Chromogenic enzyme substrates based on [2-(nitroaryl)ethenyl]pyridinium and quinolinium derivatives for the detection of nitroreductase activity in clinically important microorganisms.

A series of [2-(nitroaryl)ethenyl]pyridinium and quinolinium derivatives have been synthesised as potential indicators of microbial nitroreductase activity. When assessed against a selection of 20 clinically important pathogenic microorganisms, microbial colonies of various colours (yellow, green, red, brown, black) were produced and attributed to nitroreductase activity. Most substrates elicited colour responses with Gram-negative microorganisms. In contrast, the growth of several species of Gram-positive microorganisms and yeasts was often inhibited by the substrates and hence coloured responses were not seen.

Beyond GalNAc! Drug delivery systems comprising complex oligosaccharides for targeted use of nucleic acid therapeutics.

Nucleic Acid Therapeutics (NATs) are establishing a leading role for the management and treatment of genetic diseases following FDA approval of nusinersen, patisiran, and givosiran in the last 5 years, the breakthrough of milasen, with more approvals undoubtedly on the way. Givosiran takes advantage of the known interaction between the hepatocyte specific asialoglycoprotein receptor (ASGPR) and N-acetyl galactosamine (GalNAc) ligands to deliver a therapeutic effect, underscoring the value of targeting moieties. In this review, we explore the history of GalNAc as a ligand, and the paradigm it has set for the delivery of NATs through precise targeting to the liver, overcoming common hindrances faced with this type of therapy. We describe various complex oligosaccharides (OSs) and ask what others could be used to target receptors for NAT delivery and the opportunities awaiting exploration of this chemical space.

Evaluation of a Novel Chromogenic Medium for the Detection of Pseudomonas aeruginosa in Respiratory Samples from Patients with Cystic Fibrosis.

Pseudomonas aeruginosa is a dominant cause of respiratory infection in individuals with cystic fibrosis (CF), leading to significant morbidity and mortality. Detection of P. aeruginosa is conducted by culture of respiratory samples but this process may occasionally be compromised due to overgrowth by other bacteria and fungi. We aimed to evaluate a novel chromogenic medium, Pseudomonas aeruginosa chromogenic agar (PACA), for culture of P. aeruginosa from respiratory samples, from patients with CF. A total of 198 respiratory samples were cultured onto PACA and three other media: CHROMID® P. aeruginosa, CHROMagar™ Pseudomonas and MacConkey agar. P. aeruginosa was recovered from 66 samples (33%), using a combination of all media. After 72 h incubation, the sensitivity of the four chromogenic media was as follows: 91% for PACA and CHROMagar™ Pseudomonas, 85% for CHROMID® P. aeruginosa and 83% for MacConkey agar. For the three chromogenic media, the positive predictive value after 72 h was as follows: 95% for PACA, 56% for CHROMagar™ Pseudomonas and 86% for CHROMID® P. aeruginosa. PACA proved to be a highly effective culture medium for the isolation and specific detection of P. aeruginosa from respiratory samples.

A flexible topo-optical sensing technology with ultra-high contrast.

Elastic folding, a phenomenon widely existing in nature, has attracted great interests to understand the math and physical science behind the topological transition on surface, thus can be used to create frontier engineering solutions. Here, we propose a topo-optical sensing strategy with ultra-high contrast by programming surface folds on targeted area with a thin optical indicator layer. A robust and precise signal generation can be achieved under mechanical compressive strains (>0.4). This approach bridges the gap in current mechano-responsive luminescence mechanism, by utilizing the unwanted oxygen quenching effect of Iridium-III (Ir-III) fluorophores to enable an ultra-high contrast signal. Moreover, this technology hosts a rich set of attractive features such as high strain sensing, encoded logic function, direct visualisation and good adaptivity to the local curvature, from which we hope it will enable new opportunities for designing next generation flexible/wearable devices.

Fluorogenic l-alanylaminopeptidase substrates derived from 6-amino-2-hetarylquinolines and 7-amino-3-hetarylcoumarins and their potential applications in diagnostic microbiology.

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.

Rigidly linking cyclometallated Ir(iii) and Pt(ii) centres: an efficient approach to strongly absorbing and highly phosphorescent red emitters.

The synthesis and photophysical properties of an unprecedented tetranuclear complex are described, in which a fac-tris-cyclometallated Ir(iii) centre is rigidly connected to three cyclometallated Pt(ii) centres. The complex absorbs strongly up to ∼600 nm and emits red light with unusually high efficiency.

The Synthesis of L-Alanyl and ß-Alanyl Derivatives of 2-Aminoacridone and Their Application in the Detection of Clinically-Important Microorganisms.

In clinical microbiology the speed with which pathogenic microorganisms may be detected has a direct impact on patient health. One important strategy used in the laboratory is the growth of cultures in the presence of an enzymatic substrate which, once transformed by the appropriate microbial enzyme, generates a detectable colour or fluorescence output. Such substrates have previously been prepared by our group and others and are available as commercial diagnostic kits, however they all suffer from some degree of diffusion when used in a solid growth medium. This diffusion complicates the detection and differentiation of species in polymicrobial cultures and so we sought to improve on our previous work. In this work we have prepared and evaluated a series of novel fluorogenic enzyme substrates based on N-substituted-2-aminoacridones. All of the prepared substrates were found to be suitable for the detection and differentiation of certain microorganisms, however those based on the 2-amino-10-benzylacridone core in particular showed no apparent diffusion when incorporated into solid growth media. On transformation these substrates generated brightly fluorescent colonies that are clearly contrasted with the background medium due to the difference in emission wavelength (λem 445-450 nm for the substrate, λem 550 nm for the product). Here we have shown that our L-alanyl aminopeptidase substrate, 2-(N-L-alanylamino)-10-benzylacridone, is particularly suited to the detection of Gram-negative bacteria, and our ß-alanyl aminopeptidase substrate, 2-(N- ß-alanylamino)-10-benzylacridone, to the detection of Pseudomonas aeruginosa and Serratia marcescens when grown on solid media incorporating these substrates. The resulting fluorophore shows no apparent diffusion from the colonies of interest, and the enhanced sensitivity offered by fluorescent emission may allow for the detection of these organisms as microcolonies using automated fluorescence microscopy.

2-(Nitroaryl)benzothiazole and benzoxazole derivatives as fluorogenic substrates for the detection of nitroreductase activity in clinically important microorganisms.

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.

A high-throughput screening method for determining the substrate scope of nitrilases.

Nitrile compounds are intermediates in the synthesis of pharmaceuticals such as atorvastatin. We have developed a chromogenic reagent to screen for nitrilase activity as an alternative to Nessler's reagent. It produces a semi-quantifiable blue colour and hydrolysis of 38 nitrile substrates by 23 nitrilases as cell-free extracts has been shown.

Synthesis of 2-arylbenzothiazole derivatives and their application in bacterial detection.

A series of 2-arylbenzothiazole derivatives have been prepared as fluorogenic enzyme substrates in order to detect aminopeptidase, esterase, phosphatase and ß-galactosidase activity in clinically important Gram-negative and Gram-positive bacteria. Substrates were incorporated into an agar-based culture medium and this allowed growth of intensely fluorescent bacterial colonies based on hydrolysis by specific enzymes. Substrate 20 targeted L-alanine aminopeptidase activity and was hydrolysed exclusively by a range of Gram-negative bacteria and inhibited the growth of a range of Gram-positive bacteria. Substrate 19a targeted ß-alanyl aminopeptidase activity and generated fluorescent colonies of selected Gram-negative species including Pseudomonas aeruginosa. Substrate 21b targeted C8-esterase activity and resulted in strongly fluorescent colonies of selected species known to harbour such enzyme activity (e.g., Salmonella and Pseudomonas). Most Gram-negative species produced colonies with an intense blue fluorescence due to hydrolysis of phosphatase substrates 24a-c and substrate 24c was also hydrolysed by strains of Staphylococcus aureus. Compounds 26b and 26c targeted ß-galactosidase activity and generated strongly fluorescent colonies with coliform bacteria that produced this enzyme (e.g., Escherichia coli).