RESUMEN
An exocellular ß-(1â6)-D-glucan (lasiodiplodan) produced by a strain of Lasiodiplodia theobromae (MMLR) grown on sucrose was derivatized by sulfonation to promote anticoagulant activity. The structural features of the sulfonated ß-(1â6)-D-glucan were investigated by UV-vis, FT-IR and (13)C NMR spectroscopy, and the anticoagulant activity was investigated by the classical coagulation assays APTT, PT and TT using heparin as standard. The content of sulfur and degree of substitution of the sulfonated glucan was 11.73% and 0.95, respectively. UV spectroscopy showed a band at 261 nm due to the unsaturated bond formed in the sulfonation reaction. Results of FT-IR and (13)C NMR indicated that sulfonyl groups were inserted on the polysaccharide. The sulfonated ß-(1â6)-D-glucan presented anticoagulant activity as demonstrated by the increase in dose dependence of APTT and TT, and these actions most likely occurred because of the inserted sulfonate groups on the polysaccharide. The lasiodiplodan did not inhibit the coagulation tests.
Asunto(s)
Anticoagulantes/química , Anticoagulantes/farmacología , Ascomicetos/química , Ácidos Sulfónicos/química , beta-Glucanos/química , beta-Glucanos/farmacología , Antitrombinas/farmacología , Ascomicetos/crecimiento & desarrollo , Pruebas de Coagulación Sanguínea , Heparina/farmacología , Humanos , Solubilidad , Sacarosa/química , Trombina/antagonistas & inhibidores , Agua/químicaRESUMEN
The exopolysaccharide botryosphaeran (EPS(GLC); a (1--> 3)(1-->6)-ß-D-glucan from Botryosphaeria rhodina MAMB- 05) was sulfonated to produce a water-soluble fraction (EPS(GLC)-S) using pyridine and chlorosulfonic acid in formamid. This procedure was then repeated twice to produce another fraction (EPSGLC-RS) with a higher degree of substitution (DS, 1.64). The purity of each botryosphaeran sample (unsulfonated and sulfonated) was assessed by gel filtration chromatography (Sepharose CL-4B), where each polysaccharide was eluted as a single symmetrical peak. The structures of the sulfonated and re-sulfonated botryosphaerans were investigated using ultraviolet-visible (UV-Vis), Fourier-transform infrared (FT-IR), and (13)C nuclear magnetic resonance ((13)C NMR) spectroscopies. EPS(GLC) and EPS(GLC)-RS were also assayed for anticoagulation activity, and EPS(GLC)-RS was identified as an anticoagulant.
Asunto(s)
Anticoagulantes/química , Glucanos/química , Anticoagulantes/metabolismo , Anticoagulantes/farmacología , Ascomicetos/química , Ascomicetos/metabolismo , Coagulación Sanguínea/efectos de los fármacos , Glucanos/metabolismo , Glucanos/farmacología , Humanos , Estructura Molecular , Relación Estructura-ActividadRESUMEN
Botryosphaeran (EPS(FRU)), an exopolysaccharide of the beta-(1-->3,1-->6)-d-glucan type with 31% branching at C-6, is produced by the fungus Botryosphaeria rhodina MAMB-05 when grown on fructose as carbon source. Botryosphaeran was derivatized by sulfonation to induce anticoagulant activity. The effectiveness of the sulfonation reaction by chlorosulfonic acid in pyridine was monitored by the degree of substitution and FT-IR analysis of the sulfonated EPS(FRU) (once sulfonated, EPS(FRUSULF); and re-sulfonated, EPS(FRURESULF)). Activated partial thromboplastin time (APTT) and thrombin time (TT) tests of EPS(FRURESULF) indicated significant in vitro anticoagulant activity that was dose-dependent. EPS(FRU) did not inhibit any of the coagulation tests.
Asunto(s)
Anticoagulantes/farmacología , Ascomicetos/crecimiento & desarrollo , Ascomicetos/metabolismo , Fructosa/metabolismo , Glucanos/química , Glucanos/farmacología , Ácidos Sulfónicos/metabolismo , Anticoagulantes/química , Anticoagulantes/metabolismo , Coagulación Sanguínea/efectos de los fármacos , Glucanos/metabolismo , Heparina/farmacología , Humanos , Espectroscopía Infrarroja por Transformada de FourierRESUMEN
Four exopolysaccharides (EPS) obtained from Botryosphaeria rhodina strains isolated from rotting tropical fruit (graviola, mango, pinha, and orange) grown on sucrose were purified on Sepharose CL-4B. Total acid hydrolysis of each EPS yielded only glucose. Data from methylation analysis and (13)C NMR spectroscopy indicated that the EPS from the graviola isolate consisted of a main chain of glucopyranosyl (1-->3) linkages substituted at O-6 as shown in the putative structure below: [carbohydrate structure: see text]. The EPS of the other fungal isolates consisted of a linear chain of (1-->6)-linked glucopyranosyl residues of the following structure: [carbohydrate structure: see text]. FTIR spectra showed one band at 891 cm(-1), and (13)C NMR spectroscopy showed that all glucosidic linkages were of the beta-configuration. Dye-inclusion studies with Congo Red indicated that each EPS existed in a triple-helix conformational state. beta-(1-->6)-d-Glucans produced as exocellular polysaccharides by fungi are uncommon.
Asunto(s)
Ascomicetos/química , Frutas/microbiología , Glucanos/química , Polisacáridos/química , beta-Glucanos/química , Ascomicetos/aislamiento & purificación , Conformación de Carbohidratos , Secuencia de Carbohidratos , Cromatografía de Gases y Espectrometría de Masas , Glucanos/metabolismo , Espectroscopía de Resonancia Magnética , Modelos Moleculares , Datos de Secuencia Molecular , Espectroscopía Infrarroja por Transformada de Fourier , Clima Tropical , beta-Glucanos/metabolismoRESUMEN
Three D-glucans were isolated from the mycelium of the fungus Botryosphaeria rhodina MAMB-05 by sequential extraction with hot-water and hot aqueous KOH (2% w/v) followed by ethanol precipitation. Following their purification by gel permeation chromatography on Sepharose CL-4B, the structural characteristics of the D-glucans were determined by FT-IR and 13C NMR spectroscopy and, after methylation, by GC-MS. The hot-water extract produced a fraction designated Q1A that was a beta-(1-->6)-D-glucan with the following structure: [Formula: see text] The alkaline extract, when subjected to repeated freeze-thawing, yielded two fractions: K1P (insoluble) that comprised a beta-(1-->3)-D-glucan with beta-D-glucose branches at C-6 with the structure: [Formula: see text] and K1SA (soluble) consisting of a backbone chain of alpha-(1-->4)-linked D-glucopyranosyl residues substituted at O-6 with alpha-D-glucopyranosyl residues: [Formula: see text]
Asunto(s)
Ascomicetos/química , Pared Celular/química , Glucanos/química , Glucanos/aislamiento & purificación , Micelio/química , Secuencia de Carbohidratos , Cromatografía de Gases y Espectrometría de Masas , Espectroscopía de Resonancia Magnética , Datos de Secuencia Molecular , Espectroscopía Infrarroja por Transformada de FourierRESUMEN
The herbicide, Scepter, whose active principle is imazaquin, is commonly used in soybean farming to combat wide-leaf weeds. The basidiomycete, Pleurotus ostreatus , and the ascomycete, Botryosphaeria rhodina , were evaluated for their growth and laccase production when cultured on basal media containing Scepter. Both fungi could grow on the herbicide when cultivated in solid and submerged liquid culture in the presence of Scepter at concentrations of 0-6% (v/v) for P. ostreatus , and up to 0-50% (v/v) for B. rhodina , and in each case produced laccases when assayed against ABTS [2,2(1)-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid)] and 2,6-dimethoxyphenol. P . ostreatus could tolerate up to 6% of Scepter before it became toxic to the fungus, while in the case of B. rhodina , 50% (v/v) Scepter was the highest amount that supported grow, and laccase activity was detectable up to 25% (v/v). An inverse relationship existed between the level of Scepter in the culture medium that supported fungal growth and laccase production. Analysis of the results showed that the fungi studied presented different behaviour towards Scepter in the culture environment.