We are setting up some hepatocyte-based MetID experiments. Our benchmark reactions are: dealkylation of 7etoxycoumarin and conjugation of 7hydroxycoumarin to glucuronide and sulfate.
Two independent incubations of 7EC and 7HC. How come we only observe conjugation? We don't have dealkylation of 7EC rat hepatocytes.
Any clue why?
Thanks.
Thanks for the answe, Jai. My question wasn't clear, sorry. We observe conjugation in 7HC incubation only. If we incubate 7EC nothing happens. No metabolites are formed. Parent peak area does not change.
The observation of Phase II metabolites from 7HC suggests your system is metabolically viable. However, if you have lost the Phase I metabolic activity but not the Phase II activity my first thoughts are whether the solvent you have dissolved your 7EC in is having an effect? Have you tried a different solvent for your 7EC or a much lower solvent concentration?
A second thought is whether your 7EC concentration is too high so you are saturating the metabolic capacity of your cells and struggling to detect minor decreases in the 7EC concentration?
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