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1.
Mar Drugs ; 21(9)2023 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-37755099

RESUMO

This work reports the detailed structure of fucoidan from Sargassum miticum (2SmF2) and its ability to potentiate the inhibitory effect of glycolysis inhibitor 2-deoxy-d-glucose (2-DG). 2SmF2 was shown to be sulfated and acetylated galactofucan containing a main chain of alternating residues of 1,3- and 1,4-linked α-l-fucopyranose, fucose fragments with monotonous 1,3- and 1,4-type linkages (DP up to 3), α-d-Gal-(1→3)-α-L-Fuc disaccharides, and 1,3,4- and 1,2,4-linked fucose branching points. The sulfate groups were found at positions 2 and 4 of fucose and galactose residues. 2SmF2 (up to 800 µg/mL) and 2-DG (up to 8 mM) were not cytotoxic against MDA-MB-231 and SK-MEL-28 as determined by MTS assay. In the soft agar-based model of cancer cell colony formation, fucoidan exhibited weak inhibitory activity at the concentration of 400 µg/mL. However, in combination with low non-cytotoxic concentrations of 2-DG (0.5 or 2 mM), 2SmF2 could effectively inhibit the colony formation of SK-MEL-28 and MDA-MB-231 cells and decreased the number of colonies by more than 50% compared to control at the concentration of 200 µg/mL. Our findings reveal the metabolically oriented effect of fucoidan in combination with a glycolysis inhibitor that may be beneficial for a therapy for aggressive cancers.


Assuntos
Melanoma , Sargassum , Humanos , Fucose , Polissacarídeos/farmacologia
2.
Int J Biol Macromol ; 225: 648-657, 2023 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-36395953

RESUMO

Six fucoidan fractions were isolated from the brown alga Alaria angusta. Structures of enzymatic hydrolysis products of the fraction 1AaF2 (Fuc:Gal ~ 1:1; 33 % of sulfates) by fucanase from Wenyingzhuangia fucanilytica were studied by chemical and instrumental (NMR spectroscopy and mass-spectrometry) methods. It was shown that 1AaF2 consisted of two structurally different fucoidans: a sulfated 1,3;1,4-α-L-fucan and an enzyme-resistant sulfated and acetylated complex fucogalactan (Fuc:Gal ~ 1:2; 19 % of sulfates) 1AaF2_HMP containing extended 1,3-linked fucose and 1,3/1,4-linked galactose fragments (up to 5 residues). The fractions 1AaF2 and 1AaF2_HMP were a non-cytotoxic, possessed dose-dependent chemopreventive effect on EGF-induced neoplastic cell transformation of mouse normal epidermal JB6 Cl41 cells and inhibited the colony formation of human melanoma SK-MEL-28 cells.


Assuntos
Antineoplásicos , Melanoma , Phaeophyceae , Animais , Camundongos , Humanos , Antineoplásicos/farmacologia , Antineoplásicos/química , Polissacarídeos/farmacologia , Polissacarídeos/química , Phaeophyceae/química , Sulfatos/química
3.
Int J Biol Macromol ; 206: 614-620, 2022 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-35219778

RESUMO

The fucoidan SdeF was isolated from brown alga Saccharina dentigera. The structure of the obtained polysaccharide was studied by chemical methods, NMR spectroscopy of the fully and partially desulfated derivatives, and mass spectrometry of the fucoidan fragments, labeled with 18O. The SdeF was shown to be sulfated (40%) 1,3-linked α-L-fucan, with branches at C2. The sulfate groups were found at positions C2 and C4. Derivatives SdeFDS and SdeFPL were obtained by solvolytic desulfation and autohydrolysis of SdeF, respectively. According to 13C NMR data, SdeFDS is 1,3-linked α-L-fucan, while SdeFPL is 4-sulfated 1,3-linked α-L-fucan. Native fucoidan SdeF was shown to be a non-toxic anticancer substance in the model of human malignant melanoma RPMI-7951, colorectal adenocarcinoma HCT-116, and small intestine adenocarcinoma HuTu 80 cells. The partial desulfation of SdeF at C2 and/or the reduction of its Mw, from 229 to 28 kDa, decreased the anticancer activity; complete removal of the sulfated groups and/or Mw reduction to 4.7 kDa further reduced the effect of this polysaccharide.


Assuntos
Adenocarcinoma , Antineoplásicos , Phaeophyceae , Antineoplásicos/química , Antineoplásicos/farmacologia , Humanos , Phaeophyceae/química , Polissacarídeos/química , Polissacarídeos/farmacologia , Sulfatos
4.
Int J Biol Macromol ; 183: 1427-1435, 2021 Jul 31.
Artigo em Inglês | MEDLINE | ID: mdl-34023368

RESUMO

The aim of this study was to establish the fine structure of fucoidan from Sargassum oligocystum and to study the radiosensitizing effect of fucoidans from three algae of genus Sargassum (S. oligocystum, S. duplicatum, and S. feldmannii) with different structures. The fucoidan SoF2 from S. oligocystum was sulfated (32%) galactofucan (Fuc:Gal = 2:1), with a Mw of 183 kDa (Mw/Mn = 2.0). Its supposed structure was found to be predominantly 1,3-linked fucose as the main chain, with branching points at C2 and C4. The branches could be single galactose and/or fucose short chains with terminal galactose residues. Sulfate groups were found at positions C3, C2, and/or C4 of fucose residues and at C2 and/or C4 of galactose residues. The radiosensitizing effect of galactofucans from S. oligocystum, S. duplicatum, and S. feldmannii against human melanoma SK-MEL-28, colon HT-29, and breast MDA-MB-231 cancer cells was investigated. The influence of all investigated polysaccharides treatments with/without X-ray radiation on colony formation of human melanoma cells SK-MEL-28 was weak. Fucoidan from S. feldmannii has been shown to be the most promising radiosensitizing compound against human colon HT-29 and breast MDA-MB-231 cancer cells.


Assuntos
Polissacarídeos/química , Sargassum/química , Linhagem Celular Tumoral , Células HT29 , Humanos , Radiossensibilizantes/química
5.
Carbohydr Polym ; 250: 116921, 2020 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-33049835

RESUMO

Polysaccharide fractions of alginate, laminarans and fucoidans were obtained from the brown alga Tauya basicrassa. Yields of alginate and laminarans were large (19.7 % and 5.62 %, respectively), whereas the content of fucoidans (0.52 %) was not significant. Alginate and laminarans had typical structures for those substances. Fucoidans were low- and medium-sulfated heterogeneous polysaccharides. The fucoidan fraction 1TbF1 was sulfated fucogalactan containing a backbone from 1,6-linked residues of ß-d-galactopyranose with branches at C3 and C4, terminal fucose and galactose residues and fragments from 1,3-; 1,4-; and 1,2-fucose residues. Sulfate groups were found at positions 2 and 4 of fucose, and positions 2, 3 and 4 of galactose residues. Laminaran 2TbL was subjected to a sulfation to obtain the derivative 2TbLS with partial sulfation (46 %) at C2, C4 and C6. It was shown that 2TbL and 2TbLS inhibited colony formation of sensitize-tested colon cancer cells HT-29 and HCT-116 to X-ray radiation.


Assuntos
Neoplasias do Colo/tratamento farmacológico , Glucanos/farmacologia , Phaeophyceae/química , Polissacarídeos/química , Radiossensibilizantes/farmacologia , Sulfatos/química , Antineoplásicos , Neoplasias do Colo/patologia , Glucanos/química , Humanos , Polissacarídeos/farmacologia , Radiossensibilizantes/química , Células Tumorais Cultivadas , Raios X
6.
Carbohydr Polym ; 229: 115518, 2020 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-31826457

RESUMO

The sulfated polysaccharide from sterile alga Mastocarpus pacificus was investigated. Partial reductive hydrolysis and NMR spectroscopy showed that the extracted polysaccharides were only carrageenans. According to FT-IR- and NMR spectroscopy this polysaccharide was a hybrid kappa/iota-carrageenan with a predominance of kappa-type units. According to MALDI-TOFMS, oligosaccharide fragments obtained by mild acid hydrolysis had a polymerization degree of 1-9, while chains built up of galactose residues were up to 3. Tandem ESI mass spectrometry together with innovative 18O-labelling method showed that the polymer chain of the carrageenan included kappa-carrabiose, kappa-carratetraose, iota-carrabiose, hybrid kappa/iota oligosaccharide units and contained minor insertions of mu-carrageenan (the precursor of kappa-carrageenan). Parallel artificial membrane permeability assay shown that the studied carrageenan inhibited bile salts permeation through an artificial membrane imitating the gastrointestinal barrier by 50 % on average compared to negative control independent of incubation time. However, its action was less pronounced than the hindering ability of cholestyramine.

7.
Carbohydr Polym ; 221: 157-165, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31227154

RESUMO

The sulfated α-l-fucans ScF and LlF were obtained from brown algae of the Laminariaceae family (Saccharina cichorioides and Laminaria longipes). According to spectroscopy NMR, the LlF fucan predominantly contained the →3)-α-l-Fucp-(2SO3-)-(1→4)-α-l-Fucp-(1→2)-α-l-Fucp-(4SO3-)-(1→ repeating units, with small amounts of disaccharide 1,4-linked fragments and 3-sulfated fucose residues. Mass spectrometric analysis revealed the presence of the following fragments in the fucan structure: α-l-Fucp-(2SO3-)-(1→4)-α-l-Fucp-(2SO3-)-(1→3)-α-l-Fucp-(4SO3-); α-l-Fucp-(2,4SO3-)-(1→3)-α-l-Fucp-(1→3)-α-l-Fucp-(4SO3-); α-l-Fucp-(2SO3-)-(1→2)-α-l-Fucp; α-l-Fucp-(2SO3-)-(1→2)-α-l-Fucp-(4SO3-); α-l-Fucp-(2SO3-)-(1→3)-α-l-Fucp; α-l-Fucp-(2,4SO3-)-(1→3)-α-l-Fucp; α-l-Fucp-(4SO3-)-(1→4)-α-l-Fucp; and α-l-Fucp-(4SO3-)-(1→4)-α-l-Fucp-(2SO3-). Both ScF and LlF fucoidans inhibited colony formation and growth of melanoma and colon cancer cells and sensitize-tested cancer cells to X-ray radiation to a comparable degree.


Assuntos
Antineoplásicos/farmacologia , Laminaria/química , Polissacarídeos/farmacologia , Radiossensibilizantes/farmacologia , Antineoplásicos/química , Sequência de Carboidratos , Linhagem Celular Tumoral , Humanos , Polissacarídeos/química , Radiossensibilizantes/química
8.
Int J Biol Macromol ; 124: 220-228, 2019 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-30496854

RESUMO

Fucoidans are valuable biologically active polysaccharides of brown algae. The aim of this study was to investigate the structure of fucoidan from Sargassum feldmannii and the anticancer effects of native and modified polysaccharides from S. feldmannii and S. duplicatum. The structure of sulfated (25.3%) galactofucan SfF2 (Fuc/Gal = 72/28 mol%) from S. feldmannii was investigated by NMR spectroscopy of desulfated derivative and mass spectrometry of fucoidan fragments labelled with 18O. SfF2 was shown to contain the main chain from 1,3-linked α-l-fucopyranose and ß-d-galactopyranose residues with fucose branches at C4 and C6 of galactose residues and C2 of fucose residues. The following fragments were also identified in SfF2: Fuc-(1,4)-Fuc, Gal-(1,3)-Gal, and Gal-(1,4)-Gal. The sulfate groups occupied positions C2, C3, and C4 of fucose residues and C2, C3, C4, and C6 of galactose residues. The galactofucans from S. feldmannii, S. duplicatum, and their derivatives exhibited no cytotoxicity in vitro. The native and deacetylated fucoidans (200 µg/mL) inhibited colony formation of human colon cancer cells (DLD-1, HT-29, and HCT-116). Both desulfated fucoidans possessed weak anticancer activity.


Assuntos
Antineoplásicos/química , Polissacarídeos/química , Sargassum/química , Antineoplásicos/isolamento & purificação , Antineoplásicos/farmacologia , Neoplasias do Colo/tratamento farmacológico , Galactose/química , Células HT29 , Humanos , Polissacarídeos/isolamento & purificação , Polissacarídeos/farmacologia , Sulfatos/química
9.
Int J Biol Macromol ; 117: 1101-1109, 2018 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-29885396

RESUMO

In the present study, three sulfated polysaccharides, two fractions of fucosylated chondroitin sulfates, and one sulfated fucan were isolated from the body wall of the Vietnamese sea cucumber Stichopus variegatus. The structure of the sulfated fucan fraction SvF3 from S. variegatus was investigated for the first time. According to NMR spectroscopy data, the sulfated fucan SvF3 contained 1,2- and 1,3-linked α-l-fucopyranose residues. Sulfate groups were found at the 2 and/or 4 positions. The structural analysis of fucoidan was assisted by tandem mass spectrometry; the recently-developed technique of autohydrolysis in heavy­oxygen water for the obtaining of selectively labeled fucoidan fragments was applied. The labeling (+2 Da mass shift at the reducing end) allowed us to assign MS/MS data unambiguously, and thus to confirm the NMR data and revealed minor sulfation at position 3. It was shown that the sulfated fucan SvF3 was not cytotoxic to human breast cancer T-47D and MDA-MB-231 cell lines, and it inhibited colony formation of those cells in vitro. SvF3 also possessed slight activity against migration of MDA-MB-231 cells in vitro.


Assuntos
Antineoplásicos/química , Antineoplásicos/farmacologia , Polissacarídeos/química , Polissacarídeos/farmacologia , Animais , Antineoplásicos/isolamento & purificação , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Hidrólise , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Peso Molecular , Polissacarídeos/isolamento & purificação , Pepinos-do-Mar/química , Relação Estrutura-Atividade , Temperatura
10.
Carbohydr Polym ; 184: 260-268, 2018 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-29352918

RESUMO

The sulfated and acetylated fucoidan fraction, containing fucose, galactose, mannose, glucose and uronic acid residues, was isolated from the brown alga Padina boryana. The structure of galactofucan part was studied after different modifications by NMR spectroscopy and mass spectrometry. It was shown that galactofucan contained the main chain of alternating 1,4-linked α-l-fucopyranose and 1,3-linked ß-d-Galactopyranose. Single fucose residues were found as branches at C4 of galactose residues. Also, fucoidan contained 1,3- or 1,4-linked Fuc-Fuc and Gal-Gal fragments. The sulfate groups occupied positions C2, C3 and C4 of both fucose and galactose residues, which was shown by tandem mass spectrometry of fragments, labeled with heavy-oxygen. The anticancer effect of native and modified fucoidan fractions was studied in vitro on the colorectal carcinoma cells DLD-1 and HCT-116. All fucoidans had no cytotoxicity under 400 µg/mL and inhibited colony formation of cancer cells at concentration of 200 µg/mL.


Assuntos
Polissacarídeos/química , Animais , Antineoplásicos/química , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Glucanos/química , Humanos , Camundongos , Phaeophyceae/química , Polissacarídeos/farmacologia , Sargassum/química
11.
Carbohydr Res ; 455: 10-13, 2018 Jan 02.
Artigo em Inglês | MEDLINE | ID: mdl-29128704

RESUMO

A procedure for the partial depolymerization of sulfated fucans and selective labeling with 18O was developed. A tandem electrospray ionization mass spectrometry (ESI MS/MS) was applied for the direct analysis of mixtures of structurally-different oligosaccharides, derived from the fucoidans of known structure. The presence of label allowed unambiguous distinguishing between the fragment ions of 0,2X0-type at m/z 287 and 2,4A-type at m/z 285, since 18O at the reducing end gave +2 mass shifting. Thus, ESI MS/MS was able to detect (1,2)-type of linkage in disaccharides from the fucoidan of brown alga S. cichorioides for the first time. It was also discovered that 2,4A-type fragments in 4-linked disaccharides that were incorrectly assigned to 0,2X-type previously, suggested, probably, substitution at C-4 in mono- and disaccharide fragments, derived from the fucoidan of the brown alga F. evanescens.


Assuntos
Polissacarídeos/química , Espectrometria de Massas em Tandem/métodos , Dissacarídeos/química , Oligossacarídeos/química , Isótopos de Oxigênio/química , Espectrometria de Massas por Ionização por Electrospray/métodos
12.
Carbohydr Polym ; 175: 547-556, 2017 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-28917899

RESUMO

The laminaran SdL and fucoidan SdF were isolated from brown algae Sargassum duplicatum. SdL was 1,3;1,6-ß-d-glucan (1,3:1,6=6:1) with a main chain, represented by 1,3-linked glucose residues, due to NMR spectroscopy data. Single glucose residues could form branches at C6. Unusual structure of fucoidan SdF was studied by chemical and enzymatic methods, NMR spectroscopy of desulfated and deacetylated polysaccharide and mass spectrometry of fucoidan fragments labeled with 18O. Fucoidan was sulfated (31.7%) and acetylated galactofucan (Fuc:Gal∼1:1) with a main chain of 1,4-linked alternating α-l-fucose and ß-d-galactose residues. Side chains were represented by extensive (DP≥5) 1,3-linked 2,4-disulfated α-l-fucose residues with branching points at C2. Fucose residues in the main chain were sulfated at C2 and less at C3, while galactose residues were sulfated at C2, C3, and less at C4, C6. The fucoidan SdF was effective against colony formation of colon cancer cells in vitro.


Assuntos
Antineoplásicos/química , Glucanos/química , Polissacarídeos/química , Sargassum/química , Antineoplásicos/farmacologia , Glucanos/farmacologia , Células HT29 , Humanos , Isótopos de Oxigênio , Polissacarídeos/farmacologia
13.
Protein Pept Lett ; 24(10): 974-981, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28741465

RESUMO

BACKGROUND: Tick-borne encephalitis poses a serious public health threat in the endemic regions. The disease treatment is restricted to symptomatic therapy, so great expectations are in the development of the prophylactic and therapeutic vaccines. The domain III of E protein of the tickborne encephalitis virus is the main antigenic domain which includes virus-specific epitopes recognized by neutralizing antibodies. OBJECTIVES: The main objective of this study was to design, express, isolate and characterize the chimeric protein based on the fusion of domain III of E protein of the tick-borne encephalitis virus and bacterial porin OmpF from Yersinia pseudotuberculosis. METHODS: The chimeric gene was obtained by the PCR based fusion method from two fragments containing overlapping linker sequences. Resulting plasmids were transformed into BL21(DE3) pLysS electrocompetent cells for subsequent heterologous protein expression. All recombinant proteins were purified using immobilized metal affinity chromatography under denaturing conditions. The identity of the chimeric protein was confirmed by MALDI-TOF mass spectrometry and immunoblot analysis. The content of antibodies against the EIII protein was estimated in mice blood serum by ELISA. RESULTS: The bacterial partner protein was used for decreasing toxicity and increasing immunogenicity of antigen. The chimeric protein was successfully expressed by the Escherichia coli cells. The purified protein was recognized with immunoblots by anti-E protein of tick-borne encephalitis virus monoclonal antibodies. Furthermore, the protein was able to elicit antibody response against domain III of E protein in immunized mice. CONCLUSION: The newly obtained chimeric antigen could be valuable for the development of the preventing tick-borne encephalitis subunit vaccines.


Assuntos
Vírus da Encefalite Transmitidos por Carrapatos/química , Porinas/química , Proteínas do Envelope Viral/química , Yersinia pseudotuberculosis/química , Animais , Anticorpos Antivirais/sangue , Feminino , Flavivirus/química , Camundongos Endogâmicos BALB C , Porinas/imunologia , Domínios Proteicos , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologia , Vacinas de Subunidades Antigênicas/imunologia , Proteínas do Envelope Viral/genética , Proteínas do Envelope Viral/imunologia , Vacinas Virais/imunologia
14.
Carbohydr Polym ; 157: 1503-1510, 2017 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-27987862

RESUMO

A fucoidan ScF from brown alga Saccharina cichorioides was extracted, purified and partially depolymerized by autohydrolysis at 37°C for 24, 48 and 72h. Supernatant (SN) and pellet (PL) fractions were obtained by ethanol precipitation of each sample. Unlike spectral data of ScF, NMR of PL derivatives clearly suggested the structure: 1,3-linked α-l-Fucp-4-OSO3- repeating unit. Molecular weights (MWs) of PL fractions were 30, 26 and 18kDa for 24, 48 and 72h of autohydrolyis, respectively. MALDI-TOFMS, size-exclusion HPLC and carbohydrate polyacrylamide-gel electrophoresis (C-PAGE) indicated a similarity of SN mixtures. They consisted mainly of a polysaccharide part (MW 6kDa, C-PAGE data) with a structure similar to PL components (NMR data) and monosaccharides α-l-Fucp-4-OSO3-, α-l-Fucp-2,4-di-OSO3-. PL fractions exhibited almost identical antiproliferative activity in vitro as native fucoidan, while an SN sample for 72h of autohydrolysis was slightly more active against colony formation of colorectal carcinoma cells HT-29.


Assuntos
Phaeophyceae/química , Polissacarídeos/química , Polissacarídeos/farmacologia , Proliferação de Células/efeitos dos fármacos , Células HT29 , Humanos
15.
Carbohydr Polym ; 153: 258-265, 2016 Nov 20.
Artigo em Inglês | MEDLINE | ID: mdl-27561495

RESUMO

Laminaran and three fucoidan fractions were obtained from the brown alga Alaria marginata. Alaria angusta, studied earlier by us, has the same polysaccharide composition. Galactofucan AmF3 from A. marginata has a main chain of →3)-α-l-Fucp-(2,4-SO3(-))-(1→residues, similar to galactofucan from A. angusta. However, the structure of the branches in fucoidan AmF3 can differ from those in the fucoidan from A. angusta. The following fragments were identified in AmF3: HexA-(1→2)-Fuc, HexA-(1→2)-Gal, Gal-(1→4)-HexA, Fuc-(1→2)-Gal-6-SO3(-), Fuc-4-SO3(-)-(1→6)-Gal, Gal-(1→2)-Gal-2-SO3(-), Gal-4-SO3(-)-(1 →6)-Gal, Gal-4-SO3(-)-(1→3)-Fuc-(1→3)-Fuc, Fuc-4-SO3(-)-(1→6)-Gal-(1→4)-Gal, Gal-(1→4)-Gal-(1→3)-Fuc, Gal-2-SO3(-)-(1→4)-Gal-(1→4)-Gal, Gal-(1→4)-Gal-6-SO3(-)-(1→2)-Gal. Chains of galactose residues (DP up to 9) were found in AmF3 fucoidan. The laminarans, galactofucans and their derivatives from both algae exhibited no cytotoxicity in vitro. Polysaccharides from A. angusta were more effective against colony formation of HT-29 cells, while those from A. marginata had a greater effect on T-47D cells. Sulfated and desulfated fucoidans possessed weak antitumor activity using SK-MEL-28 cells.


Assuntos
Glucanos/química , Glucanos/farmacologia , Phaeophyceae/química , Polissacarídeos/química , Polissacarídeos/farmacologia , Antineoplásicos/química , Antineoplásicos/isolamento & purificação , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Glucanos/isolamento & purificação , Células HT29 , Humanos , Polissacarídeos/isolamento & purificação , Relação Estrutura-Atividade , Ensaio Tumoral de Célula-Tronco
16.
Carbohydr Polym ; 151: 523-534, 2016 Oct 20.
Artigo em Inglês | MEDLINE | ID: mdl-27474596

RESUMO

Gelling sulfated polysaccharide from the cystocarpic plants of Ahnfeltiopsis flabelliformis was studied. According to FT-IR and NMR spectroscopy data, the polysaccharide was found to be iota/kappa-carrageenan with iota- and kappa-type units in a 2:1 ratio containing beta-carrageenan units and minor amounts of nu- and mu-carrageenans. The HPLC and ESI MS/MS data of enzymatic hydrolysis products revealed that the main components of the polymer chain are iota-carrabiose, iota-carratetraose and hybrid tetra- and hexasaccharides consisting of kappa- and iota-units. Xylose was a substituent of a hydroxyl group at C-6 of 1,3-linked ß-d-galactose in the total polysaccharides. It was shown that the ability of carrageenans to increase the synthesis of cytokines depended on their molecular weight. The polysaccharide induced the synthesis of the anti-inflammatory cytokine IL-10, whereas oligosaccharides increased the synthesis of both pro- and anti-inflammatory cytokines at high concentrations.


Assuntos
Carragenina , Interleucina-10/biossíntese , Rodófitas , Fator de Necrose Tumoral alfa/biossíntese , Carragenina/química , Carragenina/isolamento & purificação , Carragenina/farmacologia , Géis , Humanos , Interleucina-10/sangue , Estrutura Molecular , Sulfatos , Fator de Necrose Tumoral alfa/sangue
17.
Carbohydr Polym ; 132: 118-25, 2015 Nov 05.
Artigo em Inglês | MEDLINE | ID: mdl-26256332

RESUMO

Laminaran and three fractions of fucoidan were isolated from brown alga Alaria angusta. The laminaran AaL was characterized as a typical 1,3;1,6-ß-D-glucan (ratio of bonds 1,3:1,6 = 10:1). Fucoidans AaF1 and AaF2 are sulfated heteropolysaccharides, containing fucose, galactose, mannose and xylose. The fraction AaF3 is sulfated and acetylated galactofucan with the main chain represented by a repeating unit → 3)-α-L-Fucp-(2,4-SO3(-))-(1 →. According the data of methylation analysis, AaF3 contains mainly 1,3-linked fucose, less 1,4-linked and 1,4,6-linked galactose residues. The autohydrolysis (37 °C) of fucoidan AaF3 allowed to obtain selectively 2-desulfaled polysaccharide fraction, built up of fucose only, and low molecular weight (LMW) fraction. The negative-ion tandem mass spectrometry of LMW fraction, further hydrolyzed by acid hydrolysis identified the following fragments: Gal-2-SO3(-)-(1 → 4)-Gal, Gal-4-SO3(-)-(1 → 4)-Gal, Gal-(1 → 2)-Gal-4-SO3(-), Fuc-2-SO3(-)-(1 → 4)-Gal, Gal-2-SO3(-)-(1 → 3)-Fuc-(1 → 3)-Fuc, Fuc-2-SO3(-)-(1 → 3)-Fuc-(1 → 4)-Gal. The laminaran AaL and the fucoidan AaF3 exhibited no cytotoxicity in vitro for HT 29, T-47D, and SK-MEL-28 cell lines. The AaF3 fraction suppressed colony formation of HT 29 and T-47D cells, AaL-only HT 29 cells.


Assuntos
Antineoplásicos/química , Glucanos/química , Phaeophyceae/química , Polissacarídeos/química , Antineoplásicos/isolamento & purificação , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Glucanos/isolamento & purificação , Glucanos/farmacologia , Humanos , Neoplasias/tratamento farmacológico , Polissacarídeos/isolamento & purificação , Polissacarídeos/farmacologia
18.
Methods Mol Biol ; 1308: 299-312, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26108514

RESUMO

The application of mass spectrometry towards the structural analysis of the most interesting sulfated biopolymers of the brown algae-fucoidans only developed relatively recently. During method development, many problems, both chemical and instrumental, have to be solved. For example, mass spectrometry has a limitation in the analysis of anionic high molecular weight (HMW) polysaccharides because of the labile nature of sulfate groups which cause the polysaccharide to desulfate rather than ionize. Thus, decomposition methods should be developed taking into account the structural features of such a complex and fragile compound. The selection of optimal instrument settings for the electrospray ionization mass spectrometry (ESIMS) and of matrix media for matrix-assisted laser desorption/ionization mass spectrometry (MALDIMS) is also required. When optimal parameters for mass spectrometric analyses are found, the application of these methods to the elucidation of structural features of fucoidans (by studying their fragments) allows researchers to rapidly obtain new and unique data, often impossible to achieve by other techniques. Herein, we describe tandem mass spectrometry of sulfated fucooligosaccharides, obtained by an autohydrolysis technique from structurally different fucoidans.


Assuntos
Phaeophyceae/química , Polissacarídeos/análise , Espectrometria de Massas por Ionização por Electrospray/métodos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Sequência de Carboidratos , Dados de Sequência Molecular , Espectrometria de Massas em Tandem/métodos
19.
Carbohydr Polym ; 121: 207-16, 2015 May 05.
Artigo em Inglês | MEDLINE | ID: mdl-25659691

RESUMO

A sulfated galactofucan SgF (MW 123kDa) was purified from the brown alga Saccharina gurjanovae. Polysaccharide was depolymerized by autohydrolysis at 25 and 60°C, and products were studied by mass spectrometry and (13)C NMR spectroscopy. According to results of investigation, the main chain of this polysaccharide is built of a repeating units →3)-α-L-Fucp-(2,4-OSO3(-))-(1→. Fucose chains could be sometimes terminated by (1→3)-linked galactose residues. Shorter (1→4)- and/or (1→6)-linked sulfated galactose chains are attached at positions C-2, C-3 of fucose residues. Sulfate groups can occupy positions C-2 and/or sometimes C-3 of Gal residues, but a sulfation at C-4 of the galactofucan could not be excluded. The SgF-AH25-H preparation (71kDa) was obtained by autohydrolysis of SgF at 25°C, which leaded to a selective desulfation at C-2 and, probably, to a cleavage of galactose chains, since structure of SgF-AH25-H represented a repeating unit →3)-α-l-Fucp-(4-OSO3(-))-(1→, which was definitely established by (13)C NMR spectroscopy. Galactofucan SgF and its derivative SgF-AH25-H exhibited no cytotoxic activity and leaded to about the same colony formation inhibition in colon cancer DLD-1 cells. Hence, structural simplification of SgF by lowering its molecular weight, desulfation at C-2 and removing of galactose residues by autohydrolysis at 25°C did not decrease its anticancer activity. This procedure allows obtaining standardized products which can be used as medical.


Assuntos
Antineoplásicos/química , Phaeophyceae/química , Polissacarídeos/química , Antineoplásicos/farmacologia , Sequência de Carboidratos , Linhagem Celular Tumoral , Humanos , Dados de Sequência Molecular , Polissacarídeos/farmacologia
20.
Carbohydr Polym ; 115: 768-75, 2015 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-25439960

RESUMO

Structural peculiarities of low molecular weight (LMW) sample obtained by mild acid hydrolysis of κ-carrageenan from Chondrus armatus was investigated by a rapid mass spectrometric method. The selected conditions allowed avoiding excess destruction of 3,6-AnGal residues that was shown by using tandem (MS/MS) mode. Main oligosaccharide fraction with molecular weight 2.3 kg/mol, obtained by mild acid hydrolysis was chosen for the analysis. It was shown that fragments with even degree of polymerization (DP) were mostly built of (-G4S-DA-)n repeating unit, n=1-5. Some fragments with odd DP were shown to contain -DA2S- insertions, being the fragments of ι-type blocks, which were randomly distributed along the polysaccharide chain as single insertions. The anti-inflammatory activity (on acetic acid-induced colitis in mice) of the initial polymer and its derivatives was studied. The anti-inflammatory effect of the polymer was observed at a dose of 5mg/kg. Polysaccharide decreased the degree of colon damage more than twice and area of damage in 40%.


Assuntos
Anti-Inflamatórios , Carragenina , Chondrus , Colite/tratamento farmacológico , Oligossacarídeos , Ácido Acético , Animais , Anti-Inflamatórios/química , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/uso terapêutico , Carragenina/química , Carragenina/farmacologia , Carragenina/uso terapêutico , Colite/induzido quimicamente , Colite/enzimologia , Colite/patologia , Colo/efeitos dos fármacos , Colo/enzimologia , Colo/patologia , Géis , Hidrólise , Masculino , Camundongos , Peso Molecular , Oligossacarídeos/química , Oligossacarídeos/farmacologia , Oligossacarídeos/uso terapêutico , Peroxidase/metabolismo , Espectrometria de Massas em Tandem
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