Resolution enhancement in multidimensional solid-state NMR spectroscopy of proteins using spin-state selection

We show that the resolution of homonuclear multidimensional solid-state NMR correlation experiments can be significantly improved using transition selection and spin-state-selective polarization transfer techniques. The selection and transfer of single states allow the removal of the J-coupling contribution from the line width in both the direct and indirect spectral dimensions. This is demonstrated with a new spin-state-selective CO-Calpha correlation experiment, applied to a microcrystalline 85-residue protein. With this new sequence, all four components of the CO-Calpha cross-peaks are separated into different spectra, obtained by linear combination of four recorded data sets. Line narrowing of up to 44% was obtained on the protein sample for the spin-state-selective CO-Calpha spectrum compared to a standard spin-diffusion experiment. The new technique also allows an easy distinction between "direct" and "relayed" transfer cross-peaks.     

Application of samarium diiodide (SmI2)-induced reduction of gamma-acetoxy-alpha,beta-enoates with alpha-specific kinetic electrophilic trapping for the synthesis of amino acid derivatives

Gamma-acetoxy-alpha,beta-enoates were easily reduced by samarium diiodide (SmI2) in THF to generate samarium dienolates which were kinetically trapped with ease at their alpha-positions by electrophiles (proton, aldehydes or ketones) to yield (E)-alkene dipeptide isosteres or gamma-amino acid derivatives in high chemical yields.     

Trace determination of gadolinium in biomedical samples by diode laser-based multi-step resonance ionization mass spectrometry

The application of high-resolution multi-step resonance ionization mass spectrometry (RIMS) to the trace determination of the rare earth element gadolinium is described. Utilizing three-step resonant excitation into an autoionizing level, both isobaric and isotopic selectivity of >10(7) were attained. An overall detection efficiency of approximately 10(-7) and an isotope specific detection limit of 1.5 x 10(9) atoms have been demonstrated. When targeting the major isotope (158)Gd, this corresponds to a total Gd detection limit of 1.6 pg. Additionally, linear response has been demonstrated over a dynamic range of six orders of magnitude. The method has been used to determine the Gd content in various normal and tumor tissue samples, taken from a laboratory mouse shortly after injection of gadolinium diethylenetriaminepentaacetic acid dimeglumine (Gd-DTPA), which is used as a contrast agent for magnetic resonance imaging (MRI). The RIMS results show Gd concentrations that vary by more than two orders of magnitude (0.07-11.5 microg mL(-1)) depending on the tissue type. This variability is similar to that observed in MRI scans that depict Gd-DTPA content in the mouse prior to dissection, and illustrates the potential for quantitative trace analysis in microsamples of biomedical materials.     

Dimethylformamide as a carbon monoxide source in fast palladium-catalyzed aminocarbonylations of aryl bromides

Dimethylformamide (DMF) acts as an efficient source of carbon monoxide and dimethylamine in the palladium-catalyzed aminocarbonylation (Heck carbonylation) of p-tolyl bromide to provide the dimethylamide. Addition of amines to the reaction mixture in excess delivers the corresponding aryl amides in good yields. The amines employed, benzylamine, morpholine, and aniline, all constitute good reaction partners. The reaction proceeds smoothly with bromobenzene and more electron-rich aryl bromides, but electron-deficient aryl bromides fail to undergo aminocarbonylation. The reactions are conducted at 180-190 degrees C for 15-20 min with microwave heating in a reaction mixture containing imidazole and potassium tert-butoxide: the latter is required to promote decomposition of the DMF solvent at a suitable rate. The beneficial effects of controlled microwave irradiation as an energy source for the rapid heating of the carbonylation reaction mixture are demonstrated. The carbonylation procedure reported herein, which relies on the in situ generation of carbon monoxide, serves as a convenient alternative to other carbonylation methods and is particularly applicable to small-scale reactions where short reaction times are desired and the direct use of carbon monoxide gas is impractical.     

Langmuir-Blodgett films of a clay mineral and ruthenium(II) complexes with a noncentrosymmetric structure

Mono- and multilayers of amphiphilic [Ru(phen)(2)(dcC12bpy)](2+) (phen = 1,10-phenanthroline, dcC12bpy = 4,4'-caboxyl-2,2'-bipyridyl didodecyl ester) hybridized with a clay mineral have been prepared by a modified Langmuir-Blodgett method, and their structures and properties have been investigated. Formation of a hybrid monolayer of the Ru(II) complex cations and the clay platelets at an air-clay suspension interface was confirmed by surface pressure-molecular area (pi-A) isotherm measurement and atomic force microscopic (AFM) observation. Multilayers were fabricated by depositing the hybrid monolayers onto glass substrates. The absorbance at 492 nm due to the Ru(II) complex cation in the multilayer increased linearly with the increase in the layer number, indicating layer-by-layer deposition of the hybrid monolayers. Because no increase in the second-harmonic generation (SHG) signal from the multilayers against the layer number was observed, the orientation of the Ru(II) complex cations in the layer would be disturbed. The hydrophilic surface of the transferred hybrid monolayer can be converted to a hydrophobic surface by dipping it in an aqueous solution of octadecylammonium chloride (ODAH(+)Cl(-)). The multilayers modified with ODAH(+) showed a quadratic relation between the SHG intensity and the layer number. This means that the Ru(II) complex cations in the multilayer are successfully oriented in a noncentrosymmetric way by the conversion of the surface property. Both a racemic mixture and an enantiomer of the Ru(II) complex cations were employed to examine the chiral effect on the film properties. The chiral contribution to the SHG signal was enhanced in the multilayer modified with ODAH(+).     

Synthesis,structure, and magnetic properties of dinuclear cobalt(II) complexes with a new phenol-based dinucleating ligand with four hydroxyethyl chelating arms

A new end-off type acyclic ligand with four hydroxyethyl arms, 2,6-bis[bis(2-hydroxyethyl)aminomethyl]-4-methylphenol [H(bhmp)], formed dinuclear cobalt(II) complexes [Co(2)(bhmp)(OAc)(2)]BPh(4) (1) and [Co(2)(bhmp)(OBz)(2)]BPh(4) (2). The complex 1.2.5CH(3)CN (C(50)H(62.5)BCo(2)N(4.5)O(9)) crystallizes in the monoclinic space group C2/c with dimensions a = 25.424(5) A, b = 13.376(2) A, c = 29.913(6) A, beta = 105.930(3) degrees, and V = 9781(3) A(3) and with Z = 8. X-ray diffraction analysis revealed a mu-phenoxo-bis(mu-acetato)dicobalt(II) core structure containing two octahedral cobalt(II) ions. Electronic spectra were investigated for 1 and 2 in the range 400-1800 nm, and the data were typical for the octahedral high-spin cobalt(II) complexes. Magnetic susceptibility was measured for 1 and 2 over the temperature range 4.5-300 K, and the data were analyzed well using our theoretical method. The best fitting parameters were kappa = 0.77, lambda = -116 cm(-1), Delta = 572 cm(-1), and J = -0.44 cm(-1) for complex 1 and kappa = 0.96, lambda = -93 cm(-1), Delta = 616 cm(-1), and J = -0.33 cm(-1) for complex 2.     

Postcolumn fluorescence as an alternative to evaporative light scattering detection for ceramide analysis with gradient elution in non-aqueous reversed-phase liquid chromatography

Ceramide analysis was developed with gradient elution in non-aqueous reversed-phase liquid chromatography with evaporative light scattering detection (ELSD) or postcolumn fluorescence detection. Fluorescence detection (excitation, 360 nm; emission, 425 nm) after postcolumn formation of mixed assemblies between eluted ceramides and 1,6-diphenyl-1,3,5-hexatriene was developed. In comparison with ELSD, fluorescence detection allows a better detection of the minor species ceramide from ceramide type III (commercial mixture of non-hydroxy fatty acid-sphingosine) and appears to be more sensitive for quantitation of ceramides at low concentrations. The fluorescence response is linear over a wide range of injected amount of ceramide III (expressed as stearoyl-phytosphingosine): 10 ng to 1000 ng. The response of ELSD is non linear but can be linearized in double logarithmic coordinates for calculations over a narrow range, e.g. between 10 to 350 ng ceramide III injected. The lower quantitation limits of these two detectors are similar: 5 ng ceramide III was injected.     

alpha-thiocyanation of carbonyl and beta-dicarbonyl compounds using (dichloroiodo)benzene-lead(II) thiocyanate

The combination reagent (dichloroiodo)benzene and lead(II) thiocyanate in dichloromethane effects oxidation of various enol silyl ethers, ketene silyl acetals, and beta-dicarbonyl compounds, thereby providing an efficient and convenient method for alpha-thiocyanation of carbonyl and beta-dicarbonyl compounds.     

Weak intramolecular proton-hydride and proton-fluoride interactions: experimental (NMR, X-ray) and DFT studies of the bis(NBH(3)) and bis(NBF(3)) adducts of 1,3-dimethyl-1,3-diazolidine

Bis(NBH(3)), bis(NBF(3)), and NBF(3)/NBH(3) adducts 1-3 were prepared from 1,3-dimethyl-1,3-diazolidine and characterized by the (1)H, (13)C, (11)B, (19)F, 2D (1)H(-13)C HETCOR and NOESY NMR spectra. The structures and conformations of the adducts were established by the variable-temperature (1)H NMR spectra, the X-ray diffraction method (adduct 2A), and density functional calculations at different theoretical levels. The experimental and theoretical data have revealed that bis adducts 1-3 prefer trans orientations of the borane groups (1A, 2A, 3A) in solution, the solid state, and the gas phase. The studies have shown that the energetic preference of trans adducts with respect to cis compounds, decreasing as 2A (2.9 kcal/mol) > 3A (2.7 kcal/mol) > 1A (1.4 kcal/mol), is dictated by spatially repulsive interactions between the CH(3), BH(3), and BF(3) groups. The results of DFT calculations agree well with an experimental trans/cis isomeric ratio of 9:1 determined in solutions of adduct 1. The calculated geometry and energy, as well as the topological analysis of electronic densities, show that trans adducts 1-3 should exist in gas phase as twist conformations T-2 stabilized by the intramolecular C-H(delta+)...(-delta)H-B or C-H(delta+)...(-delta)F-B interactions. These interactions are characterized as closed-shell. The energy of one proton-hydride and proton-fluoride intramolecular contact, estimated as 1.9 (1A-T-2) and 0.7 (2A-T-2) kcal/mol, respectively, classifies the "elongated" intramolecular interactions CH(delta+)...(-delta)HB and CH(delta+)...(-delta)FB as weak ones. It has been established that, on going from gas phase to a condensed phase (solution and solid), the twist-conformations T-2 transform to conformations T-1, probably by intermolecular dipole-dipole interactions. The data presented in this work show that despite a weakness of the "elongated" proton-hydride and proton-fluoride interactions, they can play a significant role in the stabilization of conformational molecular states, especially when cooperativity is in action.     

A new study of the degradation of hyaluronic acid by hyaluronidase using quartz crystal impedance technique

A new quartz crystal impedance sensing technique for the assay of hyaluronidase (HAse) activity is presented. It is based on the changes in viscosity and density during the enzymatic hydrolysis of hyaluronic acid (HA) by HAse. The variations of equivalent circuit parameters of piezoelectric quartz crystal (PQC) during the enzymatic degradation are discussed. The motional resistance shift curves indicate that the viscosity of the test solutions decreases during the hydrolysis process. The initial hydrolysis rates of HA are obtained from changes in viscosity and density as a function of incubation time. Kinetic parameters (the Michaelis constant K(m) and the maximum hydrolysis rate V(max)) of the degradation reaction are estimated by using a linear Lineweaver-Burk plot in this work. The K(m) was 0.44+/-0.03 mg.ml(-1) and the V(max) was -(5.29+/-0.36)x10(-3) kg.m(-2).s(-1/2).min(-1).