Type of the bonded phase

So far we discussed primarily C18 type of bonded ligands. This is because it was historically the first used. Octadecyldimethylclorosilane is readily available sililation agent.

The main purpose of silica surface modification is shielding of the polar silanol groups and transformation of the surface into the hydrophobic one.

Now there are a number of different types of bonded phases exists on the market. Namely, C18, C14, C8, C4, C1, Hexyl-Phenyl, Phenyl, Cyano, NH2, and few others specific types.

They are shown in the order of decreasing of their hydrophobicity. The longer the bonded alkyl chain the thicker the bonded layer, and the better shielding of polar silica surface. Last four ligand types are more specific. Phenyl ring adds some polarisability to the surface; therefore polar groups of the analyte molecules may have some type of specific interactions with that surface. In the certain cases it is useful for the separation of closely related components.

The difference between the Phenyl and Phenyl-Hexyl phases is that the first one has propyl spacer between silica and phenyl ring, and the last has hexyl. The shorter the chain between the polar surface and phenyl ring, the higher the influence of silanoles on the polarization of phenyl. For the propyl-Phenyl the ligand actually acts as a transmitter of the polarity of underlying silica on the top of the bonded phase. The last two phases are the most polar ones, although cyano-phase is somewhat less specific since it could not participate in the hydrogen bonding.

Amino-phase is the most polar one, it also can act as weak anion-exchanger, being protonated at low pH.

Selection of the proper, the most suitable phase is still the trial and error process. The first choice is definitely the C18 phase, since it is the most hydrophobic and is shows a true reversed-phase behavior. If you could not get good separation on C18 phase you might look for the more specific one.

Slide 11