Ismael Nilo Lino de Queiroz

Hospital Universitário Clementino Fraga Filho, Rio de Janeiro, Rio de Janeiro, Brazil

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Publications (15)29.2 Total impact

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    ABSTRACT: Sulfated fucans from sea urchin egg jelly express well-defined chemical structures that vary with species. This species specificity regulates the sperm acrosome reaction, a critical step to assure intra-specific fertilization. In addition, these polysaccharides are involved in other biological activities such as anticoagulation. Although sulfation patterns are relevant to the levels of response in both activities, conformation and dynamics of these glycans are also contributing factors. However, data about these features of sulfated fucans are very rare. To address this, we have employed nuclear magnetic resonance (NMR) experiments combined with molecular dynamics (MD) on structurally defined oligosaccharides derived from two sulfated fucans. The results have indicated that the oligosaccharides are flexible in solution. Ring conformation of their composing units displays just the (1)C4 chair configuration. In a particular octasaccharide, composed of two tetrasaccharide sequences, inter-residual hydrogen bonds play a role to decrease dynamics in these repeating units. Conversely, the linking disaccharide [-3)-α-L-Fucp-2(OSO3 (-))-(1-3)-α-L-Fucp-4(OSO3 (-))-(1-] located right between the two tetrasaccharide units has amplified motions suggested to be promoted by electrostatic repulsion of sulfates on opposite sides of the central glycosidic bond. This conjunction of information about conformation and dynamics of sulfated fucan oligosaccharides provides new insights to explain how these glycans behave free in solution and influenced by sulfation patterns. It may also serve for future studies concerning structure-function relationship of sulfated fucans, especially those involving sea urchin fertilization and anticoagulation. © The Author 2014. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.
    Glycobiology 12/2014; DOI:10.1093/glycob/cwu184 · 3.75 Impact Factor
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    ABSTRACT: Biosimilar enoxaparins have been available for clinical use in Brazil since 2009. Although their use has reduced costs of treatment expenses, their implementation still raises some concerns about efficiency, safety, regularity and reproducibility of batches. We undertook structural and functional analyses on over 90 batches of pharmaceutical-active ingredient, and 330 ones of the final products of biosimilar enoxaparins available in the Brazilian market between 2009 and 2014. Besides a nationwide-scale analysis, we have also employed methods that go beyond those recommended by the standard pharmacopeias. We have used high-resolution 2D NMR, detailed assessment of the anticoagulant and antithrombotic properties, check of side effects in experimental animals after continuous administration, and analyses of individual composing oligosaccharides. The 1D 1H NMR spectra of all batches of biosimilar enoxaparins are fairly coincident, and the resultant average spectrum is quite identical to that from the original drug. This structural equality was also assured by highly resolved 2D NMR spectra. The anticoagulant activity, determined by diverse assays and the in vivo antithrombotic and bleeding effects of the biosimilar version were confirmed as equal as of the parental enoxaparins. Structure and function of the composing oligosaccharides were identical in both enoxaparin types. No side effect was observed after continuous subcutaneous administration to rats for 30 days at the dose of 2 mg kg-1 body weight. Biosimilar enoxaparins available in Brazil fulfilled the requirement of the five items defined by FDA-USA for approval of this type of drug.
    Thrombosis and Haemostasis 09/2014; 112(6). DOI:10.1160/TH14-05-0411 · 5.76 Impact Factor
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    ABSTRACT: Sulfated galactans are polysaccharides with heterogeneous structures that frequently show anticoagulant activity. Their anticoagulant mechanisms are complex and distinct from those observed for heparin. Sulfated galactans act through a combination of effects involving serpin-dependent and serpin-independent mechanisms. Interestingly, these polymers can also induce blood coagulation due to activation of factor XII. The structure of a complex sulfated galactan from the red alga Acanthophora muscoides was characterized by solution NMR. This polysaccharide and another previously characterized algal sulfated galactan from Botryocladia occidentalis were each used in in vitro and in vivo anticoagulant and antithrombotic assays to understand the possible structural determinants of their functional effects. The serpin-dependent anticoagulant effects and factor XII-related procoagulant effects of the sulfated galactans decreased in parallel with the molecular size. The serpin-independent anticoagulation also correlated with the chemical structure of the sulfated galactans. The sulfated galactan from A. muscoides, which showed mostly serpin-independent anticoagulant activity and reduced activation of factor XII, drastically reduced arterial thrombus formation. However, the sulfated galactans produced opposite effects on venous thrombosis; this difference appears to result from the tenuous balance between the various effects on coagulation, including serpin-dependent and serpin-independent anticoagulation and factor XIIa-dependent procoagulation. This study of novel sulfated polysaccharides with distinct effects on coagulation and thrombosis helps to establish the minimal structural-function relationship required for the development of antithrombotic drugs. This article is protected by copyright. All rights reserved.
    Journal of Thrombosis and Haemostasis 11/2013; DOI:10.1111/jth.12448 · 5.55 Impact Factor
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    ABSTRACT: Background: Sulfated polysaccharides from red marine algae have presented a variety of potentially therapeutic biological effects, however, their antinocicpetive and anti-inflammatory properties are not well understood. Methods: Male Swiss mice were pretreated with a sulfated polysaccharidic fraction obtained from the marine alga Acanthophora muscoides (AmII) (1, 3 or 9 mg/kg, iv) 30 min prior to either receiving an injection of 0.8% acetic acid or 1% formalin or prior to a thermal stimulus. AmII (1, 3 or 9 mg/kg, sc) was evaluated on carrageenan-, dextran- bradykinin-, histamine- and serotonin-induced rat paw edema models. AmII (500 μg, sc) was also injected into the paw. Additionally, mice were treated with the total sulfated polysaccharides from A. muscoides (Am-TSP) (20 mg/kg, ip) for 14 days. Results: AmII reduced the number of acetic acid-induced writhes and licking time in the second phase of the formalin test, but it did not alter the response latency in the hot plate test, suggesting that its antinociceptive action occurs through a peripheral mechanism. AmII did not reduce carrageenan-induced paw edema and MPO activity. However, it reduced dextran-, histamine- and serotonin-induced paw edemas, but not bradykinin-induced edema, suggesting that histamine is the major target of AmII anti-edematogenic activity. AmII injected into the paw did not evoke local edema. Furthermore, Am-TSP induced no consistent signs of systemic damage, as revealed by body mass, organs wet weight and by biochemical, hematological and histopathological analyses. Conclusion: AmII has important antinociceptive and anti-inflammatory properties and represents an important therapeutic agent warranting future studies.
    Pharmacological reports: PR 05/2013; 65(3):600-13. DOI:10.1016/S1734-1140(13)71037-5 · 2.17 Impact Factor
  • Acta Scientiarum Biological Sciences 09/2012; 34(4). DOI:10.4025/actascibiolsci.v34i4.8976
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    ABSTRACT: Red and brown algae sulfated polysaccharides (SPs) have been widely investigated as antinociceptive and/or anti-inflammatory agents; however, no description of these biological properties concerning green algae SPs have been reported. Caulerpa curpressoides (Chlorophyta) presents three SPs fractions (Cc-SP1, Cc-SP2, and Cc-SP3). Anticoagulant (in vitro) and anti- and pro-thrombotic (in vivo) effects of Cc-SP2 had been recently reported. We evaluated the effects of Cc-SP2 using models of nociception and acute inflammation in vivo. Male Swiss mice received Cc-SP2 (iv) 30 min prior to receiving 0.6% acetic acid (10 ml/kg, ip), 1% formalin (20 μl, sc) or were subjected to thermal stimuli (51 ± 1 °C). Cc-SP2 was injected sc to male Wistar rats in a peritonitis model or a paw edema model using carrageenan (ip or ipl, 500 μg). To analyze the systemic effects, Cc-SP2 (27 mg/kg, sc) was administrated to both genders mice before waiting for 14 days. Cc-SP2 (3, 9 or 27 mg/kg) reduced (p < 0.05) the number of writhes induced by acetic acid by 57, 89.9 and 90.6%, respectively, the licking time in the first (9 or 27 mg/kg with 42.47 and 52.1%, respectively) and the second (3, 9 or 27 mg/kg with 68.95, 82.34 and 84.61%, respectively) phases. In the hot-plate test, the antinociceptive effect of Cc-SP2 (9 mg/kg) was primarily observed at 60 min (26.7 ± 1.2 s), with its effect reversed by naloxone (8.6 ± 1.3 s), suggesting the involvement of the opioid system. Cc-SP2 (3, 9 or 27 mg/kg, sc, p < 0.05) showed anti-inflammatory effects by decreasing neutrophils migration by 64, 69 and 73%, respectively, and potently reduced the paw edema, especially at the second (0.16 ± 0.02, 0.16 ± 0.03 and 0.12 ± 0.05 ml) and third (0.16 ± 0.03, 0.18 ± 0.02 and 0.14 ± 0.04 ml) hours, respectively. Cc-SP2 did not cause hepatic or renal alterations or affect body mass or the macroscopy of the organs examined (p > 0.05). Histopathological analyses of the liver and kidney showed that both organs were affected by Cc-SP2 treatment, but these effects were considered reversible. The results indicate that the analgesic and anti-inflammatory effects of Cc-SP2 could be of biomedical applicability as a new, natural tool in pain and acute inflammatory conditions.
    Pharmacological reports: PR 03/2012; 64(2):282-92. DOI:10.1016/S1734-1140(12)70766-1 · 2.17 Impact Factor
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    ABSTRACT: Sulfated polysaccharides (SPs) have attracted growing interest for various biotechnological applications. We evaluated the efficiency of two methods of drying SPs (M I and II) extracted from Halymenia sp Rhodophyceae in order to compare the yield, purification and anticoagulant activity. The total SPs (TSPs) were first extracted with papain in 100 mM sodium acetate (pH 5.0) containing cysteine and EDTA (5 mM). The TSPs obtained were dried in an oven (M I) or lyophilized (M II) and then examined by ion exchange chromatography (DEAE-cellulose) using the NaCl gradient technique. The fractions were analyzed by 0.5% agarose gel electrophoresis and the in vitro anticoagulant activity was evaluated by the activated partial thromboplastin time test using normal human plasma and compared to heparin (HEP) (193.00 IU mg -1). There was a difference in TSP yield of 19.05% and similar chromatographic SP profiles. Electrophoresis revealed fractions with distinct resolutions. The fractions eluted with 0.75 M of salt (M I and II) were the most active, measuring 27.40 and 72.66 IU mg -1 , respectively, when compared to HEP. Therefore, obtaining SP with anticoagulant activity from Halymenia sp. is more efficient by freeze-drying. Análise de dois métodos de secagem sobre o rendimento e atividade de polissacarídeos sulfatados extraídos da rodofícea Halymenia sp. RESUMO. Os polissacarídeos sulfatados (PS) têm despertado interesse crescente para diversas aplicações em biotecnologia. Avaliou-se a eficiência de duas metodologias de secagem de PSs (M I e II) extraídos da rodofícea Halymenia sp. a fim de se comparar o rendimento, a purificação e a atividade anticoagulante. Inicialmente, os PS totais (PSTs) foram extraídos com papaína em tampão acetato de sódio 100 mM (pH 5,0) contendo cisteína e EDTA (5 mM). Os PSTs foram secados em estufa (M I) ou liofilizados (M II), seguido por cromatografia de troca iônica (DEAE-celulose) utilizando um gradiente de NaCl. As frações foram analisadas por eletroforese em gel de agarose a 0,5% e a atividade anticoagulante "in vitro" avaliada por meio do teste do tempo de tromboplastina parcial ativada usando plasma humano normal e comparadas à heparina (HEP) (193.00 UI mg -1). Verificaram-se diferença no rendimento de PSTs (19,05%) e semelhantes perfis cromatográficos de PS, revelando, por eletroforese, frações de PS distintas em grau de resolução entre os métodos. As frações eluídas com 0,75 M de sal (M I e II) foram as mais ativas, com valores de atividade da ordem de 27,40 e 72,66 UI mg -1 , respectivamente, em relação à HEP. Portanto, a secagem por liofilização seria uma forma mais eficaz na obtenção de PSs com atividade anticoagulante de Halymenia sp. Palavras-chave: Halymeniales, macromoléculas sulfatadas, métodos de secagem, coagulação sanguínea.
    Acta Scientiarum Biological Sciences 01/2012; 34(1). DOI:10.4025/actascibiolsci.v34i1.6961
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    Pharmacological reports: PR 01/2012; 64. · 2.17 Impact Factor
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    ABSTRACT: Seaweeds have attracted special interest as good sources of sulphated polysaccharides (SP) for use in pharmaceutical industries and biotechnology. In this study, we evaluated the effects of SP from the red seaweed Gracilaria cornea (Gc-TSP) in nociceptive and inflammatory models. In mice, Gc-TSP (3, 9 or 27 mg/kg) significantly reduced nociceptive responses, as measured by the number of writhes, at all tested doses. In a formalin test, Gc-TSP significantly reduced licking time in both phases of the test at a dose of 27 mg/kg. In a hot-plate test, the antinociceptive effect was observed only in animals treated with 27 mg/kg of Gc-TSP, suggesting that the analgesic effect occurs through a central action mechanism at the highest dose. Gc-TSP (3, 9 or 27 mg/kg) caused only a slight reduction in neutrophil migration in the rat peritoneal cavity. However, lower doses of Gc-TSP (3 and 9 mg/kg) significantly inhibited paw oedema induced by carrageenan, especially at 3 hr after treatment. Reduction in oedema was confirmed by myeloperoxidase activity in the affected paw tissue. In addition, treatment (s.c.) of animals with different doses of Gc-TSP inhibited paw oedema induced by dextran within the first hour in all doses tested. After 14 consecutive days of intraperitoneal administration of Gc-TSP (9 mg/kg), we measured the wet weight of the liver, kidney, heart, spleen and thymus and performed biochemical, haematological and histopathological evaluations. No systemic damage was found. These results indicate that Gc-TSP possesses analgesic and anti-inflammatory effects and is a potentially important tool worthy of further study.
    Basic & Clinical Pharmacology & Toxicology 10/2011; 110(4):335-41. DOI:10.1111/j.1742-7843.2011.00811.x · 2.29 Impact Factor
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    ABSTRACT: Heparin (HEP) is known due to their side effects and the red seaweed Halymenia floresia (Hf) sulfated polysaccharides (SP) are heparinoids. In this study we purified the Hf-SP obtained from an aqueous extract and evaluated their anticoagulant activities. Hf-SP(1) (25 degrees C), Hf-SP(2) (80 degrees C) and Hf-SP(3) (80 degrees C) were sequentially isolated. Hf-SP(3) had the highest sulfate content (37.45%). Hf-SP(3) was fractionated by ion exchange chromatography on a DEAE-cellulose column using a NaCl gradient. Fractions were lyophilized and submitted to 0.5% agarose gel electrophoresis. The anticoagulant activity was evaluated by the activated partial thromboplastin time using rabbits plasma and expressed in international units per mg of SP using standard HEP (193 IU mg(-1)). The chromatographic procedure separated into four different SP fractions (F I, F II, F III and F IV) eluted at concentrations of 0.50, 0.75, 1.00 and 1.25 M of NaCl, respectively, reveling among them different marked on charge density, when compared by electrophoresis. F III had the highest anticoagulant activity (10.72 IU mg(-1)), suggesting that the sulfate is important in this process. In conclusion, our results suggest that sequential extractions of Hf-SP are an important biotechnological tool for identification of novel anticoagulants and studies of structural characterization are already in progress.
    Acta Scientiarum Technology 09/2011; 33(4). DOI:10.4025/actascitechnol.v33i4.9143 · 0.32 Impact Factor
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    ABSTRACT: This work reports the effects of a sulfated polysaccharide (SP-Sf), isolated from the seaweed Solieria filiformis and characterized by Fourier transformed infrared (FT-IR), on nociception and inflammation. Male Swiss mice were pretreated with SP-Sf 30 min before receiving an injection of 0.8% acetic acid, 1% formalin or 30 min prior to a thermal stimulus. We observed that SP-Sf (1, 3 or 9 mg/kg) significantly reduced the number of writhes. SP-Sf also reduced the second phase of the formalin test and did not cause a significant antinociceptive effect in the hot plate test, suggesting that its antinociceptive action occurs through a peripheral mechanism. SP-Sf (1, 3 or 9 mg/kg) did not show a significant anti-inflammatory effect in Wistar rats when administrated by the systemic route 1 h before testing using carrageenan or dextran. Finally, SP-Sf (9 mg/kg) did not show significant signs of toxicity when administrated in mice.Highlights► SP-Sf showed antinociceptive action through a peripheral mechanism. ► SP-Sf did not show a significant anti-inflammatory effect. ► SP-Sf did not show significant signs of toxicity when administrated in mice. ► The FT-IR spectra of SP-Sf (F I) showed characteristic of κ-carrageenan.
    Carbohydrate Polymers 08/2011; 86(3):1207-1215. DOI:10.1016/j.carbpol.2011.06.016 · 3.92 Impact Factor
  • Acta Scientiarum Technology 07/2011; 33(3). DOI:10.4025/actascitechnol.v33i3.9400 · 0.32 Impact Factor
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    ABSTRACT: This study aimed to isolate, fractionate and evaluate the in vivo toxicity of sulfated polysaccharides (SPs) from Hypnea musciformis (Rhodophyta), when obtained by three extraction methods (M I, M II and M III). SPs were extracted with papain in 100mM sodium acetate (pH 5.0) containing cysteine and EDTA (5mM) (M I) or water (25-80°C (M II), 80°C (M III)), and then their chemical composition of total carbohydrates, free sulfate (FS) and contaminant proteins (CPs) was determined. SPs were submitted to ion-exchange chromatography (DEAE-celulose) using a sodium chloride gradient, being the degree of homogeneity and charge density evaluated by agarose gel electrophoresis of the fractions obtained and compared to heparin. The in vivo assay was performed using groups (n=6) of male and female Swiss mice (24-33g), which received: SPs (9mg kg-1, i.p.) absence of CPs (M I) and 0.9% saline (0.1mL 10g-1, i.p.), for 14 consecutive days. On the 15th day, collect blood and organs for biochemical dosages and corporal mass correlation, respectively, from the animals anesthetized and sacrificed were performed. The sulfate content of FS (31.05±0.53%) (P<0.05) and the fractionation by DEAE-cellulose showed M I more effectiveness in obtaining SPs compared to M II and M III. The animals were tolerable to SPs from M I, and it wasn't observed hepatic or renal alteration (P>0.05)
    Ciência Rural 07/2011; 41(7):1211-1217. DOI:10.1590/S0103-84782011005000094 · 0.40 Impact Factor
  • Acta Scientiarum Biological Sciences 05/2011; 33(2). DOI:10.4025/actascibiolsci.v33i2.6243
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    ABSTRACT: Red algae sulfated polysaccharides (SPs) have been widely described as anticoagulant and antithrombotic agents; however no description of antithrombotic activity regarding green algae SPs has been reported. Caulerpa cupressoides (Chlorophyta) has three different SPs fractions (SP1, SP2 and SP3). We investigated the effects of SP2 on thrombin activity by antithrombin and in an experimental model of venous thrombosis in rats. The inhibition of thrombin assay was evaluated using antithrombin (AT) in the presence of SP2 and the antithrombotic activity was investigated in rats with thromboplastin as the thrombogenic stimulus. The anticoagulant effects of SP2 are suggested be due to the potentiation of thrombin inhibition by antithrombin (IC50 ~ 10.0µg mL-1) and this mechanism of interaction is different when compared to other studied Caulerpa polysaccharides. SP2 exhibited antithrombotic effects at doses of 1.0 and 2.0mg kg-1 body weight, but at higher doses (>2.0mg kg-1 body weight) this polysaccharide revert the antithrombotic property. No hemorrhagic effect (2.0mg kg-1) was observed. As occurs with red algae SPs, these results indicate that green algae SPs are also capable of exhibiting different in vivo properties.
    Ciência Rural 04/2011; 41(4):634-639. DOI:10.1590/S0103-84782011000400014 · 0.40 Impact Factor