Structure requirements for anaerobe processing of azo compounds: implications for prodrug design

This Letter generalizes the metabolism of the azo class of compounds by Clostridium perfringens, an anaerobe found in the human colon. A recently reported 5-aminosalicylic acid-based prednisolone prodrug was shown to release the drug when incubated with the bacteria, while the para-aminobenzoic acid (PABA) based analogue did not. Instead, it showed a new HPLC peak with a relatively close retention time to the parent which was identified by LCMS as the partially reduced hydrazine product. This Letter investigates azoreduction across a panel of substrates with varying degrees of electronic and steric similarity to the PABA-based compound. Azo compounds with an electron donating group on the azo-containing aromatic ring showed immediate disproportionation to their parent amines without any detection of hydrazine intermediates by HPLC. Compounds containing only electron withdrawing groups are partially and reversibly reduced to produce a stable detectable hydrazine. They do not disproportionate to their parent amines, but regenerate the parent azo compound. This incomplete reduction is relevant to the design of azo-based prodrugs and the toxicology of azo-based dyes.