and its application to functional analysis". J Proteomics 72 (2009) 325 – 336.
[20] A. Panossian, G. Wikman, "Pharmacology of Schisandra chinensis Bail.: An overview of Russian research and uses in medicine". J Ethnopharmacology 118 (2008) 183-212.
[21] Y. Ito, "Development of Countercurrent Chromatography". Anal.
Chem.(1984), (56), 534-551.
[22] Y. Ito, N. B. Mandava, "Countercurrent Chromatography : Theory and Practice ". Marcel Dekker, Inc., New York, (1998).
[23] W. D. Conway, "Countercurrent Chromatography : Theory and Applications". VCH Publishers, Inc., (1990).
[24] Y. Ito, W. D. Conway, "High-Speed Countercurrent Chromatography". John Wiley & Sons, Inc., (1996).
[25] E. Delannay, A. Toribio, L. Boudesocque, J. M. Nuzillard, Z. H.
Monique, E. Dardennes, G. L. Dour, J. Sapi, J. H. Renault, "Multiple dual-mode centrifugal partition chromatography, a semi-continuous development mode for routine laboratory-scale purifications". J.
Chromatogr. A, 1127 (2006) 45–51.
[26] E. Delannay, A. Toribio, L. Boudesocque, J. M. Nuzillard, M.
Hanrot, E. Dardennes, G. L. Dour, J. Sapi, J. H. Renault, "Multiple dual-mode centrifugal partition chromatography, a semi-continuous development mode for routine laboratory-scale purifications". J.
Chromatogr. A 1127 (2006) 45–51.
[27] E. Gavioli, N. M. Maier, C. Minguillo´n, and W. Lindner,
"Preparative Enantiomer Separation of Dichlorprop with a Cinchona-Derived Chiral Selector Employing Centrifugal Partition Chromatography and High-Performance Liquid Chromatography: A Comparative Study". Anal. Chem. (2004), 76, 5837-5848.
[28] J. B. Friesen, G. F. Pauli, "Rational development of solvent system families in counter-current chromatography". J. Chromatogr. A, 1151 (2007) 51–59.
[29] Y. Ito, "Golden rules and pitfalls in selecting optimum conditions for high-speed counter-current chromatography". J. Chromatogr. A, 1065 (2005) 145–168.
[30] W. D. Conway, Y. Ito, "Resolution in Countercurrent Chromatography". J. Liq. Chromatogr. (1985), 8, (12), 2198-2207.
[31] J. C. Delaunay, C. Castagnino, C. Che`ze, J. Vercauteren,
"Preparative isolation of polyphenolic compounds from Vitis vinifera by centrifugal partition chromatography". J. Chromatogr. A, 964 (2002) 123–128.
[32] N. Rubio, C. Minguillon, "Preparative enantioseparation of (±)-N-(3,4-cis-3-decyl-1,2,3,4-tetrahydrophenanthren-4-yl)-3,5-din itrobenzamide by centrifugal partition chromatography". J.
Chromatogr. A, 1217 (2010) 1183-1190.
[33] G. C. Yen, S. C. Wu, P. D. Duh, "Extraction and Identification of Antioxidant Components from the Leaves of Mulberry (Morus alba L.)". J. Agric. Food Chem., (1996), 44.
[34] H. Y. Lin, L. S. Lai, "Isolation and viscometric characterization of hydrocolloids from mulberry (Morus alba L.) leaves". Food Hydrocolloids 23 (2009) 840–848.
[35] T. Katsube, N. Imawaka, Y. Kawano, Y. Yamazaki, K. Shiwaku, Y.
Yamane, "Antioxidant flavonol glycosides in mulberry (Morus alba L.) leaves isolated based on LDL antioxidant activity". Food Chem.
97 (2006) 25–31.
[36] W. Zhi, Q. Deng, "Purification of salvianolic acid B from the crude extract of Salvia miltiorrhiza with hydrophilic organic/salt-containing aqueous two-phase system by counter-current chromatography". J. Chromatogr. A, 1116 (2006) 149–152.
[37] N. Rubioa, S. Ignatovac, C. Minguillona, I. A. Sutherlandc,
"Multiple dual-mode countercurrent chromatography applied to chiral separations using a (S)-naproxen derivative as chiral selector". J. Chromatogr. A, 1216 (2009) 8505-8511.
[38] B. Pinel, G. Audob, S. Mallet, M. Lavault, F. D. L. Poype, D.
S´eraphin, P. Richommea, "Multi-grams scale purification of xanthanolides from Xanthium macrocarpum Centrifugal partition chromatography versus silica gel chromatography". J. Chromatogr.
A, 1151 (2007) 14–19.
[39] K. Azuma, M. Nakayama, M. Koshioka, K. Ippoushi, Y. Yamaguchi, K. Kohata, Y. Yamauchi, H. Ito, H. Higashio, "Phenolic Antioxidants from the Leaves of Corchorus olitorius L". J. Agric.
Food Chem. (1999), 47, 3963-3966.
[40] S. Deng, Z. Deng, Y. Fan, Y. Peng, J. Li, D. Xiong, R. Liu,
"Isolation and purification of three flavonoid glycosides from the leaves of Nelumbo nucifera (Lotus) by high-speed counter-current chromatography". J. Chromatogr. B, 877 (2009) 2487-2492.