One-step preparation of enantiopure L- or D-amino acid benzyl esters avoiding the use of banned solvents

The enantiomers of amino acid benzyl esters are very important synthetic intermediates. Many of them are currently prepared by treatment with benzyl alcohol and p-toluenesulfonic acid in refluxing benzene or carbon tetrachloride, to azeotropically remove water, and then precipitated as tosylate salt by adding diethyl ether. Here, we report a very efficient preparation of eight l- or d-amino acid benzyl esters (Ala, Phe, Tyr, Phg, Val, Leu, Lys, Ser), in which these highly hazardous solvents are dismissed using cyclohexane as a water azeotroping solvent and ethyl acetate to precipitate the tosylate salt. With some work-up modifications and lower yield, the procedure can be applied also to methionine. Chiral HPLC analysis shows that all the benzyl esters, including the highly racemizable ones such as those of phenylglycine, tyrosine and methionine, are formed enantiomerically pure under these new reaction conditions thus validating the solvents replacement. Contrariwise, toluene cannot be used in place of benzene or carbon tetrachloride because leading to partially or totally racemized amino acid benzyl esters depending on the polar effect of the amino acid \u3b1-side chain as expressed by Taft\u2019s substituent constant (\u3c3*)

Comparative investigations on soluble proteins and malate dehydrogenase of four grape varieties by isoelectric focusinq

Soluble proteins and multiple forms of malate dehydrogenase (E.C. 1.1.1.37) in both skin and juice of four grape varieties, have been compared with respect to heterogeneity and isoelectric point. The protein preparations resolved into 17 to 28 very distinct components. Seven (skins) and eight (juices) of these components were of common occurrence with all varieties, as judged by the same pI value. The presence of two to seven peculiar bands in both parts of berry was also observed. In the grape the NAD-malate dehydrogenase activity was associated with 9 to 13 isoelectric forms, apparently assembled into three sets of isoenzymes. Each of these showed a rather similar dislocation in the gels; but they contained a different number of isoenzymes of distinct pI value in the skin as well as in the juice of different varieties. A higher number of variant forms was found in the skins in comparison with juices. Between two and four isomalate dehydrogenases of the same net charge were found in both parts of berries of the same variety, whereas the remaining isoenzymes appeared to be non-identical and, thus, presumably the product of distinct genes.Vergleichende Untersuchung über die löslichen Proteine und die Malat-Dehydrogenasevon vier Rebsorten mittels isoelektrischer FokussierungDie löslichen Proteine und multiplen Formen der Malat-Dehydrogenase (E.C. 1.1.1.37) sowohl in der Schale als auch im Saft der Beeren von vier Rebsorten wurden in bezug auf Heterogenität und isoelektrischen Punkt verglichen. Die Proteinpräparate ließen sich in 17 bis 28 deutlich unterscheidbare Komponenten auftrennen. Aufgrund übereinstimmender pI-Werte besaßen alle Sorten in der Schale sieben und im Saft acht gleiche Komponenten. Auch wurden in den beiden Teilen der Beere zwei bis sieben charakteristische Proteinbanden beobachtet.Das Muster der NAD-Malat-Dehydrogenase in der Beere setzt sich aus 9 bis 13 isoelektrischen Formen zusammen, die offensichtlich in drei Gruppen von Isoenzymen angeordnet sind. Die einzelnen Gruppen waren in den Gelen ziemlich übereinstimmend lokalisiert; aber die einzelnen Sorten besaßen sowohl in der Schale als auch im Saft eine unterschiedliche Anzahl von Enzymen eines bestimmten pI-Wertes. In den Schalen wurde eine vergleichsweise größere Anzahl unterschiedlicher Isoenzyme gefunden als in den Säften. In den beiden Beerenteilen einer Sorte wurden zwei bis vier Isomalat-Dehydrogenasen mit übereinstimmendem pI-Wert ermittelt, während die restlichen Isoenzyme anscheinend nicht identisch und somit wohl auf ungleiche Gene zurückzuführen waren

Microbial biodiversity of the liquid fraction of rumen content from lactating cows

Host and dietary interactions with the rumen microbiome can affect the efficacy of supplements, and their effect on the composition of the bacterial population is still unknown. A 16S rRNA metagenomic approach and Next-Generation Sequencing (NGS) technology were used to investigate the bacterial microbiome composition in the liquid fraction of the rumen content collected via stomach tubing. To investigate biodiversity, samples were taken from three groups of four lactating dairy cows given a supplement of either 50 g of potato protein (Ctrl group), or 50 g of lyophilized Saccharomyces cerevisiae (LY group) or 50 g of dried S. cerevisiae (DY group) in a potato protein support. Rumen samples were collected after 15 days of dietary treatments and milk production was similar between the three groups. Taxonomic distribution analysis revealed a prevalence of the Firmicutes phylum in all cows (79.76%) and a significantly ( P<0.05) higher presence of the genus Bacillus in the DY group. Volatile fattyacid concentration was not significantly different between groups, possibly because of relatively high inter-animal variability or limited effect of the treatments or both, and the correlation analysis with bacterial taxa showed significant associations, in particular between many Firmicutes genera and butyrate. Limited differences were observed between dietary treatments, but the lack of microbiome data before yeast administration does not allow to draw firm conclusions on the effect of dietary treatments

GPS-based CERN-LNGS time link for Borexino

We describe the design, the equipment, and the calibration of a new GPS based time link between CERN and the Borexino experiment at the Gran Sasso Laboratory in Italy. This system has been installed and operated in Borexino since March 2012, and used for a precise measurement of CNGS muon neutrinos speed in May 2012. The result of the measurement will be reported in a different letter.Comment: 13 pages, 11 figure

Nonlinear valuation with XVAs: two converging approaches

When pricing OTC contracts in the presence of additional risk factors and costs, such as credit risk and funding and collateral costs, the starting “clean price” is modified additively by valuation adjustments (XVAs) that account for each factor or cost in isolation, while seemingly ignoring the combined effects. Instead, risk factors and costs can be jointly accounted for ab initio in the pricing mechanism at the level of cash flows, and this “adjusted cash flow" approach leads to a nonlinear valuation formula. While for practitioners this made more sense because it showed which discount factor is used for which cash flow (recall the multi-curve environment post-crisis), for academics, the focus was on checking that the resulting nonlinear valuation formula is consistent with the theoretical arbitrage-free “replication approach” that we also analyse in the paper. We formulate specific reasonable assumptions, which ensure that the valuation formulae obtained by the two approaches coincide, thus reinforcing both academics’ and practitioners’ confidence in adopting such nonlinear valuation formulae in a multi-curve setup

The Discovery of a Companion to the Very Cool Dwarf Gl~569~B with the Keck Adaptive Optics Facility

We report observations obtained with the Keck adaptive optics facility of the nearby (d=9.8 pc) binary Gl~569. The system was known to be composed of a cool primary (dM2) and a very cool secondary (dM8.5) with a separation of 5" (49 Astronomical Units). We have found that Gl~569~B is itself double with a separation of only 0".101$\pm$0".002 (1 Astronomical Unit). This detection demonstrates the superb spatial resolution that can be achieved with adaptive optics at Keck. The difference in brightness between Gl~569~B and the companion is $\sim$0.5 magnitudes in the J, H and K' bands. Thus, both objects have similarly red colors and very likely constitute a very low-mass binary system. For reasonable assumptions about the age (0.12~Gyr--1.0~Gyr) and total mass of the system (0.09~M$_\odot$--0.15~M$_\odot$), we estimate that the orbital period is $\sim$3 years. Follow-up observations will allow us to obtain an astrometric orbit solution and will yield direct dynamical masses that can constrain evolutionary models of very low-mass stars and brown dwarfs

MgC1q, a novel C1q-domain-containing protein involved in the immune response of Mytilus galloprovincialis

9 páginas, 6 figuras, 1 tablaIn this study, we present the characterization of a newly identified gene, MgC1q, revealed in suppression subtractive hybridization and cDNA libraries from immunostimulated mussels. Based on sequence homology, molecular architecture and domain similarity, this new C1q-domain-containing gene may be classified as a member of the C1q family and, therefore, part of the C1q–TNF superfamily. The expression of MgC1q was detected all along the mussel ontogeny, being detectable within 2 h post-fertilization, with a notable increase after 1 month and continuing to increase until 3 months. Measurable transcript levels were also evident in all analyzed tissues of naïve adult mussels, and the hemocytes showed the highest expression levels. Experimental infection of adult mussels with Gram positive or Gram negative bacteria significantly modulated the MgC1q expression, and confirmed it as an immune-related gene. Intra- and inter-individual sequence analyses revealed extraordinary diversity of MgC1q at both the DNA and cDNA levels. While further research is needed to define its function, our data indicate that MgC1q is a pattern recognition molecule able to recognize pathogens during innate immune responses in Myitilus galloprovincialis. The high sequence variability suggests that somatic diversification of these nonself recognition molecules could have occurred.This work has been funded by the EU Integrated Project FOOD-CT-2005-007103 and AGL2008-05111/ACU from the Spanish Ministerio de Ciencia e Innovación. Camino Gestal wishes to acknowledge additional funding from the Spanish Ministerio de Educación y Ciencia through the “Ramón y Cajal” Contract.Peer reviewe

Effect of a proteolitic enzyme preparation (alcalase) on whey proteins

When acid whey samples were adjusted to pH 7.5 and incubated with a commercial proteolytic enzyme preparation (alcalase), the turbidity of the solution gradually increased until a soft coagulum, composed of small-sized aggregates, formed after about 1 h. Under the same conditions, sweet whey produced no coagulated material unless CaCl2 was added. Aggregate formation was enhanced by CaCl2 addition in both wheys, but especially in acid whey. However, maximum aggregate formation in terms of protein recovery was about 28%. The precipitated material obtained with various amounts of Ca++ in both sweet and acid wheys resolved into two fractions when it was subjected to nondissociating gel electrophoresis. Sodium dodecyl sulfate gel electrophoresis and HPLC studies showed that the polymerized product consisted of protein fragments having an approximate molecular weight of 7000 to 10,000. A possible mechanism for enzyme-induced aggregation of these peptides is presented

Identification of five picorna-like viruses associated with the endangered cave-dwelling bivalve Congeria kusceri (Bole, 1962)

Congeria kusceri is a bivalve mollusk species endemic to the Dinaric Karst, which displays unique adaptations that have allowed its survival in the subterranean environment with small morphological changes compared with its fossil relatives. Anthropic activities have recently impacted the surface flow of the Neretva river, impairing the seasonal water cycle that has characterized the habitat of this species for hundreds of thousands of years. The lack of an adequate water supply, together with pollution from agricultural and farm water runoff, are posing a serious threat to C. kusceri, as evidenced by the sharp population decline observed in several locations during the past few decades. Due to the limited knowledge available about the basic biology of this filter-feeding species, the precise factors that may affect its health status and reproduction and therefore represent a hazard for its conservation are still unclear. Here, through a transcriptomic approach, we describe the nearly-complete genomes of five C. kusceri-associated RNA viruses belonging to the Picornaviridae family and phylogenetically related with picorna-like viruses previously described in other Mollusca. Although it is presently unknown whether these viruses may have a detrimental effect on bivalve health, we observed a significant increase of viral load during the summer seaso

PLANiTS: a curated sequence reference dataset for plant ITS DNA metabarcoding

DNA metabarcoding combines DNA barcoding with high-throughput sequencing to identify different taxa within environmental communities. The ITS has already been proposed and widely used as universal barcode marker for plants, but a comprehensive, updated and accurate reference dataset of plant ITS sequences has not been available so far. Here, we constructed reference datasets of Viridiplantae ITS1, ITS2 and entire ITS sequences including both Chlorophyta and Streptophyta. The sequences were retrieved from NCBI, and the ITS region was extracted. The sequences underwent identity check to remove misidentified records and were clustered at 99% identity to reduce redundancy and computational effort. For this step, we developed a script called 'better clustering for QIIME' (bc4q) to ensure that the representative sequences are chosen according to the composition of the cluster at a different taxonomic level. The three datasets obtained with the bc4q script are PLANiTS1 (100\u2009224 sequences), PLANiTS2 (96\u2009771 sequences) and PLANiTS (97\u2009550 sequences), and all are pre-formatted for QIIME, being this the most used bioinformatic pipeline for metabarcoding analysis. Being curated and updated reference databases, PLANiTS1, PLANiTS2 and PLANiTS are proposed as a reliable, pivotal first step for a general standardization of plant DNA metabarcoding studies. The bc4q script is presented as a new tool useful in each research dealing with sequences clustering. Database URL: https://github.com/apallavicini/bc4q; https://github.com/apallavicini/PLANiTS