LipidSearch 5.0: A New Software for Processing Data from Direct Infusion and LC-MS High Resolution Mass Spectrometry Based Lipidomics Workflows

Importance of the Topic

Lipidomics is essential for understanding the role of lipids in health, disease and biological pathways. High-resolution accurate mass (HRAM) spectrometry workflows, including direct infusion and LC-MS, generate complex data sets that require integrated software tools for efficient and standardized lipid identification and quantitation. Advances in automated data processing are critical to expand high-throughput untargeted lipidomics applications and to harmonize reporting standards across laboratories.

Study Objectives and Overview

This work introduces Thermo Scientific LipidSearch 5.0 software, designed to streamline data analysis from direct infusion and LC-MS HRAM workflows. The primary goals were to create an integrated environment for precursor and product ion searches, support custom database configuration, and enable reliable merging of MS and MS/MS results for comprehensive lipid annotation.

Methodology and Instrumentation

• Sample Preparation: A mixture of 32 deuterated and native lipid standards was combined with SPLASH internal standards and infused in isopropanol/methanol/chloroform with ammonium formate.
• Mass Spectrometry: Data were acquired on an Orbitrap Fusion Lumos MS (120K–500K resolution) and additionally on a Q Exactive HF (240K/120K resolution). Full MS scans were followed by data-dependent MS/MS (ddMS2) with stepped HCD collision energy.

Results and Discussion

The software successfully identified all 32 target lipids at the sum-composition level and confirmed the majority at the molecular species level. Out of 33 standard components, two isomeric ceramide species co-eluted and a false positive cholesterol fragment was filtered by MS/MS validation. Seven lipids lacked confirmation due to incomplete database entries. Merging of positive and negative ion data improved annotation confidence and minimized false discoveries.

Benefits and Practical Applications

• Integrated Workflow: Combines precursor and product ion searches in a single interface, reducing manual curation.
• Customizable Database: Users can edit class-specific lipid entries and define adducts, modifications and fragmentation rules.
• High Throughput: Automated merging and filtering streamline large-scale lipidomics studies in research and QC settings.

Future Trends and Opportunities

Ongoing developments include expansion of database coverage for rare lipid classes, improved algorithms for positional isomer discrimination, and integration of normalized quantitation modules. Cloud-based implementations and AI-driven spectral interpretation may further accelerate untargeted lipidomics analysis.

Conclusion

LipidSearch 5.0 provides a robust, user-friendly platform for HRAM-based lipidomics, achieving high accuracy in lipid identification and supporting streamlined data processing. Its flexibility and comprehensive workflows address key bottlenecks in untargeted lipid analysis and set the stage for broader adoption in life science research.

Reference

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