52 November / December 2016
Chromatography Today Help Desk Silica gone bad The Chromatography Help Desk
The help desk in this issue will be looking at a common problem that is observed with older generation silica columns primarily, but is occasionally seen with some newer generation silica based columns. The issue is around the batch to batch reproducibility and trying to get an understanding of the issues that separation scientist face, and the challenges that the manufacturers face in trying to ensure consistent product quality. It is one of the most frustrating aspects of separation science, that on occasion a method is developed on a particular column, and subsequent columns with the same part number do not perform the separation in the same manner. There are a range of reasons why this occurs, some of which the manufacturers have thankfully addressed, but some of which are inherent issues with the technology that is used.
Labelling issues
In the early days of column production, there was very little automation and so consequently manual handling issues were a problem, which could result in columns occasionally being mislabelled. This has resulted in chromatography users developing methods on effectively unknown columns. The helpdesk has come across examples where 50 mm columns have been labelled as a 100 mm column, which is relatively easy to identify as a labelling error, however this is not so easy if the stationary phase has been mislabelled. There are examples in the helpdesk inbox, which would suggest that the column used to develop an assay is not the same as that used for the subsequent validation studies, despite having the same part number. Fortunately, the use of electronic logging systems, bar codes and greater use of automation means that this is no longer a significant issue.
Effect of silica variability
Of greater significance for the separation scientist is the variability that is inherent within the manufacturing process. The primary reason for the variability is the substrate material that is commonly used, namely silica. The types of silica manufactured and the effect that silica can have on a separation has been discussed on several occasions in the helpdesk articles. It is also important to be aware of the effect that the packing process can have on the performance of a column, since column packing is still more of an art than a fully understood science
Why two nominally equivalent stationary phases, say C18, have very different retention mechanisms has been addressed in many articles previously, however it is always useful to go through the underlying theory to explain the situation. Ultimately it relates to
two phenomena;
• The different forms of silanol groups that can exist at the surface of a silica particle
• The inability to completely cover the substrate surface, which results in the substrate being involved in the separation
mechanism.
Figure 1. Schematic diagram of the different modes of interactions caused by the different surfaces of silica
Figure 1 is a schematic of the different forms of silica that can exist at the surface. Each of these forms of silica has different chemical properties, specifically different levels of acidity or degree of interaction with basic compounds. Manufacturers have realised that this causes issues and so with the newer phases, much effort is put into ensuring that the surface is relatively homogeneous. There are a variety of process that manufacturers can employ to reduce the variability of the silica surface including chemical treatment and heat treatment, however it could be suggested that despite the best efforts of the manufacturers a completely inert substrate material has still not been developed. The move to smaller particles presents some further challenges as some of the approaches that have been employed to overcome the physical stability issues of porous silica actually result in an increased activity of the substrate material.
The majority of the first type of spherical silica manufactured was derived from the polymerisation of a metal silicate, which resulted in an acidic surface due to a relatively high metal content. This process is still employed for many of the earlier generation of silica particles that are manufactured, and these silicas, often referred to as type 1 silica are often associated with increased tailing when they are used to separate basic compounds. Current
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