Liquid Chromatography (LC/HPLC)

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In this month's installment of "LC Troubleshooting," John Dolan takes a look at solvent strength and sees how we can use it to make predictable changes in the peak movement for reversed-phase LC separations.

Globally, 2005 sales of pharmaceuticals have been estimated at approximately $550 billion. A significant fraction of this amount was due to the top pharmaceutical companies in the world. The top five companies alone were responsible for total revenues of $168 billion, or 30% of the entire market.

Testing helps you identify the soft spots in the method, select the best conditions for operation and develop troubleshooting strategies to correct problems when they do occur.

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This article looks at current practices in bioanalytical chemistry by examining and critically assessing the various parameters that can be altered to achieve high-speed results with high resolution in LC–MS applications. The decision to opt for gradient or isocratic elution is also discussed.

John Dolan ponders te question of whether or not you will be prepared when that method eventually fails.

Ronald E. Majors presided over "HPLC: Optimization and Chemistry," an oral session held Tuesday afternoon that focused on the optimization of analytical- and preparative-scale LC separations.

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Check valves can be replaced with new ones, but often a simple cleaning procedure is sufficient to restore normal operation.

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Capillary gas chromatography (GC) is a well-established separation technique. Rick Parmely reviews some of the capillary column basics by investigating a standard test mixture and observing some of the chromatographic effects that can affect peak response, peak shape and column bleed. He provides guidelines for obtaining the best performance from a capillary column. He discusses peak tailing, column overload, ghost peaks and column bleed and speculates on "when to give up." He concludes with a set of fundamental steps to be used in achieving better GC analyses.

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Separation and retention of both polar and nonpolar compounds by the same stationary phase can be a useful approach for analyses of complex samples with a broad range of chemical properties. Typical stationary phases are designed for retention of either polar or nonpolar compounds so that multiple steps are required when designing a separation strategy. Hydride silica–based stationary phases are new materials with properties that allow for the simultaneous retention of both polar and nonpolar compounds over a range of aqueous–organic mobile phase compositions. Adjustment of the aqueous–organic ratio will determine whether polar or nonpolar compounds have greater retention.

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This article assesses the extent to which some familiar HPLC detectors can provide reliable quantitative analytical measurements when no suitable primary reference standard exists. The advantages and limitations of two candidate detector schemes are discussed ? evaporative light-scattering detection (ELSD) and chemiluminescent nitrogen detection (CLND). It was found that when the ERETIC (Electronic REference To access In-vivo Concentrations) method in proton NMR was used as a "gold standard" reference procedure the ability of CLND to provide reliable single calibrant quantification is superior to ELSD. Furthermore, ELSD showed bias towards underestimation of chromatographically-resolved impurities, resulting in an overestimation of analyte purity.

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LCGC North America and its companion publication, LCGC Europe, are targeted for chromatographers in the United States and Europe, respectively. Sometimes I wonder whether or not the same types of problems are encountered by workers in liquid chromatography (LC) on both sides of the Atlantic. As I write this installment of "LC Troubleshooting," I have just completed three weeks of teaching LC short courses in Europe. I can assure you that the content of the questions I get in these classes is the same, no matter where they are taught. This column is based upon questions gleaned from the most recent courses - you can see that there are no national borders for LC problems.