ISC 2026 - Day 3

Tuesday, September 8, brought another packed day to ISC 2026 in Prague, with the scientific program spanning everything from new stationary-phase technologies and food analysis to ion mobility, metabolomics, clinical applications, chiral separations, and lipidomics. The morning sessions highlighted advances in HILIC, SFC, multidimensional chromatography, LC-HRMS, and emerging approaches for studying complex biological and environmental samples. During the midday break, activity shifted to the exhibition and poster area, while the Job Fair and vendor seminars from Restek, Thermo Fisher Scientific, and Advanced Materials Technology connected scientific research with practical laboratory solutions and career opportunities. The afternoon continued with sessions on microfluidics, chip-based separations, technological innovations, and bioanalytical workflows, before artificial intelligence in separation science became the focus of a dedicated discussion session. The day concluded on a lighter note with the ISC Tube & Quiz, bringing participants together after another intensive day of science, technology, and networking.

Tuesday, September 8th

08:30–10:00 FUN 04: Advances in Stationary Phases

Chairs

• Gert Desmet, Belgium    
• Deirdre Cabooter, Belgium

ISC: ISC 2026 - Day 2: FUN 04 Advances in Stationary Phases chair Deirdre Cabooter

08:30 - 09:00 Non-porous nanoparticle columns for ultra-high-performance LC

• Ken Broeckhoven, Vrije Universiteit Brussel, Chemical Engineering CHIS, Brussels, Belgium.

The continuous demand for higher performance, high-throughput separations in the field of proteomics and metabolomics to resolve several hundred samples a day, requires further pushing the limits of separation performance in LC. Considering the upper limit of separation performance for a fully optimized system (Knox-Saleem limit), improved column packing quality (lower hmin), higher operating pressure (Pmax) and more permeability column structures (reduced flow resistance ) are the most straightforward options for further advancement. However, as the current limitations of 3D printing resolution currently do not allow to obtain perfectly ordered and open structures with sufficiently small feature size, the use of packed bed columns will remain the mainstay in LC for the foreseeable future. To achieve superior performance to contemporary LC systems (Pmax=1500bar, dp=1.5µm), lower particles sizes than those currently available, i.e. in the sub-micron range, will be required, especially when the separation of large (bio)molecules is considered due to their inherent mass transfer limitations (slow diffusion). In addition, the use of even higher operating pressures (2000 bar and above) than currently commercially available becomes highly advantageous. Based on the kinetic plot theory, for example, a 10-fold reduction in analysis time for N=100,000 can be achieved when using 500 nm nanoparticles operated at 3000 bar. An additional advantage of the use of nanoparticles is their tendency towards self-assembly into highly-ordered crystalline-like structures. However, a trade-off exists for these highly ordered, but dense packings (low porosity) due to their high flow resistance. The required very high operating pressures for use in short columns (<5-10 cm) also dictates the use of capillary column formats to alleviate the effects of viscous heating. In addition, a redesign of UHPLC instrumentation/set-up to eliminate extra-column-band broadening might be essential for preserving the efficiency due to the extreme small peak volume of these high-performance nanoparticle column.

To achieve this leap in separation performance, both the synthesis and packing of sub-µm sized non-porous particles in capillary columns was investigated. By tuning slurry (colloid) stability, optimizing packing procedures and packing conditions, highly ordered columns were obtained through evaporation packing. Using a novel particle immobilization technique, the bed structure was visualized and compared to its column flow characteristics. Successful grafting of a C18 stationary phase on a 750 nm nanoparticle column allowed proof-of-principle tests on a 4.5cm column with a BSA digest separation, showing good run-to-run repeatability (RSD of residence times < 0.2%).

ISC: ISC 2026 - Day 3: Ken Broeckhoven, Vrije Universiteit Brussel, Chemical Engineering CHIS, Brussels, Belgium.

9:00 - 9:20 MOF-based liquid chromatography: A new platform for polymer separation and purification

• Nobuhiko Hosono, The University of Tokyo, Department of Applied Chemistry- School of Engineering, Tokyo, Japan.

Molecular separation remains one of the most energy-intensive and technically challenging operations in the chemical industry, particularly in the case of polymeric compounds. Synthetic polymers are often produced as mixtures containing subtle structural variations arising from side reactions and incomplete conversions during polymerization. These include differences in terminal functionalities, chain topology, monomer connectivity, and local sequence irregularities/variations that are exceedingly difficult to resolve using conventional separation and analytical techniques.

To address this issue, our research group has developed a novel separation strategy based on metal–organic frameworks (MOFs), a class of crystalline porous materials constructed from metal nodes and organic linkers [1,2]. MOFs offer precisely tunable pore sizes and chemical environments, enabling selective interactions with target molecules. We have demonstrated that polymers can be adsorbed into MOF nanopores with high selectivity, facilitating the discrimination of minute structural differences that are otherwise difficult in standard chromatographic methods. This includes the successful separation of polymers differing only in end groups [3], backbone topology [4], regioregularity [5], or even single-unit isomeric variations [6,7].

These findings have led to the development of MOF-based liquid chromatography (LC) as a practical and scalable technique for polymer separation [2-6]. By packing MOF crystals into chromatographic columns and operating under flow conditions, we have achieved high-resolution separations of the structurally similar polymers. This LC-based approach enables precise tuning of interactions between polymer chains and MOF nanopores, facilitating efficient and structure-selective fractionation.

In this presentation, the mechanisms of selective polymer adsorption into MOF nanopores will be discussed, along with the broader prospects of MOF-based platforms in polymer separation, purification, and advanced analytical methodologies.

9:20 - 9:40 Governing selectivity in HILIC and mixed-mode column technology

• Alla Chernobrovkina, Lomonosov Moscow State University, Chemistry Department, Moscow, Russian Federation.

To meet the growing demand for fast, multi‑component analysis in complex industrial matrices, advanced chromatographic techniques are required. HILIC offers a robust solution for the separation of polar analytes, accommodating a wide range of molecular structures and physicochemical characteristics. The design of stationary phases with embedded hydrophobic domains introduces mixed‑mode interactions, combining different retention mechanisms. This innovation significantly broadens the scope of HILIC, making it possible to retain and resolve both polar and non‑polar analytes in a single run.

The present work establishes a systematic framework for evaluating the performance of HILIC and mixed-mode stationary phases and identifying the key factors that influence their chromatographic properties. A library of more than 100 home‑made adsorbents was synthesized to explore the impact of surface chemistry and substrate. Substrates included silica and poly(styrene-divinylbenzene), each modified with functional layers of varying structure: zwitterionic groups, amides, diols, macromolecules, and polymeric networks containing amines or acids. A combination of advanced surface engineering techniques was applied to create the layer architecture, including linker chemistry, hyperbranching, cross‑linking, and various hydrophilization techniques, enabling detailed analysis of structure–performance relationships.

Specific contribution of each functional fragment to chromatographic performance was quantified through detailed investigation of stationary phase properties using selectivity tests and model mixtures of compounds differing in hydrophilicity and charge. This approach revealed key trends for optimizing column performance and laid the foundation for the targeted task‑specific stationary phase design. Practical demonstrations of this targeted synthesis approach and regulating column technology will be presented.

9:40 - 10:00 Boosting polar resolution: Zwitterionic HILIC meets superficially porous particles

• Petra Lewits, Merck Life Sciences KGaA, Analytical Chemistry, Darmstadt, Germany.

Zwitterionic stationary phases represent one of the most effective HILIC chemistries for the retention and high-resolution separation of polar analytes. Their internally balanced positive and negative charges create strong dipolar interactions while maintaining overall electrical neutrality, enabling high selectivity, robust retention of highly polar compounds, and often improved peak shape for challenging matrices.

Extending grafted zwitterionic chemistries from fully porous particles (FPP) to superficially porous particles (SPP) can further enhance chromatographic performance. While FPP provide high surface area and strong interaction capacity, they may be associated with longer analysis times, increased susceptibility to fouling/clogging, and elevated backpressure, particularly for sub–2 µm formats. In contrast, sub–3 µm SPP can deliver FPP-like efficiency with lower backpressure, shorter run times, and improved packed-bed robustness, supporting more rugged operation without sacrificing resolution.

This presentation summarizes recent advances in grafted zwitterionic HILIC phases on superficially porous particles, with applications in HPLC and LC–MS, and evaluates how particle morphology influences chromatographic behavior. Comparative fundamental data are presented on equilibration, separation efficiency, volume and mass loadability, and injection-to-injection stability for FPP versus SPP formats. In addition, practical considerations related to column inertness are discussed, supported by representative application examples.

ISC: ISC 2026 - Day 3: Petra Lewits, Merck Life Sciences KGaA, Analytical Chemistry, Darmstadt, Germany.

08:30–10:00 BIO 04: Analysis of Food and Natural Products

Chairs

• Gertrud Morlock, Germany    
• Valérie Pichon, France

08:30 - 09:00 Comprehensive chemical characterization and neuroprotective evaluation of green extracts obtained from different natural matrices

• Alejandro Cifuentes, CSIC, Foodomics Lab- CIAL, Madrid, Spain.

This work presents a multi-analytical strategy to assess the neuroprotective potential of various natural green extracts enriched in bioactive compounds obtained from two plant sources: Pentaclethra macroloba (Willd.) Kuntze (known as pracaxi) and Ferula persica var latisecta. To obtain health-beneficial compounds, advanced environmentally friendly extraction techniques, such as compressed fluids including supercritical fluid extraction (SFE) and pressurized liquid extraction (PLE), were used. A comprehensive chemical characterization of the extracts was conducted using advanced analytical techniques, including gas chromatography and liquid chromatography coupled to high-resolution tandem mass spectrometry (GC-Q-TOF MS and LC-Q-TOF MS/MS). Over 220 compounds were identified in the pracaxi oil after SFE-CO₂ extraction, and new compounds including triterpenoid saponins and spermidine phenolamides, were tentatively identified in the pracaxi cake. The same analytical techniques were used to investigate the chemical composition of F. persica extracts, where 222 compounds belonging to 66 chemical subclasses (hydroxycoumarins, flavones, methoxyphenols, monoterpenoids, O-methylated flavonoids, and sesquiterpenoids) were identified regardless of the plant part (aerial or root). The neuroprotective potential of these extracts was investigated using in-vitro assays (ORAC, ROS, AChE, BChE, LOX), cell studies (SH-SY5Y) and blood brain barrier models (PAMPA-BBB and HBMEC-BBB).

09:00 - 09:20 Liquid chromatography strategies for metabolomic profiling and preparative purification of metabolites from complex plant matrices

• Natasha Damiana Spadafora, Italy

Plant systems are characterized by an extraordinary chemical diversity, where primary and secondary metabolites coexist, presenting wide dynamic ranges and highly heterogeneous physicochemical properties [1,2]. This intrinsic complexity poses significant analytical challenges, especially when the goal is to achieve comprehensive metabolomic coverage while preserving the integrity of structurally delicate compounds. In parallel, scalable preparative LC methods are employed to isolate metabolites of interest from complex extracts, enabling downstream structural elucidation, functional assays, and the generation of reference materials.

This contribution explores liquid chromatography strategies tailored to address the multifaceted nature of plant matrices. Emphasis is placed on maximizing the detection and characterization of secondary metabolites, which are key drivers of plant physiology, ecological interactions, and bioactivity. By integrating complementary chromatographic modes and optimizing selectivity for different constituents, the proposed approach enhances the depth and reliability of metabolomic profiling [3].
In addition, recent results in the isolation of high-revenue bioactive compounds from complex plant matrices by using green solvents and continuous purification techniques will be presented [4].
The combination of analytical and preparative chromatography approaches demonstrates how strategic method design can overcome matrix‑driven interferences, improve metabolite recovery, and support a more nuanced understanding of plant chemical space.

Overall, this work highlights the central role of chromatography in navigating the biochemical richness of botanical systems and underscores its value for advancing metabolomics and natural product research.

9:20 - 9:40 Chemical fingerprinting of shea kernels: Quality, antioxidants and health benefits

• Nikoline Juul Nielsen, University of Copenhagen, Dept. Plant and Environmental Sciences, Frederiksberg C, Denmark.

Kernels of the shea tree is a source of ingredients for plant-based foods and for medicinal and cosmetic skin care. Shea butter, constituting half of the kernel mass, is a sustainable and healthy solid fat increasingly in demand as alternative to dairy-based butter and palm oil. The sustainability of the shea tree is attributed to its wild-growing nature, its role in reducing soil degradation, its provision of essential income for female shea processors, and its considerable capacity for carbon sequestration.

Traditionally, kernel and butter quality have been defined by the ratio of desirable lipids (triacylglycerols) to their degradation products (mono and diacylglycerols and free fatty acids). However, future valorization will depend on mechanical processability at industrial scale (linked to cell integrity and polar membrane lipids), oxidative stability (antioxidants), and potential health promoting attributes such as cardio vascular benefits or antioxidative protection (phenolics, tocopherols, sterols and triterpenoids).

Here we present the comprehensive chemical fingerprinting of the shea kernel including: 60 neutral and polar lipids by MTBE: methanol extraction, NP-SPE and RPLC-HRMS [1], 50 polar secondary metabolites, primarily phenolics, by aqueous methanolic extraction, RPLC-HRMS [2] and RPLC×HILIC-cyclic IMS-HRMS [3], and 30 lipidic secondary metabolites including tocopherols, sterols and triterpenoids by hexane-extraction and GC-MS [4, 5].

NP-SPE allowed isolation of the hundredfold less abundant polar lipids from neutral lipids prior to LC-HRMS. Useful for identification of phenolics, RPLC×HILIC resulted in fine-structured group-type separation, while the IMS separation increased DIA mass spectral purity. The GC-MS platform provided one-run information on poor-quality indicator free fatty acids, beneficial antioxidants and main health promoting lipidic specialized metabolites. Of these, 18 triterpenoid esters could be accurately quantified using few standards combined with quantifier ion/TIC-corrected response factors.

Through extensive sampling across Burkina Faso and Ghana, combined with preservation experiments reflecting diverse West African processing practices, we investigated how the chemical fingerprint responds to key variables: region of origin, time from fruit drop to preservation, preservation method (smoking, parboiling, boiling, or soil pit fermentation), sun drying duration, shelling time, and post preservation storage duration. We show that the chemical profile of West African shea kernels is largely determined by the suite of events that constitute kernel preservation.

ISC: ISC 2026 - Day 3: Nikoline Juul Nielsen, University of Copenhagen, Dept. Plant and Environmental Sciences, Frederiksberg C, Denmark

9:40 - 10:00 From rapid to high-resolution separation of carbohydrates

• Jean-Pierre Chervet,  Antec Scientific, Research, Alphen a/d Rijn, Netherlands.

Recent advances in High-Performance Anion-Exchange Chromatography (HPAEC) column technology have enabled both ultrafast and ultra-high-resolution separations of carbohydrates. In particular, the development of columns packed with highly uniform monodisperse resin particles has significantly improved chromatographic efficiency while maintaining moderate system back pressure. In this work, separations were performed using SweetSep™ HPAEC columns from Antec Scientific, designed for comprehensive carbohydrate analyses ranging from monosaccharides to complex oligosaccharides and polysaccharides.

The columns are based on monodisperse crosslinked poly(divinylbenzene-co-ethylvinylbenzene) particles (3 and 5 µm) coated with quaternary amine–functionalized latex nanoparticles to provide strong anion-exchange capacity. The uniform particle size distribution and optimized surface functionalization yield reduced plate heights as low as 1.7 and column efficiencies exceeding 30,000 theoretical plates for 30 cm long columns packed with 5 µm particles, ideally suited for high-resolution profiling of complex oligo- and polysaccharides.

Applications will be presented for the separation of fructans and pectin-derived oligosaccharides, where previously unresolved oligosaccharide heterogeneities could be detected. Furthermore the columns allow for separation of oligosaccharides with degrees of polymerization (DP) up to 90, demonstrating its suitability for advanced glycomic and food analysis applications.

Another novelty is the use of HPAEC columns packed with monodisperse particles as small as 3 µm, allowing for ultrafast carbohydrate separations within a few hundred seconds, enabling high-throughput analysis. As an example, rapid determination of lactose in lactose-free labelled products will be shown, supporting quality control and authenticity verification to protect lactose-intolerant consumers.

Overall, the availably of short columns packed with 3 µm and long columns packed with 5 µm HPAEC particles, respectively, bridges ultrafast screening and ultra-high-resolution carbohydrate analysis, making them highly suitable for applications in food authentication, plant and biomass research, and glycoprotein characterization.

ISC: ISC 2026 - Day 3: Jean-Pierre Chervet, Antec Scientific, Research, Alphen Rijn, Netherlands.

08:30–10:00 HYP 04: Ion Mobility and Multidimensional Separation Approaches

Chairs

• Gérard Hopfgartner, Switzerland    
• Danila La Gioia, Italy

08:30 - 09:00 Rapid high-dimensional phenomics using LC-ion mobility-MS

• John McLean, Vanderbilt University, Department of Chemistry, Nashville, USA.

The human genome project is recognized as one of the most successful big science projects in modern history. One of the primary motivational underpinnings to undertake the HGP was to better understand what made us human and healthy - and how to use this code to improve the human condition by better understanding disease and potential treatment. While the frontiers of our knowledge expanded dramatically, we also uncovered profound biological complexity that we could not understand. This led to the current frontier in the measurement science of molecular phenomics, to catalog the broad-scale changes in the molecular inventory in cells, tissues, and biological fluids at a specific biological state, or in response to exposures and lifestyle choices. In phenomics, we seek to characterize the comprehensive molecular basis of biology (including DNA, RNA, proteins, lipids, carbohydrates, metabolites, and all of their nuances), in both space (e.g. at a cell, tissue, and organismal level) and time (e.g. healthy versus disease state). This places enormous demands on measurement technologies (including minimal sample preparation, fast measurements, high concentration dynamic range, low limits of detection, and high selectivity) and computational approaches to organize the millions of potential species present in vanishingly small spatial coordinates. The interplay between phenomic datasets and bioinformatics forms the nexus of translating phenomics data into actionable information and understanding.

Advances in computational biology rely heavily on the experimental capacity to make omics measurements, i.e. integrated proteomics, metabolomics, lipidomics, glycomics, among many others. Ion mobility-mass spectrometry (IM-MS) provides rapid (ms) gas-phase electrophoretic separations on the basis of molecular structure and is well suited for integration between rapid liquid chromatography (min) and mass spectrometry (us) detection techniques. This report will describe recent advances in LC-IM-MS integrated omics measurement strategies in the analyses of complex biological samples of interest in systems, synthetic, and chemical biology. These techniques will be highlighted through selected examples ranging from the creation of microfluidic human-organs-on-chip to replace animal testing in drug development workflows to probing the outcomes of fast genetic editing experiments (using CRISPR) in the optimization of synthetic biology for fine and commodity chemical production. While enormous challenges remain, the promise is immense – comprehensive diagnostics and predictive capabilities for health and medicine of importance to society and beyond.

ISC: ISC 2026 - Day 3: John McLean, Vanderbilt University, Department of Chemistry, Nashville, USA.

9:00 - 9:20 Unravelling tannin complexity using 2D-LC, ion mobility spectrometry and high resolution MS

• André de Villiers, Stellenbosch University, Department of Chemistry and Polymer Science, Stellenbosch, South Africa.

Tannins are large polyphenolic secondary plant metabolites which play an important role in plant protection. Their ability to bind proteins is exploited in various industrial applications (e.g. leather tanning), and together with their antioxidant activity is responsible for their importance in the human diet and as essential food and beverage constituents. Chemically, tannins can broadly be divided into two classes: hydrolysable tannins comprised of a central glucose core with gallic acid or hexahydroxydophenic acid groups, and condensed tannins, which are oligomeric and polymeric condensation products of flavanols. Both of these classes encompass very large numbers of structurally related compounds, which pose a significant analytical challenge. Indeed, the complete molecular characterisation of complex tannin mixtures is currently not possible, which hampers research into their occurrence and biological roles.

In this presentation, an up-to-date perspective on the potential of 1- and 2-dimensional liquid chromatography hyphenated to ion mobility spectrometry (IMS) and high resolution MS (HR-MS) for advanced tannins analysis will be presented. Examples of the benefits of the incorporation of IMS into 1D-LC-HR-MS workflows utilising both reversed phase LC (RP-LC) and hydrophilic interaction chromatography (HILIC) for hydrolysable and condensed tannins will be discussed. Furthermore, emphasis will be placed on the utility of comprehensive 2D-LC (LC×LC) for tannin analysis. It will be demonstrated how thorough optimisation of on-line LC×LC methods using an in-house developed predictive algorithmic approach enables the high-resolution separation of tannins. As application, the analysis of red wine tannins in a set of single-vineyard wines of consecutive vintages to investigate the evolution of tannins with wine age will be demonstrated. We will show how hyphenation of the optimised HILIC×RP-LC separation with IMS-HR-MS allows the detection of new wine tannin species. Finally, the application of the predictive software to derive optimal conditions for the off-line RP-LC×HILIC-IMS-HR-MS analysis of tannins will be discussed. This talk will illustrate how detailed information on tannin molecular composition may be obtained by judicious selection of optimised 1D and 2D-LC separation methods in combination with HR-MS and IMS.

ISC: ISC 2026 - Day 3: André de Villiers, Stellenbosch University, Department of Chemistry and Polymer Science, Stellenbosch, South Africa.

9:20 - 9:40 Design and optimization the interface of gas chromatography and photoionization ion mobility spectrometry aided by multi-physics field dynamic convolution

• Haiyang Li, Dalian Institute of Chemical Physics- CAS, Center for Advanced Mass Spectrometry, Dalian, China.

Photoionization ion mobility spectrometry (PIMS) hyphenated with gas chromatography (GC) exhibits outstanding selectivity and high sensitivity in the detection of trace volatile organic compounds (VOCs). The analytical performances of GC-PIMS are considerably affected by the interface between GC and the ionization chamber of PIMS. The sensitivity of PIMS depends on the good overlap of photoionization optical fields with the concentration distribution of effluent from GC, which is also affected by the gas flow in the ion mobility spectrometer. In this talk, a computational model based on multi-physics dynamic convolution will be introduced to evaluate the spatial concentration distribution of sample gas on the photoionization yield of PIMS. The simulation results indicate that lateral injection of GC capillary column into the gas flow of the ion mobility spectrometer could enhance the sensitivity by increasing the overlap of effluent and the VUV light in the ionization region of PIMS. The optimal capillary insert depth was increased when decreasing the carrier gas flow rate, which was confirmed by numerical simulations and experimental results. The optimal insert depth was 7.5 mm (the central axis of the ionization source) under conditions of a carrier gas flow rate of 15 mL·min-1, the limit of detection for aromatic compounds was reached 2.5 ppbv, achieving a 4-fold improvement of sensitivity over zero insertion. The GC-PIMS was tested to assess the VOCs emissions from spray-coated surfaces, and trace levels of toluene (45 μg·m-3) and xylene (60 μg·m-3) were still detectable even 14 days later, which highlights the potential of GC-PIMS apparatus for monitoring trace environmental pollutants.

ISC: ISC 2026 - Day 3: Haiyang Li, Dalian Institute of Chemical Physics- CAS, Center for Advanced Mass Spectrometry, Dalian, China.

9:40 - 10:00 Multidimensional chromatography and LC-HRMS for non-target screening of environmental contaminants: Strategies for quantification, prioritization, and identification

• Jan H Christensen, University of Copenhagen, Plant and Environmental Sciences, Copenhagen- Capital Region of Denmark, Denmark.

Environmental monitoring faces a growing analytical challenge as the number of chemicals released into the environment continues to increase. More than 100,000 industrial chemicals are currently registered for use in Europe, with over 1,000 new substances introduced annually, while only a small proportion have been comprehensively assessed for environmental persistence or toxicity. This expanding chemical space limits the effectiveness of traditional targeted monitoring approaches and has accelerated the adoption of non-target screening (NTS) using liquid chromatography coupled with high-resolution mass spectrometry (LC-HRMS).

However, the complexity of environmental matrices and the diversity of anthropogenic compounds place significant demands on both separation power and data interpretation. To address these challenges, this presentation explores the use of complementary multidimensional chromatographic platforms—including comprehensive two-dimensional gas chromatography (GC×GC-MS), two-dimensional liquid chromatography (LC×LC-MS), and liquid chromatography–supercritical fluid chromatography (LC×SFC-MS), combined with LC-HRMS detection. These multidimensional approaches increase peak capacity and reduce co-elution, enabling improved characterization of chemically diverse contaminant mixtures that often remain unresolved using one-dimensional separations.

Because NTS datasets typically contain thousands of detected features, prioritization strategies are essential for identifying environmentally relevant compounds. Several complementary prioritization approaches will be presented, including suspect screening, reliability-based filtering, chemistry-driven prioritization, process-related prioritization, endpoint-driven strategies linking chemical fingerprints to biological effects through virtual effect-directed analysis (vEDA), predictive prioritization using quantitative structure–property and toxicity relationships (QSPR), and image-based prioritization using pixel- and tile-based analysis of chromatographic datasets.

In addition to compound discovery, quantitative interpretation remains one of the major limitations of NTS workflows. Recent advances in response-factor prediction and machine-learning-based quantification approaches offer promising routes toward improved semi-quantitative and quantitative assessment without the need for extensive libraries of reference standards.

Examples from environmental applications, including wastewater, surface water, and sediment samples, demonstrate how the combination of multidimensional chromatography, LC-HRMS, and systematic prioritization enables the detection and identification of contaminants of emerging concern (CECs) and provides a stronger analytical basis for environmental risk assessment and regulatory decision-making.

Together, these developments highlight how advanced separation techniques combined with modern data analysis strategies can significantly extend the analytical capabilities of NTS for environmental monitoring and chemical risk evaluation.

10:00–10:30 Coffee Break

10:30–12:00 FUN 05: Separations with Supercritical and Subcritical Fluids

Chairs

• Isabelle François, Belgium    
• Kateřina Plachká, Czech Republic

ISC: ISC 2026 - Day 3: FUN 05 Separations with Supercritical and Subcritical Fluids chairs (Isabelle François, Belgium, Kateřina Plachká, Czech Republic)

10:30 - 11:00 One fluid, many molecules: Expanding SFC across the chemical space of natural products

• Caroline West,  University of Orleans, Institute of Organic and Analytical Chemistry, Orléans, France.

In the recent years, supercritical fluid chromatography (SFC) has been confirmed as a powerful complementary technique to liquid chromatography (LC) for the analysis of complex mixtures, especially when it is hyphenated to mass spectrometry (MS). Using supercritical CO₂ as the primary mobile phase, SFC combines low viscosity and high diffusivity with tunable solvent strength through organic co-solvent and additives. These physicochemical properties enable fast separations, high efficiency, and reduced solvent consumption compared with conventional liquid chromatography. In addition, the use of diverse achiral and chiral stationary phases allows fine control of selectivity across a wide range of chemical diversity.

These characteristics make SFC particularly attractive for the analysis of natural products, which encompass a chemically diverse array of metabolites ranging from highly lipophilic (triacylglycerols, terpenoids, alkaloids, furocoumarins…) to highly polar compounds, including ionizable ones (iridoids, glycosylated flavonoids, anthocyanins, amino acids…). Furthermore, the compatibility of SFC with supercritical fluid extraction enables the development of integrated workflows for natural product extraction and analysis.

In this presentation, we illustrate how methodological and technological adaptations in SFC can be used to address the analytical challenges posed by different classes of natural products. Through selected case studies, strategies for the analysis of non-polar, polar, and ionizable compounds will be discussed, demonstrating how SFC can be tailored to accommodate the remarkable chemical diversity encountered in natural product research.

ISC: ISC 2026 - Day 3: Caroline West, University of Orleans, Institute of Organic and Analytical Chemistry, Orléans, France.

11:00 - 11:20 From molecular structure to SFC Selectivity: AI-guided mapping of Retention mechanisms

• Jean Christophe Garrigues, CNRS - Toulouse University, SOFTMAT laboratory, Toulouse, France.

Supercritical fluid chromatography (SFC) has gained importance in pharmaceutical, cosmetic and environmental analysis, particularly when coupled with mass spectrometry [1]. However, SFC method development remains complex due to optimisation of mobile phases. Interactions between modifiers and residual silanol groups on stationary phases may induce progressive selectivity changes through silyl ether formation. The use of appropriate conditions, including the addition of water or ammonia in methanol, can improve stationary phase stability.

In previous work, we investigated selectivity variations in gradient mode across 10 stationary phases and 107 analytes [2,3,4]. Earlier studies mainly focused on eluted compounds, while non-eluted analytes were not considered, although they ranged from 6 to 41 depending on the conditions. In this study, to better understand elution capability and retention mechanisms in SFC, we developed an artificial intelligence (AI)-driven-molecular-modelling strategy structured in two stages. The first stage consisted of predicting analyte elution across stationary phases under three mobile phase conditions. The second stage focused on modelling gradient retention mechanisms to estimate representative retention factors. All analytes were modelled in Molecular Operating Environment in multiple charge states corresponding to apparent pH between 4-8 under MeOH, MeOH-10mM NH₃ and MeOH-2% H₂O conditions. Conformational analysis was performed, followed by calculation of 500 two-dimensional and three-dimensional descriptors per conformer, representing nearly 5000 descriptors per analyte. The dataset was processed using generative and predictive AI-approaches. Genetic algorithms automatically optimised neural network architectures to model elution probability and numerical retention factors. In this framework, genetic algorithms inspired by evolutionary processes iteratively evolve populations of candidate solutions encoded as chromosomes. Here, chromosomes represent subsets of descriptors, including conformational, interatomic distances and descriptors reflecting protonation states. Cross-validation demonstrated predictive performance with r² training > 0.8 and r² test > 0.6. This approach enables prediction, from molecular structure alone, of elution probability, theoretical retention factor, optimal stationary phase and relevant gradient conditions. The integration of genetic algorithms and neural networks provides a detailed mapping of stationary phases within an expanded experimental space, which will be compared with traditional LSER models [5], establishing a predictive framework for robust SFC method development.

11:20 - 11:40 Comprehensive SFC and NPLC chiral screening and optimization of tacrine‑based metabolites: A step toward a safe, highly active drug

• Ondrej Horacek, Farmaceutická fakulta v Hradci Králové UK, Department of Pharmaceutical Chemistry and Pharmaceutical analysis, Hradec Králové, Czech Republic.

Supercritical fluid chromatography (SFC) is widely used in pharmaceutical workflows, particularly for rapid and efficient chiral separations, due to straightforward method development and typically shorter analysis times compared with liquid chromatography. Because chiral resolution is crucial in early drug discovery, SFC offers a fast and robust platform for separating racemic drug candidates, including tacrine‑based metabolites. Tacrine is a commonly explored structural motif in the search for new treatments for Alzheimer’s disease, and structural modifications typically aim to preserve its cholinesterase‑inhibitory activity while reducing undesirable effects. Recent studies have shown that several racemic tacrine‑based metabolites exhibit reduced toxicity while retaining inhibitory potency. However, their enantiomers may differ in both toxicity and cholinesterase‑inhibitory activity, and currently, no enantioselective synthetic routes or established chiral separation methods are available. Therefore, effective chiral methods are essential for evaluating enantiomeric purity and isolating individual enantiomers from racemates.

In this study, we carried out comprehensive chiral screening of chiral stationary phases using reversed-phase liquid chromatography (RPLC), normal-phase liquid chromatography (NPLC), and SFC, followed by detailed optimization of the mobile-phase composition and column-oven temperature. Chiral separations were more successful in SFC and NPLC compared to RPLC, achieving baseline separation of 9 out of 10 racemic metabolites using the chiral column Whelk O1. This chiral stationary phase was further examined through molecular modeling and computational studies. The role of hydrogen bonding and π–hydrogen bonding in the interactions between the selectand and the selector was emphasized, and the position of the hydroxyl group of the selectand was found to have the most significant impact on chiral recognition.

The methods developed herein were used for semipreparative isolation of enantiomers of the most active racemate, 9-amino-6-chloro-1,2,3,4-tetrahydroacridin-1-ol. Thanks to chiral methods in NPLC and SFC, the absolute configurations of the enantiomers were determined by single-crystal X-ray diffraction. The bioactivity assays showed that the R-enantiomer was the eutomer, with higher acetylcholinesterase-inhibitory potency than the racemate and tacrine. These findings were enabled by chiral SFC and NPLC methods, highlighting their significance in the early stages of drug discovery.

11:40 - 12:00 Accelerating forensic analysis: A rapid SFC–MS workflow for novel regioisomeric fluorofentanyl derivatives and related compounds

• Jennifer Field, Shimadzu UK, Technical, Milton Keynes, United Kingdom.

The proliferation of synthetic opioids, notably fentanyl analogues, has intensified public health risks due to wide potency variation among regioisomers. Ortho-, meta- and para-fluorofentanyl exemplify this challenge: meta‑ and para‑isomers exhibit roughly 0.5× potency relative to parent fluorofentanyl, whereas the ortho‑isomer is 2.5–3.5× more potent. Reliable separation and unequivocal identification of these regioisomeric species are therefore critical in forensic, clinical and toxicological contexts to prevent misidentification and erroneous interpretation.

This study presents a novel, sustainability-focused analytical workflow for rapid, unambiguous identification of 18 fluorofentanyl structural isomers and related analogues. Central to this work is an SFC–MS/MS method developed as a fast, targeted, and greener alternative to conventional LC-MS. Method development included systematic screening of stationary phase chemistries and organic modifiers to maximise speed and isomer resolution while minimising organic solvent consumption. Targeted Multiple Reaction Monitoring (MRM) transitions, combined with chromatographic separation, provided unambiguous confirmation of analyte identity.

This approach was critically evaluated against two complementary chromatographic strategies - high‑pH reversed phase UHPLC, and hydrophilic interaction liquid chromatography (HILIC)—to optimise selectivity across diverse regioisomeric families.

The SFC-MS approach was assessed on seized street samples, demonstrating the SFC workflow’s ability to rapidly and sustainably confirm the presence and identity of illegal fluorofentanyl analogues in real‑world matrices. By delivering faster analysis times, reduced organic solvent use, and robust isomer discrimination, the proposed SFC–MS approach advances forensic toxicology toward higher throughput and improved environmental performance without sacrificing analytical confidence.

10:30–12:00 BIO 05: Metabolomics

Chairs

• Michael Lämmerhofer, Germany    
• Tomáš Pluskal, Czech Republic

ISC: ISC 2026 - Day 3: BIO 05 Metabolomics chairs (Michael Lämmerhofer, Germany and Tomáš Pluskal, Czech Republic)

10:30 - 11:00 From metabolomics to exposomics: Examining health from small molecules based on liquid chromatography-mass spectrometry

• Guowang Xu, Dalian Institute of Chemical Physics- Chinese Academy of Sciences, Division of Biotechnology, Dalian, China.

The intricate relationship between human health and disease is profoundly reflected in the profile of small molecules within the body, which encompasses endogenous metabolites, microbial metabolic products, and exogenous exposures. Advances in high-resolution mass spectrometry (HRMS) enable us to probe this complex chemical space. However, metabolomics faces the significant challenge of "dark matter"-the vast majority of spectral features that remain unannotated. To address this, we have developed advanced computational tools and databases, such as MetExDB and novel structural association networks (SGMN, E-SGMN), which dramatically improve annotation coverage and accuracy by making use of molecular structure and retention/fragment rules. A typical clinical application example is in non-alcoholic fatty liver disease (NAFLD) intervened by resistant starch.Extending from the endogenous metabolome to the exogenous exposome is crucial for a holistic understanding of health. Our key methodological advancements include the development of two-dimensional (2D) LC-HRMS method for broad-spectrum screening of over 1,200 exogenous chemicals. Based on the untargeted screening results, GC-MS/MS and LC-MS/MS targeted methods were developed for accuracy quantitation. Population-based studies with 10,000 persons revealed significant regional, age, and gender differences in chemical exposure levels. The integration of exposomics with metabolomics and epidemiological data provides a systems-level understanding of disease mechanisms. Further, to improve the analytical throughput, two novel methods have been developed for the simultaneous acquisition of metabolome in HRMS and exposome in multiple reaction monitoring (MRM) modes in one injection. In conclusion, the deep integration of metabolomics and exposomics, empowered by advanced analytical platforms, multi-omics data, and large cohorts, is essential for elucidating disease mechanisms, it is very helpful for precision medicine and effective public health interventions.

ISC: ISC 2026 - Day 3: Guowang Xu, Dalian Institute of Chemical Physics- Chinese Academy of Sciences, Division of Biotechnology, Dalian, China.

11:00 - 11:20 Exploring targeted polar metabolomics by Porous Graphitic Carbon (PGC)-based LC-HRMS: A class-focused comparison with HILIC and C18 approaches

• Danila La Gioia,  University of Salerno, Department of Pharmacy, Fisciano- Salerno, Italy.

The separation of extremely polar metabolites remains a significant analytical challenge in the field of LC-MS–based metabolomics. Metabolite classes such as TCA cycle intermediates, polyamines, sugar phosphates, nucleotides, cofactors, and neurotransmitters are biologically relevant yet remain critical for C18 phases. Whilst HILIC-based workflows are successfully employed for the analysis of polar metabolome providing increased coverage, several challenges arise from ionic species, isomers and phosphorylated metabolites, that require extensive method development, resulting often in careful selection of buffers for acidic for basic metabolites, and furthermore, lack of repeatability and stability over time. Thus, there is still interest for orthogonal approaches for targeted analysis of key polar metabolites.

In this work, we performed benchmarking of a porous graphitic carbon (PGC) column as a complementary stationary phase to Z-HILIC and C18 for highly polar metabolites. Study design was based on selection of 90 metabolites in the low logP range, covering the most hydrophilic regions of the metabolome. The different stationary phases were evaluated under acidic, neutral, and basic mobile phase conditions.

The chromatographic performance was evaluated in terms of retention, resolution, peak symmetry, and tailing, with a specific focus on class-dependent selectivity. As expected PGC exhibited increased retention with respect to C18 and reduced co-elution for the most hydrophilic metabolites with simple mobile phases composed of water and acetonitrile with 0.1%HCOOH v/v. CCritically, organicacids, sugar phosphates and nucleotides were successfully retained, also providing isomer separation capability. Compared to Z-HILIC, a different behavior was obtained, resulting in enhanced response for class specific compounds, such as TCA intermediates, polyamines, polyols and glycol-derivates, thus highlighting limitations of HILIC-based approaches for specific metabolite classes.

The optimized conditions were applied for a broadly targeted metabolomics using parallel reaction monitoring (PRM) by hyphenation with a quadrupole-Orbitrap analyzer, for the analysis of class-specific compounds in liver and cellular extracts prepared using different protocols: trichloroacetic acid (TCA) precipitation, Folch and solid phase extraction (SPE). Fold-change at the metabolite subclass level revealed that TCA extraction with 0.1%HCOOH phases on PGC resulted in the comprehensive coverage of extremely polar metabolites, including polyamines, sugar phosphates, and organic acids and orthogonal coverage to Z-HILIC. Lastly, high temperature separation was explored in PGC with potential reduction of acetonitrile employment.

Overall, these results pave the way to implementation of PGC for class-driven and application-oriented targeted metabolomics as complementary and easier approach to HILIC for accessing the most hydrophilic regions of the metabolome.

ISC: ISC 2026 - Day 3: Danila La Gioia, University of Salerno, Department of Pharmacy, Fisciano- Salerno, Italy.

11:20 - 11:40 Towards plant pan-metabolome: Metabolic diversity of 3000 plant species

• Markéta Macho, Max Planck Institute of Molecular Plant Physiology, Laboratory of Central Metabolism, Potsdam, Germany.

Plants are well-known for their ability to synthesize a large spectrum of small molecules known as secondary metabolites. These compounds play important roles in plant growth and development and mediate interactions with the environment, including responses to biotic and abiotic stress. As a result, metabolite levels vary across tissues, developmental stages, environmental conditions, and ecological contexts. However, this variation explains only part of a plant’s chemical profile, as the ability to produce specific compounds also depends on the presence of corresponding biosynthetic pathways encoded in the genome and regulated at multiple levels, including transcriptomic and environmental regulation. Understanding why and how plant phytochemical diversity evolved, how it is maintained, and which ecological and phylogenetic factors shape it remains a central question in plant chemical ecology.

To date, over 300,000 plant metabolites have been reported, and estimates suggest the total number may reach up to 15 million. Yet, the full extent of plant chemical diversity remains largely unexplored, partly due to the limited chemical characterization of most plant species. Of the ~400,000 accepted species, only a fraction has been studied in detail, with research biased toward medicinal, crop, or industrial plants. Many plant compounds have valuable properties, ranging from nutritional and pesticidal to pharmaceutical, and a substantial proportion of modern drugs are plant-derived. Expanding our knowledge of plant secondary metabolism therefore provides opportunities for discovering novel bioactive compounds, biopesticides, or improving crop resilience.

In this study, we aimed to broaden the understanding of plant chemical diversity by sampling a wide spectrum of previously underexplored plant taxa across the plant kingdom. In collaboration with several international botanical gardens and institutes, we collected over 4,400 samples, primarily leaf tissue, from nearly 3,000 plant species representing 14 major classes of the phylum Embryophyta, including over 280 families in 85 orders. The dataset includes mosses, ferns, conifers, monocots, and eudicots, and covers rare and endemic species as well as plants with traditional uses in food and medicine. Samples were analysed using untargeted UPLC-MS/MS metabolomic profiling, resulting in a dataset comprising over 6,000 MS/MS spectra.

This study provides one of the most extensive comparative plant metabolomics datasets to date and enables the investigation of phylogenetic patterns in specialized metabolite production, the links between ecology and chemical function, and the identification of potentially bioactive compounds. Ultimately, this work contributes to a more comprehensive understanding of plant chemical diversity and supports biodiversity research and conservation efforts.

11:40 - 12:00 Tracing microbial rosmarinic acid metabolism in a colon model applying a validated UHPLC-qTOF method and 13C-labelling

• Laura Falk, University of Bonn, Institute of Nutritional and Food Sciences, Bonn, Germany.

Rosmarinic acid (RA) is a bioactive polyphenol associated with various health-promoting properties, including antiviral, antibacterial, anti-inflammatory, and antioxidant effects [1]. A detailed understanding of its metabolism is essential to elucidate its mode of action in the human organism. However, previous studies have been limited by insufficient substrate traceability, as several reported metabolites may already be present in the food matrix or originate from bacterial primary metabolism, such as 4-hydroxyphenylacetic acid [2–4]. Consequently, these compounds cannot always be unequivocally assigned as RA-derived metabolites. In addition, only incomplete recovery of the administered substrate in the form of metabolites has been reported after gastrointestinal passage and in vitro experiments [2]. Here, we present an efficient UHPLC-qTOF method for the analysis of RA and its microbial metabolites. The approach is based on a small-scale in vitro colon model consisting of a complex microbiota isolated from human feces. The analytical method enables the detection and quantification of 23 small molecular phenolic compounds and was optimized and validated for application in this fermentation system [5]. After optimization and validation, the system allows precise, time-resolved quantification of microbially formed RA metabolites while accounting for matrix effects. Our experiments demonstrate that distinguishing RA-derived metabolites by 13C-labelling is essential for reliable metabolite assignment in complex fermentation matrices like colon fermentation systems.

ISC: ISC 2026 - Day 3: Laura Falk, University of Bonn, Institute of Nutritional and Food Sciences, Bonn, Germany.

10:30–12:00 HYP 05: Clinical Analysis

Chairs

• Mario Thevis, Germany    
• Steven R. H. Wilson, Norway

10:30 - 11:00 Enantioselective and multi-dimensional HPLC analysis of amino acids and related compounds for the screening of novel biomarkers and functional molecules

• K. Hamase, Kyushu University, Graduate School of Pharmaceutical Sciences, Fukuoka, Japan.

Higher animals predominantly utilize L-amino acids, and the stereoisomers, D-amino acids, are considered to be rare in the tissues and physiological fluids. However, by the recent advancement of enantioselective and sensitive analytical methodologies, a variety of D-amino acids were found in mammals including humans. Currently, several dipeptides including D-amino acids have also been discovered in mammals. Although these D-amino acids and D-form dipeptides are increasingly gathering attention as new biologically active substances and/or biomarkers, their determination is frequently interfered with major L-amino acids and numerous intrinsic L-form peptides. Therefore, highly sensitive and selective analytical methods are essential, and the enantioselective multi-dimensional HPLC systems are the straightforward options. In the present study, the amino acids and peptides were derivatized with 4-fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F), and the NBD-amino acids and dipeptides were separated by two-dimensional (2D) LC-MS/MS and three-dimensional (3D) LC systems. For the first dimension of the LC separation, the reversed-phase mode was frequently integrated, where the target amino acids and peptides were separated by their hydrophobicity. The final dimension of the LC separation is the enantioselective mode, where the chiral discrimination of amino acids and peptides was performed. Concerning the 3D-LC system, an additional separation mode (anion-exchange or mixed mode) different from those of first and third dimensions was integrated; the target compounds were separated as their scalemic mixtures using the first and second dimensions. In the last chiral recognition dimension, an enantioselective column was integrated. By using Pirkle type columns having N-(3,5-dinitrophenylaminocarbonyl)-L-leucine, L-2-aminobutyric acid and L-norvaline as their chiral selectors, all of the proteinogenic amino acids plus allo-forms were enantiomerically separated. The present 2D LC-MS/MS and 3D-LC systems could be applied to clinical, biological and food/beverage samples and screening of novel biomarkers and functional molecules is continuously ongoing.

11:00 - 11:20 From organoids to exposure markers: LC-MS approaches for PFAS-related health effects

• Hanne Røberg-Larsen, University of Oslo, Department of Chemistry, Oslo, Norway.

Per- and polyfluoroalkyl substances (PFAS) are known as “forever chemicals” and may adversely affect human health. Although several PFAS are now banned or restricted, a deeper understanding of their biological impact remains incomplete. These compounds are present in a wide range of consumer products and consequently accumulate in human and animal tissue. Studying their health effects in traditional animal models is challenging due to difficulties in obtaining true PFAS-free control samples, limited environmental control, and physiological differences from humans.

New approach methodologies (NAM), including organoids and microphysiological systems (MPS), are emerging as powerful human-relevant tools for replacing animal models in toxicology and disease research. Moreover, these systems enable controlled exposure studies using human cells within physiologically relevant in vitro environment. However, organoids can be inherently heterogeneous and limited in size (e.g., liver organoids contain approximately 20 000 cells), which challenges established analytical strategies for assessing responses to e.g., drugs or environmental pollutants.

To meet these challenges, we have developed liquid chromatography-mass spectrometry (LC-MS)-based analytical workflows tailored specifically to MPS- and organoid-derived samples, with emphasis on high sensitivity, high throughput, and high reproducibility. Our analytical toolbox includes validated sterol analysis in single organoids and gastruloids using optimized derivatization (guided by experimental design)1, FDA-validated assays for small-molecule drugs and metabolites2, both with automated sample clean-up, and LC-MS strategies for quantifying stress and exposure markers following PFAS treatment. These target workflows will be integrated with robust inter-sample normalization procedures and global metabolomics and proteomics to provide deeper insight into the biological effects of selected PFAS in human-relevant in vitro systems.

11:20 - 11:40 Comprehensive plasma lipidomics by RP-UHPLC-MS/MS reveals lipid alterations in paediatric cancers

• Laiyu Zhao, University of Pardubice, Department of Analytical Chemistry, Pardubice, Czech Republic.

Paediatric cancer survivors frequently experience late adverse effects affecting multiple organs and physiological systems, largely attributed to premature cellular ageing induced by chemotherapy, radiotherapy or their combination. Alterations in membrane lipid dynamics are known to occur in cancer cells, suggesting that lipidomic markers may provide insights into cancer-related biological processes and potentially predict the treatment effects in survivors. Therefore, there is an urgent need for systematic studies to support targeted surveillance and early identification of individuals at high risk of late complications.

In this study, an optimised reversed-phase ultrahigh performance liquid chromatography tandem mass spectrometry (RP-UHPLC/MS/MS) method based on multiple reaction monitoring (MRM) was developed for the quantification of 498 lipid species at the fatty acyl/alkyl level across 26 lipid subclasses. Chromatographic separation was achieved using an Agilent 1290 Infinity UHPLC system equipped with a BEH C18 column (150 × 2.1 mm, 1.7 μm). Lipid initial identification was supported by the retention dependences in logical series for particular classes.

Plasma samples were collected from 32 paediatric cancer patients (≤18 years), 190 survivors (>19 years), and 56 healthy controls. The cohort included multiple cancer types, including lymphomas, sarcomas, germ cell tumours, central nervous system (CNS) tumours and others. Clear group separation between paediatric patients and age-matched healthy controls was observed in both unsupervised and supervised statistical analyses. Lipid alterations were dominated by the downregulation of phospholipids and sphingolipids. Specifically, phospholipid subclasses (e.g., LPC, PE, PE O, and PE P) showed significant downregulation in species containing polyunsaturated fatty acyl chains, particularly 18:2 and 22:6. In addition, sphingolipids (e.g., SM, Cer, SHexCer) containing 18:1 or 18:2 sphingoid bases with very long-chain saturated fatty acyls (≥C22) also displayed downregulation. In contrast, comparisons between survivors and adult healthy controls showed no clear separation. Neither gender nor the time since treatment influenced the lipidomic patterns. Limited predictive performance was also observed across cancer types, with only minor separation found in sarcoma survivors.
This work provides comprehensive quantitative coverage of plasma lipid subclasses in paediatric cancer patients and survivors. Further studies could go deeper to identify lipids at a more detailed structural level to elucidate lipid alterations in cancer survivors.

This work was supported by the ERC Adv grant No. 101095860 (European Research Council) and Project no. CZ.02.01.01/00/22_008/0004644 - Saving lives through research in early cancer detection and prevention: Molecular, genomic and societal factors, (SALVAGE) sponsored by the Ministry of Education, Youth and Sports, Czech Republic.

11:40 - 12:00 Gut microbiota-mediated transformation of bisphenol pollutants and associated metabolic disruptions

• Guangbo Qu, Research Center for Eco-Environmental Sciences- Chinese Academy of Sciences, State Key Laborotary of Envrionmental Chemistry and Environmental Toxicology, Beijing, China.

The gut microbiota is increasingly recognized as an important factor affecting the transformation of environmental pollutants, yet its role in the biotransformation of structurally diverse bisphenol contaminants remains insufficiently understood. This work presents a liquid chromatography coupled with mass spectrometry (LC–MS) based analytical strategy for characterizing gut microbiota-mediated transformation of representative bisphenol pollutants and evaluating the resulting toxicity changes.

A high-resolution LC–MS workflow was used to investigate the transformation of tetrabromobisphenol A (TBBPA) and related analogs, as well as a broader panel of 21 bisphenols, by representative human gut bacterial strains under anaerobic conditions. The combination of nontarget screening, kinetic analysis, and structural characterization enabled the identification of multiple previously unrecognized transformation products and revealed clear species-specific differences in pollutant metabolism.

For TBBPA and its analogs, nine rhamnosylated or debrominated transformation products were detected, and rhamnosylation was identified as a distinctive microbial transformation pathway. Kinetic analysis further indicated rapid bioaccumulation and intracellular transformation in Clostridium manihotivorum. For the broader bisphenol panel, bisphenol A-glycerol methacrylate (Bis-GMA) showed the most pronounced microbial transformation. Eight species-specific products, including acetylated, ester-hydrolyzed, and palmitoylated derivatives, were identified across different bacterial strains. In vivo experiments further confirmed the formation of bisphenol A bis (2,3-dihydroxypropyl) ether (Bis-HPPP), consistent with the in vitro results.

Beyond product identification, the analytical workflow was combined with mechanistic and toxicological evaluation. Genomics, molecular docking, and dynamic metabolomics supported the identification of a candidate rhamnosyltransferase involved in TBBPA transformation. In the Bis-GMA case, comparison of the parent compound and its microbial transformation product showed that Bis-HPPP had lower cytotoxicity, whereas the parent compound caused more severe effects on human intestinal organoids, including changes in apoptosis, proliferation, and epithelial integrity.

Overall, this research establishes a LC–MS based analytical platform for resolving gut microbiota-mediated transformation of environmental bisphenol pollutants. It highlights the value of LC–MS in identifying previously unrecognized metabolic pathways, characterizing transformation products, and supporting exposure and health risk assessment.

12:00–14:30 Lunch – Exhibition – Poster Sessions

12:30 - 14:00 Industry Session: Activity 3: Job Fair

13:00 - 14:00 Industry Session: Restek vendor seminar - Virtual chromatographic modelling software for optimising separation of isobaric compounds in GC-MS methods

There is a constant balance for high-throughput laboratories between method run times for large panel analyses and resolution of analytes across these panels to allow sufficient data quality. Developing such methods is often time consuming and expensive for laboratories, as dedicated resources are required to test methods iteratively. Chromatographic modelling tools are becoming increasingly popular for method setup and refinement in high-throughput laboratories looking to minimise instrument and analyst downtime. These tools traditionally offer column phase and method condition suggestions based on the separation of a list of analytes, which can then be modified to improve specific separations as necessary.

When developing gas chromatography (GC) methods, shortening cycle time often comes at the cost of compounds co-eluting, with the expectation that a mass selective detector, such as a mass spectrometer (MS) will allow resolution through ion-selective quantitation. An issue arises when the method attempts to analyse isobaric compounds, which cannot be distinguished by MS, and as a result labs must rely upon physical separation via column phase chemistry.
As GC-MS techniques continue to be adopted by laboratories, there is a need for more tools that consider mass data for analytes when optimising large panel separations. The virtual chromatographic modelling tool used in this study is built with mass spectrum data for compounds included in libraries. When operating in “MS mode”, the software identifies isobaric compounds and prioritises their separation when making recommendations regarding column phase and method conditions. For those using non-mass selective detection methods, the tool is easily toggled to “FID mode” for users interested in resolving all compounds, regardless of specific mass, allowing for flexibility in instrumentation. Results help labs save time and resources in method development and refinement by considering the instrumentation involved in the model.

13:00 - 14:00 Industry Session: Thermo Fisher Scientific vendor seminar - ASOs, ADCs, and other therapeutics: Advanced characterization with novel reversed-phase columns and integrated LC–MS platform

Biotherapeutics are becoming increasingly structurally diverse, placing greater demands on analytical platforms that can support efficient development and regulatory readiness. Reversed-phase chromatography (RPC) remains a widely adopted and adaptable technique, supporting applications that range from oligonucleotide analysis to detailed protein characterization and post-translational modification assessment.

This seminar explores the integration of a macroporous, monodisperse reversed-phase column with a next-generation bioinert LC system for the analysis of two emerging classes of biotherapeutics. Antisense oligonucleotides were examined under optimized chromatographic conditions, using a design-of-experiments strategy to evaluate how ion-pairing reagents influence both LC separation and downstream MS performance. The platform was further applied to antibody–drug conjugates, combining RPC with Orbitrap mass spectrometry to support multi-level characterization. Intact, sub-unit, and bottom-up analyses enabled drug-to-antibody ratio determination alongside detailed PTM profiling — including glycosylation, oxidation, and deamidation — under native and stress conditions. Collectively, these studies demonstrate a streamlined and adaptable workflow for comprehensive biotherapeutic characterization.

13:00 - 14:00 Industry Session: Advanced Materials Technology vendor seminar - Advantages of superficially porous particle columns for HPLC and LC-MS

Superficially porous particle (SPP) columns have existed in one form or another since the late 1960s. However, the benefits of the particle with the solid silica core surrounded by the porous silica shell were not realized for small molecules until 2006 when the first sub-3-µm superficially porous particle columns were introduced. This particle technology enabled efficiencies similar to sub-2-µm fully porous particles (FPPs) without the need for ultra-high pressure capable instrumentation. As a result of the high efficiencies and lower back pressure, the columns may be run at faster flow rates to enable high throughput separations. Additionally, the SPP columns are rugged and offer long column lifetimes. This presentation will highlight comparisons between SPP columns and FPP columns for separations of pharmaceuticals, pesticides, and oligonucleotides.

14:30–16:00 FUN 06: Chiral Separations

Chairs

• Bezhan Chankvetadze, Georgia    
• Katarína Maráková, Slovakia

ISC: ISC 2026 - Day 3: FUN 06 Chiral Separations chairs (Bezhan Chankvetadze, Georgia and Katarína Maráková, Slovakia)

14:30 - 15:00 Enantiomers and other isomers in bioanalytical applications: Still an analytical challenge?

• Michael Lämmerhofer, University of Tuebingen-, Institute of Pharmaceutical Sciences, Tuebingen, Germany.

Biological systems are built from a myriad of constitutional and stereoisomers. Without methods that can distinguish these isomers, the true structural complexity of metabolites, lipids, peptides and drugs remains hidden. Modern analytical chemists therefore need a multidimensional toolbox that combines chromatographic separations (conventional LC, chiral stationary phases (CSPs), two-dimensional LC), ion‑mobility spectrometry (IMS; orthogonal gas‑phase separation), orthogonal fragmentation modalities (e.g. collisional-induced dissociation (CID) and electron-activated dissociation (EAD) to generate complementary product‑ion spectra, targeted derivatization with chemoselective tags or chiral derivatization. Together these strategies enable the resolution of all classes of isomers encountered in bio‑analysis, metabolomics, lipidomics and pharmaceutical research.

This presentation highlights applications from the field of pharmaceutical analysis, bioanalysis, metabolomics and lipidomics will be employed to demonstrate the importance of isomer separations and showcase tools that can be applied for isomer-selective analysis. Amino acid enantiomer analysis is important in metabolomics and pharmaceutical analysis. A new column, VAAST, enables the efficient and fast enantiomer separation of all proteinogenic amino acids in a single run. Accurate quantification of trace D-levels in presence of large excess of L-AAs is a challenge for fast and robust LC-MS/MS analysis. Some strategies how to achieve accurate quantification by LC-MS/MS in pharmaceutical formulations and biological matrices will be discussed. Peptide stereochemistry is commonly determined after total hydrolysis on the amino acid level, but destroys sequence information, making D‑AA localisation impossible. A stereopeptidomics approach with partial hydrolysis and stereoselective LC‑MS retains positional information; identifies D‑AA sites in non‑ribosomal and synthetic peptides. Lipid double‑bond and sn‑isomers, e.g. n‑3 vs. n‑6, sn1/sn2‑phospholipid isomers, are indistinguishable by conventional LC‑MS. Derivatization concepts (e.g. Paterno Büchi, epoxidation) or electron-activated dissociation enable location of double bond positions by specific fragments. Oxidation of lipids with polyunsaturated fatty acid side chains provide a wealth of possible isomers which can be distinguished by specific fragments. In general, complex mixtures of constitutional isomers overwhelm CID‑only workflows. IMS-MS adds an extra separation dimension and richer structural fingerprints. By integrating chiral LC, ion‑mobility, orthogonal MS fragmentation, and smart derivatization, analysts can finally untangle the full isomeric landscape of biological samples—delivering the structural detail required for accurate pharmaceutical, metabolomic, and lipidomic investigations.

ISC: ISC 2026 - Day 3: Michael Lämmerhofer, University of Tuebingen-, Institute of Pharmaceutical Sciences, Tuebingen, Germany.

15:00 - 15:20 Elucidating molecular interactions governing enantiorecognition in polysaccharide-based chiral HPLC using descriptor-guided AI analysis

• Carlos Pardo-Cortina, Universitat de València, Química Analítica, Valencia, Spain.

Understanding the molecular interactions responsible for chiral recognition remains a central challenge in chromatographic enantioseparations. Although polysaccharide-based chiral stationary phases (CSPs) are widely used in high-performance liquid chromatography (HPLC), the mechanisms governing enantiorecognition are still not fully clarified due to the complex interplay of noncovalent interactions involved.

In this work, we propose a data-driven strategy aimed at identifying and hierarchizing the molecular interactions responsible for enantioseparation through the analysis of molecular descriptors selected during artificial neural network (ANN) modeling. The study was carried out using a dataset of 78 structurally diverse chiral compounds belonging to 20 pharmacological and agrochemical families, chromatographed on a Lux® Cellulose-1 polysaccharide-based chiral stationary phase under nine aqueous–acetonitrile mobile-phase compositions.

Chromatographic performance was evaluated using the Efficient Enantioseparation (EES) parameter [1], which integrates resolution and retention behavior to provide a practical measure of separation efficiency. ANN models were optimized using a chaotic competitive-learning neural network algorithm, which simultaneously determines network topology and performs feature selection among 62 structural molecular descriptors describing steric, electronic, hydrophobic, and chiral-center environments.

Beyond predictive capability, the analysis of descriptor selection frequency and consistency across different ANN modeling strategies enabled the identification of the molecular features most strongly associated with efficient enantioseparation. These descriptors were subsequently interpreted in terms of fundamental interaction types involved in chiral recognition.

The results suggest that hydrogen bonding capacity, π–π interactions involving aromatic moieties, and steric complementarity around the chiral center play a dominant role in the studied chromatographic system, while electrostatic and hydrophobic contributions appear to modulate retention behavior depending on mobile-phase composition.

This descriptor-guided approach provides a chemically interpretable framework for elucidating enantiorecognition mechanisms in polysaccharide-based CSPs, offering insights that may contribute to a more rational design of chiral separations and to the development of predictive chromatographic methodologies.

15:20 - 15:40 Revealing the chirality of boron clusters: New chemical entities in drug development

• Radim Kucera, Charles University- Faculty of Pharmacy, Pharmaceutical Chemistry and Pharmaceutical Analysis, Hradec Králové, Czech Republic.

Twelve-vertex boron clusters represent an intriguing group of highly stable abiotic compounds. Replacing BH(-) units with CH(-) units results in icosahedral carboranes. These compounds and their derivatives have recently gained attention in medicinal chemistry due to their favorable properties, including increased hydrophobicity and stability within natural enzymatic systems. The compounds either exhibit their own biological activity or are studied as nonclassical bioisosteres of a phenyl ring. Although many of these compounds are chiral, this feature has been largely overlooked over the past six decades. However, regulatory authorities now require detailed information on the fate and activity of both enantiomers. Therefore, understanding how experimental conditions influence chiral discrimination is critically important in the drug development process.

We explored the possibilities for rapid and reliable chiral method development, especially for anionic boron clusters, which are more challenging analytes than zwitterionic species. Various chiral selectors were tested using HPLC, SFC, and CZE. Based on extensive chiral screening, a strategy for rapid and effective method development was proposed. Concerning the chiral selectors, cyclodextrins are useful in RPLC and non-aqueous CZE. The polysaccharide-based stationary phases demonstrated favorable enantiomer-discriminating abilities under RPLC and SFC conditions. Notably, chlorine-containing selectors proved effective for rapid chiral baseline separations.

Our work shows differences in interactions between anionic and zwitterionic species; we achieved chiral discrimination of some molecules for the first time and thus confirmed their chirality; we proposed a strategy for developing chiral methods for anionic carboranes in drug development. Additionally, the results could be applied in any field where the chirality of these interesting molecules is relevant.

ISC: ISC 2026 - Day 3: Radim Kucera, Charles University- Faculty of Pharmacy, Pharmaceutical Chemistry and Pharmaceutical Analysis, Hradec Králové, Czech Republic.

15:40 - 16:00 Combining per-aqueous and chiral reversed-phase separation modes towards a comprehensive two-dimensional LC platform for amino acid analysis

• José Meneses, Ghent University, Organic and Macromolecular Chemistry, Ghent, Belgium.

The analysis of amino acids is crucial due to their role in biochemical and pharmaceutical applications such as chiral drug development and metabolic profiling. Traditionally, comprehensive amino acid characterization requires two analytical approaches with different objectives. Reverse-phase liquid chromatography (RP-LC) is commonly used to identify and quantify amino acids in complex mixtures, enabling characterization in a single run. In contrast, chiral analysis focuses on resolving enantiomers but typically requires prior isolation of each amino acid or prior knowledge of its identity. These methods rely on one-dimensional chromatography and mobile phases with high organic solvent content, which involve environmental and economic drawbacks due to solvent demand. Developing techniques that minimize solvent consumption represents a more sustainable alternative.

A potential solution is comprehensive two-dimensional chromatography (2D-LC), which enhances resolution by coupling two chromatographic dimensions. The effluent from the first dimension is divided into small fractions and sequentially transferred to the second-dimension column for further separation, enabling analysis of complex mixtures with increased peak capacity and selectivity compared to one-dimensional separations [1].

In this work, 2D-LC with Per Aqueous Liquid Chromatography (PALC) in the first dimension on a hydrophilic interaction (HILIC) column and fast chiral RP-LC in the second dimension was implemented for amino acid analysis, allowing identification of amino acids and separation of their enantiomers in a single analysis.

In the first dimension, amino acids were separated on a commercial HILIC column operated under PALC conditions. Unlike conventional HILIC, which relies on high acetonitrile concentrations, PALC employs water-rich eluents, reducing solvent use and providing a more environmentally friendly approach [2]. A gradient from 90% aqueous mobile phase to 30% ACN was applied over 60 minutes before enantioselective analysis in the second dimension.

A key advantage of PALC is that its aqueous eluent simplifies the transition to comprehensive 2D-LC, allowing direct injection into the second dimension and avoiding solvent-mismatch effects that can compromise analyte focusing and peak shape [3]. In contrast, conventional HILIC or RP-LC produce eluents with high organic solvent content, often requiring heart-cutting approaches with dilution or solvent exchange.

After first-dimension separation, the effluent was fractionated and injected into a chiral reverse-phase column, enabling separation of amino acid enantiomers. The method successfully resolved and identified all target amino acids, providing a more sustainable and cost-effective approach for chiral analysis with potential application to complex real-world samples.

ISC: ISC 2026 - Day 3: José Meneses, Ghent University, Organic and Macromolecular Chemistry, Ghent, Belgium

14:30–16:00 BIO 06: Lipidomics

Chairs

• Takeshi Bamba, Japan    
• Josef Cvačka, Czech Republic

14:30 - 15:00 Clinical lipidomics for precision cardiovascular risk prediction: Development of the L.G.CVDRisk score and translation to a high-throughput clinical platform

• Peter Meikle, Australia

Cardiovascular disease (CVD) risk assessment in clinical practice relies largely on traditional lipid measures and demographic risk factors, which incompletely capture the underlying biology of atherosclerosis. High-resolution lipidomics provides the opportunity to quantify hundreds of molecular lipid species that reflect diverse metabolic pathways linked to cardiometabolic disease. We developed a lipidomic risk framework (L.G.CVDRisk) and translated the underlying analytical methodology into a scalable clinical lipidomics platform suitable for implementation in routine pathology laboratories.

The L.G.CVDRisk score integrates detailed plasma lipidomic profiling with established clinical risk algorithms. In population cohorts including the Australian Diabetes, Obesity and Lifestyle Study (AusDiab) and the Busselton Health Study, targeted LC-MS/MS lipidomics quantified hundreds of individual lipid species from small plasma volumes. Using regularised regression modelling of 689 lipid species, a lipidomic score capturing molecular signatures of CVD risk was derived and integrated with the AusCVDRisk clinical model and a coronary artery disease polygenic risk score. This combined framework improved discrimination and substantially enhanced reclassification of individuals at intermediate clinical risk, correctly reclassifying a large proportion of future CVD events into higher-risk categories. These findings demonstrate that molecular lipid profiles capture complementary biological information not reflected in conventional clinical markers.

To enable translation of these discoveries into clinical practice, we developed a Clinical Lipidomics Platform (CLP), a high-throughput LC-MS/MS assay designed specifically for clinical laboratories. The CLP quantifies 270 lipid analytes (248 lipid species and 22 internal standards) spanning 37 lipid classes using dynamic multiple reaction monitoring on a triple quadrupole mass spectrometer with a six-minute runtime. Lipid species were selected from the larger research lipidomics platform to retain maximal biological and predictive information while enabling automated data processing and high analytical throughput. Quantification incorporates stable isotope-labelled internal standards, in addition to post-column infusion of standards, and normalisation using the NIST Standard Reference Material plasma (SRM 1950) to ensure cross-cohort comparability and analytical reproducibility.

Validation in the BioHEART-CT cohort demonstrated strong concordance between lipid measurements obtained using the clinical platform and the original research lipidomics method, with lipid-derived risk scores showing excellent agreement between platforms and improved prediction of coronary artery calcium burden compared with traditional risk scores.

Together, these developments establish a pathway from discovery lipidomics to scalable clinical implementation. Clinical lipidomics enables molecularly informed cardiovascular risk assessment and provides a foundation for precision prevention strategies in cardiometabolic disease.

ISC: ISC 2026 - Day 3: Peter Meikle, Australia

15:00 - 15:20 Lipid alterations across six PDAC-related cohorts characterized by targeted RP-UHPLC/MS/MS

• Zuzana Lásko, University of Pardubice, Departement of Analytical Chemistry, Pardubice, Czech Republic.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive cancers, with a 5-year survival around 13% (and only 3% at the metastatic stage). Late diagnosis remains the major limitation, as current diagnostic approaches (including imaging methods and CA 19-9) lack sensitivity for early-stage disease and often involve invasive procedures. Metabolic reprogramming in PDAC is reflected in circulating lipids, which can be profiled using minimally invasive blood lipidomics.

We applied a targeted RP-UHPLC/MS/MS workflow to lipidomic profiling of >450 human plasma/serum samples from six PDAC-related cohorts: (a) healthy controls without cancer history, (b) patients after total pancreatectomy for non-malignant disorders, (c) high-risk individuals, (d) patients with pancreatic precancerous lesions, (e) PDAC patients, and (f) long-term PDAC survivors (≥5 years without recurrence). Lipids were extracted using an MTBE-based protocol and spiked with a mixture of internal standards. Separations were performed on an Agilent 1290 Infinity II LC system using a bioinert Acquity Premier BEH C18 column (Waters). A 25 min reversed-phase gradient using acetonitrile/water (60/40, v/v) and acetonitrile/isopropanol (10/90, v/v) mobile phases, both supplemented with 5 mM ammonium formate and 0.1% formic acid, provided separation of lipid species based on hydrophobicity. This approach also improved selectivity for fatty acyl chain length and degree of unsaturation and increased the resolution of isomeric species. Detection was performed using Agilent 6495 triple quadrupole mass spectrometer operated in multiple reaction monitoring mode, providing quantification of more than 640 lipid species across 30 lipid subclasses.

Statistical analysis and data visualization revealed significant lipidomic alterations in PDAC samples. In supervised OPLS-DA analysis, PDAC and healthy controls showed clear separation, while long-term survivors clustered within the PDAC group. These findings indicate that metabolic alternations in the lipidome persist even during long-term complete remission and may limit the use of this method for monitoring treatment response or recurrence. Among the most affected lipid classes were sphingolipids with very long acyl chains, consistent with previously published data. No lipidomic changes associated with PDAC were observed in high-risk individuals, whereas patients with precancerous lesions showed clear positivity. These results support the potential of lipidomic profiling for early PDAC detection and its possible application in cancer screening.

ISC: ISC 2026 - Day 3: Zuzana Lásko, University of Pardubice

15:20 - 15:40 Benzoyl chloride derivatization enhances separation and quantitation of multiple lipid classes

• Ondřej Peterka, University of Pardubice, Department of Analytical Chemistry, Pardubice, Czech Republic.

The chemical derivatization of lipid functional groups can improve extraction efficiency, chromatographic separation, and sensitivity. Benzoyl chloride (BzCl) is a highly reactive derivatization agent for several functional groups found in lipids. The derivatization reaction was thoroughly optimized, resulting in a reaction time of 60 min at ambient temperature. Benzoylation increased the sensitivity by 2- to 100-fold, mainly for MG, SPB, and DG, compared to the non-derivatized approach. The untargeted analysis of human plasma led to the detection of 169 lipids from 11 lipid classes using RP-UHPLC/MS [1].

Benzoylation increases the non-polar character of the analytes, which was exploited for the selective liquid–liquid extraction of six lipid classes (TG, DG, MG, SE, ST, and FA) from human plasma into hexane. This strategy enables sensitive profiling of these classes by RP-UHPSFC/MS using two C18 columns (100+150×3.0 mm; 1.8μm) in series, which provides high chromatographic resolution, sufficient to separate isomeric species, including cis/trans and positional isomers of double bonds in FA. The method was validated and 147 less-polar lipids were quantified in human plasma in single 18-min analytical run [2].

The benzoyl moiety serves as a universal tag, enabling selective and sensitive MRM transitions. This is particularly advantageous for lipids that lack specific fragmentation pathways in their native form (e.g., MG, DG, SPB, ST). In total, 450 lipids from 19 lipid subclasses were identified using combination of derivatization and RP-UHPLC/MS/MS. The validated method was used for targeted analysis of serum samples and for the comparison of the lipid profiles of pancreatic cancer patients and healthy controls. The lipidomic profiles show differences primarily in sphingolipids and phospholipids, but our approach reveals new findings, including significant upregulation of MG and SPB [3].

Furthermore, the derivatization can help with easier identification and accurate quantitation using H/D pooling strategy. Individual samples are derivatized with non-labelled BzCl (H), while standardized pooled plasma is derivatized with its deuterated analogue (D). Mixing H- and D-derivatives in the same ratio creates an internal standard (D-labeled lipid) for each derivatized lipid, resulting in isotopic doublet for every analyte in mass spectrum. This methodology ensures coelution of analyte and standard, which effectively corrects matrix effects and ionization variability.

ISC: ISC 2026 - Day 3: Ondřej Peterka, University of Pardubice, Department of Analytical Chemistry, Pardubice, Czech Republic.

15:40 - 16:00 Toward low-input lipidomics: A microbore RP-UHPLC-4D-TIMS strategy for organoids analysis

• Fabrizio Merciai, University of Salerno, Department of Pharmacy, Salerno, Italy.

Low-input biological systems are increasingly employed in clinical and translational research, where analytical platforms must combine high sensitivity with high throughput despite the limited availability of biological material. Among these models, patient-derived organoids (PDOs) have emerged as physiologically relevant tools for investigating disease mechanisms and evaluating drug responses. PDOs are commonly cultured in bulk conditions, which represent a well proved biological sample for conventional LC–MS lipidomics workflows. However, the growing adoption of high-throughput experimental designs, such as multiwell formats, is driving the need for analytical strategies compatible with significantly reduced sample amounts (~10³ cells).

In this study, we investigated the implementation of a 1.0 mm internal diameter column coupled with trapped ion mobility mass spectrometry (micro-4D-TIMS), to enhance analytical sensitivity for lipidomic profiling of PDOs cultured in a 96-well array format. The microbore configuration provided substantial improvements in analytical performance, including increased signal intensity, enhanced signal-to-noise ratio, and markedly improved detection sensitivity. On average, the method achieved a 12-fold reduction in limits of detection (LOD), with improvements reaching up to 68-fold for specific lipid species, such as LPC 18:1. To demonstrate the applicability of the workflow in biologically relevant samples, we compared lipidomic profiles of colorectal cancer PDOs treated with two chemotherapeutic agents under both bulk culture conditions and 96-well miniaturized formats.

Lipidomic profiling revealed highly consistent treatment-induced alterations between the two culture systems. In particular, phosphatidylcholines (PCs) and phosphatidylethanolamines (PEs) represented the most significantly modulated lipid subclasses in response to treatment, confirming the biological comparability of the miniaturized platform. Furthermore, relative to a conventional 2.1 mm I.D. LC setup, the micro-4D-TIMS configuration enabled the detection of 103 additional lipid species across 17 lipid subclasses, including hexosylceramides, phosphatidylserines and phosphatidylglycerols that were not observed with the standard configuration. Overall, these results demonstrate that reducing column internal diameter represents a robust strategy to significantly improve sensitivity in low-input lipidomics workflows. This approach provides a practical analytical solution to support high-throughput lipidomic investigations of organoid models, facilitating their application in precision medicine and drug response studies.

ISC: ISC 2026 - Day 3: Fabrizio Merciai, University of Salerno, Department of Pharmacy, Salerno, Italy.

14:30–16:00 HYP 06: Microfluidic and Chip-Based Separations in MS

Chairs

• František Foret, Czech Republic    
• Christian Neusüß, Germany

ISC: ISC 2026 - Day 3: HYP 06 Microfluidic and Chip-Based Separations in MS chairs (František Foret, Czech Republic and Christian Neusüß, Germany)

14:30 - 15:00 Best of both worlds: Modular microfluidics for micro-separation–mass spectrometry coupling

• Detlev Belder, Leipzig University, Analytical Chemistry, Leipzig, Germany.

Highly integrated lab-on-a-chip systems have demonstrated remarkable potential in analytical sciences, including micro-scale separation techniques. However, the development of increasingly complex and fully integrated devices remains technically challenging. An attractive alternative is the modularization of microfluidic systems using a microfluidic breadboard concept. Modularization simplifies technical implementation while providing the key advantage of standardized components.

Here, we present a modular microfluidic approach that combines the strengths of capillary-based and chip-based microfluidics. While capillaries offer easy accessibility, broad commercial availability, and a wide range of dimensions, microfabricated chips enable the integration of complex functionalities and, as we have recently demonstrated, dead-volume–free connections between different modules.

Functional chip modules are fabricated using selective laser etching and incorporate structures such as particle frits, flow splitters, MS emitters, and electrochemical or photochemical detection cells. These modules can be seamlessly connected via capillaries, enabling flexible and dead-volume–free system configurations.

This modular platform allows the implementation of a wide range of micro-scale separation techniques coupled to mass spectrometry or ion mobility spectrometry. Demonstrated examples include micro- and nano-scale SFC–MS, micro- and nano-HPLC–MS, and CE–MS using sheathless MS emitters. In addition, the platform enables the straightforward integration of further chemical functionalities, such as microreactors containing immobilized catalysts, opening new possibilities for integrated microchemical workflows.

15:00 - 15:20 Study of the analytical performance of ultra-miniaturized preconcentrators based on MEMS technology for the analysis of planetary samples

• Lylia Skander, LATMOS- IPSL- UVSQ Université Paris-Saclay- CNRS- Guyancourt- France, Solar System, Guyancourt, France.

Preconcentrators are widely used in analytical chemistry, particularly in gas chromatograph developed for space missions, to preconcentrate and focalize trace analytes prior to analysis. Preconcentration step is essential in space analysis, as samples are pyrolyzed and introduced into the system over a given period of time. The preconcentrator enables the analytes to be focalized, which allows a precise injection at the head of the separation column. In this framework, it is essential to optimize energy consumption and instrument weight and volume. Consequently, we are developing lab-on-chip preconcentrators based on MEMS (Micro Electro Mechanical Systems) technology and integrated onto a printed circuit boards (PCB) allowing the thermal heating and temperature monitoring of the chip. To evaluate the analytical performance of MEMS preconcentrators filled with TENAX TA® as an adsorbent, analytical tests were carried out by coupling the preconcentrator to a non-polar commercial column (30m×0.25mm×0.25µm, Rxi-5Sil, Restek) and a commercial gas chromatograph. The preconcentration factor (PF) was used to characterize the efficiency of the preconcentrator, defined as the ratio between the maximum concentration measured during desorption and the initial concentration of the analytes. The adsorption and desorption conditions were optimized for nonane by adjusting the adsorption and desorption time as well as the thermal desorption temperature. Preliminary results with liquid injection of nonane show a direct effect of temperature on the amount of analyte desorbed, showing an increase of about 24% between 150°C and 230°C. However, an incomplete desorption was also observed and is probably due to a breakthrough of the trap. Additional tests will be carried out using gas injection with lower concentrations in order to investigate the breakthrough volume of the preconcentrator and then to analyze more complex mixtures of alkanes, aromatics, and nitriles, as well as natural/synthetic analogous samples. The use of new adsorbent materials for the sample preconcentrator will also be investigated in order to extend the range of compounds that can be tested.

ISC: ISC 2026 - Day 3: Lylia Skander, LATMOS- IPSL- UVSQ Université Paris-Saclay- CNRS- Guyancourt- France, Solar System, Guyancourt, France

15:20 - 15:40 Bridging mass spectrometry and SERS via microfluidics: A step toward integrated peptide characterization

• Jakub Novotny, Institute of Analytical Chemistry of the Czech Academy of Sciences- v.v.i., Department of Bioanalytical Instrumentation, Brno, Czech Republic.

The structural characterization of therapeutic peptides is crucial in modern pharmaceutical development. While mass spectrometry (MS) serves as the analytical standard for assessing peptide composition and stability, capturing the full in-solution conformation of these peptides often requires orthogonal techniques. Among these, surface-enhanced Raman spectrometry (SERS) represents a promising approach. SERS provides insight into spatial conformation, whereas MS delivers definitive mass-to-charge and compositional data. However, their simultaneous implementation remains challenging, since MS requires fluidic solutions to be free of the plasmonic nanostructures that SERS depends on. Thus, pioneering strategies for SERS-MS detection depend heavily on static, non-fluidic setups - such as depositing an analyte onto a nanostructured solid emitter.

Our research aims to develop a custom microfluidic glass chip enabling continuous simultaneous SERS-MS detection. As a necessary intermediate optimization step, we have designed a dual-purpose microfluidic device in which SERS and MS functionalities are deliberately decoupled, allowing independent optimization of each modality prior to their integration into a unified platform.

By integrating fused-silica capillary emitters, 3D-printed frames, and fluidic connections, the chip enables the formation of a controlled and adaptable liquid environment, supporting robust nanoelectrospray ionization (nanoESI) and allowing fine-tuning of nanoESI-MS analyses. The same glass chip architecture enables in-flow SERS detection as well. The required plasmonic nanostructures are introduced in the form of suspended microbeads coated with silver nanoparticles (AgNPs). The interaction of these composite particles with the target peptide establishes the crucial close contact (less than 10 nm) required for Raman signal enhancement. To integrate this into a continuous flow, we utilize acoustofluidic forces. By applying ultrasound at low MHz frequencies, we generate an acoustic field inside the microchannel to manipulate and locally concentrate these AgNP-coated microparticles before the laser excitation of the Raman signal.

Our results demonstrate that the combination of a custom-built microfluidic glass chip and the associated technologies can secure a reliable nanoESI interface as well as in-flow SERS compatibility. Our work has laid the essential groundwork for a multiplexed analytical platform capable of structural characterization of therapeutic peptides in solution.

ISC: ISC 2026 - Day 3: Jakub Novotny, Institute of Analytical Chemistry of the Czech Academy of Sciences- v.v.i., Department of Bioanalytical Instrumentation, Brno, Czech Republic.

15:40 - 16:00 Application of 3D printing in environmental analysis

• Dingyi Wang, Research Center for Eco-Environmental Sciences- Chinese Academy of Sciences, State Key Laboratory of Environmental Chemistry and Ecotoxicology, Beijing, China.

3D printing technology, characterized by high design freedom and the ability to achieve integrated forming of complex structures, has been widely applied in scientific research. Over the past decade, our team has focused on the application of 3D printing technology in the development of customized environmental analysis devices, particularly for chromatography–mass spectrometry (MS) coupled systems.

We established a novel development paradigm for analytical devices based on “simulation-assisted design—precise 3D printing—rapid experimental validation.” A customized ICP-MS sampling interface was developed and 3D-printed for the analysis of heterogeneous samples, such as single particles and single cells, enabling efficient coupling with high-performance liquid chromatography, size-exclusion chromatography, and capillary electrophoresis. Through the integration of a microfluidic droplet separation module and an optimized nebulizer and spray chamber design, sample transport efficiency was improved from <10% to >60%, significantly enhancing the accuracy of heterogeneous sample analysis [1]. Additionally, a columnar gel electrophoresis device and elution interface were designed and 3D-printed for coupling with ICP-MS, achieving high-efficiency separation and real-time online analysis of metalloproteins in bacterial and cellular samples, providing robust support for metalloproteomics studies [2]. Furthermore, a sampling and detection integrated chromatographic column device was developed. By directly 3D printing porous or micro-column array structures, precise control over pore size, porosity, and architecture was achieved, enabling the selective enrichment and in situ online detection of different forms of mercury contaminants in water.

These results advance the application of 3D printing in environmental analysis, providing a foundation for the rapid development of customized and miniaturized analytical devices, and promoting innovation in chromatography–mass spectrometry instrumentation.

16:00–16:30 Coffee Break

16:30 - 18:00 Industry Session: Activity 4: Discussion session: Artificial Intelligence in separation science - From concept to practice

16:30–18:00 FUN TY 2: Technological Innovations in Chromatography

Chair

• Jiří Urban, Czech Republic    
• Bram Huygens, Belgium

16:30 - 17:15 The great (separation) power of two-dimensional liquid chromatography

• Peter Schoenmakers, University of Amsterdam, HIMS, Amsterdam, Netherlands.

In two-dimensional liquid chromatography (2D-LC) one (“heart-cut”), several (“multiple heart-cut”), or all fractions from a first-dimension LC separation are subsequently subjected to a second separation, which (almost always) offers a substantially different selectivity. Heart-cut 2D-LC is increasingly used for applications ranging from sample preparation (isolating a group or class of analytes for detailed separation) to compound isolation for analytical (e.g. peak-purity testing) or preparative (analyte purification) purposes. The additional selectivity offered by the different separation mechanisms is the single great advantage of this method. Comprehensive two-dimensional liquid chromatography (LC×LC) offers the additional benefit of a greatly increased separation power (peak capacity) in comparison with conventional 1D-LC, which makes it an attractive method for the separation of very complex samples.

In this tutorial lecture we will discuss the advantages and limitations of 2D-LC in detail. The implementation of 2D-LC methods will be discussed, with specific focus on challenges arising from the interfacing of very different separation mechanisms. Column selection is vital with regards to selectivity, but column dimensions and operating conditions also need careful attention, especially in LC×LC. Developing 2D-LC methods may be the greatest bottleneck for the proliferation of powerful two-dimensional separation methods.

All these concepts will be illustrated with practical applications of 2D-LC.

17:15 - 17:30 MEMS column for space missions: Design optimization of fluidic interface and column inlet-outlet geometry to combine test reliability and maximum efficiency

• Malak Bigourd, LATMOS/IPSL- UVSQ Université Paris-Saclay- Sorbonne Université- CNRS- Guyancourt- France, Solar System, Guyancourt, France.

In space exploration missions dedicated to planetology and exobiology studies, GC is used to perform in situ analysis of organic molecules in order to improve our knowledge of prebiotic chemistry and the origins of life on Earth and in the universe. The miniaturization of GC instruments is becoming possible using the recent development in MEMS/NEMS (Micro/Nano-Electro-Mechanical System) and micro-fluidic technologies that will allow a real jump of scale on many levels. While the only MEMS columns [1] developed in laboratory for space exploration are still using fragile interfacing with glued capillaries, we developed a patented fluidic interface [2,3] to reliable integrate MEMS preconcentrator, column and detector considering the space requirements in term of mechanical, fluidic and thermal constraints. Besides, this interface with screwed connections offers the possibility to change easily capillaries without damaging MEMS components. The same fluidic interface is used to coat the column with stationary phase following a patented method [4,5]. A fine optimization was performed on this fluidic interface in order to minimize extra-column volume, allow reliable experimental measurements and raise the range of achievable temperature in order to extend the panel of analyzed molecules. Besides, the inlet and outlet column geometry were optimized to enhance the column performance. Obtained results showed a significant gain in measured efficiency (up to 74%). The coupling of our MEMS column to Mass spectrometry was successfully achieved and a large panel of organic compounds of interest for exobiology such as alkanes, FAMEs and amino-acids were analyzed. This work shows the importance of optimized fluidic interface and column geometry in analyzing complex mixtures with high reliability.

17:30 - 17:45 In silico optimization of two-dimensional liquid chromatography in proteomics

• Kateřina Hrůzová, Masaryk University, Department of Chemistry, Brno, Czech Republic.

Two-dimensional liquid chromatography (2D-LC) provides enhanced separation power for complex samples, particularly when implemented in an online comprehensive mode. However, its broader application remains limited due to challenges in method development and optimization. In this study, we present a predictive and optimization framework for online comprehensive 2D-LC that systematically guides the workflow from column selection to method parameter optimization, aiming to maximize peak capacity.

Scouting gradients were performed on reversed-phase (RP) and hydrophilic interaction liquid chromatography (HILIC) columns using a tryptic digest of proteins. Peptides consistently detected across all experiments were used to establish retention models of elution volumes. The linear solvent strength model was applied to RP separations, while an adsorption-based model was used for HILIC separations. A key aspect of this work is the incorporation of column dimension optimization, where combinations of column lengths and internal diameters were evaluated.

The developed model enables simultaneous optimization of gradient conditions, flow rates, modulation times, and sampling parameters. Peak capacity in each dimension was calculated based on the separation window and average peak width. Experimental peak widths obtained from scouting runs were converted to plate-height equivalents, revealing a strong polynomial relationship that enabled the prediction of peak broadening behavior. Furthermore, the calculation of peak capacity in the second dimension was adapted to account for modulation and first-dimension gradient effects, allowing both dimensions to be optimized in a fully integrated manner.

The framework was used to simulate 2D-LC separations and generate predicted chromatographic distributions, which were subsequently compared with experimental measurements to assess model performance. This approach provides a comprehensive and practical tool for rational method development in comprehensive online 2D-LC, facilitating efficient optimization while reducing turnaround time.

17:45 - 18:00  Molecular dynamics simulations of mixed-mode chromatography: Generation and solvation of a reversed-phase/anion-exchange stationary phase

• Daniel Frerichs, Germany

As an initial step towards predicting retention of small, hydrophilic, ionizable analytes in mixed-mode liquid chromatography (MMLC) separations, we designed a slit-pore model of a silica-based, endcapped, reversed-phase/anion-exchange (RP/AEX) stationary phase. Molecular dynamics simulations were performed to investigate its solvation by a binary water‒acetonitrile (W‒ACN) mobile phase under conditions where the AEX function is either neutral or positively charged. In the latter case, sodium and chloride ions were added as co-ions and counterions, respectively, to the 60/40 (v/v) W/ACN mobile phase. The silica surface functionalization, designed to reflect an existing column technology, yielded a unique surface topology, where bonded-phase islands formed by octadecylsilyl chains around a central tertiary alkylamine group are interspersed with hydroxylated silica patches that are exposed to the mobile phase. Both MMLC systems were analyzed regarding the solvent density, structure, and orientation with respect to the density distribution of hydrophobic and hydrophilic bonded-phase moieties (alkyl groups and nitrogen atoms, respectively) at varying distances from the silica surface. Effects originating from the surface functionalization were identified by comparison with a conventional RPLC stationary phase, whereas effects arising from the presence of charge were quantified by comparing the two simulated MMLC systems (neutral vs. charged surface). The analysis of ion density distributions and ion contact profiles suggested that the surface topology of the MMLC stationary phase shields the AEX function from contact with hydrophilic counterions and favors the accumulation of analyte compounds with hydrophobic and hydrophilic structural elements.

16:30–18:00 BIO TY 02: Chromatographic Methods in Bioanalysis

Chairs

• Petr Chocholouš, Czech Republic    
• Marie Pardon, Belgium

16:30 - 17:15 Enantioseparations in high-performance liquid chromatography

• Bezhan Chankvetadze, Iv. Javakhishvili Tbilisi State University, Chemistry, Tbilisi, Georgia.

In this tutorial lecture I am going to cover the aspect of so called “chiral HPLC” that could be of interest for a broad audience involved in enantioseparations. These would comprise on one hand the researchers involved in a development of new chiral stationary phases and on the other hand researchers performing separations of high societal importance such as analysis of novel drugs of abuse. Thus, we will start with a design of non-covalent interactions (physical chemistry) [1], go to the synthesis of targeted polysaccharide-based chiral selectors (synthetic organic and polymer chemistry) [2, 3], combining chiral selectors with silica for (materials science) [4, 5], column packing and evaluation (analytical chemistry), uncommon effects in separation science (analytical and physical chemistry) and end with the most recent studies in forensic analysis and clinical toxicology [6-9]. Besides the aspects mentioned above, the emphases will be made on method selection and development with the focus on enantiomer elution order, greenness and sustainability.

17:15 - 17:30 Ion-pair-free HILIC-HRMS/MS for multi-level characterization of intact modified oligonucleotides

• Khaoula Adouairi, University of Lyon- Institute of Analytical Sciences, Rhône 69, Villeurbanne, France.

Chemical modifications of oligonucleotides, notably N6-methyladenosine (m6A), are increasingly recognized as key determinants of small RNA stability and maturation in viral and cellular contexts (e.g., the 22-mer miR-K11). Yet, routine characterization of such modifications on intact sequences remains challenging, particularly for complex synthesis batches containing shortmer impurities, multiple global methylation states, and positional isomers that share identical methyl counts.

Here, we present an ion-pair-free HILIC-HRMS/MS method for the analysis of intact modified miRNA analogues of up to 22 nucleotides. The approach combines amide-based HILIC separation with high-resolution top-down CID MS/MS and provides multi-level characterization within a single 20-minute analysis. First, the HILIC dimension enables MS-friendly separation of synthesis-related shortmers from the intact full-length 22-mer. Second, the chromatographic dimension separates intact oligonucleotides according to the number of methylation modifications. Finally, positional m6A isomers are distinguished by top-down CID HRMS/MS through specific diagnostic fragments, allowing confident localization of methylation sites based on the presence or absence of diagnostic ions.

This work establishes ion-pair-free HILIC-HRMS/MS as a powerful analytical method for the rapid, selective and multi-level characterization of modified oligonucleotides, opening new opportunities for quality control and structural verification of methylated miRNA analogues in a single run. In parallel, the development of more sustainable analytical strategies is becoming a major issue in oligonucleotide analysis, a direction we have already begun to explore with promising preliminary results for greener HILIC-based approaches.

17:30 - 17:45 Green solvent alternatives for RP-HPLC–PDA–MS analysis of polyphenols in natural products

• Carmelo Coppolino, Foundation Prof. Antonio Imbesi, University of Messina, Messina, Italy; Messina Institute of Technology, Department of Chemical- Biological- Pharmaceutical and Environmental Sciences- University of Messina, Messina, Italy.

In recent years, the implementation of green chemistry principles has stimulated considerable efforts to replace hazardous and environmentally harmful solvents with safer and more sustainable alternatives across nearly all areas of chemistry. In this context, dimethyl carbonate (DMC), a biodegradable and low-toxicity solvent, was evaluated as a green alternative to acetonitrile (ACN) as organic modifier in reversed-phase high-performance liquid chromatography (RP-HPLC). Despite its favorable environmental profile, the main limitation of DMC compared to ACN is its limited miscibility with water. To address this issue, the miscibility of various binary mixtures of DMC and ethanol (EtOH) at different water contents was investigated, thereby simulating solvent behavior throughout the entire LC gradient. Once the DMC/EtOH miscible mixture was selected, the flow rate was subsequently optimized using an HPLC–PDA–MS system equipped with an Ascentis® Express C18 column (15 cm × 2.1 mm I.D., 2.7 μm d.p.) Merck Life Science (Merck KGaA, Darmstadt, Germany). Under the optimized conditions, an almost complete separation of a polyphenol standard mixture was achieved, with peak widths comparable to those obtained using conventional ACN-based methods. Importantly, UV and MS spectral quality and sensitivity were preserved, demonstrating that the substitution of ACN with a DMC/EtOH system does not compromise polyphenol identification. Finally, the developed method was successfully applied to the characterization of polyphenols in natural products, confirming its potential as a viable green alternative for RP-HPLC analyses, in line with Green Analytical Chemistry (GAC) principles.

17:45 - 18:00 Development and validation of UHPLC methods for melamine detection and aerobic biodegradability assessment in wastewater

• H. Aghakhanlou, Wood K plus - Competence Center for Wood Composites & Wood Chemistry, Linz, Austria.

16:30–18:00 HYP TY 2: Hyphenated Techniques

Chairs

• Ondřej Peterka, Czech Republic    
• Fabrizio Merciai, Italy

16:30 - 17:15 Bridging the gap: Interfacing electro-driven and microfluidic separations with mass spectrometry

• Jorg P. Kutter, University of Copenhagen, Dept. of Pharmacy, Copenhagen, Denmark.

Electro-driven separation techniques — from capillary electrophoresis (CE) to microfluidic chip-based platforms — offer compelling advantages in speed, sample consumption, and miniaturization. Yet their coupling to mass spectrometry (MS), arguably the most powerful detection modality available to the separation scientist, remains far less routine than the equivalent liquid chromatography–MS workflow. This tutorial will attempt to highlight why that gap exists, what interface designs have emerged to bridge it (including commercial offers), and where the proposed solutions still fall short.

The fundamental tension at the heart of CE–MS and microfluidic–MS coupling is electrochemical and fluidic in nature: maintaining a stable electrospray ionization (ESI) plume demands a stable liquid flow and a defined electrical contact close to the spray tip, while the separation itself depends on a carefully controlled, often near-zero bulk flow and an unperturbed electric field across the capillary or channel. Satisfying both constraints simultaneously is non-trivial, and every interface design represents a different compromise.

This tutorial will give a brief overview over the principal interface architectures in current use. The sheath-liquid interface — the longest-established approach — provides robust electrical contact and stable spray at the cost of dilution and suppression of analyte signal. Sheathless designs, including the porous-tip and etched-capillary formats, preserve sensitivity but demand tighter fabrication tolerances and are more susceptible to clogging and spray instability. Junction-at-the-tip and pressurized-flow approaches offer intermediate trade-offs. For microfluidic platforms, the additional challenges of chip material compatibility, channel geometry, dead volumes, connectors, and integrated electrode design introduce further complexity, and we will discuss both the promises and the practical frustrations of fully integrated chip–MS systems.

Beyond the spray interface itself, remaining underlying challenges include managing buffer depletion effects, achieving adequate concentration sensitivity for trace-level analytes without sacrificing separation efficiency, and ensuring long-term robustness — particularly relevant for clinical and regulated environments. Emerging strategies to be discussed include online sample concentration via isotachophoresis or field-amplified stacking, nanospray-optimized emitter geometries, and the use of surface coatings to decouple EOF control from analyte interaction.

We will conclude by identifying the most tractable near-term problems and the interface configurations most likely to see broader adoption as CE–MS and microfluidic–MS mature from specialist techniques into routine analytical tools.

17:15 - 17:30 Seeing more in TLC: How multichannel detection improves complex mixture analysis

• Lukasz Pieszczek, University of Silesia in Katowice, Faculty of Science and Technology, Katowice, Poland.

Samples examined in analytical laboratories are most often complex multicomponent mixtures that must undergo preliminary preparation to isolate the compounds of interest. Despite chemical heterogeneity being a natural characteristic of such mixtures, it is not often considered a central analytical parameter. Paradoxically, in recent decades, a wide range of instrumental methods and data‑analysis approaches has emerged specifically to detect and exploit chemical inhomogeneity within samples. Among these are multispectral imaging techniques and spectroscopic methods, both of which are widely used for characterizing solid materials.

Liquid mixtures, in turn, can be preliminarily separated chromatographically, for example by thin-layer chromatography (TLC), and subsequently analyzed spectroscopically, typically by densitometry. In most studies, however, TLC plates are scanned at a single illumination wavelength. Such an approach may limit the accurate characterization of chromatographic bands and hinder the detection of their chemical heterogeneity. Although modern densitometers are capable of multichannel measurements, this capability is less widely used in TLC analysis, mainly due to the complexity of the resulting datasets and the challenges associated with their processing.

This study discusses the benefits of multichannel measurements and proposes data-processing strategies for TLC datasets to enhance the identification of complex mixtures. In addition, a low-cost solution enabling multichannel detection of chromatographic bands is introduced [1]. The developed device is an HPTLC measurement chamber in which the sophisticated and costly components typical of professional TLC densitometry systems - such as monochromators and collimators - are replaced with a set of inexpensive light sources emitting in different spectral ranges (UV-A, UV-C, and visible light). Detection is carried out using a CMOS camera integrated into a smartphone. For this measurement setup, a workflow is proposed for HPTLC plate imaging, data aggregation, and data processing, enabling the resulting datasets to be analyzed using methods commonly applied in multi- and hyperspectral image analysis.

17:30 - 17:45 A multicomponent MAE–SPE–HPLC–MS/MS workflow for the analysis of contaminants in sediments and microplastics

• Janina Heim, KNAUER wissenschaftliche Geräte GmbH, Application and Academy, Berlin, Germany.

Aquatic ecosystems are facing increasing challenges due to growing antrophogenic contamination with a wide range of chemicals. Among others, pharmaceuticals as well as personal care products are constantly emitted into the environment and can have negative effects on many species. These contaminants are not only present in the water itself but also adsorb to sediments and other particles. Microplastics are of special concern because polymer particles not only pose a risk on their own, but also strongly accumulate other contaminants and may transfer them into the food chain when accidentally taken up by aquatic organisms. Nevertheless, knowledge about the contamination of sediments and microplastics in the environment remains limited. Multicomponent analysis is needed to comprehensively assess risks. However, analysis is often challenging due to the low concentrations of target compounds and the strong matrix effects associated with environmental samples.

In this study, we developed a workflow for the analysis of more than 20 exemplary contaminants, including antibiotics, analgesic drugs, anticonvulsants, beta blockers, stimulants, insect repellents and plasticizers, in sediments and microplastics. Samples were collected from beaches, coasts, fjords, and lakes in Germany, Denmark, and Norway. Microplastics and sediment were first separated by density using an overflow method, followed by microwave-assisted solvent extraction (MAE). MAE has not yet been widely applied for this purpose, but it offers several advantages due to its fast, efficient, and parallelized procedure. For enrichment and clean-up of the extracts, a solid phase extraction (SPE) method was optimized regarding target recovery and solvent consumption. Quantification was performed using an HPLC-MS/MS method using an internal standard. The workflow was validated with spiked blanks before environmental samples were analyzed.

17:45 - 18:00 Towards circular wind energy: Sample preparation strategies in combination with GC×GC-TOFMS for the characterization of wind turbine blade recycling products

• Giulia Giacoppo, University of Ferrara, Department of Chemical- Pharmaceutical- and Agricultural Sciences, Ferrara, Italy.

The rapid expansion of renewable energy technologies has intensified the need for sustainable end‑of‑life strategies for wind turbine blades (WTBs), whose heterogeneous composite structure complicates recycling efforts. In this study, two distinct recycling pathways — solvolysis and pyrolysis — were explored, each requiring a tailored sample preparation workflow prior to detailed molecular characterization of the resulting products.

For solvolysis products, microwave-assisted extraction (MAE) was performed using a hexane–methanol mixture (10:3 ratio), followed by water addition (2.5 ratio) and centrifugation to achieve phase separation. Pyrolysis products, in parallel, were subjected to solid-phase extraction (SPE), which enabled the fractionation of the complex mixture into distinct chemical families.

Subsequently, the MAE extracts and the fractions obtained by SPE were analyzed using comprehensive two-dimensional gas chromatography coupled with time-of-flight mass spectrometry (GC×GC-TOFMS). Compound identification was based on mass spectral electron ionization (EI) database matching at 70 eV (≥800/1000) and the Linear Retention Index (LRI) (±20 range). Additionally, the structured distribution of analytes across the 2D-GC plane was considered.

In total, around 120 compounds belonging to different chemical classes such as hydrocarbons, aromatics, and heteroatom-containing molecules were tentatively identified in the solvolysis products. Regarding the SPE fractions of pyrolysis-derived products, the use of a normal-phase sorbent with eluting solvents of increasing polarity allowed for the separation of hydrocarbon-rich fraction from one enriched in oxygen- and nitrogen-containing compounds.

18:30–19:30 ISC Tube and Quiz

Chairs

• Lucie Nováková, Czech Republic    
• Jiří Urban, Czech Republic

ISC: ISC 2026 - Day 3: ISC Tube chairs Lucie Nováková and Jiří Urban

ISC: ISC 2026 - Day 3: ISC band

ISC: ISC 2026 - Day 3: ISC Quiz winners

ISC: ISC 2026 - Day 3: ISC Tube winner