Radiometric assay of ghrelin hydrolase activity and 3H-ghrelin distribution into mouse tissues

Biochem Pharmacol. 2015 Dec 15;98(4):732-9. doi: 10.1016/j.bcp.2015.10.016. Epub 2015 Oct 26.

Abstract

A high-throughput radiometric assay was developed to characterize enzymatic hydrolysis of ghrelin and to track the peptide's fate in vivo. The assay is based on solvent partitioning of [(3)H]-octanoic acid liberated from [(3)H]-octanoyl ghrelin during enzymatic hydrolysis. This simple and cost-effective method facilitates kinetic analysis of ghrelin hydrolase activity of native and mutated butyrylcholinesterases or carboxylesterases from multiple species. In addition, the assay's high sensitivity facilitates ready evaluation of ghrelin's pharmacokinetics and tissue distribution in mice after i.v. bolus administration of radiolabeled peptide.

Keywords: Butyrylcholinesterase; Di-isopropyl fluorophosphates (PubChem CID: 5936); Enzyme kinetics; Ghrelin; Human desacyl-ghrelin (PubChem CID: 71728433); Human ghrelin (PubChem CID: 91668172); Pharmacokinetics; Radiometric assay; Toluene (PubChem CID: 1140); iso-OMPA (PubChem CID: 5420).

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Enzyme Activation / physiology
  • Ghrelin / metabolism*
  • Humans
  • Hydrolases / metabolism*
  • Male
  • Mice
  • Mice, Inbred C57BL
  • Radiometry / methods
  • Tissue Distribution / physiology
  • Tritium / metabolism*

Substances

  • Ghrelin
  • Tritium
  • Hydrolases