In vivo 'turn on' fluorescence detection of free cysteine in zebrafish kidney and liver.

Cysteine is directly associated with a wide range of biological processes. Besides its essential role in protein synthesis, cysteine undergoes a variety of post-translational modifications which modulate several physiological processes. Dysregulated cysteine metabolism is associated with several neurodegenerative disorders. Accordingly, restoring cysteine balance has therapeutic benefits. It is therefore essential to detect the presence of endogenous free cysteine in order to understand different physiological modes of action inside the cell. Here, a carbazole-pyridoxal conjugate system (CPLC) has been developed to detect endogenous free cysteine in the liver and kidney of an adult zebrafish. In consequence, we have also determined the fluorescence intensity statistics of zebrafish kidney and liver images. CPLC interacts in a very fascinating way with two cysteine molecules through chemodosimetric and chemosensing approaches which are conclusively proved by different spectroscopic analyses (UV-vis, fluorescence, NMR) and theoretical calculations (DFT). The detection limit of CPLC towards cysteine is 0.20 µM. Moreover, this preliminary experiment has been done using HuH-7 cell line to check the permeability of CPLC, interaction with cysteine intracellularly, and assessment of the toxicity of CPLC, if any, before performing details in-vivo experiments in zebrafish model.

Identification of a novel quinoline-based UV-protective pigment from a psychrotrophic Arctic bacterium.

AIMS: Psychrotrophs are extremophilic microorganisms that grow optimally in low temperature having many unique bioactive molecules of biotechnological applications. In this study, we characterized a pigment from an arctic bacterium with protective activity towards UV exposure. METHODS AND RESULTS: The present research reports isolation and characterization of a psychrotrophic bacteria, RSAP2, from the soil sample of NyAlesund (78°56"N, 11°54"E), Svalbard, Norway. The strain showed closest 16S rRNA gene sequence similarity (99.9%) with Kocuria indica NIO-1021. RSAP2 is a Gram-positive, coccoid aerobe which produces a yellow pigment. The optimal parameters for pigment production while grown in LB medium were 3% (w/v) NaCl and 4 days of incubation of the culture at 20°C and pH 9 with shaking (180 rpm). The pigment was extracted in methanol and acetone (2:1) and further purified through column chromatography. It was characterized by mass spectrometry, UV-visible, fluorescence, IR, 1 H NMR, 13 C NMR spectroscopy and CHNS/O analysis. The pigment has a molecular weight of about 258 daltons and the molecular formula was determined as C15 H18 N2 O2 and is a quinoline derivative. We show that the pigment can protect Escherichia coli against UV-mediated mutagenesis. We further demonstrate that the pigment displays a significant antimicrobial effect and in sublethal concentrations it impairs biofilm formation ability of the model organism Staphylococcus aureus. CONCLUSIONS: The pigment of a psychrotrophic Arctic bacterium, most likely a strain of K. indica, was purified and its chemical structure was determined. The quinoline-based pigment has the ability to protect live cells from UV induced damage. SIGNIFICANCE AND IMPACT OF STUDY: Analysis and characterization of this newly isolated quinoline-based pigment is a potential candidate for future application in skin care products.

Reliable fluorescence technique to detect the antibiotic colistin, a possible environmental threat due to its overuse.

Colistin, considered a drug of last resort as it is effective towards multidrug-resistant Gram-negative bacterial infections. Oral administration of colistin in the poultry industry is a common practice, not only to prevent and reduce bacterial infections, but also as a rapid-growth promoter. Long-term exposure to any antibiotic will eventually lead to the development of bacterial resistance towards all antibiotics through various mechanisms in the physiological system and environment. Chicken is the most consumed source of animal protein for humans throughout the world. In addition, the manure of poultry, containing traces of the used antibiotics, is being used in farming. Exposure to excess amounts of colistin causes a great concern not only to the humans but to the environment as a whole. In the present contribution, colistin has been detected in chicken hepatocyte cells through in vivo confocal microscopy. In addition, the amount of colistin in the chicken excrements has been estimated. A simple chemosensor NAF, a dye-based on napthaldehyde furfural, was developed for the detection of colistin, supplemented with experimental evidence and theoretical calculations.

An antibacterial compound pyrimidomycin produced by Streptomyces sp. PSAA01 isolated from soil of Eastern Himalayan foothill.

Selective isolation of soil Actinobacteria was undertaken to isolate a new class of antibiotics and bioactive molecules. A Streptomyces sp. PSAA01 (= MTCC 13,157), isolated from soil of Eastern Himalaya foothill was cultivated on a large scale for the production of the antimicrobial SM02. It has been found that the maximum amount of SM02 produced while PSAA01 was grown in ISP-2 medium (pH 7.0) for 7 days at 30 °C in shaking (180 rpm) condition. A significant zone of inhibition against Staphylococcus aureus MTCC 96 has been found with the crude cell-free culture media (50 µL) of 7 days grown PSAA01. After the purification and chemical structural characterization, we found that SM02 is a new antimicrobial having 746 dalton molecular weight. The compound SM02 contains pyrimidine moiety in it and is produced by a species of Streptomyces and thus we have named this antibiotic pyrimidomycin. The antimicrobial spectrum of pyrimidomycin has been found to be restricted in Gram-positive organisms with a MIC of 12 µg/mL. SM02 was found active against Mycobacterium sp. and also multi-drug resistant Gram-positive bacteria with similar potency and found to disrupt the bacterial cell wall. Pyrimidomycin also showed significant impairment in the biofilm formation by S. aureus. Furthermore, pyrimidomycin showed synergy with the most used antibiotic like ampicillin, vancomycin and chloramphenicol. Pyrimidomycin did not have cytotoxicity towards human cell lines indicating its limited activity within bacteria.

A handy and accessible tool for identification of Sn(II) in toothpaste.

An easily accessible colorimetric probe, a carbazole-naphthaldehyde conjugate (CNP), was successfully prepared for the selective and sensitive recognition of Sn(II) in different commercially-available toothpaste and mouth wash samples. The binding mechanism of CNP for Sn2+ was confirmed by UV-Vis, 1H, and 13C NMR titrations. The proposed sensing mechanism was supported by quantum chemical calculations. Selective detection of Sn(II) in the nanomolar range (85 nM), among other interfering metal ions, makes it exclusive. Moreover, Sn2+ can be detected with a simple paper strip from toothpaste, which makes this method handy and easily accessible. The potential application of this system for monitoring Sn2+ can be used as an expedient tool for environmental and industrial purposes.

Development of a new fluorescent probe for cysteine detection in processed food samples.

Cysteine is a crucial amino acid, found in a huge amount in protein-rich foods. We focused our research to determine the amount of free cysteine consumed highly in foods such as pork, beef, poultry, eggs, dairy, red peppers, soybeans, broccoli, brussels sprouts, oats, and wheat germs. A newly designed carbazole-pyridine-based fluorescent probe (CPI) has been introduced for quantitative estimation of cysteine (Cys) with a "turn on" fluorescence in some popular processed food samples chosen from our daily diet. CPI shows both naked eye and UV-visible color changes upon interaction with cysteine. The binding approach between CPI and Cys at biological pH has been thoroughly explored by UV-visible and fluorescence spectroscopy. From Job's plot analysis, 1:1 stoichiometric reaction between CPI and Cys is observed with a detection limit of 3.8 µM. NMR, ESI mass spectrometry, and time-dependent density functional theory (TD-DFT) study enlightens the formation of more stable product CPI-Cys. The "turn on" response of the probe CPI occurs due to the interruption of intra-molecular charge transfer (ICT) process upon reacting with cysteine. Moreover, CPI is a very stable, cost-effective compound and exhibits excellent real-time selectivity towards Cys over all other comparative biorelevant analytes. Interestingly, our proposed method is much advantageous as it is able to estimate cysteine predominantly by screening out other comparative biocomponents found in different protein-rich foods.

Consumption of H2S from Our Daily Diet: Determination by a Simple Chemosensing Method.

A unique method has been developed for comparative analysis of H2S produced from food samples from our daily diet, both qualitatively and quantitatively. The selective detection of H2S has been executed by introducing a simple chemodosimeter (PN-N 3 ) that gives response on the basis of intramolecular charge transfer. UV-vis, fluorimetric, and NMR titrations were performed to demonstrate the sensing mechanism and electronic environment of PN-N 3 in the presence of H2S. Density functional theory calculations were performed to validate the mechanism of azide (PN-N 3 ) reduction to amine (PN-NH 2 ) by the strong reducing power of H2S. The potentiality of this chemosensing method is that it could be treated as a simple, less-time-consuming, and cost-effective method for determining H2S in biological samples in the nanomolar range.

2'-Deoxy-5-(hydroxymethyl)cytidine: estimation in human cancer cells with a simple chemosensor.

A pyrrole-based rhodamine conjugate (CS-1) has been developed and characterized for the selective detection and quantification of 2'-deoxy-5-(hydroxymethyl)cytidine (5hmC) in human cancer cells with a simple chemosensing method.