LCGC Europe May 2017

May 2017 | Volume 30, Issue 5
Cover Story
Fluorescence polarization (FP) is a highly regarded technique for studying drug–protein interactions, but has limited value regarding protein mixtures. As a novel approach to drug target discovery, the possibility of combining FP with liquid chromatography (LC) was explored. Nondenaturing protein LC principles such as size-exclusion chromatography (SEC), hydrophobic interaction chromatography (HIC), and ion exchange chromatography (IEX) were found to be orthogonal and compatible with FP because the mobile phases used do not negatively affect detection. For simple protein mixtures, the SEC/HIC/IEX–FP approach was able to identify tankyrase as the target of a triazole-based inhibitor of the Wnt signaling pathway, which is heavily associated with colon cancer. However, the total peak capacity of the three LC dimensions was not sufficient to resolve at cell-proteome level, calling for higher resolution of intact proteins to enable stand-alone drug target discovery with LC and FP.
Hot Topics in Separation Science
A recent argument was raised in the scientific press that in pursuit of greater speed and separation resolution, ultrahigh performance liquid chromatography (UHPLC) is faced with practical limitations and will struggle with its own version of Moore’s law.
Lipidomics, the analysis of lipids by mass spectrometric methods, revolutionized lipid science (1). It provides detailed quantitative information on hundreds of lipid species and opens new possibilities to gain an insight into lipid biology. This helps not only to explain the vital role of lipid species as membrane building blocks, but also to unravel their bioactive functions. Thus, lipid species can act as signaling molecules and modulate membrane properties, which are essential for organelle and membrane protein function. Moreover, the first examples demonstrated their potential as novel biomarkers to monitor human health.
The HPLC symposium series is recognized as “the forum” where new developments in liquid phase separations and their hyphenation to mass spectrometry (MS) for the analysis of (bio)pharmaceutical compounds and their metabolites are presented.
The importance of glycosylated structures in modern biology and medicine has been beyond dispute for many years, but there are still gaps in biochemical understanding. The current realization that virtually all major human diseases have been associated with glycosylation changes demands in-depth structural studies of these highly complex glycobiomolecules. Glycoscience with its many directions and a broad scope in both prokaryotic and eukaryotic systems is currently securing its place at the centre stage of modern biological research.
Environmental analyses of food, soil, and water have changed dramatically over the last decade. Topics such as pesticides, food additives, and natural products have become important as food products are globally grown and distributed (1). Monitoring their quality is critical to international business. Pharmaceuticals, fluorinated surfactants, and endocrine disruptors in water are major new topics, where not only parent compounds are unknown but also their metabolites and degradation products are often more important or more abundant than the parent compound (2). New environmental issues, such as hydraulic fracturing and its wastewater, have captured our attention as the production of oil and gas has increased exponentially in the past decade (3). With this technology comes the problem of wastewater disposal and groundwater contamination. These environmental issues have greatly benefited from the combination of ultrahigh‑performance liquid chromatography (UHPLC) mated to high resolution mass spectrometry (HRMS
Some members of the separation science community are still not yet convinced of the value of comprehensive two‑dimensional liquid chromatography (LC×LC). They feel that the large increase in separation power (that is, in peak capacity: the number of component peaks that may possibly be separated) may be compromised by losses in sensitivity and robustness of the separation. However, the chairmen of HPLC 2017 will have seen a great number of abstracts come their way from scientists who want to change this perception.
In the biomedical research of molecular bases of both normal and pathological biological processes, it is currently necessary not only to detect, identify, and quantify individual compounds, but also to study their interactions with endo- and exogenous compounds. Obviously, for these purposes it is crucial to develop new advanced high‑performance analytical methods providing high sensitivity, high selectivity, and high throughput. These challenging requirements are well met by capillary electromigration (CE) methods. They have developed in the last three and half decades into high‑performance separation techniques suitable for the analysis of a wide spectrum of both low- and high‑molecular mass bioactive compounds.
LC Troubleshooting
Several variables can be used to change selectivity in a liquid chromatography (LC) separation. Here we compare the variables in an effort to prioritize which experiments will be most effective.
GC Connections
In this instalment, John Hinshaw presents an annual review of new developments in the field of gas chromatography (GC) seen at Pittcon 2017 and other venues in the past year.
Multidimensional Matters
Comprehensive two-dimensional liquid chromatography (LC×LC) is evolving and becoming more commonly used in practice, but there are some specific problems still present that hamper the widespread use of this technology. One key aspect is the coupling of an on-line LC×LC system to a mass spectrometer. Generally, on-line LC×LC is based on a very fast second dimension separation to achieve low cycle times. This often results in flow rates that are far above the optimum for electrospray ionization mass spectrometry (ESI-MS). This month’s “Multidimensional Matters” looks at the benefits of miniaturization in the first and second dimension for coupling with a high-resolution mass spectrometer (HRMS) and describes an environmental analysis application.
Events
By LCGC Editors
The International Symposium for High-Performance Thin-Layer Chromatography (HPTLC 2017) will take place in Berlin, Germany, 4–8 July 2017.
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