A continuous spectrophotometric assay for monoamine oxidase and related enzymes in tissue homogenates.
A continuous, peroxidase-linked spectrophotometric assay is described which is suitable for measuring monoamine and diamine oxidase and semicarbazide-sensitive amine oxidase activities in tissue homogenates. In the assay, 4-aminoantipyrine is oxidized and then condenses with vanillic acid to give a red quinoneimine dye. The absorbance at 498 nm is proportional to the amount of hydrogen peroxide released in the amine oxidase reaction. The molar absorption coefficient of the dye at pH 7.6 was 4654 M-1 cm-1. The method is suitable for use with any amine oxidase substrate which has a higher oxidation-reduction potential than does 4-aminoantipyrine. Following preincubation of rat liver homogenates with selective monoamine oxidase (MAO)-A and -B inhibitors, kinetic constants were obtained for metabolism of the mixed substrate, p-tyramine. Inhibition of MAO in rat liver homogenates was also measured following administration of the antidepressant, phenelzine. This inexpensive assay which employs reagents with low toxicity can thus be used to determine the degree of inhibition of MAO elicited by potential antidepressant and anti-parkinsonian agents. Drugs, their metabolites, and environmental toxins can also be screened as possible amine oxidase substrates or inhibitors, and kinetic constants for turnover of novel substrates can be determined.
Holt
A
A
Department of Chemistry, University of Alberta, Edmonton, Canada.
Sharman
D F
DF
Baker
G B
GB
Palcic
M M
MM
D000042:Absorption; D000818:Animals; D002733:Chlorophenols / Q000494:pharmacology; D007700:Kinetics; D008099:Liver / Q000201:enzymology; D008297:Male; D008995:Monoamine Oxidase / Q000032:analysis*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013053:Spectrophotometry / Q000379:methods*; D014439:Tyramine / Q000378:metabolism
D002733:Chlorophenols; D008995:Monoamine Oxidase; C004762:2,4-dichlorophenol; D014439:Tyramine