Analysis of plasma tocopherols alpha, gamma, and 5-nitro-gamma in rats with inflammation by HPLC coulometric detection

Reactive nitrogen oxide species (RNOS) have been implicated as effector molecules in inflammatory diseases. There is emerging evidence that gamma-tocopherol (gammaT), the major form of vitamin E in the North American diet, may play an important role in these diseases. GammaT scavenges RNOS such as peroxynitrite by forming a stable adduct, 5-nitro-gammaT (NGT). Here we describe a convenient HPLC method for the simultaneous determination of NGT, alphaT, and gammaT in blood plasma and other tissues. Coulometric detection of NGT separated on a deactivated reversed-phase column was linear over a wide range of concentrations and highly sensitive (approximately 10 fmol detection limit). NGT extracted from blood plasma of 15-week-old Fischer 344 rats was in the low nM range, representing approximately 4% of gammaT. Twenty-four h after intraperitoneal injection of zymosan, plasma NGT levels were 2-fold higher compared to fasted control animals when adjusted to gammaT or corrected for total neutral lipids, while alpha- and gammaT levels remained unchanged. These results demonstrate that nitration of gammaT is increased under inflammatory conditions and highlight the importance of RNOS reactions in the lipid phase. The present HPLC method should be helpful in clarifying the precise physiological role of gammaT.

S-0 and S-1 State Structure, Methyl Torsional Barrier Heights, and Fast Intersystem Crossing Dynamics of 5-Methyl-2-hydroxypyrimidine

We report the analysis of the SI So rotational band contours of jet-cooled 5-methyl-2-hydroxypyrimidine (5M2HP), the enol form of deoxythymine. Unlike thymine, which exhibits a structureless spectrum, the vibronic spectrum of 5M2HP is well structured, allowing us to determine the rotational constants and the methyl group torsional barriers in the S-0 and S-1 states. The 0(0)(0), 6a(0)(1), 6b(0)(1), and 14(0)(1) band contours were measured at 900 MHz (0.03 cm(-1)) resolution using mass-specific two-color resonant two-photon ionization (2C-R2PI) spectroscopy. All four bands are polarized perpendicular to the pyrimidine plane (>90% c type), identifying the S-1 <- S-0 excitation of 5M2HP as a 1n pi* transition. All contours exhibit two methyl rotor subbands that arise from the lowest 5-methyl torsional states 0A '' and 1E ''. The S-0 and S-1 state torsional barriers were extracted from fits to the torsional subbands. The 3-fold barriers are V-3 '' = 13 cm(-1) and V3' = SI cm(-1); the 6-fold barrier contributions V-6 '' and V-6' are in the range of 2-3 cm(-1) and are positive in both states. The changes of A, B, and C rotational constants upon S-1 <- S-0 excitation were extracted from the contours and reflect an "anti-quinoidal" distortion. The 0(0)(0) contour can only be simulated if a 3 GHz Lorentzian line shape is included, which implies that the S-1(1n pi*) lifetime is similar to 55 ps. For the 6a(0)(1) and 6b(0)(1) bands, the Lorentzian component increases to 5.5 GHz, reflecting a lifetime decrease to similar to 30 ps. The short lifetimes are consistent with the absence of fluorescence from the 1n pi* state. Combining these measurements with the previous observation of efficient intersystem crossing (ISC) from the Si state to a long-lived T-1((3)n pi*) state that lies similar to 2200 cm(-1) below [S. Lobsiger, S. et al. Phys. Chem. Chem. Phys. 2010, 12, 5032] implies that the broadening arises from fast intersystem crossing with k(ISC) approximate to 2 x 10(10) s(-1). In comparison to 5-methylpyrimidine, the ISC rate is enhanced by at least 10 000 by the additional hydroxy group in position 2.

Replacement of histidine in position 105 in the alpha(5) subunit by cysteine stimulates zolpidem sensitivity of alpha(5)beta(2)gamma(2) GABA(A) receptors

Zolpidem is a positive allosteric modulator of GABA(A) receptors with sensitivity to subunit composition. While it acts with high affinity and efficacy at GABA(A) receptors containing the alpha(1) subunit, it has a lower affinity to GABA(A) receptors containing alpha(2), alpha(3), or alpha(5) subunits and has a very weak efficacy at receptors containing the alpha(5) subunit. Here, we show that replacing histidine in position 105 in the alpha(5) subunit by cysteine strongly stimulates the effect of zolpidem in receptors containing the alpha(5) subunit. The side chain volume of the amino acid residue in this position does not correlate with the modulation by zolpidem. Interestingly, serine is not able to promote the potentiation by zolpidem. The homologous residues to alpha(5)H105 in alpha(1), alpha(2), and alpha(3) are well-known determinants of the action of classical benzodiazepines. Other studies have shown that replacement of these histidines alpha(1)H101, alpha(2)H101, and alpha(3)H126 by arginine, as naturally present in alpha(4) and alpha(6), leads to benzodiazepine insensitivity of these receptors. Thus, the nature of the amino acid residue in this position is not only crucial for the action of classical benzodiazepines but in alpha(5) containing receptors also for the action of zolpidem.

Structural Studies of Transition Metal Complexes with 4,5-Bis(2-pyridylmethylsulfanyl)-4‘,5‘-ethylenedithiotetrathiafulvalene: Probing Their Potential for the Construction of Multifunctional Molecular Assemblies

Three divalent transition metal complexes of 4,5-bis(2-pyridylmethylsulfanyl)-4‘,5‘-ethylenedithiotetrathiafulvalene have been prepared and crystallographically characterized. The isostructural Co(II) and the Ni(II) complexes show octahedral geometries around the metal ions with the coordination sites occupied by the pyridyl nitrogen atoms and the thioether sulfur atoms of the ligand and cis coordination of the halide ions. Cyclic voltammetry reveals that the complexation leads to a small anodic shift in the first oxidation potential of the TTF system.