title: Novel chromogenic aminopeptidase substrates for the detection and identification of clinically important microorganisms.
abstract: A series of amino acid derivatives 8-10, 42 and 43 have been prepared as chromogenic enzyme substrates in order to detect aminopeptidase activity in clinically important Gram-negative and Gram-positive bacteria. Enzymatic hydrolysis liberates the amino acid moiety and either a 4-aminophenol or a 4-dialkylaminoaniline derivative which undergoes oxidative coupling with 1-naphthol or a substituted 1-naphthol giving an indophenol dye. Substrates and 1-naphthols were incorporated into an agar-based culture medium and this allowed growth of intensely coloured bacterial colonies based on hydrolysis by specific enzymes. Red/pink coloured colonies were produced by the substrates 8-10 and blue coloured colonies were formed by the substrates 42 and 43. The L-alanyl aminopeptidase substrates 8 targeted L-alanyl aminopeptidase activity and gave coloured colonies with a range of Gram-negative bacteria. Substrates 9 targeted β-alanyl aminopeptidase activity and generated coloured colonies with selected Gram-negative species including Pseudomonas aeruginosa. Three substrates for L-pyroglutamyl acid aminopeptidase (10a, 10c and 43) were hydrolysed by enterococci and Streptococcus pyogenes to generate coloured colonies. Two yeasts were also included in the study, but they did not produce coloured colonies with any of the substrates examined.
authors.affiliation: Research & Development Microbiology, bioMérieux SA, 3 route de Port Michaud, 38 390 La-Balme-les-Grottes, France.
authors.forename: Marie
authors.initials: M
authors.lastname: Cellier
authors.affiliation: Department of Applied Sciences, Northumbria University, Newcastle upon Tyne NE7 7DN, UK.
authors.forename: Arthur L
authors.initials: AL
authors.lastname: James
authors.affiliation: Research & Development Microbiology, bioMérieux SA, 3 route de Port Michaud, 38 390 La-Balme-les-Grottes, France.
authors.forename: Sylvain
authors.initials: S
authors.lastname: Orenga
authors.affiliation: Department of Microbiology, Freeman Hospital, Newcastle upon Tyne NE7 7DN, UK.
authors.forename: John D
authors.initials: JD
authors.lastname: Perry
authors.affiliation: Department of Applied Sciences, Northumbria University, Newcastle upon Tyne NE7 7DN, UK.
authors.forename: Ari K
authors.initials: AK
authors.lastname: Rasul
authors.affiliation: Department of Applied Sciences, Northumbria University, Newcastle upon Tyne NE7 7DN, UK.
authors.forename: Shaun N
authors.initials: SN
authors.lastname: Robinson
authors.affiliation: Department of Applied Sciences, Northumbria University, Newcastle upon Tyne NE7 7DN, UK. Electronic address: steven.stanforth@northumbria.ac.uk.
authors.forename: Stephen P
authors.initials: SP
authors.lastname: Stanforth
mesh_terms: D000626:Aminopeptidases / Q000737:chemistry / Q000378:metabolism*; D001419:Bacteria / Q000201:enzymology* / Q000378:metabolism; D002863:Chromogenic Compounds / Q000737:chemistry* / Q000378:metabolism; D006868:Hydrolysis; D015394:Molecular Structure; D013379:Substrate Specificity
chemical_list: D002863:Chromogenic Compounds; D000626:Aminopeptidases
keywords: Aminopeptidase; Bacteria detection; Chromogenic substrates