Synthesis and application of isotope-labeled carnosine in LCMS/MS

Carnosine is an endogenous dipeptide, composed of \u3b2-alanine and L-histidine, and is highly concentrated in skeletal muscle and other excitable tissues. Its physiological roles, based on its biochemical properties, include pH-buffering, metal-ion chelation and antioxidant capacity as well as the ability to protect against the formation of advanced glycation and lipoxidation end-products.1 For these reasons, besides its nutritional ergogenic application in the sport community,2 carnosine supplementation offers a therapeutic potential for the treatment of numerous diseases in which ischemic or oxidative stress is involved.1 Quantitation of carnosine in biological matrices appears to be crucial for these applications, and LC-MS procedures with isotope-labeled internal standards are the state-of-the-art approach for this analytical need.3 The use of these standards allows to account for variations during the complex sample preparation process, different matrix effects between patient samples, and variations in instrument performance. Figure 1 In this work, we present a fast and highly efficient synthetic route to obtain a deuterated carnosine analogue (Figure 1) starting from the trideuterated L-histidine (\u3b1-d1, imidazole-2,5-d2). Moreover, the use of Carnosine-d3 in the validation of a multiple reaction monitoring (MRM) LC-MS/MS method for the analytical quantitation of carnosine in a biological matrix will be reported. References 1. Boldyrev, A. A.; Aldini, G.; Derave, W. Physiol. Rev. 2013, 93, 1803\u20131845. 2. Brisola, G.; Zagatto, A. J. Strength Cond. Res. 2019, 33, 253-282. 3. Stokvis, E.; Rosing, H.; L\uf3pez-L\ue1zaro, L.; Schellens, J. H. M.; Beijnen, J. H. Biomed. Chromatogr. 2004, 18, 400-402

Erectile dysfunction and angiographic extent of coronary artery disease in type II diabetic patients

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Response of Foraminifera to Anthropogenic Nicotine Pollution of Cigarette Butts: An Experimental Approach

The most often dispersed environmental pollutants that are released both directly and indirectly into the environment that may eventually reach aquatic ecosystems and contaminate aquatic biomes are cigarette butts (CBs). Toxicants such as nicotine, dangerous metals, total particulate matter, and recognized carcinogens can be introduced and transported via CBs into aquatic ecosystems. The examination of the effects of synthetic nicotine on three different species of cultured benthic foraminifera was the focus of this study. Three foraminiferal species from three distinct biomineralization pathways were specifically examined for viability and cellular ultrastructure, including the calcareous perforate Rosalina globularis, the calcareous imperforate Quinqueloculina spp., and the agglutinated Textularia agglutinans. The survival rate, cellular stress, and decalcification were used to assess the toxicological effects of synthetic nicotine. We were able to analyze the reaction of major macromolecules and calcium carbonate to this pollutant using FTIR (Fourier Transform Infrared) spectroscopy. High Performance Liquid Chromatography (HPLC) study was performed to increase our understanding of nicotine bioavailability in the medium culture. Different acute experiments were performed at different dates, and all indicated that synthetic nicotine is acutely hazardous to all three cultured foraminiferal taxa at lethal and sublethal concentrations. Each species responded differently depending on the type of shell biomineralization. Synthetic nicotine enhances shell decalcification and affects the composition of cytoplasmic macromolecules such as lipids and proteins, according to the FTIR spectroscopy investigations. The lipid content rose at lethal concentrations, possibly due to the creation of vesicles. The proteins signal evidences general cellular dyshomeostasis. The integration among the acute toxicity assay, synchrotron, and chemical HPLC analyses provided a valuable approach for the assessment of nicotine as a biomarker of exposure to the toxicants associated with smoking and the impact of this emerging and hazardous material on calcifying marine species

USE OF NANOCOMPOSITE HYDROGEL WITH N-UREA IN THE PRODUCTION OF EGGPLANT SEEDLINGS

The use of quality seedlings of eggplant is directly related to the success of their production, with polymers added to the substrate, which work as water conditioners, increase the water retention capacity, and provide better seedling quality. The study aimed to evaluate the use of nanocomposite hydrogel enriched with different proportions of N-urea in the production of eggplant seedlings. The experiment was conducted at the State University of Mato Grosso do Sul, Cassilândia, MS, Brazil, from June to August 2019, under sombrite® 30%. Five treatments were evaluated, using the commercial substrate, Carolina Soil®: 1) commercial substrate without hydrogel; 2) commercial substrate with 0.075g of pure hydrogel (0.00g of N-urea)/15 mL of a substrate; 3) commercial substrate with 0.075g of hydrogel and 10% N-urea/15 mL of a substrate; 4) commercial substrate with 0.075g of hydrogel and 20% N-urea/15 mL of a substrate, and 5) commercial substrate with 0.075g of hydrogel and 40% N-urea/15 mL of a substrate. The experiment was conducted with four replications of 25 seedlings. The emergence speed index, percentage of emergence, height, number of leaves, stem diameter, shoot dry matter, root dry matter, and total dry matter were evaluated, as well as the Dickson Quality Index. The data were subjected to analysis of variance (SPEEDSTAT statistical software) and grouping test of means. A regression analysis was performed to adjust equations for some of the variables. The best seedlings can be obtained using the dosage of 28.83% N-urea with 0.075g of hydrogel per 15 ml of the substrate, according to the DQI adjustment, which includes several traits of the seedlings, thus reflecting on its quality

Design, synthesis and preliminary biological evaluation of 3-cyclopropyl-4-phenoxy-1H-pyrazole derivatives as small molecular ligands of RAGE

Receptor for advanced glycation end products (RAGE) is a multiligand receptor belonging to the immunoglobulin superfamily and plays a crucial role in the development of many human diseases such as neurodegenerative diseases, diabetes, cardiovascular diseases and cancer.1 RAGE is involved in a number of cell processes such as neuroinflammation, apoptosis, proliferation and autophagy, and therefore it is of considerable interest as a promising drug target for innovative therapeutic approaches. It consists of an extracellular region, a short hydrophobic transmembrane spanning region, and a highly charged amino acid cytoplasmatic tail. The extracellular region contains a signal peptide, followed by one N-terminal V-type immunoglobulin domain and two C-type (C1 and C2) immunoglobulin domains.2 RAGE is able to interact with a large number of pro-inflammatory and regulatory molecules, such as advanced glycation end-products (AGEs), quinolinic acid, beta amyloid (A\u3b2), high mobility group box 1 (HMGB1), S100/calgranulin family proteins.3,4 However, due to the structural heterogeneity of these endogenous ligands, little is known about the key pharmacophore elements for ligand-RAGE interaction and the specific mode of binding. On these grounds, we aimed at designing new small molecules able to bind the VC1 extracellular domains of RAGE, in order to clarify the structural features that account for RAGE affinity and activation, and to identify new drug-like compounds. Following a process of structural simplification of known pyrazole-5-carboxamide RAGE ligands,1 we planned a set of novel derivatives characterized by a variously functionalized 3-cyclopropyl-4-phenoxy-1H-pyrazole scaffold (Figure 1). The design and synthesis of the new putative RAGE ligands will be presented and discussed, together with the results of their in vitro screening by means of a surface plasmon resonance (SPR)-based assay to estimate their binding ability to the RAGE extracellular domain. References 1. Bongarzone S., Savickas V., Luzi F., Gee A. D. J. Med. Chem. 2017, 60, 7213-7232. 2. Hudson B. I., Carter A. M., Harja E., Kalea A. Z., Arriero M., Yang H., Grant P. J., Schmidt A. M. FASEB J. 2008, 22, 1572-1580. 3. Xue J., Rai V., Singer D., Chabierski S., Xie J., Reverdatto S., Burz D. S., Schmidt A. M., Hoffmann R., Shekhtman A. Structure 2011, 19, 722\u2013732. 4. Koch M., Chitayat S., Dattilo B. M., Schiefner A., Diez J., Chazin W. J., Fritz, G. Structure 2010, 18, 1342-1352

A better immune reaction to Erbb-2 tumors is elicited in mice by DNA vaccines encoding rat/human chimeric proteins.

The Erbb-2 (neu in rat and Her-2 in humans) tyrosine kinase receptor is an oncoantigen (i.e., a tumor- associated molecule directly involved in cancer progression). Because oncoantigens are self-tolerated mole- cules, to trigger a response circumventing tolerance, we generated two plasmids (RHuT and HuRT) coding for chimeric neu-Her-2 extracellular and transmembrane proteins that are expressed on the cell membrane of the transfected cells and recognized by monoclonal antibodies reacting against neu and Her-2. RHuT encodes a protein in which the 410 NH2-terminal residues are from the neu extracellular domain and the remaining residues from Her-2. Almost symmetrically, HuRT encodes for a protein in which the 390 NH2-terminal resi- dues are from Her-2 and the remainder from neu. The ability of RHuT and HuRT to elicit a protective response to neu and Her-2 in wild-type mice and in transgenic mice tolerant to neu and Her-2 proteins was compared with that of plasmids coding for the fully rat or fully human extracellular and transmembrane domains of the Erbb-2 receptor. In most cases, RHuT and HuRT elicited a stronger response, although this chimeric benefit is markedly modulated by the location of the heterologous moiety in the protein coded by the plasmid, the immune tolerance of the responding mouse, and the kind of Erbb-2 orthologue on the targeted tumor

Novel pharmacological tools which activate mAChRs: a question of "dualsterism"

Muscarinic acetylcholine receptors (mAChRs) represent an excellent model system to study orthosteric and allosteric interactions. The high sequence homology shown by orthosteric sites of mAChRs has hampered the development of subtype selective agonists. On the other hand, allosteric recognition sites are less conserved among the various mAChR subtypes. We synthesized a series of hybrid ligands designed to simultaneously interact with both orthosteric and allosteric sites (\u201cdualsteric\u201d compounds) by fusing orthosteric activators with M2-selective allosteric fragments (W84 and Naphmethonium). In particular, among the oxotremorine-like orthosteric agents, iperoxo emerged as a potent agonist with supraphysiological efficacy but devoid of subtype selectivity.1 To explore the whole chemical space of the binding region, we modified the structure of the three component parts (orthosteric and allosteric moieties and spacer) of dualsteric ligands.2 These ligands permitted to prove for the first time that GPCR\u2019s allosteric vestibule is able to control the extent of receptor movement to govern a hierarchical order of G-protein coupling.3 In addition, they were found to dynamically switch between two distinct binding orientations, engendering both active and inactive populations of receptors bound by a given ligand.4 More recently, some of these ligands (notably N-8-IPER) revealed interesting antinociceptive properties and good tolerability.5 The synthetic approaches together with relevant results and implications of the biological investigation will be presented. References 1. Schrage R et al. Agonists with supraphysiological efficacy at the muscarinic M2 ACh receptor. Br J Pharmacol 2013;169(2):357-70. 2. Disingrini T et al. Design, synthesis, and action of oxotremorine-related hybrid-type allosteric modulators of muscarinic acetylcholine receptors. J Med Chem 2006;49(1):366-72; Antony J et al. Dualsteric GPCR targeting: a novel route to binding and signaling pathway selectivity. FASEB J 2009;23(2):442-50. 3. Bock A et al. The allosteric vestibule of a seven transmembrane helical receptor controls G-protein coupling. Nat Commun 2012;3:1044. 4. Bock A et al. Dynamic ligand binding dictates partial agonism at a G protein-coupled receptor. Nat Chem Biol 2014;10(1):18-20. 5. Matera C et al. Bis(ammonio)alkane-type agonists of muscarinic acetylcholine receptors: Synthesis, in vitro functional characterization, and in vivo evaluation of their analgesic activity. Eur J Med Chem 2014;75:222-232

ESTIMATION OF REAL PER CAPITA CONSUMPTION OF MEAT IN ITALY

Food consumption refers to the amount of food available for human consumption. The knowledge of food consumption is crucial to set production and food supply policies, to compare eating habits with other countries, to assess the nutritional status of a population and to study the relationship between diet and health. In the last years all these aspects have taken an increasingly important interest because epidemiological studies have indicated a possible association between high consumption of meat and an risk of several forms of cancer as well as metabolic and cardiovascular diseases. Unfortunately meat consumption is often estimated by methods that are inappropriate for this use because they do not represent the actual amount of meat consumed or, better, eaten by the consumers. The actual food consumption may be lower than the quantity shown as food availability depending on the magnitude of wastage and losses of food during the slaughtering, in the household, e.g. during storage, in preparation and cooking, as plate-waste or quantities fed to domestic animals and pets, thrown or given away The consumption estimated by FAO and by statistical offices of the various countries through the national food balance sheets does not indicate the amount of meat, ie the weight of the skeletal muscles of animals with included or adherent tissues, but the amount of the weighted carcass at the slaughterhouse, including bones, tendons, connective tissues and fat. This paper discusses a method of estimating the real per capita consumption of meat in Italy with accuracy comparable to that of individual consumption, developed by the Study Commission of Animal Science and Production Association (ASPA). This action responds to the need of producing statistical indicator related to health food, as recommended by many international organizations (FAO, Eurostat)

Dipole Anisotropy in the COBE DMR First-Year Sky Maps

We present a determination of the cosmic microwave background dipole amplitude and direction from the COBE Differential Microwave Radiometers (DMR) first year of data. Data from the six DMR channels are consistent with a Doppler-shifted Planck function of dipole amplitude Delta T = 3.365 +/-0.027 mK toward direction (l,b) = (264.4 +/- 0.3 deg, 48.4 +/- 0.5 deg). The implied velocity of the Local Group with respect to the CMB rest frame is 627 +/- 22 km/s toward (l,b) = (276 +/- 3 deg, 30 +/- 3 deg). DMR has also mapped the dipole anisotropy resulting from the Earth's orbital motion about the Solar system barycenter, yielding a measurement of the monopole CMB temperature at 31.5, 53, and 90 GHz, to be 2.75 +/- 0.05 K.Comment: Post Script (4 figures) Ap J 419, 1-6 (1993