Liquid Chromatography (LC/HPLC)

The use of dual polarization interferometry (DPI) to measure the behaviour of alkyl-modified surfaces under different solvent regimes is described. The empirically observed and modelled behaviour of grafted long alkyl chain adlayers on a silica surface were in good agreement. The empirical data was also compared with data using analogous systems and these also matched well. This approach has the potential to gain much greater insight into the behaviour of reversed-phase (RP) columns and could help to design column surfaces that are resistant to phase collapse.

In 2001, the second glossary of common and not-so-common terms and "buzz words" for reference to HPLC columns and column technology was published. It is time for an update since new terms have arisen or, in some cases, their original meanings have expanded or changed.

High performance liquid chromatography (HPLC) enhances throughput in countless processes, including environmental analysis, dissolution testing, separation of biomolecules, and total drug analysis. To take advantage of the benefits of HPLC technology, samples and solvents must be cleared of particulate contaminants. Doing so protects instrument components and minimizes maintenance costs. This article outlines HPLC technology, explains the importance of sample preparation, and reviews the factors that must be considered when preparing HPLC samples by membrane filtration. The data from experiments to test these factors show that the physicochemical characteristics of membrane filters can affect HPLC results.

In this installment of "Column Watch," Ron Majors examines the various approaches to increasing the speed of high performance liquid chromatography (HPLC) separations.

How to avoid an expensive shot in the dark.

The retention behaviour of several compounds has been compared for their selectivity using reversed-phase high performance liquid chromatography with binary water mobile phases composed of methanol, acetonitrile or tetrahydrofuran as modifiers.

BMC Pediatrics reports on a study characterizing total homocysteine (tHcy) levels at birth in preterm and term infants and identifing associations with intraventricular hemorrhage (IVH) and other neonatal outcomes such as mortality, sepsis, necrotizing enterocolitis, bronchopulmonary dysplasia, and thrombocytopenia.

Gerald Drager and colleagues from University of Hannover in Leibniz, Germany, have developed a system that enables immobilization and purification of enzymes in the same reactor, which can then be used for a variety of enzymatic syntheses. Using this technique it is possible to obtain ready-to-use enzyme reactors from crude protein mixtures within minutes.

A new study by Frost & Sullivan shows that the liquid chromatography (LC) market in the U.S. will grow from an estimated $596.3 million in 2006 to $1.018 billion by 2013, thanks to increasing interest and use from both the commercial and research communities.

Nanostream (Pasadena, Calif.) has launched a portal for its customers to deliver real-time access to company and product information. The portal will also act as a way to facilitate troubleshooting for the company's micro parallel liquid chromatography (uPLC) products and provide product updates.

For the last six months, "LC Troubleshooting" has been concentrating on the process for the development of isocratic methods (those for which the mobile phase composition is constant throughout the run). The use of a gradient scouting run to speed isocratic method development is the subject of this month?s "LC Troubleshooting."

Proteins can be separated according to their isoelectric points on an ion-exchange column using pH gradients in the first dimension and according to their hydrophobicity on an RP column in the second dimension.

How do I get the most out of my efforts?

This month's instalment of "MS in Practice" provides a slightly different view of how practitioners employ the skills of interpretation that have been the focus in recent columns.

In the previous installments of this series on efficient development of LC methods (1?5), we have concentrated on improving resolution by modifying the mobile phase, choosing a different stationary phase, or changing some other condition, such as column temperature. In this month?s "LC Troubleshooting" installment, we take a look at trading some of that resolution for a faster separation.

In this installment of "Column Watch," columnist Ron Majors examines the role of pressure in high performance liquid chromatography (HPLC) from two viewpoints: the impact of the ultrahigh pressures encountered in ultrahigh-pressure liquid chromatography (UHPLC) on chromatographic parameters and increases in column pressure encountered in normal daily use. The latter is of more practical consequence to HPLC users since increased back pressure usually implies that something has gone wrong with the column. Pressure increases due to physical and chemical contamination are explored and practical approaches to solve these problems are suggested.

Basic information on stereochemistry is provided in this article to help readers develop a better understanding of the separation mechanisms that come into play in various separation methods used for chiral compounds. This knowledge can allow readers to select a desirable chiral separation method, based upon the molecular structure of the chiral compound of interest. Logical reasons for the selection process are discussed later in this article.

Faster isn't always better.

High performance liquid chromatography–solid phase extraction–nuclear magnetic resonance (HPLC–SPE–NMR) is a novel hyphenation technology that concentrates single chromatographic peaks to elution volumes matching those of NMR flow probes. The SPE unit facilitates the solvent exchange from the mobile phase of the optimized HPLC assay to a deuterated NMR solvent. The well-defined NMR solvent conditions make spectra comparisons feasible, which means databases and spectra catalogues can be used to swiftly identify analytes. The ability to accumulate analytes on the SPE cartridges by multiple trapping reduces the need to perform residual solvent suppression experiments and allows heteronuclear NMR experiments to be performed overnight. Structure elucidation of natural products directly from crude extract HPLC samples has become the key application of this technique.

This month, Chromatography Online's Technology Forum looks at the topic of HPLC/TLC and the trends and issues surrounding it. Joining us for this discussion are Eike Reich from Camag, Gerda Morlock, from University of Hohenheim, and Colin poole, from Wayne University.

Polymer-based columns still are widely used in size-exclusion chromatography (SEC) and ion chromatography-ion-exchange chromatography - they have always been used in these modes, even back in 1987.

The unique features of silica hydride-based columns are described with a focus on how these columns can expand a laboratory's capabilites in HPLC and LC-MS. Of particular interest is the ability to retain both polar and nonpolar compounds and provide an orthogonal method of analysis for currently existing procedures.

Shimadzu Scientific Instruments (Columbia, Maryland) and Groton Biosystems (Boxborough, Massachusetts) recently announced a collaboration effort to interface Groton’s online sampling monitors with Shimadzu’s high performance liquid chromatography (HPLC) system to form a closed-loop solution for online analysis.

Gilson, Inc. (Middleton, Wisconsin), an industry leader in purification systems for high performance liquid chromatography (HPLC), solid phase extraction (SPE), and gel permetation chromatography (GPC) clean-up, now offers direct support and sales in the United States.

Thermo Fisher Scientific Inc., a leader in the separation sciences and chromatography industries, announces the launch of its enhanced partner program. The program is designed to support the expansion of the companies' informatics activity via its partners around the globe, the world class program will utilize business partners to better meet customer needs in multiple industries. Thermo Scientific?s informatics offerings include laboratory information management systems, chromatography data systems, and spectroscopy software.

This article describes the factors that affect the selection of columns for two-dimensional (2D) LCÃ-LC separations. The maximum increase in peak capacity compared with single-dimension (1D) separations is obtained by using "orthogonal" systems employing various combinations of separation mechanisms to provide as different separation selectivities as possible for the sample compounds in the first and in the second dimension systems. To obtain best results, matching the chemistry of the stationary phase, column dimensions and mobile phases in the first and in the second dimensions is essential for successful separations, especiall for comprehensive LCÃ-LC.

Getting started on the right foot is important for efficient method development.

How to avoid an expensive shot in the dark.

An ultrahigh-pressure liquid chromatography (UHPLC) method was developed to separate paroxetine from several of its related compounds using a systematic screening protocol that monitors combinations of selectivity factors including column chemistry, organic modifier, and pH. When the best combination of these factors was selected, the method was optimized by varying gradient slope and temperature.

HPLC 2007 was held in Ghent, Belgium in June. Last month, columnist Ron Majors summarized some the important column developments as well as other Symposium highlights. This month, he winds up coverage with additional highlights in the areas of technology and applications. Among the topics covered are stationary phase preparation and characterization, multi-dimensional and comprehensive LC, temperature studies, detectors and an application overview.