International Journal of Biological Macromolecules 67 (2014) 303–311
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Water soluble sulfated-fucans with immune-enhancing properties from Ecklonia cava Rong-An Cao a,b , YongJin Lee c,∗ , SangGuan You b,∗ a
College of Food Science, Heilongjiang Bayi Agricultural University, Daqing 163319, China Department of Marine Food Science and Technology, Gangneung-Wonju National University, 120 Gangneung Daehangno, Gangneung, Gangwon 210-702, South Korea c Chuncheon Bioindustry Foundation, Chuncheon, Gangwon 200-957, South Korea b
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Article history: Received 5 December 2013 Received in revised form 4 March 2014 Accepted 11 March 2014 Available online 21 March 2014 Keywords: Ecklonia cava Sulfate-fucans Immune-enhancing NF-B MAPK
a b s t r a c t Water-soluble sulfated fucans isolated from Ecklonia cava were fractionated using an anion-exchange chromatography to investigate their molecular characteristics and immunomodulating activities. The crude fucoidan extract and puriﬁed fractions (EF1 , EF2 , and EF3 ) consisted mostly of different ratios of neutral sugars, proteins, sulfates, uronic acids, and their monosaccharide compositions were also signiﬁcantly different. The backbone of the most immunoenhancing fraction, EF2 , was mainly linked by (1→3)-linked fucopyranosyl and (1→4)-linked mannopyranosyl residues with sulfates at C-4 of fucopyranosyl units. The molecular weights of the crude fucoidan extract and puriﬁed fractions ranged from 8.3 × 103 to 442.6 × 103 g/mol. The crude extract, EF1 and EF2 stimulated RAW264.7 cells to produce considerable amounts of nitric oxide and cytokines. The treatment of cells with the sulfated fucans induced the degradation of I-B and the phosphorylation of MAPK in RAW264.7 cells, implying that they might stimulate RAW264.7 cells through the activation of NF-B and MAPK pathways. © 2014 Elsevier B.V. All rights reserved.
1. Introduction Ecklonia cava (E. cava) is a perennial brown alga belonging to the family Laminariaceae, widely distributed along the coasts of South Korea and Japan. E. cava is a rich source of natural compounds having antioxidant, anticancer and anticoagulant activities . Among these components, phlorotannins, a class of compounds with polymerized phloroglucinol units, are responsible for the strong antioxidant activity of E. cava. In addition, an increasing amount of evidence of in vitro and in vivo model studies has demonstrated that the phlorotannins exhibit anti-HIV , hypnotic effects , anti-melanogenesis , matrix metalloproteinase inhibitory , anticancer , anti-inﬂammatory , anti-allergic , antimetastatic , ␣-glucosidase and ␣-amylase inhibitor , and antidiabetic  activities. The yield of the phlorotannins for E. cava is from 0.35% to 9.13% [5,9,11] depending on the extraction methods, therefore, around 90% residue is simply wasted and/or used as
∗ Corresponding authors at: Gangneung Daehangno, Gangneung, Gangwon, 210702, Republic of Korea. Tel.: +82 33 640 2853/+82 33 258 6161; fax: +82 33 640 2340/+82 33 258 6170. E-mail addresses: [email protected]
(Y. Lee), [email protected]
(S. You). http://dx.doi.org/10.1016/j.ijbiomac.2014.03.019 0141-8130/© 2014 Elsevier B.V. All rights reserved.
a fertilizer or an animal feed. The residue also contains the bioactive soluble and insoluble components such as fucoidans, alginates, and dietary ﬁbers commonly found in brown algae. Fucoidans from brown algae have been extensively studied because of their numerous bioactivities including antioxidation , antivirus , anticoagulation  and anticancer . The biological activities of fucoidans have been reported to be closely related to their sulfate content and molecular weight . Oversulfated fucoidans led to higher stimulation of the glutamic–plasminogen activation in comparison with native fucoidan . It was also found that oversulfated fucoidan possessed higher amyloglucosidase inhibition activity than native fucoidans, and thus could effectively inhibit starch digestibility . On the other hand, partially desulfated fucoidans with sulfate contents less than 20% showed drastic decreases in both anticoagulant and anticancer activities . It was reported that the molecular weight of fucoidan polymers was related to their anticoagulant and anticancer activities [20,21]. Lee et al. found that fucoidan polymers might have optimum ranges of molecular weights for the potent anticoagulant or anticancer activities . Several researches have been performed on fucoidans from E. cava, revealing that they have anti-inﬂammatory , anticoagulative  and antiproliferative activities . However, relatively limited information is available on the molecular structures of fucoidan from E. cava. Basic understanding of both the primary and
R.-A. Cao et al. / International Journal of Biological Macromolecules 67 (2014) 303–311
secondary structures of the polysaccharides may lead to successful interpretation of their bioactivities. In this study, fucoidan was extracted from E. cava using a dilute acidic solution and fractionated by an anion-exchange chromatography. The purpose of this study was to determine the chemical and molecular characteristics of the fractionated fucoidans and their immunomodulatory activities. 2. Materials and methods 2.1. Materials The brown seaweed E. cava was collected from the coast of Jeju Island, South Korea. The seaweed was thoroughly washed with tap water and air-dried at 60 ◦ C. The dried raw material was milled using a blender, sieved (