Separation of polar compounds on polar stationary phases with partly aqueous eluents is by no means a new separation mode in LC. The first HPLC applications were published more than 30 years ago, and were for a long time mostly confined to carbohydrate analysis. In the early 1990s new phases started to emerge, and the practice was given a name, hydrophilic interaction chromatography (HILIC). Although the use of this separation mode has been relatively limited, we have seen a sudden increase in popularity over the last few years, promoted by the need to analyze polar compounds in increasingly complex mixtures. Another reason for the increase in popularity is the widespread use of MS coupled to LC. The partly aqueous eluents high in ACN with a limited need of adding salt is almost ideal for ESI. The applications now encompass most categories of polar compounds, charged as well as uncharged, although HILIC is particularly well suited for solutes lacking charge where coulombic interactions cannot be used to mediate retention. The review attempts to summarize the ongoing discussion on the separation mechanism and gives an overview of the stationary phases used and the applications addressed with this separation mode in LC.

The porosity and flow characteristics of macroporous polymer monoliths that may be used to prepare separation media, flow-through reactors, catalysts, or supports for solid-phase chemistry can be controlled easily during their preparation. Key variables such as temperature, composition of the pore-forming solvent mixture, and content of cross-linking divinyl monomer allow the tuning of average pore size within a broad range spanning 2 orders of magnitude. The polymerization temperature, through its effects on the kinetics of polymerization, is a particularly effective means of control, allowing the preparation of macroporous polymers with different pore size distributions from a single composition of the polymerization mixture. The choice of pore-forming solvent is also important, larger pores being obtained in a poor solvent due to an earlier onset of phase separation. Increasing the proportion of the cross-linking agent present in the monomer mixture not only affects the composition of the final monoliths but also decreases their average pore size as a result of early formation of highly cross-linked globules with a reduced tendency to coalesce. The synergy of different effects has also been observed under specific polymerization conditions using two monomer pairs, styrene−divinylbenzene and glycidyl methacrylate−ethylene dimethacrylate polymerized in close molds. Mercury intrusion porosimetry measurements, inverse size exclusion chromatography, and back pressure measured at different flow rates with the macroporous monoliths were used for the characterization of the porous properties. A good correlation between pore size and flow resistance that follows the Hagen−Poiseuille equation used previously to describe flow through a straight tube has been found.

A model system has been developed for in situ photopolymerization of glycidyl methacrylate and trimethylolpropane trimethacrylate, leading to macroporous monolithic sorbents. This model system allows the preparation of continuous porous objects intended for applications such as detection, separation, and catalysis. The ease of the preparation, the short time needed for reaction, and the possibility of running the reaction at a low temperature are some of the main advantages of the photoinitiated in situ polymerization compared to a thermally initiated polymerization. Important system variables acting upon the porous properties and flow characteristics of the monoliths have been investigated in an experimental 2 3 full factorial design. The porous properties of the monoliths are a direct consequence of the quality of the porogenic solvent, as well as the percentage of cross-linking monomer and the ratio between the monomer and porogen phases. The presence of interactive effects between these reaction conditions were verified using multivariate analysis. It was concluded that the pore formation mechanism in an in situ photopolymerization follows rules similar to those found earlier for thermally initiated in situ polymerization of poly(glycidyl methacrylate-co-ethylenedimethacrylate) and poly(styrene-co-divinylbenzene).

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Hyperphosphorylation at tyrosine is commonly observed in tumor proteomes and, hence, specific phosphoproteins or phosphopeptides could serve as markers useful for cancer diagnostics and therapeutics. The analysis of such targets is, however, a challenging task, because of their commonly low abundance and the lack of robust and effective preconcentration techniques. As a robust alternative to the commonly used immunoaffinity techniques that rely on phosphotyrosine(pTyr)-specific antibodies, we have developed an epitope-imprinting strategy that leads to a synthetic pTyr-selective imprinted polymer receptor. The binding site incorporates two monourea ligands placed by preorganization around a pTyr dianion template. The tight binding site displayed good binding affinities for the pTyr template, in the range of that observed for corresponding antibodies, and a clear preference for pTyr over phosphoserine (pSer). In further analogy to the antibodies, the imprinted polymer was capable of capturing short tyrosine phosphorylated peptides in the presence of an excess of their non-phosphorylated counterparts or peptides phosphorylated at serine.

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