Advances in the understanding of stationary phases for HILIC

Oct 01, 2012

A recent survey that was conducted by LCGC in late 2011 showed that since 2008, hydrophilic interaction liquid chromatography (HILIC) has more than doubled its application. HILIC is finding more and more applications for polar compounds that are either unretained or weakly retained on reversed-phase chromatographic columns. The technique has also been used for the separation of biomolecules. This article reviews the newer stationary phases intended for HILIC and studies performed to categorize columns for optimized separations based on their selectivities.

Hydrophilic interaction liquid chromatography (HILIC) has become an established chromatographic technique in widespread use in most application areas. This great interest has led to an accompanying increase in the number of new stationary phases explored, some of which have become commercially available. For the user, this multiplication of phases is beneficial, but it also presents some challenges. Where to start? What column to choose? How can one get optimal selectivity for a specific application? Therefore, approaches to classify different HILIC stationary phases into categories have emerged. This article will summarize the progress in this direction since a previous article was published on this topic in the 2008 North American supplement (1).

Majors recently reported in an LCGC reader survey (2) that the use of HILIC mode had doubled since the year 2007. This means that roughly 25% of chromatographers are using HILIC today. I had the pleasure to be responsible for the market introduction of the column brand ZIC-HILIC in January 2002, which was immediately well received by researchers at AstraZeneca. At that time, there were about 70 papers on HILIC in the scientific literature published since the first paper by Alpert in 1990 (3). Back then few chromatographers had a complete picture of the retention mechanism of HILIC or which column to use, but there was a multitude of unsolved application problems that were delaying research in several branches of science. Most important, the orthogonal selectivity of HILIC to reversed-phase chromatography was driving the users interest.

Figure 1: The number of HILIC papers per year (43).
Later on, in April 2008 when the previous review in this series was written by McCalley (1), he had to consider in total 350 HILIC papers. Today, this number has risen to almost 1300 papers, with more than 300 new contributions only last year (see Figure 1). Now, scientific literature has become a valuable source of separation applications using HILIC. The question of whether separation is possible can often be answered based on the published work already done. More focus can be given to how it is done and to other aspects of analysis such as sample preparation, throughput and detection. Readers are referred to special journal issues on HILIC (4–6), to a textbook with various chapters addressing successful applications (7), and to reviews on different application areas such as pharmaceutical analysis (8), proteomics (9), metabonomic and metabolomic analysis (10), glycomics (11) and bioanalysis (12).

It is clear that the improved understanding of HILIC and growing interest for selectivity, in general, also has led to better sample handling by complementary pretreatment protocols and, in many methods, to an overall faster throughput.