A high-throughput-compatible assay for determining the activity of fatty acid amide hydrolase

Anal Biochem. 2003 Jul 15;318(2):270-5. doi: 10.1016/s0003-2697(03)00217-3.

Abstract

Fatty acid amide hydrolase (EC 3.5.1.4.) is the enzyme responsible for the rapid degradation of lipid-derived chemical messengers such as anandamide, oleamide, and 2-arachidonoylglycerol. The pharmacological characterization of this enzyme in vivo has been hampered by the lack of selective and bioavailable inhibitors. We have developed a simple, radioactive, high-throughput-compatible assay for this enzyme based on the differential absorption of the substrate and its products to activated charcoal. The assay was validated using known inhibitors. It may be applied for the identification of new inhibitors from a compound library.

MeSH terms

  • Absorption
  • Amidohydrolases / analysis*
  • Amidohydrolases / antagonists & inhibitors
  • Amidohydrolases / metabolism*
  • Arachidonic Acids / chemistry
  • Arachidonic Acids / metabolism
  • Cell Line, Tumor
  • Charcoal
  • Endocannabinoids
  • Enzyme Inhibitors / isolation & purification
  • Enzyme Inhibitors / pharmacology
  • Humans
  • Kinetics
  • Molecular Structure
  • Polyunsaturated Alkamides
  • Radioactivity
  • Reference Standards
  • Reproducibility of Results

Substances

  • Arachidonic Acids
  • Endocannabinoids
  • Enzyme Inhibitors
  • Polyunsaturated Alkamides
  • Charcoal
  • Amidohydrolases
  • fatty-acid amide hydrolase
  • anandamide