Page 12 - Subyek Encyclopedia - Encyclopedia of Separation Science

Sepsci*1*TSK*Venkatachala=BG
I / AFFINITY SEPARATION 7

Figure 2 Triazine coupling. (A) Coupling of human serum albumin (HSA) to ready-acitivated supports as a funciton of pH. (B) Time
course of coupling of human IgG to ready-activated supports at 43C. , CNBr-activated agarose 4XL; , triazine-activated agarose
4XL.

complex biological macromolecules or assemblies, Fortunately there are a vast number of biological
mostly or exclusively consisting of amino acids enti- ligands that can interact with more than one macro-
ties linked together in a speciRc manner. This com- molecule and consequently group-speciRc ligands are
plexity of structure provides many opportunities to commonplace. Since group-speciRc adsorbents retain
exploit the physicochemical differences between a range of ligates with similar binding requirements,
the target molecule and the ligand to be used. Each a single adsorbent may be used to purify a number of
structure contains the four basic intermolecular bind- ligates. Group-speciRc ligates can be used when the
ing forces } electrostatic, hydrogen bonding, hydro- desired ligate is present in high concentration, but this
phobic and van der Waals interactions } spread implies that some preprocessing has taken place and
throughout the structure in an exactly deRned spatial a concentration step interposed. The use of non/
manner. The degree of accessibility and spatial pre- group-speciRc adsorbents can only offer partial
sentation within the pore of the medium, and the separations. This results in having to apply several
strength of each force relative to each other, dictate stages in series, each only capable of removing a pro-
whether these forces are utilized to effect the portion of the impurities. In contrast a highly selec-
separation. The biological recognition between spe- tive ligand can exclusively remove the target in one
cies is a reSection of the sum of the various mo- step, but often the resulting complexes are very tight-
lecular interactions existing between them, and ly bound, have low binding capacities, are easily
this summation is Rxed for the ligate. However, denatured and are expensive to produce. Until re-
various ligands may be found that emulate some cently these adsorbents were restricted to technically
or all of the available binding forces to various difRcult isolations. Today the use of computer-
degrees. assisted molecular modelling systems provides oppor-
AfRnity adsorbents are therefore assigned to tunities to investigate relationships between designed
one of three broad ligand categories: nonspeciRc, ligands and relevant protein structures. For the Rrst
group speciRc or highly speciRc. NonspeciRc ligands time logical design approaches can be applied and
have only a superRcial likeness to biological ligands consequently stable inexpensive ligands have now
and binding is usually effected by just one of the become available.
four binding processes described above. Although ion
exchange materials can be used in a similar manner to Analytical Scale-Up
afRnity adsorbents, they only exhibit the single
force of electrostatic binding. They are thus limited to Modern biotechnology uses two different types
relatively indiscriminate binding. In this case the only of chromatography. Analytical separations require
criterion for binding is that of an overall charge. that run time is minimized, while resolution and