HPLC Column Selection Guide

Carbon Load

Bonding Type

OH

Bonded Phase Bonded Phase Bonded Phase Bonded Phase Bonded Phase Bonded Phase

Bonded Phase Bonded Phase Bonded Phase

OH + X Si R CH3

Monomeric Bonding

O Si R + HX

X
+ X Si R

OH

Polymeric Bonding

CH3 R + 2HX X Si

Carbon Load refers to the amount of bonded phase attached to the base material. It is used to compare C18, C8, and phenyl bonded phases. Carbon load is not generally useful for shorter bonded phases (CN, NH2, C4, C2, ion exchange) because the weight percent of bonded phase is too small to be a useful comparative tool.

Effect on Chromatography
For C18, C8, or phenyl packings the carbon load is a good indication of the column capacity. Normal carbon load values range from approximately 3% to 20% or greater. Higher carbon loads will generally give higher column capacities, greater resolution, and longer run times. More organic will be required in the mobile phase to elute the same sample on columns with higher carbon load. Low carbon loads will shorten analysis times and may provide selectivity differences because of the greater exposure of the base material. For the same carbon load, polymerically bonded packings (see Type of Bonding) will give greater capacity and longer run times than monomeric phases.

Bonding Type refers to how the bonded phase is attached to the base material. There are two methods of attaching the bonded phase to the base material often referred to as monomeric bonding and polymeric bonding. Monomeric bonding is characterized by a single point of attachment of each bonded phase molecule to the base material. This results in a brush type phase so named because the bonded phase is thought to extend perpendicular from the surface of the base material similar to bristles on a brush. Polymeric bonding is characterized by multiple attachment points of each bonded phase molecule to the base material. Bonded phase molecules may also bond to each other leading to a somewhat denser bonded phase layer and better coverage of the base material.

Effect on Chromatography
Monomeric phases generally provide predictable chromatography and good mass transfer leading to high efficiency columns; however, they show somewhat limited stability in highly aqueous mobile phases. Polymerically bonded packings exhibit greater sample retention for a given carbon load than do monomeric phases. Polymeric bonding also provides a more stable phase especially under acidic conditions which can hydrolyze bonded phases. A selectivity change is often seen when switching from one bonding type to the other.

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How to Choose

Choose high carbon loads for complex samples which require the maximum degree of separation. Choose low carbon loads to give shorter analysis times for simple sample mixes and for samples which require a high water content for solubility or stability.

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How to Choose

Choose monomeric bonding for rapid mass transfer and high column efficiency when chemically similar molecules need to be resolved. Choose polymeric packings for maximum column lifetime, for high sample loading capacity, and when highly aqueous mobile phases are used.

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CH3

CH3