Evaluation of a Multi-Mode ODS Column for the Separation of Polar Metabolites
The analysis of polar, ionic metabolites is important for drug discovery and biomarker research. Often times, it is necessary to separate these metabolites from each other and biological matrixes prior to MS detection (1).
The analysis of polar, ionic metabolites is important for drug discovery and biomarker research. Often times, it is necessary to separate these metabolites from each other and biological matrixes prior to MS detection (1).
Figure 1
Experimental and Results
All data was generated with semi-micro HPLC system equipped with UV or ELS detection. Solutes were separated using LC–MS compatible conditions on Unison UK-C18 (conventional ODS) and Scherzo SM-C18 (Multi-Mode ODS consisting of ODS + anion + cation ligands, Figure 1). Figure 2 shows the separation of a steroid hormone from its metabolites. 17b-estradiol (neutral) showed similar retention on both phases — indicating reversed-phase is the main mode of retention. The glucuronide and disulfate metabolites are poorly retained on conventional ODS, but show improved retention on Scherzo SM-C18 (due to anion exchange). Figure 3 shows the separation of mevalonic acid and mevalonolactone. Mevalonic acid shows improved retention on Scherzo SM-C18 due to anion exchange. Figure 4 shows the separation of glutathiones (both reduced and oxidized forms). These compounds are difficult to separate on conventional ODS, but show improved retention/separation on the Multi-Mode ODS (cation + anion exchange).
Figure 2
Figure 3
Conclusion
Scherzo SM-C18 (Multi-Mode ODS) shows improved selectivity for polar metabolites over conventional ODS. This column should be useful for scientists who work with polar metabolites, as well as those who require an ODS column with different selectivity.
Figure 4
References
(1) "Using Mass Spectrometry for Drug Metabolism Studies;" Korfmacher, W.A., Ed.; CRC Press: Boca Raton; Fl, 2005.
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SEC-MALS of Antibody Therapeutics—A Robust Method for In-Depth Sample Characterization
June 1st 2022Monoclonal antibodies (mAbs) are effective therapeutics for cancers, auto-immune diseases, viral infections, and other diseases. Recent developments in antibody therapeutics aim to add more specific binding regions (bi- and multi-specificity) to increase their effectiveness and/or to downsize the molecule to the specific binding regions (for example, scFv or Fab fragment) to achieve better penetration of the tissue. As the molecule gets more complex, the possible high and low molecular weight (H/LMW) impurities become more complex, too. In order to accurately analyze the various species, more advanced detection than ultraviolet (UV) is required to characterize a mAb sample.
