Ligand immobilization

Porous glass (PG) modified with covalently adsorbed poly(p-nitrophenyl acrylate), as described in Sect. 4.1, turned out to be a highly suitable carrier for immobilization of various biospecific ligands and enzymes. When the residual active ester groups of the carrier were blocked by ethanolamine, the immobilized ligands when bound to the solid support via hydrophilic and flexible poly(2-hydroxyethyl acrylamide). The effective biospecific binding provided by the ligands... 

Glos and Reiser [23] introduced aza-bis(oxazolines) as new chiral ligands for copper and palladium catalysts. Because of the structural flexibility of these compounds they also prepared an immobilized ligand by covalent grafting to methoxypoly(ethyleneglycol) (structures 14 and 15 in Scheme 9). 

Successful affinity chromatography requires that the protein interact with an immobilized ligand tightly enough to be captured from solution,... 

Nonspecific protein binding to the solid phase complicates the method and is a selective pressure driving its evolution. The adaptive response has been the development of intrinsically comparative methods in which specific binding to an immobilized ligand is blocked in one out of two otherwise identical samples. When the respective protein components of the samples are compared, specifically bound proteins are present in one but severely depleted in the other. To allow relative quantitation, the two samples can be made isotopically distinct by a chemical or metabolic process and then mixed for an analytical step that avoids intersample variability [15]. 

Prepare the sulfhydryl column by washing with a disulfide reducing agent. Apply 10 ml of freshly made lOmM DTT solution (15.4mg of DTT dissolved in 10ml of equilibration buffer). This treatment converts the immobilized ligands into a fully reduced form (free —SH groups). 

Even under the most inert atmosphere conditions, the 31CP/MAS spectrum of the immobilized ligand showed a major signal at 6 = 42 ppm (wrt 85% H3POO characteristic of phosphine oxide rather than phosphine. This could be quantitatively reduced by HSiCl3 and this surface reaction monitored by NMR but the subsequent exchange reaction (equation [5]) generated substantial quantities of phosphine oxide and a number of different isomeric complexes were f ormed. 

The signal is measured in terms of resonance units (RU) and the mathematical treatment is as described above. An increase in RU signal is proportional to the concentration of complex AB. The maximum of the specific change in signal then corresponds to a complete saturation of the immobilized ligand. 

The advantages that heterogeneous catalysts have is that they are easily separable from the product, and can be recycled. A number of studies have been conducted in which ligands have been attached or bound to polymeric material to provide an immobilized ligand, and these include polyacrylate and silica [27], polyurea [28], polythiourea [29], polyether [30, 31] and dendritic [32] systems. Upon metal coordination, the immobilized catalysts have retained most of the activity and selectivity, but they now provide the advantage of simple separation and recycling. For exam-... 

A similar type of immobilization was obtained by reacting the phosphonylated 2,2 -bipyridine ligand depicted in Figure 42.10 with excess titanium alkoxide. Rhodium and iridium complexes of this immobilized ligand showed activity for... 

Analysis of Macro molecular Interactions Using Immobilized Ligands, I. Chaiken, S. Rose, R. Karlsson, Analytical Biochemistry 1992, 201,197-210. 

Fig. 2 Determination of Bt values (amount of functional immobilized ligand in the column) for the immobilized Erythrina cristagalli agglutinin. / -Nitrophenyl, (pNP)-lactose, diluted to various concentrations (8 to 50 pM), was used for concentration-dependence analysis. (A) The solid and dotted lines demonstrate elution profiles of pNP-lactose and control sugar (pNP-mannose), respectively. (B) Woolf-Hofstee-type plot was made by using V-V0 values. Adapted from 47 with permission.

Figure 5.17 Structure of the ligand, linker precursor and immobilized ligand assembl)r for silica supports and subsequent functionalization with [Rh(COD)2]Bp4. (Redrawn from Pugin and Blaser [49].)...

Affinity chromatography M Immobilized ligand on column matrix... 

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