Dextrans Sephadex

The materials originally used as stationary phases for GPC were the xerogels of the polyacrylamide (Bio-Gel) and cross-linked dextran (Sephadex) type. However, these semi-rigid gels are unable to withstand the high pressures used in HPLC, and modern stationary phases consist of microparticles of styrene-divinylbenzene copolymers (Ultrastyragel, manufactured by Waters Associates), silica, or porous glass. 

Commonly employed water-insoluble supports for the covalent attachment of enzymes include synthetic supports such as acrylamide-based polymers, maleic anhydride-based polymers, methacrylic acid-based polymers, styrene-based polymers, and polypeptides, and natural supports such as agarose (Sepharose), cellulose, dextran (Sephadex), glass, and starch (Zaborsky, 1973). 

G. Dirheimer and J. P. Ebel (1964a). Separation des acides ribonucleiques et des polyphosphates inorganiques. IV. Separation par filtration sur gel de dextrane (Sephadex G-200). Bull. Soc. Chim. Biol., 46, 396—4-02. 

The media most commonly used in gel chromatography (otherwise termed gel filtration or molecular sieve chromatography) are the cross-linked dextrans (Sephadex), bead-form polyacrylamides (Bio-Gel P) and the bead form agaroses (Sepharose, Bio-Gel A, and Indubiose). It is on these gels that most data have been accumulated and the majority of interactions of interest occur. 

Axen R, Myrin P, and Janson J. Chemical fixation of chymotrypsin to water-insoluble crosslinked dextran (Sephadex) and solubihza-tion of the enzyme derivatives by means of dextranase. Biopolymers 1970 9 401 13. 

A clean separation of the 4 main nucleosides (either ribo- or deoxyribo-) on cross-linked Dextran (Sephadex G-10) has been described by Braun (1967b). He used a 1.5x90 cm column eluted with 0.01 M citric acid Na2HP04, pH 3.5 at 25 ml/hr. The sample was loaded in 0.5 ml water and contained 0.1 mg of each nucleoside. The elution took 10 hr and each nucleoside was eluted in a volume of 10-20 ml in the order, C, U, A, G or C, T, A, G for the... 

Without doubt, concanavalin A (Con A) is the most celebrated and has proven to be one of the most useful of the plant lectins. Its physical chemical properties and carbohydratebinding properties are well documented in previous reviews [3,8]. Suffice it to note that it was first isolated and crystallized by Sumner and Howell in 1936, who showed it to require metal ions for its activity [74]. By virtue of its interaction with branched a-D-glucans, it is readily prepared by affinity chromatography on crossed-linked dextran (Sephadex) [75, 76]. A homotetramer at pH 7 (subunit M, = 26 500 Da) of Con A has been sequenced [77] and its crystal structure determined both in its native form [38,39] and complexed with methyl a-D-mannopyranoside [40] and Man(al-3)[Man(al-6)]Man[49]. 

For many years rehydratable granulated dextran (Sephadex) or polyacrylamide (Bio-Gel) gels have been used for gel chromatography and in the form of hydrated gel layers for analytical and preparative isoelectric focusing (l t, 15). Recently, rehydratable layers of granulated gels on a plastic film have been developed ( ). The presence of carrier ampholytes or addition of 1-2 % glycerol was necessary... 

Figure 4.5 Partial structure of a cross-linked dextran (Sephadex).

Several types of crosslinked gels [e.g. dextrans (Sephadex), poly(acrylamide) (Bio-Gel P), and poly(acryloy Imorpholine) (Enzacryl)] are used for the separation of biopolymers by thin-layer chromatography. These materials must be swollen in an appropriate buffer/solvent system before spreading as a layer. Laboratory-made layers require modified equipment for their use and afford only modest performance characteristics. 

Newer 2-diethylaminoethyl ethers, such as 0-( 2-diethylaminoethyl )-agarose gels," are useful as media for electrophoresis and immunoelec-trophoresis, and the 2-diethylaminoethyl derivative of O-(2-hydroxy-propyl ) cross-linked dextran (Sephadex LH20) is an effective, lipophilic anion-exchanger. The relative stiffness of the molecular chain of 0-(2-diethylaminoethyl) dextran has been estimated from measurements of viscosity at various ionic strengths, and compared with that of other polysaccharide polycations and polyanions. 0-(2-Diethylaminoethyl) dextran itself is claimed to be toxic." ... 

Chloro-2-hydroxypropyl derivatives of such polysaccharides as cellulose and cross-linked dextran (Sephadex) may be prepared by treatment with epichlorohydrin in boron trifluoride etherate. These 3-chloro-2-hydroxypropylated polysaccharides may, in turn, react with ammonia, or primary, secondary, or tertiary amines, and the resultant derivatives of cellulose and Sephadex LH-20, particularly an 0-[3-(di-butylamino)-2-hydroxypropyl] derivative of the latter, are useful anion-exchangers. In the amination procedure, between 50 and 100% of the chlorine atoms are utilized, except in the reaction with tertiary... 

Insoluble streptococcal dextran has been used for the affinity chromatographic purification of the extracellular D-glucosyltransferase from Streptococcus mutans 6715, and Lathyrus odoratus (sweet pea) lectins have been purified by affinity chromatography on crosslinked dextran (Sephadex ). ... 

The gels used in molecular-sieve chromatography are made of cross-linked dextrans (Sephadex type), polyacrylamide, and agar and agarose (Bio-gel type). 

Figure 9-18. Elution volume Vc as a function of the molar mass M of dilute salt solutions of saccharose and various proteins on cross-linked dextrane (Sephadex G-75) (according to P. Andrews).

In Uppsala, J. Porath and P. Flodin developed an extremely important method for the separation of peptides and proteins gel permeation chromatography with crosslinked dextran (Sephadex). One of Porath s former associates, Ulf Ragnarsson contributed with innovative modifications to the solid phase method of peptide synthesis and in more recent years introduced the principle of complete protection of the amine function by diacylation. Another Porath student, Gunnar Lindeberg, explored the properties of vasopressin analogs. 

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