Mass spectrometry combined with affinity probes for the identification of CP4 EPSPS in genetically modified soybeans

Sample preparation methods used for genetically modified organisms (GMOs) analysis are often time consuming, require extensive manual manipulation, and result in limited amounts of purified protein, which may complicate the detection of low‐abundance GM protein. A robust sample pretreatment method prior to mass spectrometry (MS) detection of the transgenic protein (5‐enolpyruvylshikimate‐3‐phosphate synthase [CP4 EPSPS]) present in Roundup Ready soya is investigated. Liquid chromatography‐multiple reaction monitoring tandem MS (nano LC‐MS/MS‐MRM) was used for the detection and quantification of CP4 EPSPS. Gold nanoparticles (AuNPs) and concanavalin A (Con A)‐immobilized Sepharose 4B were used as selective probes for the separation of the major storage proteins in soybeans. AuNPs that enable the capture of cysteine‐containing proteins were used to reduce the complexity of the crude extract of GM soya. Con A‐sepharose was used for the affinity capture of β‐conglycinin and other glycoproteins of soya prior to enzymatic digestion. The methods enabled the detection of unique peptides of CP4 EPSPS at a level as low as 0.5% of GM soya in MRM mode. Stable‐isotope dimethyl labeling was further applied to the quantification of GM soya. Both probes exhibited high selectivity and efficiency for the affinity capture of storage proteins, leading to the quantitative detection at 0.5% GM soya, which is a level below the current European Union's threshold for food labeling. The square correlation coefficients were greater than 0.99. The approach for sample preparation is very simple without the need for time‐consuming protein prefractionation or separation procedures and thus presents a significant improvement over existing methods for the analysis of the GM soya protein.     

Synthesis,Characterization, DNA Binding Studies,Photocleavage, Cytotoxicity and Docking Studies of Ruthenium(II) Light Switch Complexes

A new ligand 3-(1H-imidazo[4,5-f][1,10]phenanthrolin-2yl)phenylboronic acid and its (IPPBA) three ruthenium(II) complexes [Ru(phen)₂(IPPBA)](ClO₄)₂ (1), [Ru(bpy)₂(IPPBA)](ClO₄)₂ (2) and [Ru(dmb)₂(IPPBA)](ClO₄)₂ (3) have been synthesized and characterized by elemental analysis, UV/VIS, IR, ¹H-NMR,¹³C-NMR and mass spectra. The binding behaviors of the three complexes to calf thymus DNA were investigated by absorption spectra, emission spectroscopy, viscosity measurements, thermal denaturation and photoactivated cleavage. The DNA-binding constants for complexes 1, 2 and 3 have been determined to be 7.9?×?10⁵ M^?1, 6.7?×?10⁵ M^?1 and 2.9?×?10⁵ M^?1. The results suggest that these complexes bound to double-stranded DNA in an intercalation mode. Upon irradiation at 365 nm, three ruthenium complexes were found to promote the cleavage of plasmid pBR322 DNA from super coiled form ? to nicked form ??. Further in the presence of Co²⁺, the emission of DNA–Ru(ΙΙ) complexes can be quenched. And when EDTA was added, the emission was recovered. The experimental results show that all three complexes exhibited the “on–off–on” properties of molecular “light switch”. The highest Cytotoxicity potential of the complex1 was observed on the Human alveolar adenocarcinoma (A549) cell line. Good agreement was generally found between the spectroscopic techniques and molecular docked model which provides further evidence of groove binding.     

Cryogenic absorption cells operating inside a Bruker IFS-125HR: First results for CH4 at 7 μm

New absorption cells designed specifically to achieve stable temperatures down to 66 K inside the sample compartment of an evacuated Bruker IFS-125HR Fourier transform spectrometer (FTS) were developed at Connecticut College and tested at the Jet Propulsion Laboratory (JPL). The temperature stabilized cryogenic cells with path lengths of 24.29 and 20.38 cm were constructed of oxygen free high conductivity (OFHC) copper and fitted with wedged ZnSe windows using vacuum tight indium seals. In operation, the temperature-controlled cooling by a closed-cycle helium refrigerator achieved stability of ±0.01 K. The unwanted absorption features arising from cryodeposits on the cell windows at low temperatures were eliminated by building an internal vacuum shroud box around the cell which significantly minimized the growth of cryodeposits. The effects of vibrations from the closed-cycle helium refrigerator on the FTS spectra were characterized. Using this set up, several high-resolution spectra of methane isotopologues broadened with nitrogen were recorded in the 1200-1800 cm⁻¹ spectral region at various sample temperatures between 79.5 and 296 K. Such data are needed to characterize the temperature dependence of spectral line shapes at low temperatures for remote sensing of outer planets and their moons. Initial analysis of a limited number of spectra in the region of the R(2) manifold of the ν₄ fundamental band of ¹³CH₄ indicated that an empirical power law used for the temperature dependence of the N₂-broadened line widths would fail to fit the observed data in the entire temperature range from 80 to 296 K; instead, it follows a temperature-dependence similar to that reported by Mondelain et al. [17] and [18]. The initial test was very successful proving that a high precision Fourier transform spectrometer with a completely evacuated optical path can be configured for spectroscopic studies at low temperatures relevant to the planetary atmospheres.     

Multispectrum analysis of the ν9 band of C2H6: Positions, intensities, self- and N2-broadened half-width coefficients

Line positions, intensities, Lorentz self- and N₂-broadened half-width coefficients have been measured for ^PQ₃, ^PQ₂, ^PQ₁, ^RQ₀, ^RQ₁, ^RQ₂, and ^RQ₃ sub-band transitions in the ν₉ fundamental band of ¹²C₂H₆. A multispectrum nonlinear least-squares fitting technique was used to fit up to 17 high-resolution (∼0.00156 cm⁻¹), room temperature absorption spectra of pure (99.99% chemical purity) natural sample of ethane and lean mixtures of the high-purity ethane diluted with N₂. A Bruker IFS 120HR Fourier transform spectrometer located at the Pacific Northwest National Laboratory (PNNL), in Richland, Washington was used to record the data. A standard Voigt line shape was assumed to fit all the data since no line mixing or other non Voigt line shapes were required to fit any of the spectra used in the analysis. Short spectral intervals (∼2-2.5 cm⁻¹) of all 17 spectra covering a specific ^PQ or ^RQ sub-band were fit simultaneously. For the first time in an ethane band, pressure-broadened half-width coefficients were determined for the torsional-split components. However, for better reliability of the retrieved coefficients for the weaker components (transitions with large intensity ratios of 4:1 or 3:1 for most K levels between the strong and weak components), constraints were used such that the half-width coefficients of both torsional-split components for a given J were identical for a specific broadening gas. No pressure-induced shift coefficients were necessary to fit the spectra to their noise level. The present study revealed for the first time the dependence of self- and N₂-broadened half-width coefficients upon the J, K quantum numbers of the transitions in ethane. A number of transitions belonging to the ν₉+ν₄−ν₄ and the ν₉+2ν₄−2ν₄ hot bands were also observed in the fitted regions and measurements were made when possible.     

High-power, continuous-wave Ti:sapphire laser pumped by fiber-laser green source at 532 nm

We report the first experimental demonstration of efficient and high-power operation of a Ti:sapphire laser pumped by a simple, compact, continuous-wave (cw) fiber-laser-based green source. The pump radiation is obtained by direct single-pass second-harmonic-generation (SHG) of a 33 W, cw Yb-fiber laser in a 30-mm-long MgO:sPPLT crystal, providing 11 W of single-frequency green power at 532 nm in TEM₀₀ spatial profile with power and frequency stability better than 3.3% and 32 MHz, respectively, over 1 h. The Ti:sapphire laser is continuously tunable across 743-970 nm and can deliver an output power up to 2.7 W with a slope efficiency as high as 32.8% under optimum output coupling of 20%. The laser output has a TEM₀₀ spatial profile with M²<1.44 across the tuning range and exhibits a peak-to-peak power fluctuation below 5.1% over 1 h.     

Design,synthesis, and antibacterial studies of potent pyrazolinyltriazoles

Research on Chemical Intermediates - A library of pyrazolinyltriazole hybrids (3a–l, 4a–l) was synthesized via azide–alkyne dipolar (Huisgen) cycloaddition of azidoacetyl...     

Diphenyltin(IV) complexes of the 5-[(E)-2-(aryl)-1-diazenyl]quinolin-8-olates: Synthesis and multinuclear NMR, Sn Mössbauer, electrospray ionization MS, X-ray characterization and assessment of in vitro cytotoxicity

A series of cis-bis{5-[(E)-2-(aryl)-1-diazenyl]quinolinolato}diphenyltin(IV) complexes have been synthesized and characterized by ¹H, ¹³C, ¹¹⁹Sn NMR, ESI-MS, IR and ^119mSn Mössbauer spectroscopic techniques in combination with elemental analysis. The structures of a ligand L⁶H (i.e., 5-[(E)-2-(4-ethoxyphenyl)-1-diazenyl]quinolin-8-ol) and three diphenyltin(IV) complexes, viz., Ph₂Sn(L¹)₂ · (CH₃)₂CO (1), Ph₂Sn(L⁴)₂ (4) and Ph₂Sn(L⁵)₂ (5) (L = 5-[(E)-2-(aryl)-1-diazenyl]quinolin-8-ol: aryl = phenyl - (L¹H); 4′-methylphenyl - (L⁴H) and 4′-bromophenyl - (L⁵H)) were determined by single crystal X-ray diffraction. In general, the complexes were found to adopt a distorted cis-octahedral arrangement around the tin atom. These complexes retain their solid-state structure in non-coordinating solvent as evidenced by ¹¹⁹Sn NMR spectroscopic results. The in vitro cytotoxicity of 1 is reported and compared with Ph₂Sn(Ox)₂ (Ox = deprotonated quinolin-8-ol) against seven well characterized human tumor cell lines.     

Dehydration of 1,4-dioxane through blend membranes of poly(vinyl alcohol) and chitosan by pervaporation

Blend membranes prepared from poly(vinyl alcohol) (PVA) and chitosan (CS) were crosslinked with glutaraldehyde and used in the pervaporation dehydration of 1,4-dioxane. Membranes were characterized by Fourier transform infrared spectroscopy (FTIR), thermogravimetric analysis (TGA), differential scanning calorimetry (DSC) and X-ray diffraction (X-RD) to assess, respectively, the intermolecular interactions, thermal stability and crystallinity. Equilibrium sorption studies were carried out in pure liquids and binary mixtures of different compositions of water + 1,4-dioxane mixtures to assess the polymer–liquid interactions. The crosslinked membrane showed a good potential in breaking the azeotrope of 82 wt.% aqueous 1,4-dioxane giving a selectivity of 117 with a reasonable water flux of 0.37 kg/m² h. The effect of operating parameters such as feed composition, membrane thickness and permeate pressure was evaluated.     

Determination of amoxapine and nortriptyline in blood plasma and serum by salt-assisted liquid-liquid microextraction and high-performance liquid chromatography

A new, selective and sensitive method has been developed for the determination of tricyclic antidepressant drugs, amoxapine and nortriptyline, in human blood plasma and serum, involving their reaction with allyl isothiocyanate and extraction of thiourea derivatives with water-miscible organic solvent acetonitrile. The phase separation was effected by addition of ammonium sulphate, a process called salt-assisted liquid-liquid microextraction. The extract was analyzed by HPLC with UV detection at 254?nm. The method has been optimized for derivatization reaction time and temperature, solvent for extraction, and salt for solvent phase separation. Under the optimal conditions, a linear calibration graph was obtained between the amount of drug and the peak area of thiourea derivatives in the range of 0.002-20?mg/L drugs. The correlation coefficient and limit of detection values for amoxapine and nortriptyline in serum/plasma samples were in the range of 0.9953-0.9999 and 0.46-0.58?μg/L, respectively. The recovery in spiking experiments ranged, respectively, 75-88% (RSD 3.4-7.2%) and 79-97% (RSD 3.7-7.9%) for the two drugs.     

Isoquinolinium bromochromate： An efficient and stable reagent for bromination of hydroxylated aromatic compounds and oxidation of alcohols

The new chromium(Ⅵ) oxidizing reagent isoquinolinium bromochromate(IQBC) was prepared and characterized.The IQBC has been found to be stable and an efficient solid reagent which can be easily prepared in good yield.It act as an efficient brominating reagent for hydroxylated aromatic compounds as well as good oxidizing reagent for the conversion of alcohols to carbonyl compounds in good to excellent yield.The synthesized isoquinolinium bromochromate is more ideal reagent,with number of specification inclu...