Rapid Analysis of Rifampicin Using Discovery LC-MS Stationary Phases

By: David Bell, Reporter EU Volume 13

David Bell dbell@sial.com


The desire for quality data in an increasingly timely manner places a significant burden on the modern analytical laboratory. In this article, the use of short LC-MS columns are shown to provide faster analysis times while retaining adequate resolution of the target analytes. The use of shorter dimension columns is often an effective choice where high throughput is required.

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There exists a great interest in the modern analytical laboratory to reduce analysis time. One approach has been to use short HPLC columns to reduce chromatographic run times. The most common column length used in HPLC is 15cm. Although such lengths are sometimes necessary for adequate resolution, shorter columns can be substituted in situations where enough resolution is retained. Furthermore, with the growing use of LC-MS systems, qualitative and quantitative information is not as dependent on chromatographic resolution because the mass spectrometer is capable of providing additional selectivity based on mass resolution.

In this study the chromatographic run time for the analysis of the antibiotic rifampicin is contrasted between traditional column geometries and new, shorter columns. Adequate resolution is retained with a concomitant reduction of up to 10 minutes per chromatographic run.

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Rifampicin (Cat. No. R3501) was run on Discovery C18 columns with internal diameters of 2.1mm and lengths of 2 and 15cm. See conditions in Figures B-C. The results were contrasted in terms of the retention of adequate resolution and the reduction in analysis time.

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The structure of rifampicin is provided in Figure A. Figure B shows a rifampicin analysis as acquired on a 15cm x 2.1mm Discovery C18 column. Two major responses are observed that are extensively resolved. Figure C depicts the same separation on a Discovery C18 2cm x 2.1mm column. As shown in this figure, the run time is reduced from approximately 12 minutes to just under 2 minutes while the resolution of the two responses is conserved. Adequate retention and resolution promises to provide a system capable of suitable qualitative and quantitative analysis. The reduced analysis time allows for a 6-fold increase in throughput.

Figure A. Rifampicin

Figure B. Rifampicin on Conventional Discovery C18 Columns

Figure C. Rifampicin on Short Discovery C18 Columns

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The modern analytical laboratory is required to provide quality analytical data in a short amount of time. Faster analyses need not, however, necessitate a compromise in data integrity. Where standard analytical column dimensions (15cm, for example) provide significant resolution of target analytes, shorter columns may be an intelligent substitute. The shorter columns often provide a means of significantly reducing run times without loss of adequate resolution. Where mass spectrometry is utilized, loss of chromatographic resolution may be offset by the mass resolving power of the mass spectrometer.

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