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ADME PK

Aldehyde Oxidase (AO) reaction phenotyping assay

Understand if your compound is metabolized by aldehyde oxidase (AO).

Aldehyde oxidase (AO) reaction phenotyping is a non-CYP mediated metabolism assay within our portfolio of in vitro ADME screening services. Cyprotex deliver consistent, high quality data with the flexibility to adapt protocols based on specific customer requirements.

Determining potential metabolism by aldehyde oxidase enzymes

  • Aldehyde oxidase is a cytosolic enzyme and is a sub family of the molybdo-flavoenzymes.
  • For catalytic activity, aldehyde oxidase requires a molybdo-pterin cofactor (molybdenum cofactor, MoCo) and flavin adenine dinucleotide.
  • It has broad substrate specificity and catalyses the oxidation of aldehydes into carboxylic acids and also hydroxylation of some N-heterocycles.
  • The main isoform in humans is AO1 which has highest activity in the liver. The enzyme has also been detected in other tissues such as the lung, gastrointestinal tract and kidney.
  • Cyprotex’s aldehyde oxidase (AO) reaction phenotyping assay determines if your compound is a substrate for aldehyde oxidase (AO).
It has been increasingly recognized in this past decade that AO, through its unique structure, distribution, and substrate recognition, has an important role to play in the metabolism of drugs.

1Pryde DC et al. (2010), J. Med. Chem., 53:8441-8460

Protocol

Aldehyde Oxidase (AO) reaction phenotyping assay protocol

Test System Human liver cytosol incubated with and without specific AO inhibitor, 100 µM menadione (other species and enzyme sources available on request)
Test Article Concentration 1 µM (different concentrations available)
Positive Control Substrate Phthalazine
Time Points 0, 10, 20, 40, 60, 120 min
Test Article Requirements 100 µL of a 10 mM DMSO solution (or equivalent amount in solid)
Analysis Method LC-MS/MS
Data Delivery % Parent compound remaining at each time point
Half life
Standard error of half life

Data

Data from the Aldehyde Oxidase (AO) reaction phenotyping assay

 
aldehyde oxidase reaction phenotyping

Figure 1
Results from human liver cytosol stability of AO substrates carbazeran, famciclovir, vanillin and phthalazine in the presence of different AO inhibitors, displayed as a percentage of the CLint of the minus inhibitor control compound.  Inset: displays substrate depletion over time in the absence of inhibitor.

References

1Pryde DC et al. (2010) Aldehyde oxidase: an enzyme of emerging importance in drug discovery. J Med Chem 53: 8441-8460

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Please give details of the assays you are interested in. Where appropriate please specify one or more species (human, rat, mouse etc.), isoforms (CYP1A1,CYP1B1, etc) or other relevant details.

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