The enhanced green fluorescent protein as a tool for the analysis of protein dynamics and localization: local fluorescence study at the single-molecule level

The green fluorescent protein (GFP) has emerged, in recent years, as a powerful reporter molecule for monitoring gene expression, protein localization and protein-protein interaction. Several mutant variants are now available differing in absorption, emission spectra and quantum yield. Here we present a detailed study of the fluorescence properties of the Phe-64-->Leu, Ser-65-->Thr mutant down to the single molecule level in order to assess its use in quantitative fluorescence microscopy and single-protein trafficking. This enhanced GFP (EGFP) is being used extensively as it offers higher-intensity emission after blue-light excitation with respect to wild-type GFP. By means of fluorescence spectroscopy we demonstrate the absence of the neutral form of the chromophore and the lack of photobleaching recovery after ultraviolet light irradiation. Furthermore, we show that the EGFP spectral properties from isolated to densely packed molecules are highly conserved. From these experiments EGFP emerges as an ideal molecule for quantitative studies of intra and intercellular tagged-protein dynamics and fluorescence-activated cell sorting, but not for monitoring single-protein trafficking over extended periods of time.     

Preparation, Structure, and Properties of V2GeO4F2—Chains of VO4F2 Octahedra in the First V(III) Metallate Fluoride

Powder samples of the new oxide fluoride V₂GeO₄F₂ have been obtained by the reaction of appropriate amounts V₂O₃, VF₃, and GeO₂ at 700°C for 18 h in an argon-filled sealed platinum tube. V₂GeO₄F₂ crystallizes in the space group Pnma with a=9.336(1), b=8.898 (1), and c=4.912 (1) Å. The structure has been refined from X-ray powder diffraction data using the Rietveld method (R_int=5.5% and R_p=9.8%). The structure of V₂GeO₄F₂ exhibits close packed layers of the anions with an ordering of O and F. The characteristic building units are discrete GeO₄ tetrahedra with Ge-O distances of 1.75-1.80 Å. The V are coordinated by four O and two F to form VO₄F₂ octahedra connected via two common edges to give zigzag chains. These chains are linked via corners to form a three-dimensional network. The temperature dependence of the magnetic susceptibility of V₂GeO₄F₂ indicates antiferromagnetic correlations.     

DNA interaction, superoxide scavenging and cytotoxicity studies on new copper(II) complexes derived from a tridentate ligand

The copper complexes [Cu(Pyimpy)(H₂O)](ClO₄)₂ (1), [Cu(Pyimpy)₂](ClO₄)₂ (2), [Cu(Pyimpy)(Cl)₂]·2H₂O (3·2H₂O), [Cu(Pyimpy)(N₃)(ClO₄)]₂ (4) and [Cu(Pyimpy)(SCN)(ClO₄)]₂ (5) were synthesized and characterized by spectroscopic techniques, crystal structures and electrochemical studies (Pyimpy: (2-((2-phenyl-2-(pyridin-2-l)hydrazono)methyl)pyridine)). The superoxide scavenging activity of the two water soluble complexes 1 and 3 was examined. DNA interaction studies by UV-Vis absorption spectral changes during a titration experiment indicated the generation of new species. These small molecule SOD mimics exhibited excellent DNA cleavage activity in the presence of H₂O₂ as well as 2-mercaptoethanol. Complexes 1-5 exhibited better cytotoxicity compared to CuCl₂·2H₂O and the ligand Pyimpy, and showed more potency than cisplatin for MCF-7, PC-3 and HEK-293 cells. Complex 3 exhibited the highest potency for MCF-7, PC-3 and HEK-293 cells compared to the other complexes.     

Poly(PEGDMA‐MAA) copolymeric micro and nanoparticles for oral insulin delivery

Poly(ethylene glycol) dimethacrylate (PEGDMA) and methacrylic acid (MAA) based micro and nanoparticles were prepared and evaluated as a carrier for oral delivery of insulin. PEGDMA was synthesized by esterification reaction of the PEG4000 with MAA in the presence of an acid catalyst. Particles of different size were prepared by emulsion polymerization reaction using different concentration of sodium lauryl sulphate (SLS) as an emulsifying agent. Synthesized copolymeric particle were characterized by attenuated total reflectance‐Fourier transform infrared spectroscopy (ATR‐FTIR), scanning electron microscopy, and acid value. The mean particle diameter of the polymeric micro and nanoparticles at various physiologically relevant pH values was measured using dynamic light scattering. Insulin loading efficiency of the particles was found to be directly proportional to the particle size and inversely proportional to the acid value of the particles. In vitro insulin release studies from various insulin loaded particles were performed by simulating the gastrointestinal tract conditions using HPLC. At pH 2.5, the release of insulin from polymeric particles was observed in the range of 5–8% while a significant higher release (20–35%) was observed at pH 7.4 during first 15 min of in vitro release. Largest size copolymeric particles of 8.3 µm also showed the highest efficiency to reduce the blood glucose level in diabetic rabbits. Copyright © 2010 John Wiley & Sons, Ltd.     

Solution processable indoloquinoxaline derivatives containing bulky polyaromatic hydrocarbons: synthesis, optical spectra, and electroluminescence

New hybrid materials featuring the dipolar fragment 6H-indolo[2,3-b]quinoxaline attached to the bulkier polyaromatic hydrocarbons such as fluoranthene, triphenylene, or polyphenylated benzene have been synthesized by a two-step procedure involving Sonogashira and Diels-Alder reactions. They were characterized by absorption, emission, electrochemical, thermal, and theoretical investigations. The electronic properties of the compounds were dominated by the 6H-indolo[2,3-b]quinoxaline chromophore, and the incorporation of polyaromatic hydrocarbons reduces the chances of nonradiative deactivation processes associated with the excited state and improves the emission properties. The compounds displayed cyan emission with moderate quantum efficiency when excited at the absorption maximum. All of the compounds exhibited an irreversible reduction process corresponding to the addition of electron at the quinoxaline segment. They showed moderate thermal stability and glass transition temperature greater than 100 °C. The presence of rigid 6H-indolo[2,3-b]quinoxaline and bulkier polyaromatic hydrocarbon segments enhances the thermal stability and glass transition temperature significantly. Finally, the dyes were successfully applied as an electron-transporting and emitting layer in multilayered organic light-emitting diodes comprising a N,N'-bis(l-naphthyl)-N,N'-diphenyl-1,1'-biphenyl-4,4'-diamine hole-transporting layer. The cyan emitting devices were characterized by moderate device performance parameters.     

Facile Reduction of Chalcones to Dihydrochalcones with NaBH4/Ni2+ System

Chalcones have been found to undergo facile reduction on treatment with sodium borohydride-nickel chloride system in dioxan-methanol medium to afford dihydrochalcones.     

Titania-silica nanoparticles ensemblies assisted heterogeneous catalytic strategy for the synthesis of pharmacologically significant 2,3-diaryl-3,4-dihydroimidazo[4,5-b]indole scaffolds

Present paper elicits the multicomponent reaction (MCR) strategy assisted by titania nanoparticles hosted on silica (TiO₂.SiO₂ NPs) as heterogeneous catalyst to synthesize a series of pharmacologically significant 2,3-diaryl-3,4-dihydroimidazo[4,5-b]indole derivatives. To the best of our information, the use of isatin as one of the precursors was hitherto unreported. The decrease in reaction time, low catalyst loading, high product yield (up to 92%), and excellent reusability of the catalyst (up to 7 cycles) put this protocol under the umbrella of green chemistry tenets. Characterization of catalysts was achieved through a number of techniques viz., energy-dispersive X-ray (EDX) spectroscopy, field emission scanning electron microscopy (FESEM), powder X-ray diffraction (XRD), fourier transform infrared (FTIR) spectra of adsorbed pyridine, temperature-programmed desorption of ammonia, and porosity measurements by nitrogen adsorption (Brunauer–Emmett–Teller [BET] method). Also, the structures of synthesized compounds were corroborated on the basis of FTIR, nuclear magnetic resonance (NMR), mass, and elemental analyses data.     

Functionalization of Pyridine and Quinoline Scaffolds by Using Organometallic Li-, Mg- and Zn-Reagents

This Review discusses the various methods for functionalizing pyridine and quinoline scaffolds, including direct selective metalation (DoM), halogen/metal exchange reactions, Li, Mg, and Zn insertion, and trans-metalation approaches, which are then followed by cross-coupling reactions of the Kumada or Negishi types. Selective deprotonation of aryl or pyridyl/quinolinyl derivatives can be performed using n-BuLi, LDA, and TMP-based different organolithium, -magnesium, and -zinc reagents. The functionalized pyridine and quinoline-based heterocyclic compounds were prepared by selectively deprotonating with presenting a directing functional group substituted pyridine/quinoline analogues in the presence of TMP-bases (TMP−Li, Mg, Zn reagents). Different aryl or alkyl Li, Mg, and Zn reagents with electron-donating and electron-withdrawing substituents undergo transition metal-catalyzed C(sp²)−C(sp²) and C(sp²)−C(sp³) types of cross-coupling reactions with pyridine/quinoline halides under mild conditions with the sustainable process producing complex N-heterocycles. Using moderate and sustainable reaction conditions, sensitive functional group tolerance, and inexpensive and low toxic chemicals, highly functionalization of pyridine and quinoline-based bioactive therapeutic scaffolds and natural products was accomplished. Therefore, in this article, we provide a succinct overview of the numerous synthetic strategies and practical methods used by various authors between 2010 and 2023 to functionalize pyridine and quinoline analogues using diverse Li, Mg, and Zn organometallic reagents.     

Novel Electrochemical Biosensing for Detection of Neglected Tropical Parasites of Animal Origin: Recent Advances

Parasitic diseases are among neglected disease of human and animals, especially in tropical and sub tropical regions. In the era of artificial intelligence, the novel biosensing diagnostic platform is needed for an early control measure implementation. This goal can be successfully achieved by onsite application of electrochemical biosensors. They are being developed towards point of care diagnostics; however commercial availability is scanty. The recent developments during last one decade in terms of the electrode surface modification for rapid diagnosis of important emerging/re-emerging parasites is presented. The information would help future improvement in the electrochemical biosensing of parasites.