Introduction of 4-chloro-alpha-cyanocinnamic acid liquid matrices for high sensitivity UV-MALDI MS

Matrix-assisted laser desorption/ionization (MALDI) is a key ionization technique in mass spectrometry (MS) for the analysis of labile macromolecules. An important area of study and improvements in relation to MALDI and its application in high-sensitivity MS is that of matrix design and sample preparation. Recently, 4-chloro-alpha-cyanocinnamic acid (ClCCA) has been introduced as a new rationally designed matrix and reported to provide an improved analytical performance as demonstrated by an increase in sequence coverage of protein digests obtained by peptide mass mapping (PMM) (Jaskolla, T. W.; et al. Proc. Natl. Acad. Sci. U.S.A. 2008, 105, 12200-12205). This new matrix shows the potential to be a superior alternative to the commonly used and highly successful alpha-cyano-4-hydroxycinnamic acid (CHCA). We have taken this design one step further by developing and optimizing an ionic liquid matrix (ILM) and liquid support matrix (LSM) using ClCCA as the principle chromophore and MALDI matrix compound. These new liquid matrices possess greater sample homogeneity and a simpler morphology. The data obtained from our studies show improved sequence coverage for BSA digests compared to the traditional CHCA crystalline matrix and for the ClCCA-containing ILM a similar performance to the ClCCA crystalline matrix down to 1 fmol of BSA digest prepared in a single MALDI sample droplet with current sensitivity levels in the attomole range. The LSMs show a high tolerance to contamination such as ammonium bicarbonate, a commonly used buffering agent

Impact of Mesh Tracks and Low-Ground-Pressure Vehicle Use on Blanket Peat Hydrology

No abstract available

Integrating case-based reasoning and hypermedia documentation: an application for the diagnosis of a welding robot at Odense steel shipyard

Reliable and effective maintenance support is a vital consideration for the management within today's manufacturing environment. This paper discusses the development of a maintenance system for the world's largest robot welding facility. The development system combines a case-based reasoning approach for diagnosis with context information, as electronic on-line manuals, linked using open hypermedia technology. The work discussed in this paper delivers not only a maintenance system for the robot stations under consideration, but also a design framework for developing maintenance systems for other similar applications

Research Briefing: Strava-using Parkrunners: A Community Study

No abstract available

Novel antioxidant and anti-inflammatory peptides from the Siamese crocodile (Crocodylus siamensis) hemoglobin hydrolysate

Novel antioxidant and anti-inflammatory peptides were isolated from hydrolysates of Siamese crocodile (Crocodylus siamensis) hemoglobin. C. siamensis hemoglobin hydrolysates (CHHs) were obtained by pepsin digestion at different incubation times (2, 4, 6, and 8 H) at 37 °C and subjected to antioxidant and anti-inflammatory activity assessment. CHH obtained by 2-H hydrolysis (2H-CHH) showed the highest anti-inflammatory activity with respect to decreasing nitric oxide (NO) production, whereas the strongest antioxidant activity was found for 6-H hydrolysis (6H-CHH) against nitric oxide radicals. To evaluate the anti-inflammatory and antioxidant activity of individual peptide components, 2H-CHH and 6H-CHH were purified by semipreparative HPLC. Peptide fraction P57 isolated from 6H-CHH was found to exhibit the highest nitric oxide radical inhibition activity (32.0%). Moreover, purification of 2H-CHH yielded peptide fraction P16, which displayed a high efficacy in decreasing NO production of macrophage RAW 264.7 cells (83.2%) and significantly reduced proinflammatory cytokines and inflammatory mediators interleukin-6 (IL-6), interleukin-1 beta (IL-1β), and prostaglandin-E2 (PGE2) production to about 2.0, 0.3, and 1.9 ng/mL, respectively. Using LTQ orbitrap XL mass spectrometry, active peptide sequences were identified as antioxidant KIYFPHF (KF7), anti-inflammatory SAFNPHEKQ (SQ9), and IIHNEKVQAHGKKVL (IL15). Additionally, CHHs simulated gastric and intestinal in vitro digestion positively contributed to antioxidant and anti-inflammatory activity. Taken collectively, the results of this work demonstrate that CHHs contain several peptides with anti-inflammatory and antioxidant properties, which may prove valuable as treatment or supplement against diseases associated with inflammation and oxidative stress

Virus Propagation in Multiple Profile Networks

Suppose we have a virus or one competing idea/product that propagates over a multiple profile (e.g., social) network. Can we predict what proportion of the network will actually get "infected" (e.g., spread the idea or buy the competing product), when the nodes of the network appear to have different sensitivity based on their profile? For example, if there are two profiles $\mathcal{A}$ and $\mathcal{B}$ in a network and the nodes of profile $\mathcal{A}$ and profile $\mathcal{B}$ are susceptible to a highly spreading virus with probabilities $\beta_{\mathcal{A}}$ and $\beta_{\mathcal{B}}$ respectively, what percentage of both profiles will actually get infected from the virus at the end? To reverse the question, what are the necessary conditions so that a predefined percentage of the network is infected? We assume that nodes of different profiles can infect one another and we prove that under realistic conditions, apart from the weak profile (great sensitivity), the stronger profile (low sensitivity) will get infected as well. First, we focus on cliques with the goal to provide exact theoretical results as well as to get some intuition as to how a virus affects such a multiple profile network. Then, we move to the theoretical analysis of arbitrary networks. We provide bounds on certain properties of the network based on the probabilities of infection of each node in it when it reaches the steady state. Finally, we provide extensive experimental results that verify our theoretical results and at the same time provide more insight on the problem

Synthesis and Testing of Inhibitors of Dihydrodipicolinate Synthase

There are two distinct biosynthetic pathways to the essential amino acid L-lysine (A). The diaminopimelate pathway to L-lysine occurs in higher plants and bacteria. The second pathway known is the alpha-aminoadipate pathway and is found to operate in fungi and yeasts. This thesis will deal with only the diaminopimelate pathway to L-lysine and in particular with the first step, which involves the condensation of L-aspartic acid beta-semialdehyde (ASA) (B) with pyruvate (C) to form L-dihydrodipicolinate (DHDP) (D). The mechanism of formation of L-DHDP (D) was studied using electrospray mass spectrometry. The synthesis and testing of potential inhibitors of dihydrodipicolinate synthase (DHDPS) was also studied. [diagram] L-ASA is a substrate of the first enzyme of the diaminopimelate pathway to L-lysine. A former co-worker in the group, Dr D. Tudor had developed a route to 1-ASA as the trifluoroacetate salt. This route was low yielding, approximately 14% for four steps, thus a higher yielding route was developed utilising the para-methoxybenzyl (PMB) ester protecting group. This material was not suitable for use in the biochemical assay as an impurity from the deprotection stage was found to absorb strongly at the wavelength used for the our enzyme assay system. An improved procedure for the synthesis of L-ASA was developed increasing the overall yield of the procedure to approximately 48% for the same four steps. The synthesis of L-ASA as its trifluoroacetate salt allowed a number of analogues of 1-ASA to be prepared with some synthetic modification of the original route. The compounds prepared this way were alkylated derivatives of L-ASA. The alpha-methyl ASA (E) was prepared by the reaction of methyl iodide with the anion generated from treating diprotected allylglycine with lithium diisopropylamide (LDA). It proved to be a poor inhibitor but initial studies suggest that it may be a reasonably good substrate for DHDPS. A number of other derivatives were prepared including beta-methyl ASA (F) which again was a poor inhibitor but was found to be a good substrate for DHDPS (beta-methyl ASA is utilised at approximately 20% of the rate that L-ASA is consumed). A number of other derivatives and analogues of L-ASA were prepared. A number of heterocyclic compounds were prepared as analogues of DHDP and were tested for inhibitory effects with DHDPS. These compounds were prepared by a 1,3-dipolar cycloaddition of a nitrile oxide onto an alkene or alkyne. The isoxazolines produced had a general structure (G). These compounds were found to be poor inhibitors of DHDPS with none showing inhibition below 1 mM. The ring opened isoxazolines (H) were prepared as analogues of pyruvate but again they proved to be poor inhibitors of DHDPS. [diagram] An attempt to synthesise glutamic acid gamma-semialdehyde (I) starting from glutamic acid was undertaken. The compound isolated from the series of reactions was found to have cyclised and was stable as the carbinolamine (J). This route was abandoned due to the cyclisation of the product. [diagram] A number of pyridinedicarboxylic acid derivatives and analogues were prepared to test for inhibitory activity. The pyridine-2,6-dicarboxylic acid N-oxide (K) and the pyridine-2,6-dinitrile (L) showed very good inhibitory activity. These compounds were studied in detail to determine the type of inhibition they showed. The two compounds (K) and (L) were found to be non-competitive inhibitors of DHDPS. A number of other saturated and unsaturated analogues of L-DHDP were prepared and tested for inhibitory action. A study of the mechanism of DHDPS was undertaken using electrospray mass spectrometry to detect enzyme bound intermediates. The electrospray mass spectrometer was able to provide evidence for a number of pyruvate analogues bound to the enzyme as Schiff's bases. No evidence for L-ASA bound to DHDPS could be found. Further to these studies was the need to preserve stocks of DHDPS used for inhibitor testing and biotransformations. A study of the immobilisation of DHDPS on Eupergit resins was undertaken to determine the feasibility of this technique as a method of obtaining reusable DHDPS. The studies found that only up to 18% of the initial sample of DHDPS was bound to the beads. This suggests that this may not be a suitable method for the immobilisation of DHDPS, however DHDPS bound to the beads was found to have long term stability

Enhancing Palliative Care for Patients With Advanced Heart Failure Through Simple Prognostication Tools: A Comparison of the Surprise Question, the Number of Previous Heart Failure Hospitalizations, and the Seattle Heart Failure Model for Predicting 1-Year Survival

Background: Score-based survival prediction in patients with advanced heart failure (HF) is complicated. Easy-to-use prognostication tools could inform clinical decision-making and palliative care delivery. Objective: To compare the prognostic utility of the Seattle HF model (SHFM), the surprise question (SQ), and the number of HF hospitalizations (NoH) within the last 12 months for predicting 1-year survival in patients with advanced HF. Methods: We retrospectively analyzed data from a cluster-randomized controlled trial of advanced HF patients, predominantly with reduced ejection fraction. Primary outcome was the prognostic discrimination of SHFM, SQ (“Would you be surprised if this patient were to die within 1 year?”) answered by HF cardiologists, and NoH, assessed by receiver operating characteristic (ROC) curve analysis. Optimal cut-offs were calculated using Youden’s index (SHFM: <86% predicted 1-year survival; NoH ≥ 2). Results: Of 535 subjects, 82 (15.3%) had died after 1-year of follow-up. SHFM, SQ, and NoH yielded a similar area under the ROC curve [SHFM: 0.65 (0.60–0.71 95% CI); SQ: 0.58 (0.54–0.63 95% CI); NoH: 0.56 (0.50–0.62 95% CI)] and similar sensitivity [SHFM: 0.76 (0.65–0.84 95% CI); SQ: 0.84 (0.74–0.91 95% CI); NoH: 0.56 (0.45–0.67 95% CI)]. As compared to SHFM, SQ had lower specificity [SQ: 0.33 (0.28–0.37 95% CI) vs. SHFM: 0.55 (0.50–0.60 95% CI)] while NoH had similar specificity [0.56 (0.51–0.61 95% CI)]. SQ combined with NoH showed significantly higher specificity [0.68 (0.64–0.73 95% CI)]. Conclusion: SQ and NoH yielded comparable utility to SHFM for 1-year survival prediction among advanced HF patients, are easy-to-use and could inform bedside decision-making

Pathogenic Potential to Humans of Bovine Escherichia coli O26, Scotland

Escherichia coli O26 and O157 have similar overall prevalences in cattle in Scotland, but in humans, Shiga toxin–producing E. coli O26 infections are fewer and clinically less severe than E. coli O157 infections. To investigate this discrepancy, we genotyped E. coli O26 isolates from cattle and humans in Scotland and continental Europe. The genetic background of some strains from Scotland was closely related to that of strains causing severe infections in Europe. Nonmetric multidimensional scaling found an association between hemolytic uremic syndrome (HUS) and multilocus sequence type 21 strains and confirmed the role of stx&lt;sub&gt;2&lt;/sub&gt; in severe human disease. Although the prevalences of E. coli O26 and O157 on cattle farms in Scotland are equivalent, prevalence of more virulent strains is low, reducing human infection risk. However, new data on E. coli O26–associated HUS in humans highlight the need for surveillance of non-O157 enterohemorrhagic E. coli and for understanding stx&lt;sub&gt;2&lt;/sub&gt; phage acquisition

E. coli O157 on Scottish cattle farms: evidence of local spread and persistence using repeat cross-sectional data

&lt;b&gt;Background&lt;/b&gt;&lt;p&gt;&lt;/p&gt; Escherichia coli (E. coli) O157 is a virulent zoonotic strain of enterohaemorrhagic E. coli. In Scotland (1998-2008) the annual reported rate of human infection is 4.4 per 100,000 population which is consistently higher than other regions of the UK and abroad. Cattle are the primary reservoir. Thus understanding infection dynamics in cattle is paramount to reducing human infections.&lt;p&gt;&lt;/p&gt; A large database was created for farms sampled in two cross-sectional surveys carried out in Scotland (1998 - 2004). A statistical model was generated to identify risk factors for the presence of E. coli O157 on farms. Specific hypotheses were tested regarding the presence of E. coli O157 on local farms and the farms previous status. Pulsed-field gel electrophoresis (PFGE) profiles were further examined to ascertain whether local spread or persistence of strains could be inferred.&lt;p&gt;&lt;/p&gt; &lt;b&gt;Results&lt;/b&gt;&lt;p&gt;&lt;/p&gt; The presence of an E. coli O157 positive local farm (average distance: 5.96km) in the Highlands, North East and South West, farm size and the number of cattle moved onto the farm 8 weeks prior to sampling were significant risk factors for the presence of E. coli O157 on farms. Previous status of a farm was not a significant predictor of current status (p = 0.398). Farms within the same sampling cluster were significantly more likely to be the same PFGE type (p &#60; 0.001), implicating spread of strains between local farms. Isolates with identical PFGE types were observed to persist across the two surveys, including 3 that were identified on the same farm, suggesting an environmental reservoir. PFGE types that were persistent were more likely to have been observed in human clinical infections in Scotland (p &#60; 0.001) from the same time frame.&lt;p&gt;&lt;/p&gt; &lt;b&gt;Conclusions&lt;/b&gt;&lt;p&gt;&lt;/p&gt; The results of this study demonstrate the spread of E. coli O157 between local farms and highlight the potential link between persistent cattle strains and human clinical infections in Scotland. This novel insight into the epidemiology of Scottish E. coli O157 paves the way for future research into the mechanisms of transmission which should help with the design of control measures to reduce E. coli O157 from livestock-related sources