Direct quantitative analysis of lysophosphatidic acid molecular species by stable isotope dilution electrospray lonization liquid chromatography-mass spectrometry

Daniel L. Baker, Dominic M. Desiderio, Duane Miller, Elizabeth Tolley, Gabor Tigyi

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172 Citations (Scopus)

Abstract

In order to better understand the role of lysophosphatidic acid (LPA) in physiology and pathophysiology, it is necessary to accurately determine the molecular species and amounts of LPA in biological samples. We have developed a stable-isotope dilution, liquid chromatography-mass spectrometry assay for the direct quantitative analysis of 1-acyl-LPA. This method utilizes a deuterium-labeled internal standard, LPA (18:0-d35), and a single liquid-liquid extraction with acidic butanol that allows > 95% recovery of LPA, followed by online normal-phase liquid chromatography-mass spectrometry. This protocol allows for the accurate, sensitive, and reproducible analysis of the individual 1-acyl-LPA species present in biological samples. The utility of the assay is demonstrated through the analysis of LPA species in plasma and serum from human volunteers. Total LPA in EDTA plasma was 0.61 ± 0.14 /mM in males and 0.74 ± 0.17 /mM in females, which increased to 0.91 ± 0.23 and 0.99 ± 0.38 /mM after incubation for 24 h at 25°C. Total LPA in serum was 0.85 ± 0.22 μM in males and 1.57 ± 0.56 μM in females, which increased to 4.78 ± 0.89 and 5.57 ± 0.73 μM after incubation for 24 h at 25°C.

Original languageEnglish (US)
Pages (from-to)287-295
Number of pages9
JournalAnalytical Biochemistry
Volume292
Issue number2
DOIs
StatePublished - May 15 2001

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Liquid chromatography
Liquid Chromatography
Isotopes
Dilution
Mass spectrometry
Mass Spectrometry
Chemical analysis
Assays
Plasmas
lysophosphatidic acid
Butanols
Liquid-Liquid Extraction
Deuterium
Physiology
Liquids
Serum
Edetic Acid
Volunteers
Recovery

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

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title = "Direct quantitative analysis of lysophosphatidic acid molecular species by stable isotope dilution electrospray lonization liquid chromatography-mass spectrometry",
abstract = "In order to better understand the role of lysophosphatidic acid (LPA) in physiology and pathophysiology, it is necessary to accurately determine the molecular species and amounts of LPA in biological samples. We have developed a stable-isotope dilution, liquid chromatography-mass spectrometry assay for the direct quantitative analysis of 1-acyl-LPA. This method utilizes a deuterium-labeled internal standard, LPA (18:0-d35), and a single liquid-liquid extraction with acidic butanol that allows > 95{\%} recovery of LPA, followed by online normal-phase liquid chromatography-mass spectrometry. This protocol allows for the accurate, sensitive, and reproducible analysis of the individual 1-acyl-LPA species present in biological samples. The utility of the assay is demonstrated through the analysis of LPA species in plasma and serum from human volunteers. Total LPA in EDTA plasma was 0.61 ± 0.14 /mM in males and 0.74 ± 0.17 /mM in females, which increased to 0.91 ± 0.23 and 0.99 ± 0.38 /mM after incubation for 24 h at 25°C. Total LPA in serum was 0.85 ± 0.22 μM in males and 1.57 ± 0.56 μM in females, which increased to 4.78 ± 0.89 and 5.57 ± 0.73 μM after incubation for 24 h at 25°C.",
author = "Baker, {Daniel L.} and Desiderio, {Dominic M.} and Duane Miller and Elizabeth Tolley and Gabor Tigyi",
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AU - Tolley, Elizabeth

AU - Tigyi, Gabor

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AB - In order to better understand the role of lysophosphatidic acid (LPA) in physiology and pathophysiology, it is necessary to accurately determine the molecular species and amounts of LPA in biological samples. We have developed a stable-isotope dilution, liquid chromatography-mass spectrometry assay for the direct quantitative analysis of 1-acyl-LPA. This method utilizes a deuterium-labeled internal standard, LPA (18:0-d35), and a single liquid-liquid extraction with acidic butanol that allows > 95% recovery of LPA, followed by online normal-phase liquid chromatography-mass spectrometry. This protocol allows for the accurate, sensitive, and reproducible analysis of the individual 1-acyl-LPA species present in biological samples. The utility of the assay is demonstrated through the analysis of LPA species in plasma and serum from human volunteers. Total LPA in EDTA plasma was 0.61 ± 0.14 /mM in males and 0.74 ± 0.17 /mM in females, which increased to 0.91 ± 0.23 and 0.99 ± 0.38 /mM after incubation for 24 h at 25°C. Total LPA in serum was 0.85 ± 0.22 μM in males and 1.57 ± 0.56 μM in females, which increased to 4.78 ± 0.89 and 5.57 ± 0.73 μM after incubation for 24 h at 25°C.

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