Higher sensitivity analysis of a large panel of lipid mediators

Targeted profiling using the SCIEX Triple Quad™ 7500 LC-MS/MS System - QTRAP® Ready

Paul Norris, Santosh Kapil Kumar Gorti, Mackenzie Pearson
SCIEX, USA

Abstract

High sensitivity analysis is key for the analysis of lipid mediators in human plasma. Here the SCIEX lipid mediator assay has been expanded and adapted to the SCIEX 7500 System. Sensitivity was compared to the QTRAP 6500+ System and an average gain in peak areas of 53 fold was observed.

12617 7500

Introduction

In biological samples, reliable detection and quantification of lipid mediators can be challenging because these molecules are locally acting, short-lived, and produced in small quantities (often in the femtogram to nanogram range). Detecting lipid mediators is also challenging because of the complexity and diversity of different tissue matrices which can suppress ionization and negatively affect lower limits of detection and quantification when using ESI-MS/MS. Various method development techniques can be employed to help improve the recovery and detection of analytes, such as rigorous extraction protocols or employing microflow chromatography, but these can be laborious, costly, and in some cases too time consuming to perform and maintain routinely.

LC-MS/MS analysis for the targeted quantification of lipid mediators has emerged as a powerful workflow to characterize and profile lipid mediators.1 Still, innovations that can unlock new levels of sensitivity are needed to advance the biological understanding and pathway interpretation of lipid mediator production in a range of physiological processes for personalized and precision medicine research. With technological improvements on the SCIEX Triple Quad 7500 System, the lower limits of quantification and detection have been improved compared to previously attained data from the QTRAP 6500+ LC-MS/MS System.1

In this report, an expanded assay panel with LLOQs down to the lower femtogram range is described. Figure 1 compares the sensitivity levels for the same sample analyzed on the QTRAP 6500+ System and on the SCIEX 7500 System. Highlighting compounds detected near their LLODs and LLOQs, and compounds simply undetected on the QTRAP 6500+ System, significant gains in sensitivity and signal to noise were observed on the SCIEX 7500 System. Several sample matrices, including NIST 1950 Metabolites in Human Frozen Plasma as well as mouse liver, brain and human serum, were analyzed to further demonstrate sensitivity improvements.

Figure 1.  Lipid mediator analysis of NIST 1950 human plasma.  Lipid mediator MRM chromatogram from the SCIEX 7500 System (pink) vs. QTRAP 6500+ System (blue) were overlaid.  Signal for leukotriene C4 and 14-HDHA, and 9(S)-HOTrE were not detected with QTRAP 6500+ System.

Key features of the SCIEX 7500 System for higher sensitivity lipid mediators profiling

  • High-throughput quantitative analysis of lipid mediators is achieved using MRM (multiple reaction monitoring) and the Scheduled MRM™ Algorithm2
    • Expanded MRM assay containing 111 lipid mediators and 16 internal standards3
  • Average peak area gains of 53x for the compounds analyzed in human plasma matrix, compared to the previous generation of instruments
  • Enables detection of compounds in the lower femtogram level on column in complex matrices with high precision and linearity
  • Calculation of the precision, accuracy, sensitivity, and linearity method for all lipid mediators included in the assay
  • SCIEX OS Software for data acquisition and processing
  • Complete method provided for easy adoption