LCGC Awards Symposium at Pittcon Highlights Recent Developments and Current Challenges

Opening the LCGC Award symposium at Pittcon 2023 with the presentation of the 2023 LCGC Lifetime Achievement Award, session chair and LCGC editorial director Laura Bush described the accomplishments that led to the addition of Peter Schoenmakers, a professor of analytical chemistry and a former scientific director of the van ’t Hoff Institute for Molecular Science at the University of Amsterdam, to the 16-year list of impressive recipients of the prestigious award. An early landmark in Schoenmakers’s career was his foundational 1978 paper entitled “Gradient Selection in Reversed-Phase Liquid Chromatography” (1). He went on to publish more than 300 papers, including important contributions on the optimization of two-dimensional liquid chromatography (2D-LC), significant advances in polymer analysis, and even fundamental works progressing to spatial 3D separations. In discussing the intersection of his polymer work and his development of multidimensional techniques, Bush quoted Pete Carr, a professor of chemistry at the University of Minnesota and himself an icon of 2D-LC, who said, “It is not an exaggeration to say that Shoemakers has revolutionized the highly important field of polymer characterization through his seminal and extensive work in 2D-LC.”

Schoenmakers: A Look at the History of LC and Where It Is Heading

Upon receiving the award, Schoenmakers acknowledged that he had to get used to the idea of being at a point in his career where he could receive a lifetime achievement award, illustrating his unease with an image of the Grim Reaper. He said he was determined that this would not mark the end of his involvement in the scientific community.

In his presentation, Schoenmakers gave his perspective on the history of liquid chromatography and the challenges that the community is still facing. He emphasized the importance of developments in column technology from the 1970s all the way into the new millennium. Thinking back to his very first LC separations, he noted how much progress has been made, through the introduction of improved stationary-phase morphologies, new porosities, and later, ultrahigh-pressure LC (UHPLC) and core–shell technology, all of which dramatically improved the capabilities of LC.

Nevertheless, he noted that the concepts of sample dimensionality, presented by Giddings, and statistical overlap theory discussed by Davis and Giddings, clearly pointed toward multidimensional separations. The first step was 2D-LC, which can be used to achieve an order-of-magnitude greater separation power. With the first-dimension effluent becoming the (diluted) sample of the second-dimension separation, Schoenmakers noted, innovations in modulation technology were critical, and allowed sensitivity and solvent-compatibility problems to be resolved. This solution, along with significant commercial developments by manufacturers, he said, has helped to make 2D-LC more accessible to a much wider range of chromatographers and enabled its use in industry. He proceeded by noting some of his own work, showcasing how he had seen opportunities for reactions (such as light, chemical, or enzymatic degradation) to be conducted within the 2D-LC framework to unlock the analysis of previously inaccessible sample properties.

Schoenmakers also is known for his work on polymer separations, and he highlighted recent developments in thermal modulation for 2D-LC and gradient elution for polymers. More recently, his group worked on the investigation and development of technology for spatial 3D separations. “We aren’t there yet, but I hope some people have the guts to pick up on where we left off,” he said. “The way I see it, this is our only shot at getting to the peak capacity of a million required to tackle those complicated ‘omics samples.”.

As for challenges for the future, Schoenmakers pointed to the need for better software to develop methods and analyze data, the need for better gradient delivery and mixing hardware, and most of all, the desire for a universal detector for LC. He closed his talk by expressing concern about the education gap, and the need to train more analysts. “We have more LC instruments than chromatographers,” he said, while also noting the importance of LCGC as a platform to promote separation science and educate users.

Pirok: Challenges in Automating Method Development and Fully Exploiting the Data

The next presentation, by Bob Pirok, an assistant professor at the University of Amsterdam, focused on current challenges in automation of 2D-LC method development. Pirok, who completed his PhD under the supervision of Schoenmakers, explained that his research line is focused on improving the accessibility of separation technology. “Separation technology advances faster than our ability to fully exploit it and its data,” he said. “We need to stop for a second and see how we can use all the great theoretical concepts in the literature—to make better use of what we have.”

Pirok explained how retention modeling and recent developments in machine learning enable fully automated method development, showing examples for peptide separations using comprehensive 2D-LC (LC×LC) that he developed in collaboration with the group of Dwight Stoll at Gustavus Adolphus College.

At the core of the automated methodology are chemometric algorithms that track analytes across chromatograms using peak detection and mass spectrometry (MS) analysis. “There is a lot of room for improvement,” Pirok acknowledged, as he referred to the “garbage in, garbage out” (GIGO) principle of computer science, which emphasizes that no algorithm can produce something good from bad data. Pirok noted that current bottlenecks are mostly related to chemometrics, giving examples of modeling and peak detection. “As chromatographers, we need to acknowledge that we need chemometricians to help us progress,” he said.

Kohler: Addressing the Challenges of Analyzing New Psychoactive Substances (NPS)

Isabelle Kohler, an assistant professor at the Vrije Universiteit Amsterdam, switched gears to focus on the application side of analytical chemistry, giving a refreshing presentation about her work on new psychoactive substances (NPS). Kohler explained that criminals almost become chemistry wizards to circumvent the law, making small changes to the chemical structures of psychoactive compounds such that they are slightly different from banned drugs and thus remain technically legal. Kohler explained the dramatic consequences of these drugs, in particular for younger generations, who often think that because the drugs are not illegal, they must be safe.

An important example are cathinones, which include positional isomers that present one of the most challenging classes of NPS in the Netherlands. Each of these positional isomers is classified differently under current regulations and thus analytical techniques must provide 100% confidence in distinguishing the positional isomers. Kohler showed how trapped ion mobility spectrometry–MS (TIMS-MS) enables confident identification of the compounds in confiscated street samples, even when the pills contained mixtures of different NPS. An additional benefit of this direct-injection method is that it is very fast, with analysis times under 4 min.

At various points, Kohler raised the question of whether she would need chromatography for any aspect of this work. It was both a scientific question and lighthearted teasing, given the chromatography focus of the session. The conclusion, during her talk and the discussion that followed, was that chromatography would most likely be needed for quantitation of these compounds.

Gionfriddo: Microextractions to Improve Environmental and Biological Analyses

Following the break, Bush presented the LCGC Emerging Leader Award to Emanuela Gionfriddo, who is currently an assistant professor of chemistry at the University of Toledo. Amongst the list of her accomplishments and important contributions to separation science, are the development and implementation of green and biocompatible microseparation technologies for extraction of small molecules in environmental and biological samples.

Gionfriddo, who had two days earlier also received the ACS Analytical Division Satinder Ahuja Award for Young Investigators in Separation Science, first presented examples of her past work in Prof. Janusz Pawliszyn’s group at the University of Waterloo, in Canada. One example was the development of biocompatible extraction phases with enhanced coating lifetimes. Gionfriddo achieved this by combining practical and fundamental approaches, including studying the mass transfer of the analyte in the coating. Her work was more than just a fundamental novelty; she applied it to the determination of pesticides, polycyclic aromatic hydrocarbons (PAHs) and polychlorinated biphenyls (PCBs) in various foodstuffs, such as avocado, soy milk, grape juice, and dry seaweed. These experiences led her to develop her own generation of solid-phase microextraction (SPME) coatings, strong enough to endure desorption both in gas chromatography (GC) and liquid chromatography (LC), and featuring improved efficiencies due to the inclusion of hydrophilic-lipophilic balance sorbent particles embedded in polytetrafluoroethylene—amorphous fluoropolymer (PTFE-AF) .

At the University of Toledo, her overarching goal is to study how contaminants partition in the environment and how they affect humans. It is her ambition to develop robust analytical methods for both environmental studies and bioanalysis, to monitor these connections. Like Pirok, she referred to the GIGO principle, but gave it a different twist, focused on the environment. “If humans introduce a lot of chemicals into the environment, the environment is going to give them back to us and then the health implications of this exchange are not going to be pleasant,” she said. She advocated studying these effects and mapping the effects of pollutants on health.

Gionfriddo noted that current challenges for sample preparation include the presence of salts, endogenous metabolites, ion suppression, low concentrations of trace analytes, degradation to enzymatic reactions, and the large numbers of samples requiring robustness and precision. She then highlighted the role of advanced extraction techniques to achieve trace-level detection of compounds. She described some of her work on the analysis of per- and polyfluoroalkyl substances (PFAS); a novel extraction phase she developed enabled PFAS determination down to the 1-ppt level. In a bioanalysis study, to evaluate the effectiveness of a smoking cessation product, she developed biocompatible SPME fibers that enabled the simultaneous extraction and chromatographic separation of nicotine and its major metabolites in rabbit plasma. This work enabled monitoring of the metabolic interconversion of nicotine after its release into the bloodstream. In all this work, Gionfriddo consistently and comprehensively investigated all aspects, from sample preparation conditions to the fundamental retention mechanisms and column comparisons.

Gionfriddo also developed a new tool for the evaluation of occupational pesticide exposure in the cannabis industry. Her comprehensive approach included an investigation for a diverse set of 80 pesticides that are regulated in cannabis, for which she developed a method to that measured these markers using SPME and liquid chromatography–tandem mass spectrometry (LC–MS/MS). The information from this study can aid the cannabis industry to build occupational safety and health programs for their workers.

Grinias: Working Toward a Capillary-Scale Parallel 2D-LC System with a Single Detector

The final speaker was Jim Grinias, an associate professor at Rowan University. After acknowledging Gionfriddo for her scientific leadership in sample preparation, he shifted to the focus of his work. He laid out his positive vision for the further development of LC×LC and stressed the need to increase speed and reduce the eluent volumes required for typical separations. In the latter context he referred to the zero-footprint goals embraced by analytical departments in the pharmaceutical industry in particular.

In his talk, presented his research toward a capillary-scale LC×LC–MS system, with parallel second-dimension columns, whose effluents would merge into the inlet of a single mass spectrometer. He was able to show very fast and highly robust (second dimension) analysis on capillary columns. For the final system, splitting the flow of the first-dimension column across multiple second-dimension columns will be a critical aspect. Grinias also noted the importance of data processing and visualization on the final LC×LC–MS data. He noticed that few 0.3-mm i.d. columns are currently available. So far, implementation of capillary LC is still limited in both 1D- and 2D-LC, but Grinias predicted a sharp increase in their application in the not-so-distant future.

Reference

1. Schoenmakers, P. J.; Billiet, H. A. H.; Tijssen, R.; de Galan, L. Gradient selection in RPLC. J. Chromatogr. A 1978, 149, 519–537. DOI: 10.1016/S0021-9673(00)81008-0