Korean Journal of Fisheries and Aquatic Sciences (한국수산과학회지)

The Korean Society of Fisheries and Aquatic Science (한국수산과학회)
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Purification and Structure of Antioxidative Substance Derived from Tetraselmis suecica

Tetraselmis suecica유래의 항산화성 물질의 정제 및 구조

  • Kim Se Kwon (Department of Chemistry, Pukyong National University) ;
  • Byun Hee Guk (Department of Chemistry, Pukyong National University) ;
  • Park Pyo Jam (Department of Chemistry, Pukyong National University) ;
  • Adachi Kyoko (Marine Biotechnology Institute, Shimizu Laboratories)
  • Published : 2002.03.01
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Abstract

Tetraselmis suecica (T. suecica) of Prasinophyta was selected because the growth rate is comparatively higher and the culturing is also easy. In order to investigate antioxidative activity, the soluble elements of T. suecica were fractionated using water and organic solvents such as methanol, hexane, chloroform, ethylacetate and butanol. The chloroform fraction of T. suecica showed strong antioxidative activity. The potential antioxidative activity was detected in hexane: ethylacetate (1:5) once used the fractions by different mixtures of organic solvents. This fraction was further purified by preparative thin layer chromatography (PTLC) and repeated reverse-phase HPLC. On the basis of chemical and spectroscopic evidences obtained by UV, FT-IR, FAB-MS and NMR, the compound purified from T. suecica was identified as pheophorbide-a. The antioxidative activity of the compound was comparable to that of $\alpha$-tocopherol and could be act as an antioxidant in foods.

해양 미세조류는 해양생물의 영양원으로서 대부분 소비되고 있으며, 그 종류도 매우 다양하다. 이들 미세조류로부터 유용물질의 회수 및 이용은 미이용 수산자원의 효율적 이용과 아울러 생물 및 식량자원으로서 그리고 새로운 기능성 소재로서 활용될 수 있을 것이다. 본 연구에서는 이러한 해양 미세조류의 효율적인 이용을 위하여 비교적 성장이 빠르고 배양이 용이한 담녹조류인 T. suecica 중의 항산화성 물질을 분리정제하여 구조를 결정하였다. 미세조류 T. suecica의 유기용매 추출물 중의 항산화성은 chloroform 획분이 가장 좋았으며, 이 획분을 silica gel column chromatography, PTLC 및 HPLC를 사용하여 항산화성 물질을 분리${\cdot}$정제하였다. Silica gel column을 사용하여 분리된 획분의 항산화성은 hexane: ethylacetate (1:5)로 용출시킨 획분에서 가장 높았다. 이 획분은 PTLC에 전개하여 8개의 획분을 얻었으며, 그 중에서 Rf값 0.40인 획분에서 활성이 높게 나타났다 HPLC에서 단일 획분으로 정제된 물질의 항산화성은$IC_{50}$ 값으로 10.2$\mu$g/mL였으며, 화학적으로 합성된 항산화제인 BHT 및 BHA보다는 낮았지만 천연 항산화제인 $\alpha$-tocopherol과 비교될 수 있었다 이 물질의 화학적 구조는 chlorophyll을 전구체로 하는 pheophorbide-a로 동정되었으며, 항산화 활성은 a-tocopherol과 비교될 수 있어 식품 및 관련분야에서 항산화제로서 이용할 수 있을 것으로 기대된다.

Keywords

References

  1. Alonso, L., E.M. Grima, J.A.S. Perez, J.L.G. Sanchez and F.G. Camacho. 1992. Fatty acid variation among different isolations of a single strain of Isochrysis galbana. Phytochemistry, 31, 3901-3904 https://doi.org/10.1016/S0031-9422(00)97550-2
  2. Aprahamian, M., S. Evrard, P. Keller, M. Tsuji, G. Balboni, C. Damge and J. Marescaux. 1993. Distribution of pheophorbide-a in normal tissues and in experimental pancreatic cancer in rats. Anti-Cancer Drug Design, 8, 101-114
  3. Bergstrom, L.C., I. Vucenik, I.K. Hagen, S.A. Chernomorsky and R.D. Poretz. 1994. In-vitro photocytotoxicity of lysosomotropic immunoliposomes containing pheophorbide-a with human bladder carcinoma cells. J. Photochem. Photobiol B: Biol., 24, 17-23 https://doi.org/10.1016/1011-1344(94)07008-3
  4. Cahyana, A.H., Y. Shuto and Y. Kinoshita. 1992. Pyropheophytin a as an antioxidative substance from the marine alga, Arame (Eisenia bicyclis). Biosci. Biotech. Biochem., 56, 1533-1535 https://doi.org/10.1271/bbb.56.1533
  5. Cannell, R.J.P., S.J. Kellam, A.M. Owsianka and J.M. Walker. 1987. Microalgae and cyanobacteria as a source of glycosidase inhibitors. J. Gen. Microbiol., 133, 1701-1705
  6. Cannell, R.J.P., S.J. Kellam, A.M. Owsianka and J.M. Walker. 1988. Results of a large scale screen of microalgae for the production of protease inhibitors. Planta Medica, 56, 10-14
  7. Chernomorsky, S.A., R.D. Poretz and A.B. Segelman. 1984. The photodynamic effect of chlorophyll derivatives on murine myeloma cells in tissue culture. Photochem. Photobiol., 39, 49S https://doi.org/10.1111/j.1751-1097.1984.tb03403.x
  8. Choi, J.S., J.H. Lee, H.J. Park, H.G. Kim, H.S. Young and S.I. Mun. 1993. Screening for antioxidant activity of plants and marine algae and its active principles from primus davidina. Kor. J. Pharmacogn., 24, 299-303
  9. Endo, Y., R. Usuki and T. Kaneda. 1985. Antioxidant effects of chlorophyll and pheophytin on the autoxidation of oils in the dark. I. Comparison of the inhibitory effects. J. Am. Chem. Soc., 62, 1375-1378 https://doi.org/10.1007/BF02545962
  10. Fabregas, J. and C. Herrero. 1985. Marine microalgae as a potential source of single cell protein (SCP). Appl. Microbial. Biotechnol., 23, 110-113 https://doi.org/10.1007/BF00938962
  11. Glinski, J.A, E. David, T.C. Warren and G. Hansen. 1995. Inactivation of cell surface receptors by pheophorbide a, a green pigment isolated from Psychotria acuminata. Photochem. Photobiol., 62, 144-150 https://doi.org/10.1111/j.1751-1097.1995.tb05250.x
  12. Goldman, J.C. 1979. Outdoor algal mass cultures-I. Applications. Water Res., 13, 1-19 https://doi.org/10.1016/0043-1354(79)90249-5
  13. Gustafson, K.R., J.H. Cardellina II, R.W. Fuller, O.S. Weislow, R.F. Kiser, K.M. Snader, G.M. Patterson and M.R. Boyd. 1989. AIDS-antiviral sulfolipids from cyanobacteria (blue-green algae). J. Natl. Cancer. Inst., 81, 1254-1258 https://doi.org/10.1093/jnci/81.16.1254
  14. Hatano, T., H. Kagawa and T. Okawa. 1988. Two new flavonoids and other constituents in licence wet: their relative astnngency and radical scavening effect Chem. Pharm. Bull., 36, 2090-2097 https://doi.org/10.1248/cpb.36.2090
  15. Kao, C.Y. 1983. New perspective on the interactions of tetrodotoxin and saxitoxin with excitable membranes. Toxicon Suppl., 3, 211-219
  16. Kita, T. and Y. Fukuyo. 1988. Description of the gonyaulacoid dinoflagellate AIexandnum hiranoi sp. nov. inhabiting tidepools on Japanese Pacific Coast. Bull. Plank. Soc. Jap., 35, 1-7
  17. Korth, O., T. Hanke and B. Roder. 1998. Photophysical investigations of Langmuir-Blodgett mono- and multilayer films of pheophorbide-a. Thin Solid Films, 320, 305-315 https://doi.org/10.1016/S0040-6090(97)00786-4
  18. Le Tutour, B., F. Benslimane, M.P. Gouleau, J.P. Gouygou, B. Saadan and F. Quemeneur. 1998. Antioxidant and pro-oxidant activities of the brown algae, Laminaria digitata, Himanthalia elongata, Fucus vesiculosus, Fucus serratus and Ascophyllum nodosum. J. Appl. Phycol., 10, 121-129 https://doi.org/10.1023/A:1008007313731
  19. Lincoln, RA., K. Strupinski and J.M. Walker. 1991. Enzyme inhibitors from algae. Biochem. Soctrans., 19, 429S https://doi.org/10.1042/bst019429s
  20. Ma, L. and D. Dolphin. 1996. Nucleophilic reaction of 1,8-diazabicyc1o[5.4.0] undec-7-ene and 1,5-diazabicyc1o[4.3.0]non-5-ene with methyl pheophorbide-a unexpected products. Tetrahedron, 52, 849-860 https://doi.org/10.1016/0040-4020(95)00944-2
  21. Menzel, K. and A. Wild. 1989. Fatty acid composition in the lipids of some marine Chlorcoccales and Eustigmatales. Z. Naturforsch. Sect. C. Biosci., 44, 9-10
  22. Miki, W. 1991. Biological functions and activities of animal carotenoids. Pure Appl. Chem., 63, 141-146 https://doi.org/10.1351/pac199163010141
  23. Moore, R.E. 1996. Cyclic peptides and depsipeptides from cyanobacteria: A review. J. Indust. Microbiol., 16, 134-143 https://doi.org/10.1007/BF01570074
  24. Murakami, M., K. Makabe, S. Okada, K. Yamaguchi and S. Konosu. 1988. Screening of biologically active compounds in microalgae. Nippon Suisan Gakkaishi, 54, 1035-1039 https://doi.org/10.2331/suisan.54.1035
  25. Nakamura, M. and T. Yasumoto. 1985. Tetrodotoxin derivatives in puffer fish. Toxicon, 23, 271-276 https://doi.org/10.1016/0041-0101(85)90149-7
  26. Nakamura, Y., A. Murakami, K. Koshimizu and H. Ohigashi. 1996. Identification of pheophorbide a and its related compounds as possible anti-tumor promoters in the leaves of Neptunia oleracea. Biosci. Biotech. Biochem., 60, 1028-1030 https://doi.org/10.1271/bbb.60.1028
  27. Roder, B., D. Nather, T. Lewald, M. Barune, C. Nowak and W. Freyer. 1990. Photophysical properties and photodynamic activity of some tetrapyrroles in vivo. Biophys. Chem., 35, 302-312
  28. Roder, B., T. Hanke, S. Oelckers and C. Symietz. 1999. Photophysicalproperties of pheophorbide a in solution and in model mem-brane systems. J. Photochem. Photobiol. B: Biol., 44, 37-44
  29. Ruckmann, I., A. Zeug, T. Von Feilitzsch, and B. Roder. 1999.Onentational relaxation of pheophorbide-a molecules in theground and in the Krst excited state measured by transientdichroism spectroscopy. Optics Communications, 170, 361-372 https://doi.org/10.1016/S0030-4018(99)00488-5
  30. Sakata, K., K. Yamamoto, H. Ishikawa, A. Yagi, H. Etoh and K. Ina.1990. Chlorophyllone-a, a new pheophorbide-a related com-pound isolated from Ruditapes philippinarum as an antioxida-tive compound. Tetrahedrom Lett., 31, 1165-1168 https://doi.org/10.1016/S0040-4039(00)88754-7
  31. Sato, M., I. Fujimoto, T. Sakai, T. Aimoto, R. Kimura and T. Murata.1986. Effect of sodium copper chlorophyllin on lipid peroxi-dation. DC. On the antioxidative components in commercialpreparations of sodium copper chlorophyllin. Chem. Pharm.Bull., 34, 2428-2434 https://doi.org/10.1248/cpb.34.2428
  32. Segelman, A.B., I.K. Hagen, S.A. Chemomorsky, K. Weadock and G.H. Sigel. 1987. Highly puriIied pheophorbide-a as a photo-sensitizer in human bladder cancer in vitro, In New Directionin Photodyaamic Therapy, D.C. Nechers, ed., SPIE, Bellingham,pp. 205-209
  33. Servel, M.O., C. Claire, A. Derrien, and L. Coiffard. 1994. Fatty acidcomposition of some marine microalgae. Phytochemistiy, 36,691-693 https://doi.org/10.1016/S0031-9422(00)89798-8
  34. Tsien, R., D.P.L. Green, S.R Levinson, B. Rudy and J.K.M. Sanders. 1975. A pharmacologically active derivative of tetrodotoxin. Proc. R. Soc. Lond. B., 191, 555-559 https://doi.org/10.1098/rspb.1975.0145
  35. Tsuda, K., S. Ikuma, M. Kawamura, R. Tachikawa, K. Sakai, C. Tamura and O. Amakasu. 1964. Tetrodotoxin. VII. On the structures of tetrodotoxin and its derivatives. Chem. Pharm. Bull., 12, 1375-1374
  36. Viso, A.C. and J.C. Marty. 1993. Fatty acids from 28 marine microalgae. Phytochemistry, 34, 1521-1533 https://doi.org/10.1016/S0031-9422(00)90839-2
  37. Volkman, J.K., S.W. Jeffrey, P.D. Nichols, G.I. Rogers and C.D. Garland. 1989. Fatty acid and lipid composition of 10 species of microalgae used in mariculture. J. Exp. Mar. Biol. Ecol., 128, 219-240 https://doi.org/10.1016/0022-0981(89)90029-4
  38. Yang, H.Y., Z.Y. Zhang, Z.H. Han and S.D. Yao. 2000. An investigation of the triplet state of pheophorbide-a using laser flash photolysis. Dyes and Pigments, 46, 139-143 https://doi.org/10.1016/S0143-7208(00)00048-6
  39. Yongmanitchai, W. and O.P. Ward. 1991. Screening of algae for potential alternative source of eicosapentaenoic acid. Phytochemistry, 30, 2963-2967 https://doi.org/10.1016/S0031-9422(00)98231-1

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