傳統的植物浸漬標本保存方法，如:福馬林或70%酒精，雖可有效保存形態，但卻無法維持標本的色澤。花青素為花朵呈色的主要色素之一，此類水溶性小分子易受環境影響(如:pH值、溫度、酵素…等)產生降解，因此保存或安定了花青素即可維持花朵的主要色澤。本研究以蘭科植物中的石斛蘭屬(Dendrobium)花朵為材料，成功配製可有效保存石斛蘭的色澤、形態以及DNA之保存液。實驗中採用高效率液相層析法(HPLC)及酸鹼值差異法(pH-differential methods)檢測花青素在保存液中降解的情形，也進一步使用DNA電泳以及聚合酶連鎖反應放大技術(PCR)，探討此保存液對葉片DNA的影響。實驗發現，此保存液雖會造成花朵之花青素部分的降解，但仍可使其穩定維持目測之色澤。此保存液對葉片DNA也無顯著破壞，特定之基因序列仍可由PCR鑑定出來。由這些結果顯示，本研究之保存液確實可有效的保存石斛蘭標本的色澤、形態以及DNA，不僅在學術上有其重要價值，在商業上亦有其發展的潛力。

Although traditional plant preservation methods (e.g. soaking in formalin or in high percentage alcohol) can effectively preserved the configuration of specimens, losses of hues is an unsolved problem. Anthocyanins, the main pigment of colorful flowers, are water-soluble, unstable, and susceptible to external environmental factors such as pH, temperature, light, and other substances like enzymes. So preservation and stabilization of anthocyanins is here the key step in preserving hues of flowers. In this research, Dendrobium orchids are used as objects of study, we successfully developed a preservation solution which is able to keep the color, morphology, and DNA of Dendrobium orchids. In the study, we modified the pH-differential method and used HPLC to carry out quantitative and qualitative analysis of anthocyanins. Electrophoresis and PCR were used to analyze the effect of the preservation solution on leaf DNA. The results reveal that the content of anthocyanins declined significantly at the first two weeks then became stable in the long-term tracking. The hues of Dendrobium orchids are basically the same. There is no significant degradation of leaf DNA, and the DNA extracted from the mersed samples could be subjected to PCR amplification. These results suggest that the preservation solution not only can keep the appearances of the flower Dendrobium orchids but also the leaf DNA. This research has important potentials for academic and applicative purposes.

1. Andersen, O.M. (2002). Anthocyanins. In: Encyclopedia of life sciences. MacMillan Publishers Ltd., London, p.597-605.
2. Araceli Castaneda-Ovando, Ma. de Lourdes Pacheco-Hernandez, Ma. Elena Paez-Hernandez, Jose A. Rodriguez, & Carlos Andres Galan-Vidal. (2009). Chemical studies of anthocyanins: A review. Food Chemistry 113: 859–871.
3. Asen, S., Stewart, R.N., & Norris, K.H. (1972). Copigmentation of anthocyanins in plant tissues and its effect on color. Phytochemistry 11: 1139-1144.
4. Boselli, E., Boulton, R.B., Thorngate, J.H., & Frega, N.G. (2004). Chemical and sensory characterization of DOC red wines from Marche (Italy) related to vintage and grape cultivars. J Agric Food Chem 52: 3843-3854.
5. Boulton, R. (2001). The copigmentation of anthocyanins and its role in the color of red wine: A critical review. Am J Enol Vitic 52: 67-87.
6. Brouillard, R. (1982). Chemical structure of anthocyanins. In: Anthocyanins as Food Colors. Pericles Markakis (ed.), Academic Press Inc., New York, p.1-38.
7. Brouillard, R. (1983). The in vivo expression of anthocyanin color in plants. Phytochemistry 22: 1311-1323.
8. Brouillard, R., Mazza, G., Saad, Z., Albrecht-Gary, A.M., & Cheminat, A. (1989). The co-pigmentation reaction of anthocyanins: a microprobe for the structural study of aqueous solutions. J Am Chem Soc 111: 2604-2610.
9. Cabrita, L., Fossen, T., & Andersen, O. M. (2000). Colour and stability of the six common anthocyanidin 3-glucosides in aqueous solutions. Food Chemistry 68(1):101-107.
10. Cesar Romero-Sierra, Bath; John C. Webb, Kingston, both of Canada. Flower preservation. United states patent 4272571
11. Clifford, M. N. (2000). Anthocyanins – nature, occurrence and dietary burden. Journal of the Science of Food and Agriculture 80(7): 1063–1072.
12. Dlter von Wettstein, Gough, S., & Kannangara, G. (1995) Chlorophyll biosynthesis. The Plant Cell 7:1039-1057.
13. Dangles, O., Saito, N., & Brouillard, R. (1993). Kinetic and thermodynamic control of flavylium hydration in the pelargonidin- cinnamic acid complexation. Origin of the extraordinary flower color diversity of Pharbitis nil. J Am Chem Soc 115: 3125-3132.
14. Dao, L.T., Takeoka, G.R., Edwards, R.H., & Berrios, J.D.J. (1998). Improved method for the stabilization of anthocyanidins. J Agric Food Chem 46: 3564-3569.
15. Darias-Martin, J., Martin-Luis, B., Carrillo-Lopez, M., Lamuela-Raventos, R., Diaz-Romero, C., & Boulton, R. (2002). Effect of caffeic acid on the color of red wine. J Agric Food Chem 50: 2062-2067.
16. Davies, A.J., & Mazza, G. (1993). Copigmentation of simple and acylated anthocyanins with colorless phenolic compounds. J Agric Food Chem 41: 716-720.
17. Fossen, T., Cabrita, L., & Andersen, O.M. (1998). Color and stability of pure anthocyanins influenced by pH including the alkaline region. Food Chem 63: 435-440.
18. Francis, F.J. (1989). Food colorants: anthocyanins. Crit Rev Food Sci Nutr 28: 273-314
19. Garzon, G.A., & Wrolstad, R.E. (2001). The stability of pelargonidin-based anthocyanins at varying water activity. Food Chemistry 75(2): 185-196.
20. Giusti, M.M., & Wrolstad, R.E. (2003). Acylated anthocyanins from edible sources and their applications in food systems. Biochem Eng J 14: 217-225.
21. Goossens, A., Claes, L., Drieghe, J., & Put, E. (1997) Antimicrobials: preservatives, antiseptics and desinfectants. Contact Dermatitis. 39: 133-134.
22. Goshorn, R.H., Degering, E.D.F., & Tetrault, P.A. (1938) Antiseptic and bactericidal action of benzoic acid and inorganic salts-effect of pH. Industrial and Engineering Chemistry 30(6): 646-648.
23. Gutierrez, I.H. (2003). Influence of ethanol content on the extent of copigmentation in a Cencibel young red wine. J Agric Food Chem 51: 4079-4083.
24. Hale, K.L., McGrath, S.P., Lombi, E., Stack, S.M., Terry, N., Pickering, I.J. et al. (2001). Molybdenum sequestration in Brassica species.A role for anthocyanins. Plant Physiology 126:1391-1402.
25. Harborne, J.B. (1964). Plant polyphenols. XI. Structure of acylated anthocyanins. Phytochemistry 3: 151-160.
26. Harborne, J.B. (1967). Comparative Biochemistry of the Flavonoids. Academic, New York
27. Huang, H.T. (1955). Decolorization of anthocyanins by fungal enzymes. J Agric Food Chem 3: 141-146.
28. Lim, S.H., Liew, C.F., Lim, C.N., Lee, Y.H. & Goh, C.J. (1997) A simple and efficient method of DNA isolation from orchid species and hybrids. Biologia Plantarum 41(2): 313-316.
29. Lowry, J.B., & Keong, S.C. (1973). A preliminary study of Malaysian orchid pigments. Malays. J. Sci. 2 (B): 115
30. Markakis, P. (1982). Stability of anthocyanins in foods. In: Anthocyanins as Food Colors. Markakis P (ed.), Academic Press Inc., New York, p.163-178.
31. Markakis, P., Livingston, G.E., & Fellers, C.R. (1957). Quantitative aspects of strawberry-pigment degradation. Food Research 22: 117-130.
32. Mazza, G., & Brouillard, R. (1987). Recent developments in the stabilization of anthocyanins in food products. Food Chem 25: 207-225.
33. Mazza, G., & Brouillard, R. (1987). Color stability and structural transformations of cyanidin 3,5-diglucoside and four 3-deoxyanthocyanins in aqueous solutions. J Agric Food Chem 35: 422-426.
34. Mazza, G., & Brouillard, R. (1990). The mechanism of copigmentation of anthocyanins in aqueous solutions. Phytochemistry 29: 1097-1102.
35. Method and composition for plant preservation without leaf curling. US Patent 5627132
36. Nebesky, E.A., Esselen, W.B. Jr., McConnell. J.E.W., & Fellers, C.R. (1949). Stability of color in fruit juices. Food Research 14: 261-274.
37. Osawa, Y. (1982). Copigmentation of anthocyanins. In: Anthocyanins as Food Colors. Markakis P (ed.), Academic Press Inc., New York, p.41-65.
38. Palamidis, N., & Markakis, P. (1978). Stability of grape anthocyanin in carbonated beverages. Semana Vitivinicola 33: 2633, 2635, 2637-2639.
39. Pifferi, P.G., & Cultrera, R. (1974). Enzymic degradation of anthocyanins. Role of sweet cherry polyphenol oxidase. J Food Sci 39: 786-791.
40. Preservation of plant material. WO 99/55152
41. Rein, M. (2005) Copigmentation reactions and color stability of berry anthocyanins. Helsinki: University of Helsinki.
42. Robert, S. Dubrow, San Carlos, Calif. ; Geoffrey Samuels, New York,
Robert J. Molinari, Wayne, Pa. Specimen display article. United states
patent 5227205
43. Saito, N., Toki, K., Uesato, K., Atsushi, S., & Honda, T. (1994). An
acylated cyanidin glycoside from the red-purple flowers of Dendrobium.
Phytochemistry 37, 245–248.
44. Saito, N., Ku, M., Tatsuzawa, F., Lu, T.S., Yokoi, M., Shigihara, A.,
& Honda, T. (1995). Acylated cyanidin glycosides in the purple-red
flowers of Bletilla striata. Phytochemistry 40, 1523–1529.
45. Starr, M.S., & Francis, F.J. (1973). Effect of metallic ions on color and pigment content of cranberry juice cocktail. Journal of Food Science 38(6):1043-1046.
46. Tatsuzawa, F., Saito, N., Yokoi, M., Shigihara, A., & Honda, T. (1995).
Acylated cyanidin 3,7,30-triglucosides in flowers of x Laeliocattleya
cv Mini Purple and its relatives. Phytochemistry 41, 635–642.
47. Tatsuzawa, F., Saito, N., Seki, H., Hara, R., Yokoi, M., & Honda, T.(1997). Acylated cyanidin glycosides in the red-purple flowers of Phalaenopsis. Phytochemistry 45, 173–177.
48. Tatsuzawa, F., Yukawa, T., Shinoda, K., & Saito, N. (2005) Acylated antho- cyanins in the flowers of genus Dendrobium section Phalaenanthe (Orchidaceae). Biochemical Systematics and Ecology 33 : 625–629.
49. Timberlake, C.F., Bridle, P. (1966) Spectral studies of anthocyanin and anthocyanidin equilibria in aqueous solution. Nature 212: 158-159.
50. Williams, C.A., Greenham, J., Harborne, J.B., Kong, J.M., Chia, L.S., Goh, N.K., Saito, N., Toki, K., & Tatsuzawa, F. (2002). Acylated anthocyanins and flavonols from purple flowers of Dendrobium cv.”Pompadour”. Biochemical Systematics and Ecology 30(7): 667-675.
51. Williams, M., & Hrazdina, G. (1979). Anthocyanins as food colorants: effect of pH on the formation of anthocyanin-rutin complexes. J Food Sci 44: 66-68.
52. Yoshida, K., Kitahara, S., Ito, D., & Kondo, T. (2006). Ferric ions involved in the flower color development of the Himalayan blue poppy. Meconopsis Grandis Phytochemistry 67(10): 992-998.
53. Zhu Wenxue, Dong Tieyou, Zhang Zhengyong. Experimental study on treanment of immortal peony flower. Luoyang Institute of Technology, Luoyang, 471039
54. 王忠民, 石秀花, 李瑾瑜(2007) 野玫瑰色素理化性質的研究 食品科學-基礎研究 28(6): 93-97.
55. 王榮祥, 許亮, 宋吉猛, 白雄明 (2007) 原色植物浸漬標本製作方法的研究 遼寧中醫藥大學學報 9(4): 163-164.
56. 林國筆 (1990) 花朵的自然生態保存及其著色之方法 國內專利 159041
57. 郭玉梅 (1999)．新養蘭學II 洋蘭．台北 : 亞泰
58. 郭建華 (2005) 植物綠色固定保色AB液的實驗研究 廣西中醫學院學報 5(1): 3-5.
59. 黃肇宇 (2006) 植物標本原色澤的保色技術研究 玉林師範學院學報 27(3):126-128
60. 薛聰賢 (2002)．家庭園藝(第九輯) 蘭花專輯-養蘭不難．新店：台灣普綠。