Micelle‐enhanced spectrofluorimetric method for determination of sitagliptin and identification of potential alkaline degradation products using LC‐MS

A novel, quick, simple and highly sensitive spectrofluorimetric method was developed and validated for the determination of sitagliptin (SG) in its pharmaceutical formulations. The proposed method is based on investigation of the fluorescence spectral behavior of sitagliptin in an SDS micellar system. In an aqueous solution of phosphate buffer pH 4.0, the fluorescence intensity of SG in the presence of SDS was greatly enhanced, by 200%, i.e. twofold enhancement. The fluorescence intensity of SG was measured at 300 nm after excitation at 270 nm. The method showed good linearity in the range 0.03–10.0 µg/mL with a good correlation coefficient (r = 0.9998). The limits of detection and quantitation values were 5.31 and 16.1 ng/mL, respectively. The proposed method was successfully applied to the analysis of SG in its single and co‐formulated commercial tablets; the results were in good agreement with those obtained using a reference method. Application of the proposed method was extended to stability studies of SG after exposure to different forced degradation conditions according to the ICH guidelines, such as acidic, alkaline, thermal, photo‐ and oxidative stress. The chemical structure of certain potential degradation products (DPs) were investigated using LC‐MS. Copyright © 2013 John Wiley & Sons, Ltd.     

Effects of arsenic on phosphorus content in different organs and chlorophyll fluorescence in primary leaves of soybean

The effect of arsenic (32 – 96 μM) on the phosphorus content and Chl fluorescence was studied in soybean (Glycine max Merril) grown in the nutrient solution with and without phosphorus. The increased concentration of As led to the decrease in P content in plant organs. Parameters of Chl fluorescence of soybean leaves in the presence of these As concentrations did not show significant changes.     

Neural networks in intestinal immunoregulation

Key physiological functions of the intestine are governed by nerves and neurotransmitters. This complex control relies on two neuronal systems: an extrinsic innervation supplied by the two branches of the autonomic nervous system and an intrinsic innervation provided by the enteric nervous system. As a result of constant exposure to commensal and pathogenic microflora, the intestine developed a tightly regulated immune system. In this review, we cover the current knowledge on the interactions between the gut innervation and the intestinal immune system. The relations between extrinsic and intrinsic neuronal inputs are highlighted with regards to the intestinal immune response. Moreover, we discuss the latest findings on mechanisms underlying inflammatory neural reflexes and examine their relevance in the context of the intestinal inflammation. Finally, we discuss some of the recent data on the identification of the gut microbiota as an emerging player influencing the brain function.     

Uptake of 5 9Fe from soluble 5 9Fe-humate complexes by cucumber and barley plants

The capability of cucumber (Cucumis sativus L., cv. Serpente cinese), a Strategy I plant and barley (Hordeum vulgaris L., cv. Europa), a Strategy II plant to use Fe complexed by a water-soluble humic fraction (WEHS) extracted from a peat, was studied. Uptake of ⁵⁹Fe from ⁵⁹Fe-WEHS by cucumber plants was higher at pH 6.0 than at pH 7.5. Roots of intact cucumber plants were able to reduce the Fe^III-WEHS complex either at pH 6.0 or 7.5, rates being higher in the assay medium buffered at pH 6.0. After supply of ⁵⁹Fe-WEHS, a large pool of root extraplasmatic ⁵⁹Fe was formed, which could be used to a large extent by Fe-deficient plants, particularly under acidic conditions. Uptake of ⁵⁹Fe from ⁵⁹Fe-WEHS by Fe-sufficient and Fe-deficient barley plants was examined during periods of high (morning) and low (evening) PS release. Uptake paralleled the diurnal rhythm of PS release. Furthermore, ⁵⁹Fe uptake was strongly enhanced by addition of PS to the uptake solution in both Fe-sufficient and Fe-deficient plants. High amount of root extraplasmatic ⁵⁹Fe was formed upon supply of Fe-WEHS, particularly in the evening experiment. Fe-deficient barley plants were able to utilize Fe from the root extraplasmatic pool, conceivably as a result of high rates of PS release. The results of the present work together with previous observations indicate that cucumber plants (Strategy I) utilize Fe complexed to WEHS, presumably via reduction of Fe^III-WEHS by the plasma membrane-bound reductase, while barley plants (Strategy II) use an indirect mechanism involving ligand exchange between WEHS and PS.     

Analysis of common CFTR polymorphisms 5T, M470V and R75Q in healthy Serbian population

Three common CFTR polymorphisms, 5T, M470V and R75Q, have been shown to be relatively frequent in Serbian patients with monosymptomatic CF disorders. Since there is a variation in distribution of common polymorphisms among different populations, it was important to compare their frequencies in patients with the frequencies in healthy population in order to assess the possible role of these polymorphisms in the monosymptomatic CF disorders. Samples obtained from 100 healthy Serbian individuals were analyzed for the presence of CFTR 5T, M470V and R75Q variants by PSM, RFLP and DGGE methods, respectively. Allele 5T was present in two individuals, giving the allelic frequency of 1% (2/200 alleles). The frequency obtained for allele M470 was 45% (90/200 alleles), while V470 allele was present with the frequency of 55% (110/200 alleles). Polymorphism R75Q was present in two individuals, with allelic frequency of 1% (2/200 alleles). Our study has shown that the frequencies of two common polymorphisms, 5T and M470V, differ significantly in Serbian population in comparison with other South European populations. Since it appears that Serbian population has a specific distribution of studied CFTR gene variants, it would also be interesting to analyze other common variants of this gene in our population. Such data can also be potentially useful as anthropogenetic markers in population studies.     

A covalently bound photoisomerizable agonist. Comparison with reversibly bound agonists at electrophorus electroplaques

After disulphide bonds are reduced with dithiothreitol, trans-3- (α-bromomethyl)-3’-[α- (trimethylammonium)methyl]azobenzene (trans-QBr) alkylates a sulfhydryl group on receptors. The membrane conductance induced by this “tethered agonist” shares many properties with that induced by reversible agonists. Equilibrium conductance increases as the membrane potential is made more negative; the voltage sensitivity resembles that seen with 50 [mu]M carbachol. Voltage- jump relaxations follow an exponential time-course; the rate constants are about twice as large as those seen with 50 μM carbachol and have the same voltage and temperature sensitivity. With reversible agonists, the rate of channel opening increases with the frequency of agonist-receptor collisions: with tethered trans-Qbr, this rate depends only on intramolecular events. In comparison to the conductance induced by reversible agonists, the QBr-induced conductance is at least 10-fold less sensitive to competitive blockade by tubocurarine and roughly as sensitive to “open-channel blockade” bu QX-222. Light-flash experiments with tethered QBr resemble those with the reversible photoisomerizable agonist, 3,3’,bis-[α-(trimethylammonium)methyl]azobenzene (Bis-Q): the conductance is increased by cis {arrow} trans photoisomerizations and decreased by trans {arrow} cis photoisomerizations. As with Bis-Q, ligh-flash relaxations have the same rate constant as voltage-jump relaxations. Receptors with tethered trans isomer. By comparing the agonist-induced conductance with the cis/tans ratio, we conclude that each channel’s activation is determined by the configuration of a single tethered QBr molecule. The QBr-induced conductance shows slow decreases (time constant, several hundred milliseconds), which can be partially reversed by flashes. The similarities suggest that the same rate-limiting step governs the opening and closing of channels for both reversible and tethered agonists. Therefore, this step is probably not the initial encounter between agonist and receptor molecules.     

Parasitofauna of freshwater fishes in the Serbian open waters: a checklist of parasites of freshwater fishes in Serbian open waters

As a result of freshwater fish parasitofauna investigations, throughout the past 75 years, the number of parasitic species is presented. This paper reviews the history, current state, and tendencies of the fish parasitofauna studies in Serbian open waters. Up to now, in total 170 parasitic species have been reported. Systematic parasitofauna investigations have been carried out on 54 freshwater fish species in waters of the Danube River Basin in Serbia, also in hill-mountanious watercourses, lakes and salmonid fishpond of Sjenicko-Pesterska plateau (south-west region of Serbia), Homolje area (east Serbia), and in Vlasina lake (south-east Serbia). This data review pointed that it is necessary to continue the investigation on fish parasites in Serbia. The data on freshwater fish parasites are important for evaluation of general influence on the community structure.     

Protocol of a prospective study on the diagnostic value of transcranial duplex scanning of the substantia nigra in patients with parkinsonian symptoms

Background  

Parkinson's disease (PD) is the second most common neurodegenerative disorder. As there is no definitive diagnostic test, its diagnosis is based on clinical criteria. Recently transcranial duplex scanning (TCD) of the substantia nigra in the brainstem has been proposed as an instrument to diagnose PD. We and others have found that TCD scanning of substantia nigra duplex is a relatively accurate diagnostic instrument in patients with parkinsonian symptoms. However, all studies on TCD so far have involved well-defined, later-stage PD patients, which will obviously lead to an overestimate of the diagnostic accuracy of TCD.     

Mapping of the KHSRP gene to a region of conserved synteny on human chromosome 19p13.3 and mouse chromosome 17

The K homology-type splicing regulatory protein, KSRP, activates splicing through intronic splicing enhancer sequences. It is highly expressed in neural cells and is required for the neural-specific splicing of the c-src N1 exon. In this study, we mapped the gene (gene symbols KHSRP and Khsrp) to human chromosome 19 by using radiation hybrid panels and to mouse chromosome 17 by studying an interspecific backcross panel. Human KHSRP is a positional candidate gene for familial febrile convulsion and Cayman type cerebellar ataxia. Comparative analysis of the human and mouse genomes indicates that the KHSRP gene is located in regions of conserved synteny between the two species.     

Cooperative assembly of an hnRNP complex induced by a tissue-specific homolog of polypyrimidine tract binding protein

Splicing of the c-src N1 exon in neuronal cells depends in part on an intronic cluster of RNA regulatory elements called the downstream control sequence (DCS). Using site-specific cross-linking, RNA gel shift, and DCS RNA affinity chromatography assays, we characterized the binding of several proteins to specific sites along the DCS RNA. Heterogeneous nuclear ribonucleoprotein (hnRNP) H, polypyrimidine tract binding protein (PTB), and KH-type splicing-regulatory protein (KSRP) each bind to distinct elements within this sequence. We also identified a new 60-kDa tissue-specific protein that binds to the CUCUCU splicing repressor element of the DCS RNA. This protein was purified, partially sequenced, and cloned. The new protein (neurally enriched homolog of PTB [nPTB]) is highly homologous to PTB. Unlike PTB, nPTB is enriched in the brain and in some neural cell lines. Although similar in sequence, nPTB and PTB show significant differences in their properties. nPTB binds more stably to the DCS RNA than PTB does but is a weaker repressor of splicing in vitro. nPTB also greatly enhances the binding of two other proteins, hnRNP H and KSRP, to the DCS RNA. These experiments identify specific cooperative interactions between the proteins that assemble onto an intricate splicing-regulatory sequence and show how this hnRNP assembly is altered in different cell types by incorporating different but highly related proteins.