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Untargeted Metabolomics from Biological Sources Using Ultraperformance Liquid Chromatography-High Resolution Mass Spectrometry (UPLC-HRMS)
Published on: May 20, 2013
Response normalized liquid chromatography nanospray ionization mass spectrometry.
Ragu Ramanathan1, Ruyun Zhong, Neil Blumenkrantz
1Department of Drug Metabolism and Pharmacokinetics, Schering-Plough Research Institute, Kenilworth, New Jersey 07033, USA. ragu_ramanathan@yahoo.com
Journal of the American Society for Mass Spectrometry
|September 4, 2007
Summary
This study introduces a response-normalized LC-NSI-MS system for accurate drug and metabolite quantification. The new method minimizes solvent effects, achieving more uniform mass spectrometry responses across diverse compounds.
Area of Science:
- Analytical Chemistry
- Mass Spectrometry
- Pharmacokinetics
Background:
- Liquid chromatography-mass spectrometry (LC-MS) faces challenges in quantitative analysis due to variable responses for different compounds.
- Nanospray ionization (NSI) offers improved MS response consistency but requires LC separation for complex biological samples.
Purpose of the Study:
- To develop and evaluate a response-normalized LC-NSI-MS (RNLC-NSI-MS) system for enhanced quantitative determination of drugs and metabolites.
- To minimize the influence of mobile phase composition on ionization efficiency in LC-MS analysis.
Main Methods:
- A novel LC-NSI-MS system was designed using two HPLC units and a chip-based NSI source coupled to a Q-TOF mass spectrometer.
- Post-column solvent addition was employed to maintain an isocratic mobile phase composition entering the NSI source throughout the HPLC run.
- The system was tested with four classes of pharmaceutical compounds and their metabolites, including vicriviroc (VCV) and desloratadine (DL).
Main Results:
- The RNLC-NSI-MS system minimized the impact of solvent environment on ionization efficiency, leading to more uniform MS responses.
- Conventional LC-ESI-MS overestimated VCV and DL responses by 6- and 20-fold compared to radiochromatograms.
- LC-NSI-MS showed improved accuracy, with VCV and DL responses within 2- to 6-fold of radiochromatographic data, and response normalization further improved uniformity.
Conclusions:
- The developed RNLC-NSI-MS system significantly improves the quantitative accuracy and consistency of drug and metabolite measurements in biological samples.
- This approach offers a more reliable alternative to conventional LC-MS methods, especially when reference standards are unavailable.
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