Methionine 77 in calmodulin can be stereospecifically oxidized to methionine sulfoxide by mammalian methionine sulfoxide reductase A. Whether this has in vivo significance is unknown. We therefore created a mutant mouse in which wild-type calmodulin-1 was replaced by a calmodulin containing a mimic of methionine sulfoxide at residue 77. Total calmodulin levels were unchanged in the homozygous M77Q mutant, which is viable and fertile. No differences were observed on learning tests, including the Morris water maze and associative learning. Cardiac stress test results were also the same for mutant and wild type mice. .However, young male and female mice were 20% smaller than wild type mice, although food intake was normal for their weight. Young M77Q mice were notably more active and exploratory than wild type mice. This behavior difference was objectively documented on the treadmill and open field tests. The mutant mice ran 20% longer on the treadmill than controls, and in the open field test, the mutant mice explored more than controls and exhibited reduced anxiety These phenotypic differences bore a similarity to those observed in mice lacking calcium/calmodulin kinase Iiα (CaMKIIα). We then showed that M77Q calmodulin was less effective in activating CaMKIIα than wild type calmodulin. Thus, characterization of the phenotype of a mouse expressing a constitutively active mimic of calmodulin led to the identification of the first calmodulin target that can be differentially regulated by the oxidation state of Met77. We conclude that reversible oxidation of methionine 77 in calmodulin by MSRA can regulate cellular function.

A rapid synthesis of chiral sulfoxide-functionalized meta-terphenyl derivatives by a 2,5-[C4+C2] ring transformation reaction of pyrylium salts with in-situ generated enantio-merically pure .alpha.-sulfinylacetaldehydes is described. This synthetic method demonstrates for the first time the use of .alpha.-sulfinylacetaldehydes in a reaction sequence initiated by nu-cleophilic attack of pyrylium salts by .alpha.-sulfinylcarbanions to generate chiral aromatic systems. The method presented shows broad applicability starting with various methyl sulfoxides and a number of functionalized pyrylium salts, furnishing meta-terphenyls with complex substitution patterns from readily accessible starting compounds.

To determine promethazine (PMZ) and its metabolites promethazine sulfoxide (PMZSO) and monodesmethyl-promethazine (Nor1PMZ) in swine muscle, fat, kidney, and liver, a sample preparation and high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis method were established and validated. The sample was extracted with 0.1% formic acid-acetonitrile, and the extract was purified with acetonitrile-saturated n-hexane. After concentration by rotary evaporation, the extract was re-dissolved in 0.1% formic acid-water and acetonitrile (80:20, v/v). The HPLC-MS/MS column used was a Waters Symmetry C18 (100 mm × 2.1 mm i.d., 3.5 μm), with 0.1% formic acid-water and acetonitrile as the mobile phase, and the target compounds were determined by positive ion scan and multiple reaction monitoring. PMZ and Nor1PMZ were quantified with deuterated promethazine (PMZ-d6) as the internal standard, while PMZSO was quantified by external standard method. The limits of detection (LOD) and limits of quantification (LOQ) of PMZ and PMZSO in muscle, liver, and kidney spiked samples were 0.05 μg/kg and 0.1 μg/kg, respectively, while those of Nor1PMZ were 0.1 μg/kg and 0.5 μg/kg, respectively. The LOD and LOQ of analytes in fat spiked sample were 0.05 μg/kg and 0.1 μg/kg, respectively. The sensitivity of this method reaches or exceeds its presented in previous reports. The analytes PMZ and PMZSO showed good linearity within the range of 0.1 μg/kg to 50 μg/kg, while Nor1PMZ showed good linearity within the range of 0.5 μg/kg to 50 μg/kg, with correlation coefficients (r) greater than 0.99. The recoveries of the target compounds in the sam-ples were between 81.73% and 107.17%, and the precision ranged from 1.78% to 10.47%. This study developed for the first time an HPLC-MS/MS method for the determination of PMZ and its metabolites PMZSO and Nor1PMZ in four swine edible tissues, comprehensively covered target tissues of monitoring object, which is applicable for monitoring veterinary drug residues in ani-mal-derived foods and ensuring food safety.

Methane (CH4), produced endogenously in animals and plants, has recently been suggested to play a role in cellular physiology, potentially influencing the signaling pathways and regulatory mechanisms involved in nitrosative and oxidative stress responses. In addition, it has been proposed that supplementation of CH4 to organisms may be beneficial for the treatment of several diseases, including ischemia, reperfusion injury, and inflammation. However, it is still unclear if and how CH4 is produced in mammalian cells without the help of microorganisms, and how CH4 might be involved in physiological processes in humans. In this study, we provide first proof of the principle that CH4 is formed non-microbially in the human body by applying isotopically labeled methylated sulfur compounds, such as dimethyl sulfoxide (DMSO) and methionine, as carbon precursors to confirm cellular CH4 formation. A volunteer applied isotopically labeled (2H and 13C) DMSO on the skin, orally, and to blood samples. Monitoring of stable isotope values of CH4 convincingly showed the conversion of the methyl groups, as isotopically labeled CH4 was formed during all experiments. Based on these results, we consider several hypotheses about endogenously formed CH4 in humans, including physiological aspects and stress responses involving reactive oxygen species (ROS). While further and broader validation studies are needed, the results may unambiguously serve as a proof of concept for the endoge-nous formation of CH4 in humans by a radical-driven process. Furthermore, these results might encourage follow up studies to decipher the potential physiological role of CH4 and its bioactiv-ity in humans in more detail. Of particular importance is the potential to monitor CH4 as an ox-idative stress biomarker if the observed large variability of CH4 in breath air is an indicator for physiological stress responses and immune reactions. Finally, the potential role of DMSO as a radical scavenger to counteract oxidative stress caused by ROS might be considered in the health sciences. DMSO has already been investigated for many years, but its potential positive role for medical use remained highly uncertain.