Ruminations on silica-based and organic polymer-based bioanalytical SPE

Robert MacNeill received his Bachelor’s degree with Honors in Chemistry from Heriot-Watt University then his MSc in Analytical Chemistry from the University of Huddersfield, both in the United Kingdom. Robert is also a Chartered Chemist and Member of the Royal Society of Chemistry. With 18 years of experience in all aspects of quantitative bioanalytical LC–MS/MS method development, nine of these years heading method development activities within HLS/Envigo, and a regular author and peer reviewer for the journal Bioanalysis, Robert is a recognized expert and innovator in the field.

In his current role Robert coordinates all LC–MS method development and associated training, takes the lead in keeping abreast of innovation and technological development in the industry, leads in-house research projects and performs technical writing.

With particular regard to the sorbents and underlying chemistries involved, it is interesting to look at the way in which the technique of SPE has evolved in the bioanalytical domain. The early days of SPE with prepackaged and disposable cartridges, in the late 1970s, involved silica-based sorbents exclusively. In the mid-1990s came the advent of crosslinked polymeric sorbent material, predominantly based on styrene and divinylbenzene monomer units. It was not long before the ease of use of this kind of material and the potential application to small molecules, of a wide range of hydrophobicities, became obvious; polymeric sorbents have since become ubiquitous and the crux of a first-stop SPE solution for many. With the relatively large surface area bestowing a similarly large capacity and with no need to condition initially with high-organic to ensure absence of pore dewetting, polymeric sorbents present a convenient ’catch-all‘ option, which can be used with conventional reversed-phase or mixed-mode protocols to wash and subsequently elute compounds of interest with high recoveries. There is also a certain additional motivation, in general terms, to use polymer-based sorbents in preference to silica-based sorbents due to a perception that the latter can and will dry out easily and lead to recovery loss and lack of reproducibility.

In earlier installments of this column, there has been a hint of playing Devil’s Advocate, writing in favor of the use of SPE, which might appear to be gradually falling out of favor. On this occasion, I would like to zoom into the SPE domain and write an endorsement of silica-based SPE in the face of the strikingly abundant use of polymeric SPE in bioanalysis.

There’s no question that polymeric SPE is a fantastic means at our disposal. It makes things easier; a rugged platform upon which extractions can be based. I sometimes like to imagine polymeric particles swelling and shrinking, as they do, with bouncing, elastic properties that constitute a metaphor for the resilience of such methodologies to adverse conditions that could affect performance and data reliability. This is despite the fact that particle swelling and shrinking is detrimental to minimization of band broadening, of course. However, silica has distinct advantages. For one thing, it is actually very difficult to dry out a silica bed. It would take a matter of hours rather than minutes with, say, a typical vacuum, so the fear of premature elution as a result of drying out after a load or wash step is very largely unfounded. As an aside, attempting to incorporate a drying-out step prior to the elution step (so that everything remaining is subsequently eluted in the following step) is probably unwise due to this same difficulty in being able to ensure complete drying of each cartridge within a reasonable time. Besides, so many key interferences are conveniently eliminated through their over-retention beyond the elution step in methods involving reversed-phase retention, so it would be ill-advised to attempt to elute all that remains.

The main advantage of silica-based SPE is the myriad of different chemistries and selectivities attainable, reachable via almost any mode of retention that you’d care to mention. This depends on the compounds being extracted and the nature of the remainder of the applied sample, the solvent composition on-sorbent and what the sorbent has previously encountered, the presence of a bonded phase or otherwise, and of course the nature of the bonded phase including how it affects the surface chemistry. In fact, one particularly rich argument for the preservation of silica-based SPE is the fact that HILIC conditions are readily attainable, whereas on a polymeric sorbent this is not the case. The propensity for HILIC is a result of the presence of the hydroxylated silica surface which is extremely conducive to the formation of a water-enriched layer, allowing the manifestation of fundamental HILIC partitioning. Even with the presence of a hydrophobic bonded phase, this will occur, albeit less reproducibly, on a silica surface. However, on a polymeric surface, this fundamental proclivity towards HILIC is not there, and it can manifest only with the surface binding of pronouncedly polar and ionic moieties.

Then there is the fact that silica bestows the best efficiency as the base material in a stationary phase. In the analogous liquid chromatography, we see peaks eluting from silica-based columns that are sharper than from polymeric columns of the same dimensions and we are reminded of the benefit in terms of efficiency, or sharpness of the eluting bands. This is conducive to having more room to optimize and to give more definition to an optimization, i.e. more leeway to apply a more voluminous and/or stronger wash and a less voluminous and/or less powerful elution solvent blend to attain high and reproducible recoveries. This takes us a step away from a ’switch-on, switch-off‘ perceived approach to SPE and makes it more akin to a migratory chromatographic process, the reality being somewhere in between, of course.

So yes, let’s celebrate what is offered to us by polymeric SPE sorbents and keep them at the forefront where routine, high-throughput applications are prominent, where method development needs to be fast and there are no special challenges or difficulties to overcome. Further to that, though, let’s also keep our repertoire of silica-based media safe. Look at the decades of use in validated methods as testament to the ruggedness offered. Where special measures may be needed to obtain selectivity, or some key fine-tuning is required, one of the many manifestations of silica-based retention may just be a direct route to a solution.

This article is part of Robert MacNeill’s (Envigo) quarterly column for Bioanalysis Zone which focuses on quantitative method design. You can read past installments of the column here. 

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