Packings silanol activity

Another approach to a reduction in silanol activity has been the use of mixed silanes. This was first practised in the design of the YMC Basic packing, but a recent and more rigorous application of this idea is the Luna packing. The surface is derivatized with a mixture of C13 and Cg silanes. This improves the surface coverage that can be achieved, and reduces the access to surface silanols. The result of this procedure is an improvement in the peak shape of basic analytes (see Table 1). 

Despite these fundamental difficulties, the character of columns can still be described in a general way. Any of the methods can tell which packings are more hydrophobically retentive and can discriminate between high and low silanol activity. However, the relative position of closely related packings may change depending on the method, and we should be... 

If we are interested in packings with about the same hydrophobicity, we should select packings that are located on a vertical line in this chart. Let us say that we would like to find packings that result in an overall hydrophobic retention similar to Spherisorb ODS-2, 73. We could select, in sequence of reduced silanol activity, the following packings Nucleosil Cig, 72, Purospher RPjg, 86, Polyencap C g, 81, Develosil ODS UG5,... 

Table 1.4 Interactions between analytes and stationar> phase packing materials. (O Primary Interaction Secondary Interaction Silanol Activity) ...

The HPLC of large biomolecules such as proteins and DNA often requires specialized columns packed with wide-pore polymer or silica-based bonded phase with extra-low silanol activity.1215 Alternate approaches are pellicular materials or very small nonporous particles. Some of these columns are packed in PEEK or titanium hardware to allow the use of high-salt mobile phase and to prevent possible protein denaturing by metallic leachates. Further details on bio-separations and application examples are discussed in Chapter 7. 

Hydrophobic and ionic characteristics of proteins, leading to their adsorption in many LC packings with active sites (i.e., acidic silanols)... 

As mentioned in Chapter 3, protein separations typically require specialized columns packed with wide-pore polymer supports or silica materials with extra-low silanol activity. Figure 7.23 shows an example of an RPC gradient separation of a protein mixture using a column packed with Vydac C4 bonded... 

We have drawn a horizontal line in Fig. 1 to differentiate between classical bonded phases, which can all be foimd above the line, and phases with embedded polar groups, which lie below the line. Other phases with very low silanol activity can also be found below the line. An example is ODPerfect, 113, which is not based on silica. Beyond the limits of the chart shown here, one finds packings with exceptionally high silanol activities, such as the classical Zorbax Cjg or Resolve Cjg, both of which lack end-capping and are based on older silicas. 

The fundamental reason for this phenomenon is the fact that the selectivity of a separation arises from a combination of the influence of the stationary phase and the influence of the mobile phase. If the composition of the mobile phase is drastically different from the test conditions, one can expect a different position of the different columns relative to each other. It remains correct that Symmetry Cj8 has a lower silanol activity than Spherisorb ODS-2, but whether a Lima Cjg(2) or a YMC-Pack Pro Cjg has a higher hydrophobicity or a lower silanol activity surely depends on the details of the measurement conditions. 

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