Polysaccharides Sepharose

Lipoteichoic acids (from gram-positive bacteria) [56411-57-5J. Extracted by hot phenol/water from disrupted cells. Nucleic acids that were also extracted were removed by treatment with nucleases. Nucleic resistant acids, proteins, polysaccharides and teichoic acids were separated from lipoteichoic acids by anion-exchange chromatography on DEAE-Sephacel or by hydrophobic interaction on octyl-Sepharose [Fischer et al. Ear J Biochem 133 523 1983]. 

Dextran gels have been utilized since the late 1950s (1) for the separation of biopolymers. First attempts on Sephadex (2-5) and Sephadex/Sepharose (6-8) systems are documented for hydrolyzed and native starch glucans. Up until now, particularly for the preparative and semipreparative separation of polysaccharides, a range of efficient and mechanically stable Sephacryl gels (9-14) have been developped. 

Agarose gels have been used for more than two decades to separate polysaccharides (17-22). In particular, Sepharose CL 2B is widely used (6-8) to separate native starch, but continuously improved mechanical and chemical stability made all of the Sepharose CL gels perfect systems for the analysis of high molecular and broad distributed polysaccharides (23-28). 

Imidazole-HCl soluble polysaccharides Most of the polysaccharides of this group didn t elute from the Q-Sepharose column with 10 mM buffer (Table 3). The glycosyl composition (Table 4) shows that these polysaccharides are homogalacturonans and ramnogalacturonans with... 

Crude polysaccharide fraction (GL-2) was prepared from the leaves of P. ginseng by hot water extraction, ethanol precipitation and dialysis, and GL-2 was fractionated by Cetavlon precipitation and weakly acidic polysaccharide fraction (GL-4) was obtained[3]. GL-4IIb2 was purified from GL-4 by DEAE-Sepharose CL-6B as described previousely [3]. In order to remove the color-materials, GL-4IIb2 was further purified by Q-Sepharose (C1 form), and the major fraction, eluted with 0.3 M NaCl, was repurifled by gel filtration on Bio-gel P-30 column to obtain purified active polysaccharide, GL-4IIb2. ... 

This protocol describes the activation of an agarose gel, e.g., Sepharose 4B. Other polysaccharide-based supports are handled analogously. 

Stipe powder of C. comatus (100 g) was extracted three times with 1 L 95% ethanol under reflux for 2 h to remove lipid, and the residue was extracted three times with 2 L distilled water for 2 h at 80 °C with intermediate centrifugation (2000 x g, 15 min). After concentrating the collected aqueous supernatants to 400 mL (reduced pressure at 40 °C), a precipitation was performed with 3 volumes of 95% ethanol. The precipitate was washed with ethanol and acetone, and then dried at 40 C, yielding crude polysaccharide material. Crude polysaccharide material was dissolved in 100 mL 0.2 M sodium phosphate buffer (pH 6.0), and after centrifugation the solution was applied to a DEAE-Sepharose CL-6B column. 

The structure of this gel is shown m Figure 3.19E. It provides for the attachment of ligands containing hydroxyl, thiol, or amino groups. The hydroxyl groups of mono-, oligo-, and polysaccharides can readily be attached to the gel. Epoxy-activated Sepharose 6B is available from Pharmacia. 

Table 3.7 Amylose percentage of starch from 16 maize genotypes determined following sepharose 2B-CL column chromatography and the peak fraction s absorbance maxima and absorptivity (a) of the polysaccharide-iodine complex3...

Lentil Lectin Sepharose Wheat Germ Lectin Lectins Membrane proteins, glycoproteins, polysaccharides Pharmacia... 

The application of insolubilized lectins in polysaccharide fractionation was demonstrated by Kennedy and Rosevear with Con A-Sepharose... 

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