Avaliação do potencial antiinflamatório, anticoagulante e hemorrágico de heparinóides presentes em "Artemia franciscana"

Heparan sulfate (HS) and Heparin (Hep) glycosaminoglycans (GAGs) are heterogeneous and highly charged polysaccharides. HS is structurally related to Hep but is much less substituted with sulfo groups than heparin and has a more varied structure (or sequence). Because of structural similiarities between these two polymers, they have been described together as heparinoids . Both chains bind a variety of proteins and mediate various physiologically important processes including, blood coagulation, cell adhesion and growth factor regulation. Heparinoids with structural characteristics similar to these described from HS and/or Hep from mammalian tissues have been isolated from different species of invertebrates, although only a few heparinoids from unusual sources have been characterized. The present study describes the presence of unusual heparinoids population from Artemia franciscana, isolated after proteolysis and fractionation by ion exchange resin and named, F-3.0M. The study model in vivo were hemostasis (rat tail scarification) and inflamatoty activity. The tests in vitro were used for coagulations assays (PT and APTT). The analyse of the heparinoids eluted with 3,0M NaCl showed electrophoretic migration in different buffer systems a single band with a behaviour intermediate between those of mammalian HEP and HS. The main products obtained from Artemia heparinoids after enzymatic degradation with heparitinases I and II from F. heparinum were N-sulphated disaccharides (∆U-GlcNS,6S/ ∆U,2S-GlcNS and ∆U-GlcNS) and N-acetylated disaccharides (∆U, GlcNAc). This heparinoid had a lower hemorrhagic effect (400μg/ml) when compared to unfractiionated heparins(25μg/ml).The results also suggest a negligible APTT activity of this heparinoid (62.2s). No action was observed on PT indicating that F-3.0M haven t action on the extrinsic pathway. The results showed that the fraction F- 3.0M have inhibitory effect on migration of leukocytes, 64.5% in the concentration of 10 μg/ml (P<0.001). The search for new heparin and/or heparan sulphates analogs devoid of anticoagulant activity is an atractive alternative and may open up a wide variety of new therapeutic applications

Atividades Biológicas De Xilana De Sabugo De Milho

The corn cob is an agricultural by-product still little used, this in part due to the low knowledge of the biotechnological potential of their molecules. Xylan from corn cobs (XSM) is a polysaccharide present in greater quantity in the structure of plant and its biotechnology potential is little known. This study aimed to the extraction, chemical characterization and evaluation of biological activities of xylan from corn cobs. To this end, corncobs were cleaned, cut, dried and crushed, resulting in flour. This was subjected to a methodology that combines the use of alkaline conditions with waves of ultrasound. After methanol precipitation, centrifugation and drying was obtained a yield of 40% (g/g flour). Chemical analysis indicated a high percentage of polysaccharides in the sample (60%) and low contamination by protein (0.4%) and phenolic compounds (> 0.01%). Analysis of monosaccharide composition indicated the presence of xylose:glucose:arabinose:galactose:mannose:glucuronic acid in a molar ratio 50:20:15:10:2.5:2.5. The presence of xylan in the sample was confirmed by nuclear magnetic resonance (¹H and ¹³C) and infrared spectroscopy (IR). Tests were conducted to evaluate the antioxidant potential of XSM. This showed a total antioxidant capacity of 48.45 EAA/g sample. However, did not show scavenging activity of superoxide and hydroxyl radical and also reducing power. But, showing a high capacity chelating iron ions with 70% with about 2 mg/mL. The ability to XSM to influence cell proliferation in culture was also evaluated. This polymer did not influence the proliferation of normal fibroblast cells (3T3), however, decreased the rate of proliferation of tumor cells (HeLa) in a dose-dependent, reaching an inhibition of about 50% with a concentration around 2 mg/mL. Analyzing proteins related to cell death, by immunoblotting, XSM increases the amount of Bax, Bcl-2 decrease, increase cytochrome c and AIF, and reduce pro-caspase-3, indicating the induction of cell death induced apoptosis dependent and independent of caspase. XSM did not show anticoagulant activity in the PT test. However, the test of activated partial thromboplastin time (aPTT), XSM increased clotting time at about 5 times with 600 μg of sample compared with the negative control. The presence of sulfate on the XSM was discarded by agarose gel electrophoresis and IR. After carboxyl-reduction of XSM the anticoagulant activity decreased dramatically. The data of this study demonstrate that XSM has potential as antioxidant, antiproliferative and anticoagulant compound. Future studies to characterize these activities of XSM will help to increase knowledge about this molecule extracted from corn and allow their use in functional foods, pharmaceuticals and chemical industries.

Estudo etnofarmacológico e fitoquímico das espécies medicinais Cleome spinosa Jacq, Pavonia varians Moric e Croton cajucara Benth

In this present work an ethnographic research was performed with 84 native medicinal specimens from the Litoral Norte Riograndense, from which two plants Cleome spinosa Jacq e Pavonia varians Moric were submitted to ethnobotanic, phytochemistry and pharmacologic investigations. Additionally, a phytopharmacological research of the medicinal specimen Croton cajucara Benth ( native plant of the Amazon region of Brazil) was improved. The obtained phytochemical results of the C. spinosa and P. varians showed the presence of flavonoids constituents, among other components. The two flavonoids (2S)-5-hydroxy-7,4 -dimethoxy-flavanone and 5,4 -dihydroxy-3,7,3 -trimethoxy-flavone were isolated from C. spinosa. The antioxidant activity of the hydroalcoholic extracts of C. spinosa and P. varians solubilized in the microemulsion systems SME-1 and SME-4, was evaluated in the DPPHmethod. The used SME systems [obtained with Tween 80: Span 20 (3:1) and isopropyl myristate (IPM)] improved the dissolution of those tested polar extracts, with higher efficacy to the SME-1 system (in which ethanol was included as cosurfactant). The CE50 values evidenced for P. varians were 114 [g/mL (SME-1) and 246 [g/mL (SME-4); for C. spinosa it was 224 [g/mL (SME-1) and 248 [g/mL (SME-4), being the system SME-1 more effective for both tested extracts. The hydroalcoholic extracts of P. varians (HAE-PV) was also submitted to pharmacological screening for antinociceptive activity in animal models. The oral administration of this extract (100, 300 and 1000 mg/kg) inhibited the acetic acid-induced writhing in mice. The higher inhibition (74%) was evidenced to the 1000 mg/kg administered dose. Its effect on the central nervous system (CNS) was investigated by tail flick and formalin-method and reveled that it has negligible antinociceptive action on the CNS. After taking consideration of HAE-PV interaction, Pavonia varians Moric could be used as a potent analgesic agent in case of peripheral algesia, without affecting the CNS. The phytochemical study of the stem bark of Croton cajucara Benth lead to the isolation of 19-nor-clerodanetype diterpenes, as well as to the separation of its fixed oil FO-CC. This non polar oil material reveled to be rich in sesquiterpenes and 19-nor-clerodanes components. The biologic effect of OF-CC was evaluated in the development in vitro of the fungis phytopatogens such as Fusarium oxysporum, Rhizoctonia solani and Sclerotium rolfsii. Significant inhibitory effect of the tested fungis (at 0,2 mg.mL-1 dosage) were comproved. A Mass Spectrometry study of clerodane-type diterpenes was developed in order to identify characteristic fragments on mass spectrometra of both clerodane and 19-nor-clerodane presenting an α,β-insaturated carbonyl moiety at ring A of the decalin-system. For that study, mass spectroscopy data were analysed for 19-nor-clerodanes [trans-dehydrocrotonin (DCTN), trans-crotonin (CTN), cis-cajucarin B (c-CJC-B), and cajucarinolide (CJCR)] and for clerodanes [isosacacarin (ISCR) and transcajucarin A (t-CJC-A)] obtained from the stem bark of C. cajucara, and also clerodane-type from other species. The trans-junction of the enone-system clerodanes was clear correlated with the presence of the characteristic ions at m/z 95, 121 e 205. Meanwhile, the characteristics ions at m/z 122 e 124 were correlated to cis-junction. The trans-junction of the enone-system 19-nor-clerodanes showed characteristics ions at m/z 161, 134 e 121. This study could be successful employed for identification of clerodane constituents from other specimens without any additional spectroscopic analyses, as well as a previously phytochemical analyzes in clerodane project search

Avaliação do potencial anti-inflamatório, anticoagulante e hemorrágico de heparinóides presentes em "Artemia franciscana"

Heparan sulfate (HS) and Heparin (Hep) glycosaminoglycans (GAGs) are heterogeneous and highly charged polysaccharides. HS is structurally related to Hep but is much less substituted with sulfo groups than heparin and has a more varied structure (or sequence). Because of structural similiarities between these two polymers, they have been described together as heparinoids . Both chains bind a variety of proteins and mediate various physiologically important processes including, blood coagulation, cell adhesion and growth factor regulation. Heparinoids with structural characteristics similar to these described from HS and/or Hep from mammalian tissues have been isolated from different species of invertebrates, although only a few heparinoids from unusual sources have been characterized. The present study describes the presence of unusual heparinoids population from Artemia franciscana, isolated after proteolysis and fractionation by ion exchange resin and named, F-3.0M. The study model in vivo were hemostasis (rat tail scarification) and inflamatoty activity. The tests in vitro were used for coagulations assays (PT and APTT). The analyse of the heparinoids eluted with 3,0M NaCl showed electrophoretic migration in different buffer systems a single band with a behaviour intermediate between those of mammalian HEP and HS. The main products obtained from Artemia heparinoids after enzymatic degradation with heparitinases I and II from F. heparinum were N-sulphated disaccharides (∆U-GlcNS,6S/ ∆U,2S-GlcNS and ∆U-GlcNS) and N-acetylated disaccharides (∆U, GlcNAc). This heparinoid had a lower hemorrhagic effect (400μg/ml) when compared to unfractiionated heparins(25μg/ml).The results also suggest a negligible APTT activity of this heparinoid (62.2s). No action was observed on PT indicating that F-3.0M haven t action on the extrinsic pathway. The results showed that the fraction F- 3.0M have inhibitory effect on migration of leukocytes, 64.5% in the concentration of 10 μg/ml (P<0.001). The search for new heparin and/or heparan sulphates analogs devoid of anticoagulant activity is an atractive alternative and may open up a wide variety of new therapeutic applications

Atividades biológicas de xilana de sabugo de milho

The corn cob is an agricultural by-product still little used, this in part due to the low knowledge of the biotechnological potential of their molecules. Xylan from corn cobs (XSM) is a polysaccharide present in greater quantity in the structure of plant and its biotechnology potential is little known. This study aimed to the extraction, chemical characterization and evaluation of biological activities of xylan from corn cobs. To this end, corncobs were cleaned, cut, dried and crushed, resulting in flour. This was subjected to a methodology that combines the use of alkaline conditions with waves of ultrasound. After methanol precipitation, centrifugation and drying was obtained a yield of 40% (g/g flour). Chemical analysis indicated a high percentage of polysaccharides in the sample (60%) and low contamination by protein (0.4%) and phenolic compounds (> 0.01%). Analysis of monosaccharide composition indicated the presence of xylose:glucose:arabinose:galactose:mannose:glucuronic acid in a molar ratio 50:20:15:10:2.5:2.5. The presence of xylan in the sample was confirmed by nuclear magnetic resonance (¹H and ¹³C) and infrared spectroscopy (IR). Tests were conducted to evaluate the antioxidant potential of XSM. This showed a total antioxidant capacity of 48.45 EAA/g sample. However, did not show scavenging activity of superoxide and hydroxyl radical and also reducing power. But, showing a high capacity chelating iron ions with 70% with about 2 mg/mL. The ability to XSM to influence cell proliferation in culture was also evaluated. This polymer did not influence the proliferation of normal fibroblast cells (3T3), however, decreased the rate of proliferation of tumor cells (HeLa) in a dose-dependent, reaching an inhibition of about 50% with a concentration around 2 mg/mL. Analyzing proteins related to cell death, by immunoblotting, XSM increases the amount of Bax, Bcl-2 decrease, increase cytochrome c and AIF, and reduce pro-caspase-3, indicating the induction of cell death induced apoptosis dependent and independent of caspase. XSM did not show anticoagulant activity in the PT test. However, the test of activated partial thromboplastin time (aPTT), XSM increased clotting time at about 5 times with 600 μg of sample compared with the negative control. The presence of sulfate on the XSM was discarded by agarose gel electrophoresis and IR. After carboxyl-reduction of XSM the anticoagulant activity decreased dramatically. The data of this study demonstrate that XSM has potential as antioxidant, antiproliferative and anticoagulant compound. Future studies to characterize these activities of XSM will help to increase knowledge about this molecule extracted from corn and allow their use in functional foods, pharmaceuticals and chemical industries.