Synthesis and Evaluation of Novel 7- and 8‑Aminophenoxazinones for the Detection of β‑Alanine Aminopeptidase Activity and the Reliable Identification of Pseudomonas aeruginosa in Clinical Samples

• Published in: Journal of medicinal chemistry Vol. 59; no. 10; pp. 4476 - 4487
• Main Authors: Bedernjak, Alexandre F, Zaytsev, Andrey V, Babolat, Michèle, Cellier, Marie, James, Arthur L, Orenga, Sylvain, Perry, John D, Groundwater, Paul W, Anderson, Rosaleen J
• Format: Journal Article
• Language: English
• Published: WASHINGTON American Chemical Society 26.05.2016; Amer Chemical Soc
• Subjects: Anti-Bacterial Agents - chemical synthesis; Anti-Bacterial Agents - chemistry; Anti-Bacterial Agents - pharmacology; CD13 Antigens - antagonists & inhibitors; CD13 Antigens - metabolism; Chemistry, Medicinal; Dose-Response Relationship, Drug; Enzyme Inhibitors - chemical synthesis; Enzyme Inhibitors - chemistry; Enzyme Inhibitors - pharmacology; Life Sciences & Biomedicine; Microbial Sensitivity Tests; Molecular Structure; Oxazines - chemical synthesis; Oxazines - chemistry; Oxazines - pharmacology; Pharmacology & Pharmacy; Pseudomonas aeruginosa - drug effects; Pseudomonas aeruginosa - enzymology; Pseudomonas aeruginosa - isolation & purification; Science & Technology; Structure-Activity Relationship
• ISSN: 0022-2623; 1520-4804
• DOI: 10.1021/acs.jmedchem.5b01591

A series of novel 8-aminophenoxazin-3-one and 7-aminophenoxazin-3-one chromogens and their corresponding β-alanine derivatives were synthesized and evaluated for their ability to detect β-alanyl aminopeptidase activity in bacteria known to hydrolyze β-alanine derivatized substrates. The results provided insight into the structural requirements for effective visualization of enzymatic activity and the mechanism of formation of phenoxazinon-3-ones. 8-Aminophenoxazin-3-one substrates 23c, 23d, and 23e were prepared in good to high overall yield and were selective for β-alanyl aminopeptidase activity in bacteria, producing a lighter agar background coloration facilitating visualization of colored colonies, with variable localization to the colonies, but had lower sensitivities for the detection of Pseudomonas aeruginosa in comparison to the analogous 7-aminophenoxazin-3-one substrates. The synthetic methodology employed here allows the preparation of a range of substrates for evaluation and the establishment of structure–activity relationships. For example, the 2-pentyl substituted aminophenoxazin-3-one 22b performed with analogous sensitivity to the corresponding 1-pentyl-7-aminophenoxazin-3-one substrate 1 used commercially, highlighting that the position of the pentyl substituent can be varied while maintaining detection sensitivity.