ABSTRACT
R-Phycoerythrin is one of the phycobiliproteins widely found in seaweeds. In this study, we have shown to extract and purify R-Phycoerythrin from the south east cost Indian red seaweed Halymenia floresia. R-Phycoerythrin was extracted in 50mM phosphate buffer (pH 7.0). The preparative native PAGE purification was employed alternative to the chromatography and therefore can be scaled up efficiently. Both the yield and the purity of R-Phycoerythrin are very effective. The purified R-Phycoerythrin showed a single band on the examination by native PAGE electrophrosis. SDS-PAGE analysis showed five bands at 16kDa, 21kDa, 30kDa, 39kDa and 47kDa which corresponds to the α, ß and γ', γ and αß subunits. This preparative method for R-Phycoerythrin purification can offer a reference for R-Phycoerythrin purification from other marine red macro algae.
Subject(s)
Electrophoresis, Polyacrylamide Gel/methods , Phycoerythrin/analysis , Phycoerythrin/chemistry , Rhodophyta/chemistryABSTRACT
The aim of this work was to study the effect of Halymenia floresia (Hf) on duodenum contractility, and on experimental protocols of gastric compliance (GC) in rats. Fraction Hf2s exhibited a concentration-dependent myocontractile effect (EC50 12.48 µg/ml), and an inhibitory effect after consecutive washing. The contractile response promoted by Hf2s in the duodenum strips was completely inhibited by verapamil, and the effects were prevented in the presence of Ca2+-free medium. The pretreatment with atropine prevented the Hf2s myocontractile effect. Hf2s was also capable to decrease the GC (from 3.8±0.06 to 3.4±0.13 ml, P<0.05), which did not return to basal levels after more 50 min of observation. These results indicated that the algal polysaccharide possessed in vitro and in vivo gastrointestinal effects.
ABSTRACT
Alternative sources of anticoagulants have arisen as a result of the increasing demand for safer anticoagulant clinical therapy, and the sulfated polysaccharides of seaweeds have gained attention in biomedicine. In this study, crude sulfated polysaccharide fractions (denominated Hf1, Hf2 and Hf3) were obtained from the red marine alga Halymenia floresia and the anticoagulant properties of a soluble crude polysaccharide fraction (Hf2s) were assayed. The three differential extractions yielded 38.6%. The polysaccharides are composed mainly of galactose, with small amounts of xylose and glucose. The anticoagulant properties of Hf2s containing 53.8% sulfate and 3% protein was also compared to those of heparin (193.0 IU mg-1) by assays of activated partial thromboplastin time (APTT) and thrombin time (TT) using normal human plasma. Hf2s showed a higher anticoagulant activity (68.4 IU mg-1) than those of Hf1s and Hf3s, whose activities were 37.6 and 36.6 IU mg-1, respectively. The compound was less active than heparin, but its anticoagulant mechanism suggested that it is dependent on cofactor heparin II to inhibit thrombin activity, but not on cofactors VIII and IX. Therefore, the polysaccharide from H. floresia interfered on coagulation cascade.
O aumento da demanda por anticoagulantes para a terapia clínica tem motivado a busca por fontes alternativas de anticoagulantes mais seguros e os polissacarídeos sulfatados de algas marinhas têm ganhado atenção na biomedicina. Objetivou-se obter frações de polissacarídeos sulfatados brutos (denominadas Hf1; Hf2 e Hf3) da alga marinha vermelha Halymenia floresia e para avaliar as propriedades anticoagulantes de uma fração polissacarídica bruta solúvel anticoagulante (Hf2s). As três extrações diferenciais renderam 38,60%. Os polissacarídeos são principalmente compostos de galactose com pequenas quantidades de xilose e glucose. As propriedades anticoagulantes da Hf2s, contendo 53,80% de sulfato e 3% de proteínas, foram também comparadas com a heparina (193,00 UI mg-1) pelo ensaio do tempo de tromboplastina parcial ativada (TTPA) e tempo de trombina (TT), usando plasma humano normal. A Hf2s apresentou maior atividade anticoagulante (68,40 UI mg-1) que Hf1s a Hf3s, cujas atividades foram 37,60 e 36,60 UI mg-1, respectivamente. O composto foi menos ativo que a heparina, mas sugere-se que o mecanismo anticoagulante seja dependente do cofator II da heparina para inibição da atividade da trombina, exceto pelos cofatores VIII a IX. Portanto, o polissacarídeo de H. floresia interferiu na cascata de coagulação.
