臺灣博碩士論文加值系統

海馬 (seahorse) 為一種硬骨海洋動物，廣泛應用於中藥材，多以泡製為藥酒之方式供人類食用。本研究以常見海馬品種之一─庫達海馬 (Hippocampus kuda, HK) 凍乾研磨為粉後，進行成分分析，得知粗蛋白 69.16% (以乾重計，DW)、粗脂肪 0.86%、灰分 28.06%、總醣類 1.00 mg/g、總酚 0.31 mg/g及總類黃酮 0.03 mg/g。再以 10 倍量 15% 及 70% 乙醇搖晃萃取 6 hr，萃取液凍乾後分別作為 15HK 及 70HK，另取部分 15HK 經蛋白酶 pronase E 處理，凍乾後作為 15PHK。其中 15HK 之蛋白質、總醣類、總酚及總類黃酮含量，測定結果分別為 29.33、13.60、9.53 mg/g DW 及未檢出 (not detected, ND)；15PHK 分別為 12.13、11.75、13.28 mg/g DW 及 ND；70HK 分別為7.97、31.53、11.87 及 2.43 mg/g DW。接著以測定 2,2-diphenyl-1-picrylhydrazyl 自由基清除活性 (DPPH radical scavenging activity)、還原力 (reducing power) 及 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) 自由基清除活性 (ABTS radical scavenging activity) 等測定 15HK、15PHK 及 70HK 抗氧化活性，顯示 15PHK 有較高之抗氧化效果，3 種抗氧化活性之測定結果分別為 43.57%、0.12 Abs. 及 25.56%。接著將 15PHK 透過快速蛋白質層析儀 (fast protein liquid chromatography; FPLC) 搭配 Hiprep DEAE FF 16/10 column 進行純化，得到 5 種 fractions，再分別測定抗氧化性，結果顯示 5 種 fractions 純化後之抗氧化效果較未純化之 15PHK 差。
其次為進行抗腫瘤之細胞實驗，將 3 種抽出物針對人類子宮頸癌細胞 Hela、人類神經母瘤細胞 SH-SY5Y、人類肺癌細胞 A549 及人類黑色素瘤細胞 A375.S2 細胞進行抗腫瘤活性實驗，結果顯示以 70HK 對 A549 細胞具較佳抑制效果，進而測定此細胞之乳酸去氫酶釋放量，結果顯示隨著樣品處理濃度越高則數值隨之增加，表示細胞損傷程度越嚴重。一氧化氮 (nitric oxide, NO) 生成量測定實驗中，隨著樣品處理濃度越高則數值隨之增加，表示細胞透過發炎反應使其產生 NO。活性氧物質 (reactive oxygen species, ROS) 生成量測定，亦有隨樣品處理濃度越高，生成量上升之現象，ROS 含量過多可使細胞快速分裂，對人體造成損傷。於粒線體膜電位 (mitochondrion membrane potential, MMP) 測定中，隨著樣品處理濃度越多，則表現量越低，是由於細胞發生凋亡現象所導致。最後進行細胞凋亡壞死分析，結果可發現細胞發生凋亡現象，且隨樣品處理濃度越高，而逐漸趨向壞死之表現。
綜上所述，海馬 3 種抽出物中，抗氧化效果以 15PHK 為最佳，而 70HK 對人類肺癌細胞 A549 可有效降低細胞活性，其機制推測為透過 NO 及 ROS 生成導致細胞發生凋亡。

Seahorse is a small marine teleost animal. It has been used as traditional medicine for thousands of years in Eastern Asia because it has positive effects on human’s health. In this study, seahorses were collected from Penghu, freezing-dried and grounded into powder. The biochemical compositions of Hippocampus kuda (HK) powder were analyzed as follows: crude protein 69.16% (DW), crude lipid 0.86%, ash 28.06%, total saccharide 1.00 mg/g, total phenolic content 0.31 mg/g, and total flavonoids 0.03 mg/g. Furthermore, seahorses were derived into three parts. Two parts were extracted with 15% and 70% ethanol, and freezing-dried as 15HK and 70HK extracts. One part was extracted by 15% ethanol and further hydrolyzed by pronase E. The hydrolysate was as 15PHK hydrolysate. The protein content was 29.33, 12.13 and 7.97 mg/g DW in terms of 15HK extract, 15PHK hydrolysate and 70HKextract, respectively, and respective total saccharide content was 13.60, 11.75 and 31.53 mg/g, respective total phenolic content was 9.53, 13.28 and 11.87 mg/g, respective total flavonoid content was ND, ND and 2.43 mg/g. Three groups were also tested for their antioxidant activities including 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity, reducing power and 2,2'-azino-bis(3-ethylbenzothiazoline -6 -sul-
phonic acid (ABTS) radical scavenging activity The results showed that 15PHK has the potential ability of antioxidation. The results of antioxidant activities were 43.57%, 0.12 Abs. and 25.56%, respectively. 15PHK was purified with Hiprep DEAE FF 16/10 column by fast protein liquid chromatography (FPLC). Five fractions were abtained and tested their antioxidant activities. The results showed that 5 purified fractions of 15PHK appeared less antioxidant activity.
Antitumor cell experiments were performed to incubate human cervix cancer cell line HeLa, human neuroblastoma cell line SH-SY5Y, human malignant melanoma cell line A375.S2, and human lung adenocarcinoma epithelial cell line A549 with 15HK, 15PHK and 70HK treatment. In MTT assay, A549 cell could be inhibited cell viability obviously by 70HK. A549 cell shrank after 24 hr 70HK incubation by observing. Moreover, lactate dehydrogenase (LDH) was released, nitric oxide (NO) was formed, and reactive oxygen (ROS) was measured corresponding with the more amount of 70HK. In contrast, mitochondrion membrane potential (MMP) was decreased. It showed that A549 cell incubated with 70HK treatment happened to apoptosis, and the more amount of 70HK treated, the more necrosis happened.

摘要 I
Abstract II
目次 III
Figure Contents V
Table Contents VI
壹、研究背景 1
一、 海洋生物之生理活性物質 2
(一) 海洋生物 2
(二) 生理活性物質 2
(三) 生理活性物質分類 3
(四) 海洋生理活性醣類 5
(五) 海洋生理活性胜肽 5
二、 海馬 7
(一) 簡介 7
(二) 海馬種類 7
(三) 營養價值及機制 7
(四) 庫達海馬 8
三、 研究動機及目的 10
貳、研究內容 11
第一章、庫達海馬乙醇萃取物之抗氧化性探討 12
一、 前言 12
二、 材料與方法 13
三、 結果 24
四、 討論 26
五、 結果表格與圖 28
第二章、庫達海馬乙醇萃取物之抗腫瘤性探討 36
一、 前言 36
二、 材料與方法 37
三、 結果 45
四、 討論 47
五、 結果表格與圖 49
參、綜合結論 57
肆、參考文獻 58
附錄一、庫達海馬之圖片 65
謝誌 66

行政院衛生福利部，2016。健康食品概說，行政院衛生福利部，臺北。
李時珍，1988。本草綱目，中醫研究所，北京。pp. 1039。
李志昭，2015。臺灣產棘冠海星之棘刺毒液和海星皂素萃取物之生物活性作用探討，臺灣海洋大學食品科學系博士論文，基隆。
許鐘鋼、陳岳川、劉素華、林金榮，2015。庫達海馬的種苗生產。行政院農業委員會水產試驗所，基隆。
鄭松之，2007。海洋天然產物的分離純化與結構鑑定。化學工業出版社，中國。
賴志行、藍惠玲、吳純衡，2004。烏賊加工副產物之生理活性物質。水試專訊 6：42-44。
Ananthi, S., Raghavendran, H. R. B., Sunil, A. G., Gayathri, V., Ramakrishnan, G.and Vasanthi, H. R. 2010. In vitro antioxidant and in vivo anti-inflammatory potential of crude polysaccharide from Turbinaria ornata (marine brown alga). Food and Chemical Toxicology 48: 187-192.
AOAC (Association of Official Analytical Chemists). 1984. Officail Methods of Analysis, 14th edition. Association of Official Analytical Chemists, Arlington, Virginia, USA.
Artan, M., Li, Y., Karadeniz, F., Lee, S. H., Kim, M. M. and Kim, S. K. 2008. Anti-HIV-1 activity of phloroglucinol derivative, 6,6′-bieckol, from Ecklonia cava. Bioorganic & Medicinal Chemistry 16: 7921-7926.
Athukorala, Y., Lee, K. W., Kim, S. K. and Jeon, Y. J. 2007. Anticoagulant activity of marine green and brown algae collected from Jeju Island in Korea. Bioresource Technology 98: 1711-1716.
Balboa, E. M., Conde, E., Moure, A., Falqué, E. and Domínguez, H. 2013. In vitro antioxidant properties of crude extracts and compounds from brown algae. Food Chemistry, 138, 1764-1785.
Baracca, A., Sqarbi, G., Solaini, G. and Lenaz, G. 2003. Rhodamine 123 as a probe of mitochondrial membrane potential: evaluation of proton flux through F(0) during ATP synthesis. Biochimica et Biophysica Acta 30: 137-146.
Berri, M., Slugocki, C., Olivier, M., Helloin, E., Jacques, I., Salmon, H., Demais, H., Goff, M. L. and Collen, P. N. 2016. Marine-sulfated polysaccharides extract of Ulva armoricana green algae exhibits an antimicrobial activity and stimulates cytokine expression by intestinal epithelial cells. Journal of Applied Phycology 28: 2999-3008.
Bradford, M. M. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry 72: 248-254.
Brown, D. G., Sun, X. M. and Cohen, G. M. 1993. Dexamethasone-induced apoptosis involves cleavage of DNA to large fragments prior internucleosomal fragmentation. Journal of Biological Chemistry 268: 3037-3039.
Burtin, P. 2003. Nutritional value of seaweeds. Electronic Journal of Environmental, Agricultural and Food Chemistry 2: 498-503.
Charoenphun, N., Cheirsilp, B., Sirinupong, N. and Youravong, W. 2013. Calcium-binding peptides derived from tilapia (Oreochomis nilticus) protein hydrolysate. European Food Research and Technology 236: 57-63.
Chen, J. H., Lim, J.-D., Sohn, E. H., Choi, Y. S. and Han, E. T. 2009. Growth-inhibitory effect of a fucoidan from brown seaweed Undaria pinnatifida on Plasmodium parasites. Parasitology Research 104: 245-250.
Chen, J., Zhao, H., Shi, Q., Zhang, D., Cheng, H., Wang, X. and Lee, F. S. 2010. Rapid screening and identification of the antioxidants in Hippocampus japonicus Kaup by HPLC-ESI-TOF/MS and on-line ABTS free radical scavenging assay. Journal of Separation Science 33: 672-677.
Chen, R. Z., Li, Y., Dong, H., Liu, Z. Q., Li, S. Z., Yang, S. M. and Li, X. 2012. Optimization of ultrasonic extraction process of polysaccharides from Ornithogalum caudatum ait and evaluation of its biological activities. Ultrasonics Sonochemistry 19: 1160-1168.
Chen, L. P., Shen, X. R., Chen, G. H., Cao, X. Y. and Yang, J. 2015. Effect of three-spot seahorse petroleum ether extract on lipopolysaccharide induced macrophage RAW264.7 inflammatory cytokine nitric oxide and composition analysis. Journal of Oleo Science 64: 933-942.
Cheung, R. C. F., Ng, T. B. and Wong, J. K. 2015. Marine peptides: Bioactivities and application. Marine Drugs 13: 4006-4043.
Cho, S. S., Lee, H. K., Yu, C. Y., Kim, M. J., Seong, E. S., Ghimire, B. K., Son, E. H., Choung, M. G. and Lim, J. D. 2008. Isolation and characterization of bioactive peptides from Hwangtae (yellowish dried Alaska pollack) protein hydrolysate. Journal of Food Science and Nutrition 13: 196-203.
Decker, T. and Lohmann-Matthes, M. L. 1988. A quick and simple method for the quantitation of lactate dehydrogenase release in measurements of cellular cytotoxicity and tumor necrosis factor (TNF) activity. Journal of Immunological Methods 115: 61-69.
Devillé, C., Gharbi, M., Dandrifosse, G. and Peulen, O. 2007. Study on the effects of laminarin, a polysaccharide from seaweed, on gut characteristics. Journal of the Science of Food and Agriculture 87: 1717-1725.
Dubois, M., Gilles, K. A., Hamilton, J. K., Rebers, P. A. and Smith, F. 1956. Colorimetric method for determination of sugars and related substances. Analytical Chemistry 28: 350-356.
Fernandes, C. E., Vasconcelos, M. A. D. S., de Almeida Ribeiro, M., Sarubbo, L. A., Andrade, S. A. C. and Filho, A. B. d. M. 2014. Nutritional and lipid profiles in marine fish species from Brazil. Food Chemistry 160: 67-71.
Finocchietto, P., Franco, M., Holod, S., Gonzalez, A., Converso, D., Arciuch, V., Serra, M., Poderoso, J. and Carreras, M. 2009. Mitochondrial nitric oxide synthase: A masterpiece of metabolic adaptation, cell growth, transformation, and death. Experimental Biology and Medicine 234: 1020-1028.
Francavilla, M., Trotta, P. and Luque, R. 2010. Phytosterols from Dunaliella tertiolecta and Dunaliella salina: A potentially novel industrial application. Bioresource Technology 101: 4144-4150.
Gamal-Eldeen, A. M., Ahmed, E. F. and Abo-Zeid, M. A. 2009. In vitro cancer chemopreventive properties of polysaccharide extract from the brown alga, Sargassum latifolium. Food and Chemical Toxicology 47: 1378-1384.
Guan, H. S., Y. Q. Han, and Feng, X. M. 2004. Progress in research of marine bioactive peptides. Journal of Ocean University of China 5: 761-766.
Han, H. S., Qiao, Y., Karabiyikoglu, M., Diffard, R. G. and Yenari, M. A. 2002. Influence of mild hypothermia on inducible nitric oxide synthase expression and reactive nitrogen production in experimental stroke and inflammation. Journal of Neuroscience 22: 3021-3928.
Hansen, M. B., Nielson, S. E. and Berg, K. 1989. Re-examination and further development of a precise and rapid dye method for measuring cell growth/cell kill. Journal of Immunological Methods 119: 203-210.
Hansen, R. A., Harris, M. A., Pluhar, G. E., Motta, T., Brevard, S., Ogilvie, G. K., Fettman, M. J. and Allen, K. G. D. 2008. Fish oil decreases matrix metalloproteinases in knee synovia of dogs with inflammatory joint disease. The Journal of Nutritional Biochemistry 19: 101-108.
Himaya, S. W. A., Ryu, B. M., Qian, Z. J. and Kim, S. K. 2012. Paeonol from Hippocampus kuda Bleeler suppressed the neuro-inflammatory responses in vitro via NF-B and MAPK signaling pathway. Toxicology in Vitro 26: 878-887.
Holdt, S. L. and Kraan, S. 2011. Bioactive compounds in seaweed: Functional food applications and legislation. Journal of Applied Phycology 23: 543-597.
Huse, Y. C., Chang, W. H., Chen, C. S., Liao, J. W., Huang, C. J., Lu, F. J., Chia, Y. C., Hsu, H. K., Wu, J. J. and Yang, H. L. 2008. Antioxidant activities of Toona sinensis leaves extracts using different antioxidant models. Food and Chemical Toxicology 46: 105-114.
Jiang, Z., Xu, Y. and Su, Y. 2014. Preparation process of active enzymolysis polypeptides from seahorse bone meal. Food Science and Nutrition 2: 490-499.
Judé, S., Roger, S., Martel, E., Besson, P., Richard, S., Bougnoux, P., Champeroux, P. and Le Guennec, J. Y. 2006. Dietary long-chain omega-3 fatty acids of marine origin: A comparison of their protective effects on coronary heart disease and breast cancers. Progress in Biophysics and Molecular Biology 90: 299-325.
Karasudani, I., Koyama, T., Nakandakari, S. and Aniya, Y. 1996. Purification of anticoagulant factor from the spine venom of the crown-of-thorns starfish, Acanthaster planci. Toxicon 34: 871-879.
Khan, S. B., Kong, C. S., Kim, J. A. and Kim, S. K. 2010. Protective effect of Amphiroa dilatata on ROS induced oxidative damage and MMP expressions in HT1080 cells. Biotechnology and Bioprocess Engineering 15: 191-198.
Kim, S. K., Kim, Y. T., Byun, H. G., Nam, K. S., Joo, D. S. and Shahidi, F. 2001. Isolation and characterization of antioxidative peptides from gelatin hydrolysate of Alaska pollack skin. Journal of Agricultural and Food Chemistry 49: 1984-1989.
Kim, S. K., Nghiep, N. D. and Rajapakse, N. 2006. Therapeutic prospectives of chitin, chitosan and their derivatives. Journal of Chitin Chitosan 11: 1-10.
Kim, M. M., Rajapakse, N. and Kim, S. K. 2009. Anti-inflammatory effect of Ishige okamurae ethanolic extract via inhibition of NF-kappaB transcription factor in RAW 264.7 cells. Phytotherapy Research 23: 628-634.
Kim, S. K. and Wijesekara, I. 2010. Development and biological activities of marine-derived bioactive peptides: A review. Journal of Functional Foods 2: 1-9.
Kim, Y. T., Kim, S. K., Jeon, Y. J. and Park, S. J. 2014. Seahorse-derived peptide suppresses invasive migration of HT-1080 fibrosarcoma cells by competing with intracellular -enolase for plasminogen binding and inhibiting uPA-mediated activation of plasminogen. BMB Reports 47: 691-696.
Kong, C. S., Kim, J. A., Ahn, B. N., Vo, T. S., Yoon, N. Y. and Kim, S. K. 2010. 1-(3′,5′-dihydroxyphenoxy)-7-(2″,4″,6-trihydroxyphenoxy)-2,4,9-trihydroxydibenzo-1,4-dioxin inhibits adipocyte differentiation of 3T3-L1 fibroblasts. Marine Biotechnology 12: 299-307.
Kong, C. S., Kim, J. A., Yoon, N. Y. and Kim, S. K. 2009. Induction of apoptosis by phloroglucinol derivative from Ecklonia cava in MCF-7 human breast cancer cells. Food and Chemical Toxicology 47: 1653-1658.
Korhonen, H. and Pihlanto-Leppala, A. 2003. Food-derived bioactive peptides: Opportunities for designing future foods. Current Pharmaceutical Design 9: 1297-1308.
Laurienzo, P. 2010. Marine polysaccharides in pharmaceutical applications: an overview. Marine Drugs 8: 2435-2465.
Li, Y., Lee, S. H., Le, Q. T., Kim, M. M. and Kim, S. K. 2008. Anti-allergic effects of phlorotannins on histamine release via binding inhibition between IgE and FcεRI. Journal of Agricultural and Food Chemistry 56: 12073-12080.
Lin, Q., Lin, J. and Wang, C. 2009. Biochemical composition of the wild and cultured seahorses, Hippocampus kuda Bleeker and Hippocampus trimaculatus Leach. Aquaculture Research 40: 710-719.
Liu, C., Li, X., Li, Y., Feng, Y., Zhou, S. and Wang, F. 2008. Structural characterisation and antimutagenic activity of a novel polysaccharide isolated from Sepiella maindroni ink. Food Chemistry 110: 807-813.
Lopes, G., Sousa, C., Silva, L. R., Pinto, E., Andrade, P. B., Bernardo, J. Mouga, T. and Valentão, P. 2012. Can phlorotannins purified extracts constitute a novel pharmacological alternative for microbial infections with associated inflammatory conditions? PLoS One 7: e31145.
Mamelona, J., Pelletier, È., Girard,-Lalancette, K., Legault, J., Karboune, S. and Kermasha, S. 2011. Antioxidants in digestive tracts and gonads of green urchin (Strongylocentrotus drobachiensis). Journal of Food composition and Analysis 24: 179-183.
Matsuda, M., Yamori, T., Naitoh, M. and Okutani, K. 2003. Structural revision of sulfated polysaccharide B-1 isolated from a marine pseudomonas species and its cytotoxic activity against human cancer cell lines. Marine Biotechnology 5: 13-19.
McPherson, J. M. and Vincent, A. C. J. 2004. Assessing East African trade in seahorse species as a basis for conservation under international controls. Aquatic Conservation: Marine and Freshwater Ecosystems 14: 521-538.
Migliore, L. and Coppedè, F. 2009. Environmental-induced oxidative stress in neurodegenerative disorders and aging. Mutation Research/Genetic Toxicology and Environmental Mutagenesis 674: 73-84.
Moon, J. H., Eo, S. K., Lee, J. H. and Park, S. Y. 2015. Quercetin-induced autophagy flux enhances TRAIL-mediated tumor cell death. Oncology Reports 34: 375-381.
Nagata, H., Nagase, K., Kawane, N., Mukae, N. and Fukuyama, H. 2003. Degradation of chromosomal DNA during apoptosis. Cell Death and Differentiaition 10: 108-116.
Nagayama, K., Iwamura, Y., Shibata, T., Hirayama, I. and Nakamura, T. 2002. Bactericidal activity of phlorotannins from the brown alga Ecklonia kurome. Journal of Antimicrobial Chemotherapy 50: 889-893.
Oktay, M., Gülcin, İ. and Küfrevioğlu, Ö. İ. 2003. Determination of in vitro antioxidant activity of fennel (Foeniculum vulgare) seed extracts. Lebensmittel- Wissenschaft Und-Technologie Food Science and Technology 36: 263-271.
Oyaizu, M. 1986. Studies on product of browning reaction prepared from glucose amine. Japanese Journal of Nutrition 44: 307-315.
Pangestuti, R. and Kim, S. K. 2011. Neuroprotective effects of marine algae. Marine Drugs 9: 803-818.
Pangestuti, R. and Kim, S. K. 2015. Peptide-derived from seahorse exerts a protective effect against cholinergic neuronal death in in vitro model of Alzheimer’s disease. Procedia Chemistry 14: 343-352.
Pangestuti, R., Ryu, B., Himaya, S. W. A. and Kim, S. K. 2013. Optimization of hydrolysis conditions, isolation, and identification of neuroprotective peptides derived from seahorse Hippocampus trimaculatus. Amino Acids 45: 369-381.
Paulo, A. S. M. 2004. Use of sulfated fucans as anticoagulant and antithrombotic agents: Future perspectives. Current Pharmaceutical Design 10: 967-981.
Pihlanto-Leppala, A. 2001. Bioactive peptides derived from bovine proteins: Opioid and ACE-inhibitory peptides. Trends in Food Science and Technology 11: 347-356.
Proskuryakov, S. Y., Konoplyannikov, A. G., Gabai, V. L. and Konoplyannikov, G. 2003. Necrosis: A specific form of programmed cell death? Experimental Cell Research 283: 1-16.
Qian, Z. J., Kang, K. H. and Kim, S. K. 2012. Isolation and antioxidant activity evaluation of two new phthalate derivatives from seahorse, Hippocampus kuda bleeler. Biotechnology and Bioprocess Engineering 17: 1031-1040.
Qian, Z. J., Ryu, B. M., Kim, M. M. and Kim, S. K. 2008. Free radical and reactive oxygen species scavenging activities of the extracts from seahorse, Hippocampus kuda bleeler. Biotechnology and Bioprocess Engineering 13: 705-715.
Re, R., Pellegrini, R., Proteggente, N., Pannala, A., Yang, A. and Rice-Evans, C. 1999. Antioxidant activity applying an improved ABTS radical cation decolorization assay. Free Radical Biology and Medicine 26: 1231-1237.
Rosenkranz, A. R., Schmaldienst, S., Stuhlmeier, K. M., Chen, W., Knapp, W. and Zlabinger, G. J. 1992. A microplate assay for the detection of oxidative products using 2’,7’-dichlorofluorescin-diacetate. Journal of Immunological Methods 156: 39-45.
Ryu, B., Qian, Z. J. and Kim, S. K. 2010a. SHP-1, a novel peptide isolated from seahorse inhibits collagen release through the suppression of collagenases 1 and 3, nitric oxide products regulated by NF-κB/p38 kinase. Peptides 31: 79-87.
Ryu, B., Qian, Z. J. and Kim, S. K. 2010b. Purification of a peptide from seahorse, that inhibits TPA-induced MMP, iNOS and COX-2 expression through MAPK and NF-κB activation, and induces human osteoblastic and chondrocytic differentiation. Chemico-Biological Interactions 184: 413-422.
Sanaye, S. V., Pise, N. M., Pawar, A. P., Parab, P. P., Sreepada, R. A., Pawar, H. B. and Revankar, A. D. 2014. Evaluation of antioxidant activities in captive-bred cultured yellow seahorse, Hippocampus kuda (Bleeker, 1852). Aquaculture 434: 100-107.
Siriwardhana, N., Lee, K. W., Jeon, Y. J., Kim, S. H. and Haw, J. W. 2003. Antioxidant activity of Hizikia fusiformis on reactive oxygen species scavenging and lipid peroxidation inhibition. Food Science and Technology International 9: 339-346.
Stix, G. 2005. A toxin against pain. Scientific American 292: 70-75.
Su, Y. and Xu, Y. 2015. Study on the extraction and purification of glycoprotein from the yellow seahorse, Hippocampus kuda Bleeker. Food Science and Nutrition 3: 302-312.
Sun, C., Wang, J. W., Fang, L., Gao, X. D., & Tan, R. X. 2004. Free radical scavenging and antioxidant activities of EPS2, an exopolysaccharide produced by a marine filamentous fungus Keissleriella sp. YS 4108. Life Sciences 75: 1063-1073.
Vermes, I., Haanen, C., Steffens-Nakken, H. and Reutellingsperger, C. 1995. A novel assay for apoptosis Flow cytometric detection of phosphatidylserine expression on early apoptotic cells using fluorescein labelled Annexin V. Journal of Immunological Methods 184: 39-51.
Wang, S., Cheng, Y., Wang, F., Sun, L., Liu, C., Chen, G., Li, Y. Ward, S. G. and Qu, X. 2008. Inhibition activity of sulfated polysaccharide of Sepiella maindroni ink on matrix metalloproteinase (MMP)-2. Biomedicine & Pharmacotherapy 62: 297-302.
Wijesinghe, W. A. J. P. and Jeon, Y. J. 2012. Enzyme-assistant extraction (EAE) of bioacitive components: A useful approach for recovery of industrially important metabolites from seaweeds: A review. Fitoterapia 83: 6-12.
Xu, D. H., Mei, X. T. and Li, B. J. 2003. The pharmacological effects of Hippocampus capsule on enhancing sexual functions of rats. Chinese Medicine 26: 807-808.
Yang, Y. J., Kim, S. K. and Park, S. J. 2012. An anti-inflammatory peptide isolated from seahorse Hippocampus kuda Bleeler inhbits the invasive potential of MG-63 osteosarcoma cells. Fisheries and Aquatic Sciences 15: 29-36.
Yuan, X. H., Chen, C. B. and Chen, Z. 2011. Study on optimization of enzymatic and the antioxidant activity to DPPH of liquid protein of three-spot seahorse. Science and Technology of Food Industry 32: 291-294.
Zhang, N., Xu, B., Mou, C., Yang, W., Wei, J., Lu, L., Zhu, J. Du, J., Wu, X., Ye, L., Fu, Z., Lu, Y., Lin, J., Sun, Z., Su, J., Dong, M. and Xu, A. 1997. Molecular profile of the unique species of traditional Chinese medicine, Chinese seahorse (Hippocampus kuda Bleeker). FEBS Letters 550: 124-134.
Zou, Y., Qian, Z. J., Li, Y., Kim, M. M., Lee, S. H. and Kim, S. K. 2008. Antioxidant effects of phlorotannins isolated from Ishige okamurae in free radical mediated oxidative systems. Journal of Agricultural and Food Chemistry 56: 7001-7009.
Zeng, M. Y., Cui, H. Y. and Li, B. F. 2005. Research progress on the marine bioactive peptides. Chinese Journal of Marine Drugs 24: 46-51.