Fungal Transformation of the Tricyclic Antidepressant Amoxapine: Identification of N-Carbomethoxy Compounds Formed as Artifacts by Phosgene in Chloroform used for the Extraction of Metabolites

Abstract

The biotransformation of the antidepressant drug amoxapine by Cunninghamella elegans formed three metabolites, 7-hydroxyamoxapine, N-formyl-7-hydroxyamoxapine, and N-formylamoxapine; two other compounds were only present when chloroform was used in the extraction process. All five of the compounds were separated by reversed-phase HPLC, then analyzed by ¹H NMR and mass spectrometry, and by ¹³C NMR when sample quantities permitted. The artifacts were identified as N-carbomethoxy-7-hydroxyamoxapine and N-carbomethoxyamoxapine. Phosgene is a decomposition product of chloroform that can form carbomethoxy compounds at the secondary nitrogen of a piperazine ring in an alcoholic solution. Since N-carbomethoxy compounds were not observed when ethyl acetate was used for extraction of the culture medium, they were considered artifacts and not metabolites. These findings suggest that chloroform should be tested for the formation of phosgene before using it to extract any compound with a piperazine ring or any other amine-containing structure.

Key Words:

• Chloroform
• Phosgene
• N-carbomethoxy compounds
• Amoxapine
• C. elegans
• Microbial biotransformation