Quantification of bioactive acylethanolamides in rat plasma by electrospray mass spectrometry

Andrea Giuffrida, Fernando Rodríguez De Fonseca, Daniele Piomelli

Research output: Contribution to journalArticle

141 Scopus citations

Abstract

We developed a high-performance liquid chromatography/mass spectrometry (HPLC/MS) method for the identification and quantification of anandamide, an endogenous cannabinoid substance, and other fatty acid ethanolamides (AEs) in biological samples. Using a mobile-phase system of methanol/water and gradient elution, we achieved satisfactory resolution of all major AEs, including anandamide, palmitylethanolamide (PEA), and oleylethanolamide (OEA). Electrospray-generated quasi-molecular species were used as diagnostic ions and detected by selected ion monitoring (SIM). Synthetic deuterium- labeled AEs were used as internal standards, and quantification was carried out by isotope dilution. A linear correlation (r2 = 0.99) was observed in the calibration curves for standard AEs over the range 0-0.5 nmol. Detection limits between 0.1 and 0.3 pmol per sample and quantification limits between 0.5 and 1.2 pmol per sample were obtained. The method was applied to the quantification of anandamide, PEA, and OEA in plasma prepared from rat blood collected either by cardiac puncture or by decapitation. After cardiac puncture, AE levels were in the low-nanomolar range: anandamide, 3.1 ± 0.6 pmol/ml; PEA, 9.4 ± 1.6 pmol/ml; OEA, 9.2 ± 1.8 pmol/ml (mean ± SE, n = 9). By contrast, after decapitation AEs were dramatically elevated (anandamide, 144 ± 13 pmol/ml; PEA, 255 ± 55 pmol/ml; OEA, 175 ± 48 pmol/ml). Thus, disruptive procedures of blood collection may result in gross overestimates in the concentrations of circulating AEs. (C) 2000 Academic Press.

Original languageEnglish (US)
Pages (from-to)87-93
Number of pages7
JournalAnalytical Biochemistry
Volume280
Issue number1
DOIs
StatePublished - Apr 10 2000

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Keywords

  • Anandamide
  • Cannabinoid receptors
  • Fatty acid ethanolamides
  • High- performance liquid chromatography
  • Mass spectrometry

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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