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A high coverage pseudotargeted lipidomics method based on three-phase liquid extraction and segment data-dependent acquisition using UHPLC-MS/MS with application to a study of depression rats

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Abstract

Pseudotargeted analysis combines the advantages of untargeted and targeted lipidomics methods based on chromatography-mass spectrometry (MS). This study proposed a comprehensive pseudotargeted lipidomics method based on three-phase liquid extraction (3PLE) and segment data-dependent acquisition (SDDA). We used a 3PLE method to extract the lipids with extensive coverage from biological matrixes. 3PLE was composed of one aqueous and two organic phases. The upper and middle organic phases enriched neutral lipids and glycerophospholipids, respectively, combined and detected together. Besides, the SDDA strategy improved the detection of co-elution ions in the lipidomics analysis. A total of 554 potential lipids were detected by the developed approach in both positive and negative modes using ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). Compared with the conventional liquid-liquid extraction (LLE) approaches, including methyl tert-butyl ether (MTBE) and Bligh-Dyer (BD) methods, 3PLE combined with SDDA significantly increased the lipid coverage 87.2% and 89.7%, respectively. Also, the proposed pseudotargeted lipidomics approach exhibited higher sensitivity and better repeatability than the untargeted approach. Finally, we applied the established pseudotargeted method to the plasma lipid profiling from the depressed rats and screened 61 differential variables. The results demonstrated that the pseudotargeted method based on 3PLE and SDDA broadened lipid coverage and improved the detection of co-elution ions with excellent sensitivity and precision, indicating significant potential for the lipidomics analysis.

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Abbreviations

MS:

Mass spectrometry

3PLE:

Three-phase liquid extraction

SDDA:

Segment data-dependent acquisition

UHPLC-MS/MS:

Ultra-high-performance liquid chromatography-tandem mass spectrometry

LLE:

Liquid-liquid extraction

MTBE:

Methyl tert-butyl ether

BD:

Bligh-Dyer

PL:

Phospholipid

LPL:

Lysophospholipid

MRM:

Multiple reaction monitoring

Q-TOF:

Quadrupole-time-of-flight

TQ:

Triple quadrupole

IS:

Internal standard

LPE:

Lysophosphatidylethanolamine

SM:

Sphingomyelin

Cer:

Ceramide

PC:

Phosphatidylcholine

TG:

Triglyceride

QC:

Quality control

S/N:

Signal to noise

RSD:

Relative standard deviation

PLS-DA:

Partial least squares-discriminant analysis

OPLS-DA:

Orthogonal partial least squares-discriminant analysis

FDR:

False discovery rate

HMDB:

Human metabolome database

MG:

Monoglyceride

DG:

Diglyceride

LPC:

Lysophosphatidylcholine

CE:

Cholesteryl ester

LOD:

Limit of determination

LOQ:

Limit of quantitation

VIP:

Variable importance of projection

AA:

Arachidonic acid

FA:

Fatty acid

PE:

Phosphatidylethanolamine

PLA:

Phospholipase A

LOX:

Lipoxygenase

COX:

Cyclooxygenase

HpETE:

Hydroperoxyeicosatetraenoic acid

LT:

Leukotriene

PG:

Prostaglandin

TX:

Thromboxane

SMase:

Sphingomyelinase

CDase:

Ceramidase

SP:

Sphingolipid

GP:

Glycerophospholipid

GL:

Glycerolipid

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Funding

This work was supported by the National Natural Science Foundation of China (No. 21775047), and the Natural Science Foundation of Guangdong Province, China (No. 2021A1515012323).

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Authors and Affiliations

  1. School of Biology and Biological Engineering, South China University of Technology, Guangzhou, 510006, China

    Danyang Liu, Jina Yang, Wenbin Jin & Ting Zhou

  2. Shimadzu (China) Corporation, Guangzhou Branch, Guangzhou, 510010, Guangdong, China

    Qisheng Zhong

Authors
  1. Danyang Liu
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  2. Jina Yang
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  4. Qisheng Zhong
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  5. Ting Zhou
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Correspondence to Ting Zhou.

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The authors declare no competing interests.

Research involving human participants and/or animals

This animal study was approved by the Committee on the Ethics of Animal Experiments of Guangdong Pharmaceutical University (No.00199407). We executed animal experiments in accordance with Regulations on Animal Experiments in Guangdong Pharmaceutical University. We did not use samples from human in this study.

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Liu, D., Yang, J., Jin, W. et al. A high coverage pseudotargeted lipidomics method based on three-phase liquid extraction and segment data-dependent acquisition using UHPLC-MS/MS with application to a study of depression rats. Anal Bioanal Chem 413, 3975–3986 (2021). https://doi.org/10.1007/s00216-021-03349-w

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  • DOI: https://doi.org/10.1007/s00216-021-03349-w

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