Packings hydrolytic stability

Apolar stationary phases suffer from hydrolytic instability at pH extremes. The use of mixed phases of long (Cg, Clg) and short (C, C3) chain alkyls produces stationary phases with increased hydrolytic stability.7,8 Crowding of the long alkyl chains does not allow the alkylsilane molecules to deposit in close packing on a smooth or flat surface. Silane molecules polymerize in vertical direction, loosing contact with the silica surface. The insertion of short chain alkyls allows horizontal polymerization of the silane molecules. Thus, alkyl chains are aligned in a parallel way. The stability of the silane layer is increased consequently (figure 8.1). 

Bonded phase chromatography (BPC) takes place either under normal phase or reverse phase conditions. In reverse phase mode the stationary phase is non-polar while the eluant is polar, e.g. methanol or acetonitrile with aqueous buffers. Bonded phase packings have superceded the classical packings where the stationary phase was distributed over the surface of the support particles and bound simply by physical forces of attraction. However, due to the problems of solvent stripping and limited hydrolytic stability, these classical systems, though developed for a few specialised applications, have been replaced by organo-bonded stationary phase materials. 

The synthetic methods used for the preparation of bonded phase materials are illustrated in Figure 6.35. One of the first reported bonded phases, the alkoxy silanes (1) also referred to as silicate esters, was prepared by the direct esterification of silanol groups with alcohols. The major disadvantage of this packing material was its hmited hydrolytic stability, as it is readily hydrolysed by aqueous alcohol eluants. 

In chapter 4 we discussed the physical properties of chromatographic adsorbents. In this chapter we will discuss their chemical properties. The most important aspect of the chemistry of a packing is the character of the adsorbing surface. But the chemistry of the packing also plays a role with respect to its hydrolytic stability or whether and to what degree it shrinks and swells in various solvents. In addition, the chemistry of the packing influences its physical strength. The chemistry of the surface influences adsorption kinetics and mass transfer. 

Maximum hydrolytic stability of silica-based reversed-phase packings is obtained at pH values around 4. At pH 2 and pH 8 one may encounter a faster aging of the column, visible through decreased hydrophobic interaction and increased silanophilic interaction. However, the actual rate depends on many different parameters, including the choice of buffer, temperature, and the organic modifier. Columns tend to be mote stable in acetonitrile-based mobile phases than in methanol-based mobile phases, and recent results indicate that dtrate or Tris buffers are preferred over phosphate buffers (32). 

For alumina, titania, and zirconia, there exists as yet no covalent bonding chemistry that is equivalent to the silanization technique used for silica. Although attempts have been made to silanize these other oxides, the hydrolytic stability of these phases does not match up to the hydrolytic stability of the support itself. Therefore alternative surface modification tet ques have been developed that do not rely on the attachment of the modifier to the surface. The coating can be simply insoluble in the intended mobile phases, or a crosslinked coating can be formed that stretches like a net around the skeleton of the particle. Both techniques are, in principle, independent of the nature of the substrate and can be applied to all inorganic or polymeric packings. 

The higher surface coverage available with polyfunctional silanes may have an advantage with respect to the hydrolytic stability of the packing, but the results ate not clearcut. At acidic pH, a reduced rate of hydrolysis was observed for Cig-type bonded phases prepared from di- and trifunctional silanes (9). 

The type of pH modifier to make a desired mobile phase pH also has an effect on the column stability, and this is indirectly related to the peak efficiency and the retention of the analyte. As an increasing number of column volumes of the mobile phase are traversed through the column, the stability of the packing material could be comprised. Rearrangement of the packing bead leads to the loss of efficiency, dissolution of silica leads to loss in efficiency and retention, and hydrolytic decomposition of the bonded phase could impact the peak shape and retention. Different compounds, such as neutral compounds, acidic compounds, and basic compounds, could show different behaviors. 

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