Analytical instruments are indispensable for modern society, industry, research and innovation. The samples subjected to analysis and the questions asked to chromatographers are increasingly complex. To assess the composition and to characterize properties or “sample dimensions” of highly complex samples, a multitude of analytical techniques are employed. However, in many cases these techniques are applied independently and the most-relevant sample-dimensions, and the relations between these, cannot be assessed.
In this research line we develop strategies to couple analytical separations to allow multiple chemical properties – and their correlation – to be characterized simultaneously. To achieve this, we combine (multi-dimensional) separation technology, with mass spectrometry and chemometric methods.
Our work also aims to fundamentally improve the coupling of otherwise-incompatible LC separation systems in the context of 2D liquid chromatography. We have contributed to studies to the use of active-modulation systems such as stationary-phase-assisted modulation, but also thermal modulation for polymer separations.
We recently collaborated with a consortium headed by Prof. Maarten van Bommel to develop two-dimensional separation methods featuring in-line light-induced degradation. To achieve this, researchers from the Vrije Universiteit Amsterdam Dr. Iris Groeneveld developed a liquid-core waveguide reactor (see prototype in the video left) with Dr. Freek Ariese and Prof. Govert Somsen. The reactor was embedded in a 2D-LC system at the CAST facilities of the University of Amsterdam and applied to the in-line degradation of food dyes. Using the reactor, molecules could be isolated from a mixture and selectivity degraded over time.
Liquid-core waveguide reactor developed by Dr. Iris Groeneveld.
This research line draws significant inspiration from the sample dimensionality theory by Giddings. In a nutshell, analyte molecules can be regarded as comprising of several molecular building blocks (“sample dimensions”). According to Giddings, a separation system can target specific sample dimensions. To fully characterize the sample, each sample dimension requires a separate system dimension. This is one of the rationales for multi-dimensional chromatography.
However, the same theory also implies that chemical properties hidden in the molecular structure (in the figure depicted as Sample Dimension 3) cannot be targeted by a separation. For instance, a nanoparticle comprises of large numbers of polymer macromolecules. As long as the nanoparticle is intact, the polymers cannot be studied.
In this research line, we develop strategies to correlate accessible chemical properties to previously-inaccessible properties using a sample transformation mechanism.
Visit the special page on the public-privately funded PARADISE project. The project aims to resolve pressing matters in industry and society using 2D-LC. The project is funded by a consortium of Covestro, DSM, Genentech, NFI, Shell and the Dutch Research Council (NWO).
Nanoparticle Analysis by Online Comprehensive Two-Dimensional Liquid Chromatography combining Hydrodynamic Chromatography and Size-Exclusion Chromatography with Intermediate Sample Transformation, B.W.J. Pirok, N. Abdulhussain, T. Aalbers, B. Wouters, R.A.H. Peters and P.J. Schoenmakers, Anal. Chem., 2017, 89 (17), 9167–9174, DOI: 10.1021/acs.analchem.7b01906
A comprehensive 2D-LC method was developed based on hydrodynamic chromatography and size-exclusion chromatography. The method was used to correlate the particle-size distribution of waterborne polymeric nanoparticles dispersed, to the molar-mass distribution (MMD) of the constituting polymers. The transformation to polymers was realized using active dissolution with a strong organic solvent.
Analysis of Charged Acrylic Particles by On-line Comprehensive Two-Dimensional Liquid Chromatography and Automated Data-Processing
B.W.J. Pirok, N. Abdulhussain, T. Brooijmans, T. Nabuurs, J. de Bont, M.A.J. Schellekens, R.A.H. Peters and P.J. Schoenmakers, Anal. Chim. Acta., 2019, 1054, 184-192, DOI: 10.1016/j.aca.2018.12.059
The earlier-developed method was extended to now also be compatible with acid-based nanoparticles which are created using more polar polymer systems. To achieve this, the active-modulation strategy was adapted. To allow the raw data to be useful to the field, a data-processing algorithm was created.
On-line Microfluidic Immobilized-Enzyme Reactors: A New Tool for Characterizing Synthetic Polymers, B. Wouters, B.W.J. Pirok, D. Soulis, R.C. Garmendia Perticarini, S. Fokker, R.S. van den Hurk, M. Skolimowski, R.A.H. Peters and Peter J. Schoenmakers, Anal. Chim. Acta., 2019, 1053, 62-69, DOI: 10.1016/j.aca.2018.12.002
This study details the development and evaluation of an immobilized-enzyme reactor (IMER). It was hyphenated with an LC separation. Polymers could be digests within minutes, which was in line with the requirements for a comprehensive 2D-LC workflow. The work was a collaboration between Dr. Bert Wouters and Dr. Bob Pirok.
Thermal modulation to enhance two-dimensional liquid chromatography separations of polymers, L.E. Niezen, B.B.P. Staal, C. Lang, B.W.J. Pirok, P.J. Schoenmakers, Chromatogr. A, 2021, 1653, 462429, DOI: 10.1016/j.chroma.2021.462429.
This work proposes the use of thermal modulators for improved 2D-LC analysis of polymers. A proof-of-principle modulator was designed for this study by Dr. Leon Niezen together with Dr. Bob Pirok and Prof. Peter Schoenmakers. This work was conducted in close collaboration with Dr. Bastiaan Staal (BASF).
Co-Polymer sequence determination over the molar mass distribution by size-exclusion chromatography combined with pyrolysis – gas chromatography, W.C. Knol, T. Gruendling, P.J. Schoenmakers, B.W.J. Pirok, R.A.H. Peters, Chromatogr. A, 2022, 1670, 462973, DOI: 10.1016/j.chroma.2022.462973
Dr. Wouter Knol developed together with Prof. Ron Peters and Dr. Till Gruendling (BASF) an interface to hyphenate SEC with Py-GC-MS. This was done to allow the polymer sequence distribution to be determined as a function of the molar-mass distribution. It was the first in a series of publications building towards a modulation strategy to hyphenate size-based liquid-phase separations with pyrolysis gas chromatography.
Assessing the feasibility of stationary-phase-assisted modulation for two-dimensional liquid chromatography separations, J. den Uijl, T. Roeland, T.S. Bos, P.J. Schoenmakers, M.R. van Bommel, B.W.J. Pirok, J. Chromatogr. A, 2022, 1679, 463388, DOI: 10.1016/j.chroma.2022.463388
Stationary-phase-assisted modulation is one of the most-used active-modulation strategies in 2D-LC. However, one problem is that analytes may prematurely elute from the trapping columns. This causes the method to fail according to the comprehensive 2D-LC criterion that all analytes must be analyzed by both dimensions. As the use of light-induced reactors creates molecules with unknown properties, a method to quickly assess the feasibility of using this modulation strategy was developed by Dr. Mimi den Uijl.
Combining photodegradation in a liquid-core-waveguide cell with multiple-heart-cut two-dimensional liquid chromatography, M.J. den Uijl, J.H.L. van der Wijst, I. Groeneveld, P.J. Schoenmakers, B.W.J. Pirok, M.R. van Bommel, Chem. 2022, 94(31), 11055-11061, DOI: 10.1021/acs.analchem.2c01928
This work describes the first development of light-induced degradation embedded in a 2D-LC setup for enhanced molecule analysis.
Characterization of dye-loaded poly(lactic-co-glycolic acid) nanoparticles by comprehensive two-dimensional liquid chromatography combining hydrodynamic and reversed-phase liquid chromatography, J. Verduin, L. Tutiš, A. Becking, A. Famili, K. Zhang, B.W.J. Pirok, G.W. Somsen, Chem. 2023, DOI: 10.1021/acs.analchem.3c03356
In this study, a method was developed to characterize PLGA-PEG nanoparticles based on their particle-size distribution and their cargo. To mimic a drug-delivery system, the nanoparticles were loaded with a dye. This work was conducted by Joshka Verduin from the Vrije Universiteit Amsterdam.
Expected Q1 2025
We have extensive experience with developing reactors for use within separation technology. Since 2015, we have studied various mechanisms for in-line sample transformation.
Nino Milani and co-workers reviewed the latest developments in 2D gas chromatography and found a surprising dynamic field for what is considered to be an established technique.
In Amsterdam, this research line was started by Prof. Peter Schoenmakers (CAST, University of Amsterdam), Prof. Ron Peters (Covestro) and Prof. Govert Somsen (VU Amsterdam) in 2015 with the Making Analytically Incompatible Approaches Compatible (MANIAC) project. The project sought to develop modulation reactors for complex samples. Dr. Niall Macdonald developed a immobilized-microbe reactor, Dr. Bert Wouters and Dr. Bob Pirok an immobilized-enzyme reactor. The project received funding from the Dutch Research Council, but also DSM, Heineken, Micronit Microfluidics and Shell. Bob Pirok defended his PhD thesis on the MANIAC project in 2019.
Elements of this project were later adopted by Prof. Maarten van Bommel in his TooCOLD project. Under his lead, a team of scientists from the VU (Dr. Freek Ariese, Prof. Govert Somsen, and Dr. Iris Groeneveld) developed the liquid-core waveguide reactor, whereas the UvA team of Prof. Peter Schoenmakers, Dr. Bob Pirok, Dr. Mimi den Uijl implemented and applied it in a 2D-LC separation system. The project was funded by the Dutch Research Council and Unilever.
Around the same time, Prof. Govert Somsen and Prof. Peter Schoenmakers started the UNMATCHED project with BASF, Covestro, DSM and Nouryon. This project focused on the development of separation systems for polymer systems.
In 2021, Dr. Bob Pirok wrote the PARADISE project and obtained funding with Prof. Arian. van Asten, Prof. Ron Peters and Prof. Govert Somsen. The team hired PhD candidates Joshka Verduin, Rick van den Hurk and Nino Milani to further develop multi-dimensional methods for complex sample analysis and complementary chemometric methods for data analysis. The project is a collaboration with Covestro, DSM, Genentech, the Dutch Forensics Institute and Shell.
Currently, Dr. Bob Pirok continues the research line in Amsterdam together with Prof. Ron Peters and Dr. Tijmen Bos.
