Application of monoliths as supports for affinity chromatography and fast enzymatic conversion

Monoliths are useful chromatographic supports, as their structure allows im proved mass transport. This results in fast separation. Once the ligand of interest has been immobilized, chromatographic separation can also be accom plished in affinity mode. Ligands with low molecular mass have been shown t o be the easiest to immobilize. Nowadays, ligands with low molecular mass a re often designed by combinatorial chemical techniques. In addition, many a pplications have been described where ligands with high molecular mass, suc h as Proteins A and G, antibodies, lectins and receptors are used. The immobilization of an enzyme on the monolithic support creates a flow-th rough reactor. Small proteins, such as carbonic anhydrase, can be directly immobilized on the support. However, in the case of large molecules, the ac tive center of the enzyme is no longer accessible at all or only to a limit ed degree, An improvement can be achieved by introducing a spacer, which al lows maximum enzymatic conversion. Fast conversion of substrates with high molecular mass has been investigated with immobilized trypsin. It was shown that in case of high-molecular-mass substrates, the conversion rate depend s very much on the flow-rate. Most applications described have been perform ed on an analytical or semi-preparative scale. However, the technical probl ems of up-scaling are close to being definitely solved, enabling enzymatic conversion on a preparative scale in the future. (C) 2001 Elsevier Science B.V. All rights reserved.