HILIC

A rapid and robust LC–MS/MS method for determining lactulose and rhamnose concentrations in blood plasma was used to determine intestinal permeability from blood plasma, which can help diagnose gastrointestinal diseases such as Crohn’s disease.

A new study has reported the preparation, characterization, and chromatographic evaluation of a nickel oxide deposited silica stationary phase in HILIC, which exhibited multiple retention mechanisms and enhanced chromatographic selectivity for analytes.

The study highlights the importance of developing new chromatographic methods for the analysis of ionizable compounds, such as BAK, and the potential of mixed-mode chromatography for pharmaceutical applications.

In this LCGC blog, we reflect on the movement to make analytical chemistry more environmentally friendly, and how to potentially better those efforts for the industry's future.

This review article discusses the novel separation and detection strategies that are considered promising in clinical metabolomics to enhance the metabolome coverage. It includes hydrophilic interaction chromatography (HILIC), supercritical fluid chromatography (SFC), multidimensional LC approaches, as well as ion-mobility mass spectrometry (IM-MS) and data-independent acquisition (DIA) analysis methods.

This article is an update on the technique of HILIC and covers recent ideas on the mechanism of separation, and how it may be manipulated to suit the separation of particular sample types. The advantages of HILIC are discussed, and also the actual and perceived disadvantages of the technique and how the latter can be overcome. Some new applications of HILIC for characterization of biopharmaceuticals, where it can even be applied to the separation of intact proteins, and to applications in metabolomics, will be discussed.

The Column spoke to Steven Janvier, a PhD student at Sciensano and Ghent University (Belgium), and Celine Vanhee, a scientist at Sciensano, about their work to develop a hydrophilic interaction chromatography (HILIC) methodology capable of detecting counterfeit polar peptide drugs available on the black market.

The Column spoke to Núria Fontanals, a senior researcher at the Department of Analytical and Organic Chemistry, Universitat Rovira i Virgili, Spain, about her work investigating the impact of artificial sweeteners as environmental pollutants using hydrophilic interaction chromatography (HILIC) and the broader role of HILIC in environmental analysis.

Certain details related to mobile-phase preparation that can be assumed in reversed-phase LC separations can be much more consequential in HILIC.

Paying attention to the details of mobile-phase preparation can have a big impact on the reproducibility of hydrophilic-interaction chromatography (HILIC) separations.

LCGC, the leading resource for separation scientists, is proud to announce that Ronald E. Majors and Zachary S. Breitbach are the winners of the 11th annual LCGC Lifetime Achievement and Emerging Leader in Chromatography Awards, respectively. Majors and Breitbach will be honored in a symposium as part of the technical program at the Pittcon 2018 conference in Orlando, Florida, on February 26, 2018.

Valentina D’Atri from the University of Geneva spoke to The Column about the evolving role of HILIC in biopharmaceutical analysis.

These are exciting times to be involved in monoclonal antibody (mAb) and biopharmaceutical analysis. Advances in instrumentation, column technology, and reagents are providing analysts with a new set of tools to broaden their understanding of the highly complex products they are studying. A good example is hydrophilic interaction chromatography (HILIC). While the technique has been used for more than 20 years to profile enzymatically released and fluorescently labelled N-glycans, the introduction of new columns (sub-2-µm and widepore) has paved the way to explore the technique further. Remarkable separations at all levels of analysis, including protein, peptide, and glycan levels, have been demonstrated. With data from the authors’ laboratories, the versatility of HILIC in mAb analysis will be demonstrated in this month’s “Biopharmaceutical Perspectives”.

Valentina D’Atri from the University of Geneva spoke to The Column about the evolving role of HILIC in biopharmaceutical analysis.

Hydrophilic interaction liquid chromatography coupled to electrospray ionization mass spectrometry (HILIC–ESI-MS) has been established as a method to separate and quantify polar and ionic analytes in a direct way for two decades. HILIC separation is based on the polarity of analytes, so the more polar analytes have stronger retention on a HILIC column.

Babies and infants experience rapid growth within a short timeframe and the nutrition that they absorb is therefore of the utmost importance. María Mateos-Vivas from the Department of Analytical Chemistry, Nutrition, and Food Science at the University of Salamanca, Spain, has used hydrophilic interaction liquid chromatography (HILIC) coupled to tandem mass spectrometry (MS/MS) to investigate the role of nucleotides on infant health. She recently spoke to The Column about this research.

The rising numbers of liquid chromatography–mass spectrometry (LC–MS)-based metabolomics applications and untargeted metabolism studies have increased the need for reliable chromatographic separations for a large quantity of molecules with widespread polarities.

The principal aim of this work was to provide a perspective with practical utility in streamlining the chromatographic method development in pharmaceutical industries based upon predicting the chromatographic retention times from molecular structures. Workflows were suggested with a focus on reversed-phase LC, IC, and HILIC as the three major techniques. Unlike HILIC, retention prediction in both reversed-phase LC and IC can benefit from the maturity of these techniques and the transparency of their retention mechanisms. In reversed-phase LC the solute coefficients in the hydrophobic subtraction model and in IC the a and b values in the linear solvent strength model can be the subject of modelling with their subsequent use in retention prediction. A workflow for HILIC can be based on the design of experiments approach, to account for all major contributors to the retention mechanism, and direct correlation of experimental retention times to the molecular descriptors.

Babies and infants experience rapid growth within a short timeframe and the nutrition that they absorb is therefore of the utmost importance. María Mateos-Vivas from the Department of Analytical Chemistry, Nutrition, and Food Science at the University of Salamanca, Spain, has used hydrophilic interaction liquid chromatography (HILIC) coupled to tandem mass spectrometry (MS/MS) to investigate the role of nucleotides on infant health. She recently spoke to us about this research.

Biopharmaceuticals offer great new possibilities and potential in treating various life threatening diseases such as cancer and autoimmune diseases. However, their complexity, resulting from their heterogeneity, represents a great challenge to the analytical chemist and a suite of techniques is required for their characterization and analysis. Reversed phase chromatography, the mainstay of traditional small molecule analysis, is employed at multiple biopharmaceutical levels due to its versatility, inherent relative efficiency and ease of hyphenation to a mass spectrometer.

An excerpt from LCGC’s e-learning tutorial on biochromatography at CHROMacademy.com

A summary of what these techniques can tell us from a qualitative and quantitative perspective

Therapeutic peptides represent one of the fastest growing segments in the pharmaceutical market. To bring these products to the market in a consistent manner, high quality is a major concern and requires stringent quality control (QC) methods. This article discusses the potential of zwitterionic chiral ion-exchangers to support peptide analysis and quality control as a flexible complementary tool to monitor the stereochemical integrity and chemical modifications.

In the example shown here we seek to provide an alternative to HILIC methods by using a novel multi-mode column called the Scherzo SS-C18. This stationary phase has both reversed phase (C18) and polar components with the addition of a high density of strong anion and cation exchange ligands. This strategy for increasing polar retention is much more predictable run to run and does not require lengthy column pre-treatments.

Characterizing Cancer Tumours using HILIC–MS

Hydrophilic interaction chromatography (HILIC) is becoming more popular. Dr Tohru Ikegami from the Department of Biomolecular Engineering at the Kyoto Institute of Technology in Kyoto, Japan, reveals more about the evolution of HILIC, important new developments, and tips on how to get the most from this technique.