Systematic re-evaluation of the bis(2-hydroxyethyl)disulfide (HEDS) assay reveals an alternative mechanism and activity of glutaredoxins

The reduction of bis(2-hydroxyethyl)disulfide (HEDS) by reduced glutathione (GSH) is the most commonly used assay to analyze the presence and properties of enzymatically active glutaredoxins (Grx), a family of central redox proteins in eukaryotes and glutathione-utilizing prokaryotes. Enzymatically active Grx usually prefer glutathionylated disulfide substrates. These are converted via a ping-pong mechanism. Sequential kinetic patterns for the HEDS assay have therefore been puzzling since 1991. Here we established a novel assay and used the model enzyme ScGrx7 from yeast and PfGrx from Plasmodium falciparum to test several possible causes for the sequential kinetics such as pre-enzymatic GSH depletion, simultaneous binding of a glutathionylated substrate and GSH, as well as substrate or product inhibition. Furthermore, we analyzed the non-enzymatic reaction between HEDS and GSH by HPLC and mass spectrometry suggesting that such a reaction is too slow to explain high Grx activities in the assay. The most plausible interpretation of our results is a direct Grx-catalyzed reduction of HEDS. Physiological implications of this alternative mechanism and of the Grx-catalyzed reduction of non-glutathione disulfide substrates are discussed.     

Development and validation of a liquid chromatography–tandem mass spectrometry method for the simultaneous quantification of tamoxifen, anastrozole, and letrozole in human plasma and its application to a clinical study

There is substantial evidence that circulating estrogens promote the proliferation of breast cancer. Consequently, adjuvant hormonal treatment strategies targeting estrogen action have been established. Such hormonal therapies include selective estrogen receptor modulators, such as tamoxifen, which interfere at the estrogen receptors directly, or non-steroidal aromatase inhibitors, such as anastrozole and letrozole, which inhibit estrogen synthesis through blocking the aromatase, a key enzyme of estrogen production. Despite considerable therapeutic success, in several cases, the use of these drugs is limited by side effects that have been described to significantly impair the adherence of patients to endocrine treatment. However, objective data concerning patient adherence and its clinical relevance are limited. One promising approach to check patient-reported adherence is drug monitoring in human plasma. Therefore, a liquid chromatography–tandem mass spectrometry method to determine the plasma concentrations of tamoxifen, anastrozole, and letrozole has been developed and fully validated according to guidelines for clinical and forensic toxicology. The validation criteria evaluated were selectivity, linearity, accuracy and precision, limit of quantification, recovery and matrix effects, sample stability, and carryover. The six-point calibration curves showed linearity over the range of concentrations from 25 to 500 ng/ml for tamoxifen, 5 to 200 ng/ml for anastrozole, and 10 to 300 ng/ml for letrozole. The intra- and inter-day precision and accuracies were always better than 15%. The validated procedure was successfully applied to a clinical study (Patient-Reported Outcomes in Breast Cancer Patients undergoing Endocrine Therapy, PRO-BETh). A major aim of PRO-BETh study is the comprehensive evaluation of adherence to treatment in pre- and post-menopausal women with breast cancer. Plasma samples of 310 breast cancer patients undergoing anti-estrogen therapy were analyzed. Eight samples did not contain a quantifiable amount of drug, strongly indicating non-adherence of the corresponding patients to adjuvant breast cancer treatment. Furthermore, plasma concentrations at the lower end of the observed plasma level distribution might represent a hint but not a confirmation for non-adherence in terms of non-daily and irregular intake of the prescribed drug.     

Squeezing in strong light scattered by a regular structure of atoms

Squeezing in the resonance fluorescence emitted by a regular structure of atoms is discussed. Intense squeezed light fields are a prerequisite, e.g., for efficient measurements with sensitivity below the standard quantum limit. Regular structures are known to offer macroscopic squeezed light due to the fixed phase relation between the light scattered by the respective atoms, but typically restrict squeezing to low incident light intensity. As our main result, we demonstrate that this restriction to low driving field intensity can be circumvented. For this, we assume a suitable modification of the surrounding electromagnetic bath, as it can be achieved, e.g., with a cavity. The modified environment leads to a redistribution of the collective dressed state populations, such that squeezing is recovered at strong driving. In such modified environments, squeezing even occurs for a resonant driving field, in contrast to the regular vacuum case.     

Adsorption studies of a microporous phthalocyanine network polymer

The adsorption/desorption of N2 at 77 K and the adsorption from aqueous solution at 298 K of four organic probe molecules of different sizes (phenol, 4-nitrophenol, orange II, naphthol green B) were studied for a phthalocyanine network polymer of intrinsic microporosity (PIM) and for an activated carbon (Darco 20-40 mesh). N2 sorption analysis gave similar surface areas for the PIM and the carbon (610 and 545 m2 g(-1), respectively) but showed differences in pore size distribution, the PIM being essentially microporous (pore size < 2 nm), with a high proportion of ultramicropores (<0.7 nm), while the carbon had a broader pore size distribution, extending into the mesopore region. The carbon acted as an adsorbent for all the organic probe molecules studied, while the PIM was more selective, adsorbing the smaller molecules but rejecting the large dye naphthol green B. The PIM offers selectivity combined with a well-defined chemical structure incorporating catalytic sites.     

Facile migratory insertion of a N-heterocyclic carbene into a ruthenium-carbon double bond: a new type of reaction of a NHC ligand

The reaction of [RuCp(IPri)(CH3CN)2]PF6 (IPri = 1,3-bis(2,6-diisopropylphenyl)imidazol-2-ylidene) with HCCR (R = COOMe, COOEt, COMe) yields the allyl carbene complexes [RuCp(=C(R)-eta3-CHC(R)CH-IPri)]PF6. This conversion involves selective head-to-tail coupling of two alkynes and an unusual migratory insertion of the N-heterocyclic carbene into the ruthenium-carbon double bond of a ruthenacyclopentatriene intermediate.     

Sodium Trinitratouranylate(VI) Na[UO2(NO3)3]

Although potassium, rubidium, and cesium uranyl nitrate have been characterized a long time ago, the crystal structure and properties of the homologous sodium compounds has not been reported yet. Na[UO₂(NO₃)₃] crystallizes in a unique structure [Pearson code cP72, cubic, P2₁3, no. 198, Z = 4, a = 10.6324(14) Å, 938 unique reflections with I_o > 2σ(I_o), R₁ = 0.0379, wR₂ = 0.0947, GooF = 1.092, T = 293 K]. The structure is characterized by an open framework built by interconnected helical chains formed by {UO₈} and {NaO₆} units. The vibrational and optical (UV/Vis absorption and luminescence) spectra match well with those of the higher alkali metal uranyl nitrates.     

Tin in silicate glasses: structure, thermodynamics and kinetics

In this work Mössbauer spectroscopy is used to investigate the oxidation states and structures of tin in silicate glasses. Thermal treatment of the glasses in atmospheres with varying oxygen partial pressure leads to the simultaneous appearance of reduction and diffusion. Experiments with varying treatment time give the opportunity to study diffusion and reduction processes in detail. Comparison of the hyperfine parameters of reference materials with measured parameter provides information about the local surroundings of the tin atoms. An octahedral surrounding for Sn^4?+? is presumed, while Sn^2?+? and three oxygen atoms form a tetrahedral coordination.     

Practice effects in healthy adults: <Emphasis Type="Italic">A longitudinal study on frequent repetitive cognitive testing</Emphasis>

Background  

Cognitive deterioration is a core symptom of many neuropsychiatric disorders and target of increasing significance for novel treatment strategies. Hence, its reliable capture in long-term follow-up studies is prerequisite for recording the natural course of diseases and for estimating potential benefits of therapeutic interventions. Since repeated neuropsychological testing is required for respective longitudinal study designs, occurrence, time pattern and magnitude of practice effects on cognition have to be understood first under healthy good-performance conditions to enable design optimization and result interpretation in disease trials.