Nanomolar Nitric Oxide Concentrations in Living Cells Measured by Means of Fluorescence Correlation Spectroscopy

Measurement of the nitric oxide (NO) concentration in living cells in the physiological nanomolar range is crucial in understanding NO biochemical functions, as well as in characterizing the efficiency and kinetics of NO delivery by NO-releasing drugs. Here, we show that fluorescence correlation spectroscopy (FCS) is perfectly suited for these purposes, due to its sensitivity, selectivity, and spatial resolution. Using the fluorescent indicators, diaminofluoresceins (DAFs), and FCS, we measured the NO concentrations in NO-producing living human primary endothelial cells, as well as NO delivery kinetics, by an external NO donor to the immortal human epithelial living cells. Due to the high spatial resolution of FCS, the NO concentration in different parts of the cells were also measured. The detection of nitric oxide by means of diaminofluoresceins is much more efficient and faster in living cells than in PBS solutions, even though the conversion to the fluorescent form is a multi-step reaction.     

Influence of laser microfabrication on silicon electrochemical behavior in HF solution

Influence of direct laser writing with femtosecond pulses on electrochemical etching of n-type low conductivity (>1,000 Ωcm) silicon is demonstrated. It has been shown that thermal 1-μm-thick SiO₂ layer on silicon surface can be used as a protective layer in the electrochemical etching process. It has been found that laser ablation changes not only the surface morphology and structure of silicon samples but also the character of their anodic etching in aqueous solution of hydrofluoric acid. Formation of microvoids and caverns of irregular shape has been observed at the laser-ablated sites. It is proposed that the change of silicon conductivity from n- to p-type takes place at the laser fabricated regions. Processes of Si anodic oxidation and electrochemical etching are discussed.     

The effect of selected surfactants on the structure of a bicellar system (DMPC/DHPC) studied by SAXS

The stabilizing or disturbing effect of different surfactants on the bicellar phase of phospholipids significantly depends on their type. The effect of different surfactants on the bicellar structure made of a mixture of phospholipids 1,2-dimyristoyl-sn-glycero-3-phosphocholine and 1,2-dihexanoyl-sn-glycero-3-phospho-choline (DMPC/DHPC) has been studied by the small angle scattering of synchrotron radiation. The study has been performed for three surfactants: dodecyldimethyl-(hexyloxymethyl)ammonium chloride, n-undecylammonium chloride and t-octylphenoxypolyethoxyethanol (Triton X-100) introduced into a bicellar solution of DMPC/DHPC (2.8:1). The bicellar phase has been disturbed in the shortest time in the presence of dodecyldimethyl-(hexyloxymethyl)ammonium chloride in this system a transition from the bicellar to lamellar structure has been directly visible. The changes have been less pronounced in the presence of undecylammonium chloride and practically not noted in the presence of Triton X-100.     

Influence of fluorine substituents on the NMR properties of phenylboronic acids

The paper presents results of a systematic NMR studies on fluorinated phenylboronic acids. All possible derivatives were studied. The experimental ¹H, ¹³C, ¹⁹F, ¹¹B, and ¹⁷O spectral data were compared with the results of theoretical calculations. The relation between the calculated natural bond orbital parameters and spectral data (chemical shifts and coupling constants) is discussed. The first examples of ¹⁰B/¹¹B isotopic effect on the ¹⁹F spectra and ⁴J_FO scalar coupling in organic compounds are reported. Copyright © 2014 John Wiley & Sons, Ltd.     

GQDs-MSNs nanocomposite nanoparticles for simultaneous intracellular drug delivery and fluorescent imaging

Although number of stimuli-responsive drug delivery systems based on mesoporous silica nanoparticles (MSNs) have been developed, the simultaneous real-time monitoring of carrier in order to guarantee proper drug targeting still remains as a challenge. GQDs-MSNs nanocomposite nanoparticles composed of graphene quantum dots (GQDs) and MSNs are proposed as efficient doxorubicin delivery and fluorescent imaging agent, allowing to monitor intracellular localization of a carrier and drug diffusion route from the carrier.Graphene quantum dots (average diameter 3.65?±?0.81 nm) as a fluorescent agent were chemically immobilized onto mesoporous silica nanoparticles (average diameter 44.08?±?7.18 nm) and loaded with doxorubicin. The structure, morphology, chemical composition, and optical properties as well as drug release behavior of doxorubicin (DOX)-loaded GQDs-MSNs were investigated. Then, the in vitro cytotoxicity, cellular uptake, and intracellular localization studies were carried out. Prepared GQDs-MSNs form stable suspensions exhibiting excitation-dependent photoluminescence (PL) behavior. These nanocomposite nanoparticles can be easily DOX-loaded and show pH- and temperature-dependent release behavior. Cytotoxicity studies proved that GQDs-MSNs nanocomposite nanoparticles are nontoxic; however, when loaded with drug, they enable the therapeutic activity of DOX via its active delivery and release. GQDs-MSNs owing to their fluorescent properties and efficient in vitro cellular internalization via caveolae/lipid raft-dependent endocytosis show a high potential for the optical imaging, including the simultaneous real-time optical tracking of the loaded drug during its delivery and release.

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Molecular dynamics of poly(L-lactide) biopolymer studied by wide-line solid-state 1H and 2H NMR spectroscopy

The molecular dynamics of poly(L-lactide) (PLLA) biopolymer was characterized through analyses of 1H and 2H NMR line-shapes and spin-lattice relaxation times at different temperatures. At low temperatures (e.g. 90 K), the methyl group rotation is dominant leading to a significant reduction in the proton second moment. Fast methyl group reorientation occurs at ca. 130 K. In additional to the fast methyl group rotation, hydroxyl groups start to reorient as the temperature increases further, eventually leading to the breakdown of the segments of the biopolymer chains above its glass transition temperature Tg of 323 K. The analyses of the 2H NMR line-shapes indicate that both the methyl and hydroxyl reorientations can be described by the so-called cone model, in which the former has three equilibrium positions with theta(C-D) = 70.5 degrees and phi = 120 degrees while the latter one exhibits two equilibrium positions with theta(O-D) = 78 degrees and phi = 180 degrees .     

Proton longitudinal NMR relaxation of poly(p-phenylene sulfide) in the laboratory and the rotating frames reference

Spin-lattice NMR relaxation times T1 in the laboratory frame and T1rho(off) as well as T1rho(off) in the rotating frame off-resonance were employed to the study of molecular dynamics of both pristine PPS and thermally treated poly(p-phenylene sulfide) (PPS). The temperature dependence of T1 was exponential in the whole temperature range studied, whereas T1rho only in low temperatures. In the high temperature range the distribution of relaxation times T1rho and correlation times tau(c) as well as activation energy Ea was observed. The distribution of activation energy determined from T1 minima at 15 and 30 MHz and from low temperature slopes of T1rho dependence as well as from spectral density functions (estimated from proton off-resonance technique) was attributed to the reorientation of phenylene groups around the sulfur-phenyl-sulfur axis in amorphous and crystalline phases of PPS. Furthermore, it is suggested that an additional relaxation mechanism related to interactions of protons with paramagnetic centers is operative in a low temperature range. After thermal treatment of PPS the low temperature minima disappeared and the relaxation times shortened in the low temperature regime. Both these facts were attributed to an increased contribution of spin diffusion in the relaxation process.     

Rheological and NMR studies of polyethylene/calcium carbonate composites

Rheometry, 13C CP/MAS NMR spectra and 1H spin-lattice relaxation times T1 and T1rho have been employed to study the structure and molecular dynamics in composites of polyethylene (LDPE) with calcium carbonate filler. It has been found that the addition of the filler into the polymer leads to an increase in composite rigidity and a decrease in mobility in its crystalline regions. The presence of the filler affects the crystallization process making the crystal structure less perfect and reduces the size of the crystallites.