ABSTRACT
A series of fluorogenic heterocyclic azides were prepared and assessed as reductase substrates across a selection of Gram-negative and Gram-positive microorganisms. The majority of these azides showed similar activity profiles to nitroreductase substrates. Microorganisms that do not produce hydrogen sulfide reduced the azides, indicating reductase activity was not linked to hydrogen sulfide production.
Subject(s)
Coumarins/chemistry , Fluorescent Dyes/chemistry , Gram-Negative Bacteria/enzymology , Gram-Positive Bacteria/enzymology , Nitroreductases/metabolism , Phthalimides/chemistry , Coumarins/chemical synthesis , Fluorescent Dyes/chemical synthesis , Molecular Structure , Phthalimides/chemical synthesis , Substrate SpecificityABSTRACT
Six novel fluorogenic enzyme substrates for detecting l-alanylaminopeptidase activity in microorganisms have been prepared and evaluated in Columbia agar media. The substrates are l-alanyl derivatives of 6-amino-2-hetarylquinolines and 7-amino-3-hetarylcoumarins. Both the quinoline and coumarin series of substrates produced fluorescence in the presence of Gram-negative microorganisms. In contrast, fluorescence generation in the presence of the Gram-positive microorganisms and yeasts was limited or absent.
Subject(s)
CD13 Antigens/chemistry , Coumarins/chemistry , Fluorescent Dyes/chemistry , Quinolines/chemistry , Enzyme Assays , Gram-Negative Bacteria/enzymology , Spectrometry, Fluorescence , Substrate SpecificityABSTRACT
A series of carboxy-substituted 2-(nitroaryl)benzothiazole derivatives and carboxy-substituted 2-(nitroaryl)benzoxazole derivatives were prepared and evaluated as potential nitroreductase substrates for the purpose of detecting clinically important microorganisms. Several of the substrates produced highly fluorescent colonies with the majority of a panel of 10 Gram-negative bacteria and also with two of a panel of 8 Gram-positive bacteria.
Subject(s)
Bacterial Proteins/metabolism , Benzothiazoles/chemistry , Benzoxazoles/chemistry , Fluorescent Dyes/chemistry , Nitroreductases/metabolism , Bacterial Proteins/analysis , Benzothiazoles/metabolism , Benzoxazoles/metabolism , Gram-Negative Bacteria/enzymology , Gram-Positive Bacteria/enzymology , Nitroreductases/analysis , Spectrometry, Fluorescence , Substrate SpecificityABSTRACT
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 ß-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.
