Ion exchange separation of biopolymers

It is a pity that these ion exchangers, which have been proved to be effective for the separation of biopolymers are not yet produced commercially. 

This packing has high capacity and has been found useful for the separation of enzymes and other proteins. The ion exchangers suitable for the separation of biopolymers are summarized in the lower part of Table 4.5.2. 

Ion exchange cellulose, polydextran and agarose derivatives are used for the LC separations of biopolymers [19,20]. In some cases (meth)acrylate derivatives can also be applied for such a purpose. LC of other organic substances (various bases and acids, amino acids and peptides, nucleosides and nucleotides, sugars, etc.) can be realized using ion exchange resins (cf.. Ref. 5, Tables 13.1-13.9, and Ref. 6, Tables... 

With the second edition of this book the focus on fine chemicals and small pharmaceutical molecules is expanded to ion-exchange chromatography and the separation of biopolymers such as proteins. In accordance with the first edition these topics are restricted to those applications that can be modeled and simulated by current methods and procedures. 

The by far largest application area of high-performance ion-exchange chromatography is the separation of biopolymers proteins and nucleic acids. Although a special term, fast protein liquid chromatography (FPLQ, is occa-... 

Available functional groups are listed in Table 7.1. Silica can also be functionalized with groups suitable for ion exchange, affinity chromatography or the separation of enantiomers. The choice of special phases for the separation of biopolymers is immense. 

Ion-exchange chromatography is routinely used for the separation of biopolymers at laboratory scale... 

Bonded silicas are fairly stable in an acidic pH range of I to 2. Such conditions are usually applied in the resolution of peptides and proteins by reversed-phase HPLC. Separation of biopolymers is performed in the pH range between 5 and 8 with buffers in ion-exchange and hydrophobic interaction chromatography. The type and concentration of buffer have a remarkable effect on the stability of the bonded silica. For instance, phosphate buffers at pH 7 are not recommended, particularly when separations are performed at elevated temperatures. The lifetime of the column in phosphate will be much shorter than in Tris buffer [31]. Buffers with a pH above 8 should not be applied due to the dissolution of the silica matrix. 

There are two major reasons for modification of pore surfaces in some polymeric resins (1) hydrophiiization of hydrophobic surfaces and (2) an increase in loading capacity and kinetics of ion-exchange resins. The former is typical of styrene-divinylbenzene copolymers that are too hydrophobic to be used directly for the separation of biopolymers in some modes. In this case, hydrophilic polymers such as dextran, poly(oxyethylene), poly(ethylene imine), and poly-(vinyl alcohol) are adsorbed and cross-linked on [35,43] or covalently linked to the pore surface to form a thin biopolymer friendly barrier on the hydrophobic surface. Regnier [44] was one of the first to develop a covalently attached hydrophilic coating that substantially decreased the nonspecific irreversible adsorption of proteins. 

Salt (ionic strength) gradients in lEC discussed in Section 5.4.3.3 are frequently used in the separation of complex peptides, proteins, and other biopolymer samples as a complementary technique to RP solvent gradient separations, often in a 2D setup [99,100]. The gradients usually start at a low salt (chloride, sulfate, etc.) concentration and typically run from 0.005 to 0.5 M. A buffer is used to control the pH acetonitrile and methanol may be added to improve the resolution and urea to improve the solubility of proteins that are difficult to dissolve. Ion exchangers with not strongly hydrophobic matrices usually prevent protein denaturation in aqueous mobile phases. 

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