Università degli Studi di Ferrara Politecnico Milano 1863 University of Milano-Bicocca

Plenary Lectures

Martin J. Kemp

Emeritus Professor of the History of Art, Oxford University, Oxford, UK


Music for Leonardo

by Professor Emeritus Martin Kemp & Conservatorio di Milano "Giuseppe Verdi"

We are presenting a programme of music on the occasion of the 500th anniversary of Leonardo's death. The music has been chosen to resonate imaginatively with particular masterpieces by Leonardo in a timeless way, rather than attempting to reconstruct music from his own period.

Martin Kemp will provide an introduction on Leonardo and music, looking at his own musical activities and the views he expressed about music as a ’science', especially in his Paragone, the comparison of the arts. Each of the pieces of music will be preceded by a brief account of the painting, and each will be accompanied by the relevant image. The paintings will be the Annunciation, the Virgin of the Rocks, the Portrait of a Man with a Sheet of Music, the Portrait of Cecilia Gallerani, the Last Supper, and (of course) the Mona Lisa. The composers are Mozart, Schubert, Puccini, Holst and Verdi.

The pieces of music, were selected in agreement with Maestro Massimiliano Baggio, vice director of the Conservatorio di Milano “Giuseppe Verdi”, and Maestro Stefania Mormone, responsible for internal external artistic activities.
Musics will be interpreted by the students of the Conservatorio.

Attila Felinger

University of Pécs, Hungary


The art of chromatography

Chromatography has strong connections with arts. The term ‘chromatography’ was first used in arts, describing a treatment of colors and pigments, and their powers in painting. Scientific reasoning and the explanation of the phenomena occurring in separation science are often facilitated using demonstrations taken form the visual arts. The aim of this presentation is to demonstrate how closely arts and separation science are related – in particular when separation science is seen through Italian or Hungarian eyes.

The presentation is going to focus on a number of recent fundamental developments that contributed to the progress of separation science or chromatography. The better understanding of the details of the separation process is of utmost importance for the development of column technology or instrumentation. In return, novel technical solutions raise new fundamental questions. Considerable progress was made in column technology over the last decade. The progress in the technology of monolithic columns, sub-2-μm particles and core-shell particles is accompanied with new problems that call for explanation. Columns packed with these particles are several times more efficient than the conventional columns were. They permit the achievement of the same separations in a markedly shorter time or of more efficient separations in the same time. The understanding the origin of the improved level of performance brought great new opportunities and important challenges. The probability for analyte molecules to move from the eluent percolating through the bed to the eluent stagnant inside the particles and eventually to the surface of the stationary phase depends on several factors that must be determined.

These advances are discussed and illustrated with emphasis on experimental results, concepts and explanations.

Koji Otsuka

Chair of HPLC 2019 Kyoto, Japan
Kyoto University, Japan


Microscale liquid phase separations using specific interactions

As a unique LC separation medium, in which a specific interaction against a biogenic compound can be occurred, a spongy-like porous polymer or spongy monolith (SPM) consisting of poly(ethylene-co-glycidyl methacrylate) has been developed [1]. The SPM is prepared simply by blending a thermoplastic resin above its melting point with water-soluble pore templates. After removal of the pore template by washing with water, the resultant SPM contains large flow-through pores (>10 μm in diameter), and a column made of the SPM facilitates separation mediated by hydrophobic interactions and/or ion exchange at a high flow rate. The SPM can also be prepared in any shapes and easily packed into a column. Therefore, we expect that the SPM containing specific functional groups, e.g., epoxy groups, would be useful for affinity chromatography and overcome the limitations of current media. Immobilization of protein A onto the SPM (ProA-SPM) enabled high-yield collection of IgG from cell culture supernatant even at a high flow rate. The immobilization of pepsin onto the SPM (Pep-SPM) enabled efficient online digestion at a high flow rate, whereas the immobilization of concanavalin A (ConA-SPM) enabled lectin affinity chromatography for the separation of glycoproteins.

To use specific interactions in liquid phase microscale separation techniques, several novel materials to be used for separation fields have been developed. Among such specific interactions, π-interactions, e.g., π−π, CH−π, and halogen−π interactions, are considered as one of major functions contributing to intermolecular affinities for ligand recognitions and ordered structures in biomolecules.

We have developed C60-fullerene (C60) bonded monolithic capillaries for LC separations and investigated their retention characteristics, especially for π-interactions. In addition to C60, C70-fullerene (C70) bonded silica monolithic capillary was also developed. Specific separations based on effective interactions were achieved for polycyclic aromatic hydrocarbons [2]. Especially, a hemispherical molecule, i.e., corannulene (Crn), was significantly retained on the C70 column [3]. According to the computer simulation of the molecules, the specific interaction might be caused by the deflection of the π-electrons-density in C70 [4]. We prepared two types of Crn-immobilized silica monolithic columns, in which the differentiation of π interactions was investigated [5].

[1] Kubota, K.; Kubo, T.; Tanigawa, T.; Naito, T.; Otsuka, K. Sci. Rep. 2017, 7: 178.
[2] Kubo, T.; Kanao, E.; Matsumoto, T.; Naito, T.; Sano, T.; Yan, M.; Otsuka, K. ChemistrySelect 2016, 1, 5900-5904.
[3] Kanao, E.; Naito, T.; Kubo, T.; Otsuka, K. Chromatography 2017, 38, 45-51.
[4] Kanao, E.; Kubo, T.; Naito, T.; Matsumoto, T.; Sano, T.; Yan, M.; Otsuka, K. J. Phys. Chem. C 2018, 122, 15026-15032.
[5] Kanao, E.; Kubo, T.; Naito, T.; Matsumoto, T.; Sano, T.; Yan, M.; Otsuka, K. Anal. Chem., in press.

Mary Wirth

Chair of HPLC 2020 San Diego, USA
Purdue University, West Lafayette, USA


Polymer brush bonded phases for MS-compatible mobile phases in protein separations

HPLC-MS is a powerful, essential combination in protein analysis, but the optimal mobile phase compositions for HPLC and MS are at odds. TFA is typically best as acidic modifier in HPLC but it suppresses ionization and gives ion-pairing in MS. Polymer brushes, formed by surface initiated polymerization on silica, can be made sufficiently thick to mediate silanol interactions. In addition, the abundance of monomers give a wide variety of bonded phases that for tailoring selectivity. Enhanced resolution and sensitivity in HPLC-MS are demonstrated for RPLC, native-RPLC, HILIC, and packed-capillary electrophoresis of intact proteins, particularly mAbs and ADCs.

Michael Lämmerhofer

Chair of HPLC 2021 Düsseldorf, Germany
University of Tübingen, Germany


Lipidomics: a window of opportunity for clinical analysis

Lipidomics approaches are nowadays widely adopted to have a comprehensive view on lipid profiles in biological samples. Alterations in lipid profiles may provide a panel of biomarkers for diagnostic and prognostic purposes of various diseases. Untargeted lipidomics workflows may generate new hypothesis for supporting biological interpretations or complementing other omics data. Targeted lipidomics approaches are often utilized to validate biomarkers found in a discovery phase or measure particular lipids associated with a certain disease or present in low abundance (e.g. lipid mediators in inflammation). Considering the importance of many lipids in signaling pathways and membrane integrity, lipidomics has gained wide interest not only amongst analytical chemists but it has also raised broad attention in the clinical research community.

In this presentation, different workflows commonly employed in lipidomics will be evaluated against each other and critical issues discussed. Shotgun lipidomics by direct infusion suffers from ion suppression, in-source fragmentation and assay specificity problems. Therefore, UHPLC-MS/MS with data-dependent acquisition (DDA) using Orbitrap or QTOF instruments is state of the art in untargeted lipidomics and lipid species separation by RPLC is widely adopted as separation method of choice. It has been criticized that internal standards (commonly one per lipid class), do not coelute with the lipid species and hence this approach is suboptimal for quantitative analysis. Alternative approaches make use of lipid class separation, e.g. by SFC-MS/MS. In lipid class separation the distinct lipid species closely elute with the lipid class specific internal standards which supposedly better correct for matrix effect making quantitative results more accurate. Lipid class separation with HILIC-MS/MS is rarely reported due to the problem that neutral lipids (TGs, DGs, MGs, CEs) elute very early close to the void volume and are barely separated causing assay specificity problems for these classes of lipids.

In this presentation, a HILIC-MS/MS method for lipid class separation and RPLC-MS/MS method for lipid species separation both combined with ESI-QTOF-MS detection in data independent acquisition (DIA) mode using SWATH (sequential window acquisition of all theoretical fragment ion mass spectra) are validated and compared to each other.

Aside from methodological aspects, the power of lipidomics for generating useful information in clinical research will be documented by a number of clinical applications.