Using the POD sampler for quantitative diffusive (passive) monitoring of volatile and very volatile organics in ambient air: Sampling rates and analytical performance.

POD diffusive samplers loaded with Carbopack X and Carbograph 5TD were exposed to certified calibration mixtures containing a total of 110 different ozone precursor and air toxic compounds. Constant sampling rates were identified for 39 ozone precursors and 33 air toxics. As 9 of these compounds were included in both mixtures, this meant a total of 63 different volatile and very volatile compounds were sampled using the POD with overall expanded uncertainties below 30 % for the sampling rate associated with the whole range of sampling times from 2 to 24 h. Carbograph 5TD exhibited superior performance for diffusive sampling of oxygenated and halogenated compounds in the air toxics mixture, while Carbopack X showed higher sampling efficiencies for aliphatic and aromatic hydrocarbons, as well as halogenated compounds derived from benzene and C2 carbon number hydrocarbons. A model has been developed and applied to estimate sampling rates, primarily for the more volatile and weakly adsorbed compounds, as a function of the collected amount of analyte and the exposure time. For an additional 9 ozone precursors on Carbopack X, and 11 air toxics on Carbograph 5TD, the expanded uncertainties of modelled sampling rates were reduced to below 30 % and have a significantly reduced uncertainty compared to those associated with an averaged sampling rate. The paper provides Freundlich's isotherm parameters for the estimated (modelled) sampling rates and defines a pragmatic approach to their application. It does so by identifying the best sampling time to use for the expected exposure concentrations and associated analyte masses. This allows for expansion of the sampling concentration range from hundreds ng m-3 to mg m-3, while avoiding saturation of the adsorbent. Finally, field measurement comparisons of POD samplers, pumped tube samplers and online gas chromatography (GC), for sampling periods of 3 and 7 days in a semi-rural background area, showed no significant differences between reported concentrations.

Modelling and optimization of factors influencing adsorptive performance of agrowaste-derived Nanocellulose Iron Oxide Nanobiocomposites during remediation of Arsenic contaminated groundwater.

Nanocellulose Iron Oxide Nanobiocomposites (NIONs) were synthesized from rice husk and sugarcane bagasse derived nanocelluloses for adsorptive removal of arsenic and associated contaminants present in groundwater samples. These NIONSs were superparamagnetic, hence magnetically recoverable and demonstrated promising recyclability. Synthesis of NIONs was confirmed by Transmission electron microscopy (TEM), X-Ray Diffraction (XRD), Fourier transform infrared spectroscopy (FTIR) and X-ray photoelectron spectroscopic (XPS). FTIR and XPS data together with adsorption kinetics provide insights into probable adsorption mechanism of Arsenic by NIONs. The experimental conditions for 10 different variants were modelled using response surface methodology (RSM) based on central composite design (CCD), considering the parameters; adsorbate dosage, adsorbent dosage, pH and contact time. The results identified the best performing variants and the optimal conditions for maximal absorption (~99%). These results were validated using a three-layer feed-forward Multilayer Perceptron (MLP) based Artificial Neural Network (ANN) model. Both RSM and ANN chemometric models were in close conformity for optimized conditions of highest adsorption by specific variants. The standardized conditions were used to expand the study to field-based arsenic contaminated groundwater samples and their performance to commercial adsorbents. NIONs show promising commercial potential for water remediation applications due to their high adsorptive performance, magnetic recoverability and recyclability.

Gene Discovery for Synthetic Biology: Exploring the Novel Natural Product Biosynthetic Capacity of Eukaryotic Microalgae.

Eukaryotic microalgae are an incredibly diverse group of organisms whose sole unifying feature is their ability to photosynthesize. They are known for producing a range of potent toxins, which can build up during harmful algal blooms causing damage to ecosystems and fisheries. Genome sequencing is lagging behind in these organisms because of their genetic complexity, but transcriptome sequencing is beginning to make up for this deficit. As more sequence data becomes available, it is apparent that eukaryotic microalgae possess a range of complex natural product biosynthesis capabilities. Some of the genes concerned are responsible for the biosynthesis of known toxins, but there are many more for which we do not know the products. Bioinformatic and analytical techniques have been developed for natural product discovery in bacteria and these approaches can be used to extract information about the products synthesized by algae. Recent analyses suggest that eukaryotic microalgae produce many complex natural products that remain to be discovered.

Air quality concerns of unconventional oil and natural gas production.

Increased use of hydraulic fracturing ("fracking") in unconventional oil and natural gas (O & NG) development from coal, sandstone, and shale deposits in the United States (US) has created environmental concerns over water and air quality impacts. In this perspective we focus on how the production of unconventional O & NG affects air quality. We pay particular attention to shale gas as this type of development has transformed natural gas production in the US and is set to become important in the rest of the world. A variety of potential emission sources can be spread over tens of thousands of acres of a production area and this complicates assessment of local and regional air quality impacts. We outline upstream activities including drilling, completion and production. After contrasting the context for development activities in the US and Europe we explore the use of inventories for determining air emissions. Location and scale of analysis is important, as O & NG production emissions in some US basins account for nearly 100% of the pollution burden, whereas in other basins these activities make up less than 10% of total air emissions. While emission inventories are beneficial to quantifying air emissions from a particular source category, they do have limitations when determining air quality impacts from a large area. Air monitoring is essential, not only to validate inventories, but also to measure impacts. We describe the use of measurements, including ground-based mobile monitoring, network stations, airborne, and satellite platforms for measuring air quality impacts. We identify nitrogen oxides, volatile organic compounds (VOC), ozone, hazardous air pollutants (HAP), and methane as pollutants of concern related to O & NG activities. These pollutants can contribute to air quality concerns and they may be regulated in ambient air, due to human health or climate forcing concerns. Close to well pads, emissions are concentrated and exposure to a wide range of pollutants is possible. Public health protection is improved when emissions are controlled and facilities are located away from where people live. Based on lessons learned in the US we outline an approach for future unconventional O & NG development that includes regulation, assessment and monitoring.

Discrimination of epimeric glycans and glycopeptides using IM-MS and its potential for carbohydrate sequencing.

Mass spectrometry is the primary analytical technique used to characterize the complex oligosaccharides that decorate cell surfaces. Monosaccharide building blocks are often simple epimers, which when combined produce diastereomeric glycoconjugates indistinguishable by mass spectrometry. Structure elucidation frequently relies on assumptions that biosynthetic pathways are highly conserved. Here, we show that biosynthetic enzymes can display unexpected promiscuity, with human glycosyltransferase pp-α-GanT2 able to utilize both uridine diphosphate N-acetylglucosamine and uridine diphosphate N-acetylgalactosamine, leading to the synthesis of epimeric glycopeptides in vitro. Ion-mobility mass spectrometry (IM-MS) was used to separate these structures and, significantly, enabled characterization of the attached glycan based on the drift times of the monosaccharide product ions generated following collision-induced dissociation. Finally, ion-mobility mass spectrometry following fragmentation was used to determine the nature of both the reducing and non-reducing glycans of a series of epimeric disaccharides and the branched pentasaccharide Man3 glycan, demonstrating that this technique may prove useful for the sequencing of complex oligosaccharides.

[A simple method of synthesis of triterpene glycosides similar to glycyrrhizic acid and their hepatoprotective activity in vitro].

A simplified method of synthesis of triterpene l,2-trans-glycosides was developed using the glycosylation of glycyrrhetic acid (GLA) and 18,19-dehydro-GLA by beta-pyranose peracetates in the presence of SnCl(4) and molecular sieves 4 A. The synthesized glycosides exhibited hepatoprotective activity toward the human hepatoma HepG2 cell line on the model of alcohol hepatitis and decreased the level of TNF-alpha protein.

Overexpression, purification, crystallization and data collection on the Bordetella pertussis wlbD gene product, a putative UDP-GlcNAc 2'-epimerase.

The Boredetella pertussis wlbD gene product is a putative uridine-5-diphosphate N-acetylglucosamine (UDP-GlcNAc) 2'-epimerase involved in Band A lipopolysaccharide biosynthesis. The wlbD gene is homologous to Escherichia coli rffE (32% identical), an established UDP-GlcNAc 2'-epimerase that is involved in enterobacterial common antigen (ECA) formation. The structure of the rffE protein reveals an unexpected role for a bound sodium ion in orientating a substrate-binding alpha-helix in the enzyme active site. Whilst key active-site residues in rffE are present in the wlbD sequence, the sodium-binding residues outside the active site are absent. This raises questions about the modulation of enzyme activity in these two enzymes. The wlbD gene from B. pertussis has been cloned and overexpressed in E. coli and the resulting protein has been purified to homogeneity. In the current study, crystals of the mutant Gln339Arg wlbD enzyme have been obtained by sitting-drop vapour diffusion. Uncomplexed Gln339Arg and UDP-GlcNAc complex data sets have been collected in-house on a rotating-anode generator to 2.1 A. Combined, the data sets identify the space group as P2(1)2(1)2(1), with unit-cell parameters a = 78, b = 91, c = 125 A, alpha = beta = gamma = 90 degrees. The asymmetric unit contains two monomers and 53% solvent.

Synthetic mannosides act as acceptors for mycobacterial alpha1-6 mannosyltransferase.

A series of synthetic mannosides was screened in a cell-free system for their ability to act as acceptor substrates for mycobacterial mannosyltransferases. Evaluation of these compounds demonstrated the incorporation of [14C]Man from GDP-[14C]Man into a radiolabeled organic-soluble fraction and analysis by thin layer chromatography and autoradiography revealed the formation of two radiolabeled products. Each synthetic acceptor was capable of accepting one or two mannose residues, resulting in a major and a minor mannosylated product. Both products from each acceptor were isolated and their mass was confirmed by fast-atom bombardment-mass spectrometry (FABMS). Characterization of each mannosylated product by exo-glycosidase digestion. acetolysis and linkage analysis by gas chromatography mass spectrometry of partially per-O-methylated alditols, revealed only alpha1-6-linked products. In addition. the antibiotic amphomycin selectively inhibited the formation of mannosylated products suggesting polyprenolmonophosphate-mannose (C15 50-P-Man) was the immediate mannose donor in all mannosylation reactions observed. The ability of synthetic disaccharides to act as acceptor substrates in this system, is most likely due to the action of a mycobacterial polyprenol-P-Man:mannan alpha1-6 mannosyltransferase involved in the biosynthesis of linear alpha1-6-linked lipomannan.

Hydrolase and sialyltransferase activities of trypanosoma cruzi trans-sialidase towards NeuAc-alpha-2,3-gal-Gal-beta-O-PNP.

NeuAc-alpha-2,3-Gal-beta-O-PNP has been synthesised and its ability to act as a substrate for the hydrolase and transferase activities of Trypanosoma cruzi trans-sialidase have been investigated. The turn-over of this compound shows marked differences from the behaviour of NeuAc-MU. In addition, distinct differences in the action of T. cruzi trans-sialidase and Clostridium perfringens neuraminidase on NeuAc-alpha-2,3-Gal-beta-O-PNP were apparent.

Palatability of wethers fed an 80% barley diet processed at different ages and of yearling wethers grazed on native range.

Seasonal availability of lamb in the Western United States contributes to a large fluctuation in lamb supply and value. However, alternatives to fall marketing may not be practical unless palatability traits are acceptable. A 3-yr study was conducted to investigate 1) the effects of slaughter age (7 to 8; 10 to 11; or 14 to 15 mo) on carcass and palatability characteristics of wethers fed an 80% barley diet (Exp. 1); and 2) the effects of finishing on range or on an 80% barley diet on carcass and palatability traits of 14- to 15-mo-old wethers (Exp. 2). In Exp. 1, no differences (P = .27) were detected in flavor intensity or longissimus muscle area among slaughter age groups, but fat depth was greater (P < .05) for 7- to 8-mo-old wethers than for 10- to 11- or 14- to 15-mo-old wethers. Year x slaughter age interactions were detected (P < .10) for hot carcass weight, Warner-Bratzler shear value, body wall thickness, and percentage kidney fat. Hot carcass weight was greater (P < .05) for 14- to 15-mo-old wethers than for both groups of younger wethers in yr 1, did not differ (P = .53) among slaughter ages in yr 2, and was greater (P < .05) for 10- to 11- than for 14- to 15-mo-old wethers in yr 3. Warner-Bratzler shear values did not differ (P > .10) among slaughter ages in yr 1 and 3, but shear values for 14- to 15-mo-old wethers were greater (P < .05) than for both younger slaughter age groups in yr 2. Percentage kidney fat was lower (P < .05) for 14- to 15- than for 7- to 8-mo-old wethers in all years. In Exp. 2, flavor intensity of the meat did not differ (P = .35) between finishing systems, but longissimus muscle area was greater (P = .02) for range-finished wethers than for wethers fed an 80% barley diet. Year x finishing treatment interactions were detected (P < .10) for shear values, body wall thickness, percentage kidney fat, and fat depth. Shear values were greater (P = .10) for range-finished wethers than for wethers fed an 80% barley diet in yr 1, but did not differ (P > .55) in yr 2 and 3. Body wall and fat measurements were greater (P < .10) for wethers fed an 80% barley diet than for range-finished wethers in all years except yr 3, when fat depth did not differ (P = .47). Overall, slaughtering wethers fed an 80% barley diet or range-finished wethers at older ages produced acceptable carcasses with desirable meat palatability traits.

RmlC, the third enzyme of dTDP-L-rhamnose pathway, is a new class of epimerase.

Deoxythymidine diphosphate (dTDP)-L-rhamnose is the precursor of L-rhamnose, a saccharide required for the virulence of some pathogenic bacteria. dTDP-L-rhamnose is synthesized from glucose-1-phosphate and deoxythymidine triphosphate (dTTP) via a pathway involving four distinct enzymes. This pathway does not exist in humans and the enzymes involved in dTDP-L-rhamnose synthesis are potential targets for the design of new therapeutic agents. Here, the crystal structure of dTDP-6-deoxy-D-xylo-4-hexulose 3,5 epimerase (RmlC, EC5.1.3.13) from Salmonella enterica serovar Typhimurium was determined. The third enzyme of the rhamnose biosynthetic pathway, RmlC epimerizes at two carbon centers, the 3 and 5 positions of the sugar ring. The structure was determined by multiwavelength anomalous diffraction to a resolution of 2.17 A. RmlC is a dimer and each monomer is formed mainly from two beta-sheets arranged in a beta-sandwich. The structure of a dTDP-phenol-RmlC complex shows the substrate-binding site to be located between the two beta-sheets; this site is formed from residues of both monomers. Sequence alignments of other RmlC enzymes confirm that this region is very highly conserved. The enzyme is distinct structurally from other epimerases known and thus, is the first example of a new class of carbohydrate epimerase.

Ash and calcium as measures of bone in meat and bone mixtures.

Bone content of mechanically recovered meat is usually controlled by setting calcium limits, but these limits may allow more bone in some products because calcium content of fresh bone is variable. Studies involving deposition of energy, nutrients, and minerals are also dependent on ash or calcium to determine bone content of carcasses. However, ash and calcium, which is 37% of bone ash, varies by age of animal, presence or absence of tissues such as marrow or cartilage that are associated with bone, and state of bone hydration. Literature that reports ash content of bone and factors associated with its variability is the focus of this review. Based on the literature reviewed, a conversion factor of 4.5 for calcium percentage to fresh cortical bone percentage from round bones of cows, fed beef, lambs, pigs and hens is recommended. A conversion factor of calcium percentage to fresh bone percentage of 5.0 is recommended for all veal and broiler bones and for flat bones.

Effect of method of analysis on iron content of beef from advanced meat recovery systems.

A field survey was conducted by the USDA, Food Safety Inspection Service (FSIS) to provide analytical data on meat obtained from beef cervical vertebrae processed by advanced meat recovery (AMR) systems. As a result, an added iron performance standard was proposed to limit the amount of marrow in AMR products. The performance standard was based on iron content of hand boned lean compared to AMR lean. Iron content was determined by a hydrochloric wet ash digestion method. The same samples were then analyzed using dry ash digestion. The objectives of the study were to determine differences in iron content of the survey samples due to the digestion method and the impact of this difference on the added iron performance standard. Iron values by the dry ash method were approximately double those of the wet ash method. The difference was a result of incomplete volatilization of the organic matrix by hydrochloric acid in the wet ash procedure. The performance standards developed from the wet and dry ash methods were 1.8 and 3.2 mg added iron 100(-1) g, respectively. Added iron levels from the dry ash method greater than 3.2 mg 100(-1) g were present in 60% of the AMR lean indicating that some marrow was present or that factors other than amount of iron in hand boned lean should be considered before a performance standard is established.

The GPI biosynthetic pathway as a therapeutic target for African sleeping sickness.

African sleeping sickness is a debilitating and often fatal disease caused by tsetse fly transmitted African trypanosomes. These extracellular protozoan parasites survive in the human bloodstream by virtue of a dense cell surface coat made of variant surface glycoprotein. The parasites have a repertoire of several hundred immunologically distinct variant surface glycoproteins and they evade the host immune response by antigenic variation. All variant surface glycoproteins are anchored to the plasma membrane via glycosylphosphatidylinositol membrane anchors and compounds that inhibit the assembly or transfer of these anchors could have trypanocidal potential. This article compares glycosylphosphatidylinositol biosynthesis in African trypanosomes and mammalian cells and identifies several steps that could be targets for the development of parasite-specific therapeutic agents.

Man alpha1-2 Man alpha-OMe-concanavalin A complex reveals a balance of forces involved in carbohydrate recognition.

We have determined the crystal structure of the methyl glycoside of Man alpha1-2 Man in complex with the carbohydrate binding legume lectin concanavalin A (Con A). Man alpha1-2 Man alpha-OMe binds more tightly to concanavalin A than do its alpha1-3 and alpha1-6 linked counterparts. There has been much speculation as to why this is so, including a suggestion of the presence of multiple binding sites for the alpha1-2 linked disaccharide. Crystals of the Man alpha1-2 Man alpha-OMe-Con A complex form in the space group P2(1)2(1)2(1) with cell dimensions a = 119.7 A, b = 119.7 A, c = 68.9 A and diffract to 2. 75A. The final model has good geometry and an R factor of 19.6% (Rfree= 22.8%). One tetramer is present in the asymmetric unit. In three of the four subunits, electron density for the disaccharide is visible. In the fourth only a monosaccharide is seen. In one subunit the reducing terminal sugar is recognized by the monosaccharide site; the nonreducing terminal sugar occupies a new site and the major solution conformation of the inter-sugar glycosidic linkage conformation is adopted. In contrast, in another subunit the non reducing terminal sugar sits in the so called monosaccharide binding site; the reducing terminal sugar adopts a different conformation about its inter-sugar glycosidic linkage in order for the methyl group to access a hydrophobic pocket. In the third subunit, electron density for both binding modes is observed. We demonstrate that an extended carbohydrate binding site is capable of binding the disaccharide in two distinct ways. These results provide an insight in to the balance of forces controlling protein carbohydrate interactions.

N-Substituted analogues of S-nitroso-N-acetyl-D,L-penicillamine: chemical stability and prolonged nitric oxide mediated vasodilatation in isolated rat femoral arteries.

Previous studies show that linking acetylated glucosamine to S-nitroso-N-acetyl-D,L-penicillamine (SNAP) stabilizes the molecule and causes it to elicit unusually prolonged vasodilator effects in endothelium-denuded, isolated rat femoral arteries. Here we studied the propanoyl (SNPP; 3 carbon side-chain), valeryl (SNVP; 5C) and heptanoyl (SNHP; 7C) N-substituted analogues of SNAP (2C), to further investigate other molecular characteristics that might influence chemical stability and duration of vascular action of S-nitrosothiols. Spectrophotometric analysis revealed that SNVP was the most stable analogue in solution. Decomposition of all four compounds was accelerated by Cu(II) and cysteine, and neocuproine, a specific Cu(I) chelator, slowed decomposition of SNHP. Generation of NO from the compounds was confirmed by electrochemical detection at 37 degrees C. Bolus injections of SNAP (10 microl; 10(-8)-10(-3) M) into the perfusate of precontracted, isolated rat femoral arteries taken from adult male Wistar rats (400-500 g), caused concentration-dependent, transient vasodilatations irrespective of endothelial integrity. Equivalent vasodilatations induced by SNVP and SNHP were transient in endothelium-intact vessels but failed to recover to pre-injection pressures at moderate and high concentrations (10(-6)-10(-3) M) in those denuded of endothelium. This sustained effect (> 1 h) was most prevalent with SNHP and was largely reversed by the NO scavenger, haemoglobin. We suggest that increased lipophilicity of SNAP analogues with longer sidechains facilitates their retention by endothelium-denuded vessels; subsequent slow decomposition within the tissue generates sufficient NO to cause prolonged vasodilatation. This is a potentially useful characteristic for targeting NO delivery to areas of endothelial damage.

Bone marrow measurements for mechanically recovered products from machines that press bones.

Meat from Advanced Meat Recovery (AMR) systems is being used in increasing amounts in meat products, but authenticity issues resulting from the incidental inclusion of bone marrow when meat is pressed from the bones have arisen. Unfortunately, no widely accepted method for determining amount of marrow in meat from AMR systems is available. Past attempts to detect and quantify the amount of marrow in mechanically recovered products is the focus of this review. Methods for quantifying red marrow include those based on the heme pigments and on iron contained in these pigments. An immunological procedure in which antibodies are raised to bone marrow proteins and the pH method can also be used as qualitative and quantitative tests, respectively, for red marrow presence. Fatty marrow, associated with the term marrow by the public, is found in the medullary cavity of the shaft of long bones. These bones are not recommended for use in meat recovery systems, so methods for detection of fatty marrow are not addressed.

Derivation of an equation to estimate marrow content of bovine cervical vertebrae.

Marrow content of bovine cervical vertebrae from Choice- and Select-grade carcasses weighing 294 to 343 kg was determined so that a method to monitor the amount of marrow in meat from advanced meat/bone separation machinery and recovery (AMR) systems could be developed. The marrow determination requires cleaning and then ashing bones. Because a large difference in ash content of bone and bone marrow exists and because cartilage content of cervical vertebrae in Choice and Select beef is relatively constant, it was possible to derive the following equation: Weight of marrow = [weight of cartilage (% ash in cartilage - % ash in bone) + % ash in bone (total weight) - (total ash)]/[(% ash in bone - % ash in marrow)]. Constants for ash in fresh bone, marrow, and cartilage were 58.51, .57, and 2.14% with SD of 2.23, .15, and .30%, respectively. A cartilage content of 9.5% along with cervical vertebrae weight and total ash weight were also used to calculate 33.9% marrow in cervical vertebrae. Means for marrow pressed or centrifuged from bovine cervical vertebrae were lower than those obtained from the equation. Therefore, pressing and centrifuging left some marrow in spongy bone. Our ashing method for determining the amount of marrow in whole cervical vertebrae should be useful for determining marrow remaining in cervical vertebrae of bone cakes from AMR systems. Percentage ash in pressed bones is higher and the calculated marrow content is lower when pressed bones are compared to cervical vertebrae that are not pressed. The amount of marrow in whole cervical vertebrae minus the amount left in cervical vertebrae from bone cakes equals the amount in meat from AMR systems.

Solution structure of the complex between the B-subunit homopentamer of verotoxin VT-1 from Escherichia coli and the trisaccharide moiety of globotriaosylceramide.

We report the solution structure of the carbohydrate-binding B subunit of verotoxin VT-1 (VTB) from enterohemorrhagic Escherichia coli in association with the trisaccharide Galalpha1-4Galbeta1-4Glcbeta1-O-trimethylsilylethyl , determined by use of stable isotope-assisted NMR techniques. In contrast to the crystal structure of the complex which predicts three binding sites per monomer, only one of these sites is observed with substantial occupancy by the trisaccharide in solution.