Comparison of methods for analysis of proteolysis by plasmin in milk

Sensitive methods that are currently used to monitor proteolysis by plasmin in milk are limited due to 7 their high cost and lack of standardisation for quality assurance in the various dairy laboratories. In 8 this study, four methods, trinitrobenzene sulphonic acid (TNBS), reverse phase high pressure liquid 9 chromatography (RP-HPLC), gel electrophoresis and fluorescamine, were selected to assess their 10 suitability for the detection of proteolysis in milk by plasmin. Commercial UHT milk was incubated 11 with plasmin at 37 °C for one week. Clarification was achieved by isoelectric precipitation (pH 4·6 12 soluble extracts)or 6% (final concentration) trichloroacetic acid (TCA). The pH 4·6 and 6% TCA 13 soluble extracts of milk showed high correlations (R2 > 0·93) by the TNBS, fluorescamine and 14 RP-HPLC methods, confirming increased proteolysis during storage. For gel electrophoresis,15 extensive proteolysis was confirmed by the disappearance of α- and β-casein bands on the seventh 16 day, which was more evident in the highest plasmin concentration. This was accompanied by the 17 appearance of α- and β-casein proteolysis products with higher intensities than on previous days, 18 implying that more products had been formed as a result of casein breakdown. The fluorescamine 19 method had a lower detection limit compared with the other methods, whereas gel electrophoresis 20 was the best qualitative method for monitoring β-casein proteolysis products. Although HPLC was the 21 most sensitive, the TNBS method is recommended for use in routine laboratory analysis on the basis 22 of its accuracy, reliability and simplicity.

Cloud-resolving simulations of deep convection over a heated mountain

Cloud-resolving numerical simulations of airflow over a diurnally heated mountain ridge are conducted to explore the mechanisms and sensitivities of convective initiation under high pressure conditions. The simulations are based on a well-observed convection event from the Convective and Orographically Induced Precipitation Study (COPS) during summer 2007, where an isolated afternoon thunderstorm developed over the Black Forest mountains of central Europe, but they are idealized to facilitate understanding and reduce computational expense. In the conditionally unstable but strongly inhibited flow under consideration, sharp horizontal convergence over the mountain acts to locally weaken the inhibition and moisten the dry midtroposphere through shallow cumulus detrainment. The onset of deep convection occurs not through the deep ascent of a single updraft but rather through a rapid succession of thermals that are vented through the mountain convergence zone into the deepening cloud mass. Emerging thermals rise through the saturated wakes of their predecessors, which diminishes the suppressive effects of entrainment and allows for rapid glaciation above the freezing level as supercooled cloud drops rime onto preexisting ice particles. These effects strongly enhance the midlevel cloud buoyancy and enable rapid ascent to the tropopause. The existence and vigor of the convection is highly sensitive to small changes in background wind speed U0, which controls the strength of the mountain convergence and the ability of midlevel moisture to accumulate above the mountain. Whereas vigorous deep convection develops for U0 = 0 m s−1, deep convection is completely eliminated for U0 = 3 m s−1. Although deep convection is able to develop under intermediate winds (U0 = 1.5 m s−1), its formation is highly sensitive to small-amplitude perturbations in the initial flow.

The dependence of contrail formation on the weather pattern and altitude in the North Atlantic

Aircraft flying through cold ice-supersaturated air produce persistent contrails which contribute to the climate impact of aviation. Here, we demonstrate the importance of the weather situation, together with the route and altitude of the aircraft through this, on estimating contrail coverage. The results have implications for determining the climate impact of contrails as well as potential mitigation strategies. Twenty-one years of re-analysis data are used to produce a climatological assessment of conditions favorable for persistent contrail formation between 200 and 300 hPa over the north Atlantic in winter. The seasonal-mean frequency of cold ice-supersaturated regions is highest near 300 hPa, and decreases with altitude. The frequency of occurrence of ice-supersaturated regions varies with large-scale weather pattern; the most common locations are over Greenland, on the southern side of the jet stream and around the northern edge of high pressure ridges. Assuming aircraft take a great circle route, as opposed to a more realistic time-optimal route, is likely to lead to an error in the estimated contrail coverage, which can exceed 50% for westbound north Atlantic flights. The probability of contrail formation can increase or decrease with height, depending on the weather pattern, indicating that the generic suggestion that flying higher leads to fewer contrails is not robust.

Elevated levels of corticotrophin-releasing factor binding protein in the blood of patients suffering from arthritis and septicaemia and the presence of novel ligands in synovial fluid.

In view of the reported inflammatory effects of corticotrophin-releasing factor (CRF) and the associated regulatory elements in the gene of its binding protein (BP), we postulate that both BP as well as novel BP-ligands other than CRF may be involved in inflammatory disease. We have investigated BP in the blood of patients with arthritis and septicaemia and have attempted to identify CRF and other BP-ligands in synovial fluid. The BP was found to be significantly elevated in the blood of patients with rheumatoid arthritis and septicaemia. There was less BP-ligand and CRF in synovial fluid from patients with rheumatoid arthritis that from those with osteo- or psoriatic arthritis. There was at least 10-fold more BP-ligand than CRF in the fluid of all three groups of patients. A small amount of immunoreactive human (h)CRF, eluting in the expected position of CRF-41, was detected after high-pressure liquid chromatography of arthritic synovial fluid; however, the bulk of material with BP-ligand binding activity eluted earlier, suggesting that synovial fluid contained novel peptides that interacted with the BP. These results would suggest that the BP and its ligands could play an endocrine immunomodulatory role in inflammatory disease.

Modification of enrofloxacin treatment regimens for poultry experimentally infected with Salmonella enterica serovar Typhimurium DT104 to minimize selection of resistance

We hypothesized that higher doses of fluoroquinolones for a shorter duration could maintain efficacy (as measured by reduction in bacterial count) while reducing selection in chickens of bacteria with reduced susceptibility. Chicks were infected with Salmonella enterica serovar Typhimurium DT104 and treated 1 week later with enrofloxacin at the recommended dose for 5 days (water dose adjusted to give 10 mg/kg of body weight of birds or equivalence, i.e., water at 50 ppm) or at 2.5 or 5 times the recommended dose for 2 days or 1 day, respectively. The dose was delivered continuously (ppm) or pulsed in the water (mg/kg) or by gavage (mg/kg). In vitro in sera, increasing concentrations of 0.5 to 8 mu g/ml enrofloxacin correlated with increased activity. In vivo, the efficacy of the 1-day treatment was significantly less than that of the 2- and 5-day treatments. The 2-day treatments showed efficacy similar to that of the 5-day treatment in all but one repeat treatment group and significantly (P < 0.01) reduced the Salmonella counts. Dosing at 2.5x the recommended dose and pulsed dosing both increased the peak antibiotic concentrations in cecal contents, liver, lung, and sera as determined by high-pressure liquid chromatography. There was limited evidence that shorter treatment regimens (in particular the 1-day regimen) selected for fewer strains with reduced susceptibility. In conclusion, the 2-day treatment would overall require a shorter withholding time than the 5-day treatment and, in view of the increased peak antibiotic concentrations, may give rise to improved efficacy, in particular for treating respiratory and systemic infections. However, it would be necessary to validate the 2-day regimen in a field situation and in particular against respiratory and systemic infections to validate or refute this hypothesis.

Proteinase-activated receptors, targets for kallikrein signaling

Serine proteinases like thrombin can signal to cells by the cleavage/activation of proteinase-activated receptors (PARs). Although thrombin is a recognized physiological activator of PAR(1) and PAR(4), the endogenous enzymes responsible for activating PAR(2) in settings other than the gastrointestinal system, where trypsin can activate PAR(2), are unknown. We tested the hypothesis that the human tissue kallikrein (hK) family of proteinases regulates PAR signaling by using the following: 1) a high pressure liquid chromatography (HPLC)-mass spectral analysis of the cleavage products yielded upon incubation of hK5, -6, and -14 with synthetic PAR N-terminal peptide sequences representing the cleavage/activation motifs of PAR(1), PAR(2), and PAR(4); 2) PAR-dependent calcium signaling responses in cells expressing PAR(1), PAR(2), and PAR(4) and in human platelets; 3) a vascular ring vasorelaxation assay; and 4) a PAR(4)-dependent rat and human platelet aggregation assay. We found that hK5, -6, and -14 all yielded PAR peptide cleavage sequences consistent with either receptor activation or inactivation/disarming. Furthermore, hK14 was able to activate PAR(1), PAR(2), and PAR(4) and to disarm/inhibit PAR(1). Although hK5 and -6 were also able to activate PAR(2), they failed to cause PAR(4)-dependent aggregation of rat and human platelets, although hK14 did. Furthermore, the relative potencies and maximum effects of hK14 and -6 to activate PAR(2)-mediated calcium signaling differed. Our data indicate that in physiological settings, hKs may represent important endogenous regulators of the PARs and that different hKs can have differential actions on PAR(1), PAR(2), and PAR(4).

Indian summer monsoon variations could have affected the early Holocene woodland expansion in the Near East

Postglacial expansion of deciduous oak woodlands of the Zagros—Anti-Taurus Mountains, a major biome of the Near East, was delayed until the middle Holocene at ~6300 cal. yr BP. The current hypotheses explain this delay as a consequence of a regional aridity during the early Holocene, slow migration rates of forest trees, and/or a long history of land use and agro-pastoralism in this region. In the present paper, support is given to a hypothesis that suggests different precipitation seasonalities during the early Holocene compared with the late Holocene. The oak species of the Zagros—Anti-Taurus Mts, particularly Quercus brantii Lindl., are strongly dependent on spring precipitation for regeneration and are sensitive to a long dry season. Detailed analysis of modern atmospheric circulation patterns in SW Asia during the late spring suggests that the Indian Summer Monsoon (ISM) intensification can modify the amount of late spring and/or early summer rainfall in western/northwestern Iran and eastern Anatolia, which could in turn have controlled the development of the Zagros—Anti-Taurus deciduous oak woodlands. During the early Holocene, the northwestward shift of the Inter-Tropical Convergence Zone (ITCZ) could have displaced the subtropical anticyclonic belt or associated high pressure ridges to the northwest. The latter could, in turn, have prevented the southeastward penetration of low pressure systems originating from the North Atlantic and Black Sea regions. Such atmospheric configuration could have reduced or eliminated the spring precipitation creating a typical Mediterranean continental climate characterized by winter-dominated precipitation. This scenario highlights the complexity of biome response to climate system interactions in transitional climatic and biogeographical regions.

Late Quaternary fire regimes of Australasia

We have compiled 223 sedimentary charcoal records from Australasia in order to examine the temporal and spatial variability of fire regimes during the Late Quaternary. While some of these records cover more than a full glacial cycle, here we focus on the last 70,000 years when the number of individual records in the compilation allows more robust conclusions. On orbital time scales, fire in Australasia predominantly reflects climate, with colder periods characterized by less and warmer intervals by more biomass burning. The composite record for the region also shows considerable millennial-scale variability during the last glacial interval (73.5–14.7 ka). Within the limits of the dating uncertainties of individual records, the variability shown by the composite charcoal record is more similar to the form, number and timing of Dansgaard–Oeschger cycles as observed in Greenland ice cores than to the variability expressed in the Antarctic ice-core record. The composite charcoal record suggests increased biomass burning in the Australasian region during Greenland Interstadials and reduced burning during Greenland Stadials. Millennial-scale variability is characteristic of the composite record of the sub-tropical high pressure belt during the past 21 ka, but the tropics show a somewhat simpler pattern of variability with major peaks in biomass burning around 15 ka and 8 ka. There is no distinct change in fire regime corresponding to the arrival of humans in Australia at 50 ± 10 ka and no correlation between archaeological evidence of increased human activity during the past 40 ka and the history of biomass burning. However, changes in biomass burning in the last 200 years may have been exacerbated or influenced by humans.

Observations of urban boundary layer structure during a strong urban heat island event

It has long been known that the urban surface energy balance is different to that of a rural surface, and that heating of the urban surface after sunset gives rise to the Urban Heat Island (UHI). Less well known is how flow and turbulence structure above the urban surface are changed during different phases of the urban boundary layer (UBL). This paper presents new observations above both an urban and rural surface and investigates how much UBL structure deviates from classical behaviour. A 5-day, low wind, cloudless, high pressure period over London, UK, was chosen for analysis, during which there was a strong UHI. Boundary layer evolution for both sites was determined by the diurnal cycle in sensible heat flux, with an extended decay period of approximately 4 h for the convective UBL. This is referred to as the “Urban Convective Island” as the surrounding rural area was already stable at this time. Mixing height magnitude depended on the combination of regional temperature profiles and surface temperature. Given the daytime UHI intensity of 1.5∘C, combined with multiple inversions in the temperature profile, urban and rural mixing heights underwent opposite trends over the period, resulting in a factor of three height difference by the fifth day. Nocturnal jets undergoing inertial oscillations were observed aloft in the urban wind profile as soon as the rural boundary layer became stable: clear jet maxima over the urban surface only emerged once the UBL had become stable. This was due to mixing during the Urban Convective Island reducing shear. Analysis of turbulent moments (variance, skewness and kurtosis) showed “upside-down” boundary layer characteristics on some mornings during initial rapid growth of the convective UBL. During the “Urban Convective Island” phase, turbulence structure still resembled a classical convective boundary layer but with some influence from shear aloft, depending on jet strength. These results demonstrate that appropriate choice of Doppler lidar scan patterns can give detailed profiles of UBL flow. Insights drawn from the observations have implications for accuracy of boundary conditions when simulating urban flow and dispersion, as the UBL is clearly the result of processes driven not only by local surface conditions but also regional atmospheric structure.

On the cause of a magnetospheric flux transfer event

We present a detailed investigation of a magnetospheric flux transfer event (FTE) seen by the Active Magnetospheric Tracer Explorer (AMPTE) UKS and IRM satellites around 1046 UT on October 28, 1984. This event has been discussed many times previously in the literature and has been cited as support for a variety of theories of FTE formation. We make use of a model developed to reproduce ion precipitations seen in the cusp ionosphere. The analysis confirms that the FTE is well explained as a brief excursion into an open low-latitude boundary layer (LLBL), as predicted by two theories of magnetospheric FTEs: namely, that they are bulges in the open LLBL due to reconnection rate enhancements or that they are indentations of the magnetopause by magnetosheath pressure increases (but in the presence of ongoing steady reconnection). The indentation of the inner edge of the open LLBL that these two models seek to explain is found to be shallow for this event. The ion model reproduces the continuous evolution of the ion distribution function between the sheath-like population at the event center and the surrounding magnetospheric populations; it also provides an explanation of the high-pressure core of the event as comprising field lines that were reconnected considerably earlier than those that are draped over it to give the event boundary layer. The magnetopause transition parameter is used to isolate a field rotation on the boundaries of the core, which is subjected to the tangential stress balance test. The test identifies this to be a convecting structure, which is neither a rotational discontinuity (RD) nor a contact discontinuity, but could possibly be a slow shock. In addition, evidence for ion reflection off a weak RD on the magnetospheric side of this structure is found. The event structure is consistent in many ways with features predicted for the open LLBL by analytic MHD theories and by MHD and hybrid simulations. The de Hoffman-Teller velocity of the structure is significantly different from that of the magnetosheath flow, indicating that it is not an indentation caused by a high-pressure pulse in the sheath but is consistent with the motion of newly opened field lines (different from the sheath flow because of the magnetic tension force) deduced from the best fit to the ion data. However, we cannot here rule out the possibility that the sheath flow pattern has changed in the long interval between the two satellites observing the FTE and subsequently emerging into the magnetosheath; thus this test is not conclusive in this particular case. Analysis of the fitted elapsed time since reconnection shows that the core of the event was reconnected in one pulse and the event boundary layer was reconnected in a subsequent pulse. Between these two pulses is a period of very low (but nonzero) reconnection rate, which lasts about 14 mins. Thus the analysis supports, but does not definitively verify, the concept that the FTE is a partial passage into an open LLBL caused by a traveling bulge in that layer produced by a pulse in reconnection rate.

An evaluation of the simulated water balance of Eurasia and northern Africa at 6000 yr B.P. using lake status data

Changes in the water balance of Eurasia and northern Africa in response to insolation forcing at 6000 y BP simulated by five atmospheric general circulation models have been compared with observations of changes in lake status. All of the simulations show enhancement of the Asian summer monsoon and of the high pressure cells over the Pacific and Central Asia and the Middle East, causing wetter conditions in northern India and southern China and drier conditions along the Chinese coast and west of the monsoon core. All of the models show enhancement of the African monsoon, causing wetter conditions in the zone between ca 10–20 °N. Four of the models show conditions wetter than present in southern Europe and drier than present in northern Europe. Three of the models show conditions similar to present in the mid-latitude continental interior, while the remaining models show conditions somewhat drier than present. The extent and location of each of the simulated changes varies between the models, as does the mechanism producing these changes. The lake data confirm some features of the simulations, but indicate discrepancies between observed and simulated climates. For example, the data show: (1) conditions wetter than present in central Asia, from India to northern China and Mongolia, indicating that the simulated Asian monsoon expansion is too small; (2) conditions wetter than present between ca. 10–30 °N in Africa, indicating that the simulated African monsoon expansion is too small; (3) that northern Europe was drier, but the area of significantly drier conditions was more localized (around the Baltic) than shown in the simulations; (4) that southern Europe was wetter than present, apparently consistent with the simulations, but pollen data suggest that this reflects an increase in summer rainfall whereas the models show winter precipitation, and (5) that the mid-latitude continental interior was generally wetter than present.

Where were the monsoon regions and arid zones in Asia prior to the Tibetan Plateau uplift?

The impact of the Tibetan Plateau uplift on the Asian monsoons and inland arid climates is an important but also controversial question in studies of paleoenvironmental change during the Cenozoic. In order to achieve a good understanding of the background for the formation of the Asian monsoons and arid environments, it is necessary to know the characteristics of the distribution of monsoon regions and arid zones in Asia before the plateau uplift. In this study, we discuss in detail the patterns of distribution of the Asian monsoon and arid regions before the plateau uplift on the basis of modeling results without topography from a global coupled atmosphere–ocean general circulation model, compare our results with previous simulation studies and available biogeological data, and review the uncertainties in the current knowledge. Based on what we know at the moment, tropical monsoon climates existed south of 20°N in South and Southeast Asia before the plateau uplift, while the East Asian monsoon was entirely absent in the extratropics. These tropical monsoons mainly resulted from the seasonal shifts of the Inter-Tropical Convergence Zone. There may have been a quasi-monsoon region in central-southern Siberia. Most of the arid regions in the Asian continent were limited to the latitudes of 20–40°N, corresponding to the range of the subtropical high pressure year-around. In the meantime, the present-day arid regions located in the relatively high latitudes in Central Asia were most likely absent before the plateau uplift. The main results from the above modeling analyses are qualitatively consistent with the available biogeological data. These results highlight the importance of the uplift of the Tibetan Plateau in the Cenozoic evolution of the Asian climate pattern of dry–wet conditions. Future studies should be focused on effects of the changes in land–sea distribution and atmospheric CO2 concentrations before and after the plateau uplift, and also on cross-comparisons between numerical simulations and geological evidence, so that a comprehensive understanding of the evolution of the Cenozoic paleoenvironments in Asia can be achieved.

Examining the predictability of the Stratospheric Sudden Warming of January 2013 using multiple NWP systems

The first multi-model study to estimate the predictability of a boreal Sudden Stratospheric Warming (SSW) is performed using five NWP systems. During the 2012-2013 boreal winter, anomalous upward propagating planetary wave activity was observed towards the end of December, which followed by a rapid deceleration of the westerly circulation around 2 January 2013, and on 7 January 2013 the zonal mean zonal wind at 60°N and 10 hPa reversed to easterly. This stratospheric dynamical activity was followed by an equatorward shift of the tropospheric jet stream and by a high pressure anomaly over the North Atlantic, which resulted in severe cold conditions in the UK and Northern Europe. In most of the five models, the SSW event was predicted 10 days in advance. However, only some ensemble members in most of the models predicted weakening of westerly wind when the models were initialized 15 days in advance of the SSW. Further dynamical analysis of the SSW shows that this event was characterized by the anomalous planetary wave-1 amplification followed by the anomalous wave-2 amplification in the stratosphere, which resulted in a split vortex occurring between 6 January 2013 and 8 January 2013. The models have some success in reproducing wave-1 activity when initialized 15 days in advance, they but generally failed to produce the wave-2 activity during the final days of the event. Detailed analysis shows that models have reasonably good skill in forecasting tropospheric blocking features that stimulate wave-2 amplification in the troposphere, but they have limited skill in reproducing wave-2 amplification in the stratosphere.

Edible oil from Tiger nut (Cyperus esculentus L.): mechanical pressing and aqueous enzymatic extraction methods

The tiger nut tuber of the Cyperus esculentus L. plant is an unusual storage system with similar amounts of starch and lipid. The extraction of its oil employing both mechanical pressing and aqueous enzymatic extraction (AEE) methods was investigated and an examination of the resulting products was carried out. The effects of particle size and moisture content of the tuber on the yield of tiger nut oil with pressing were initially studied. Smaller particles were found to enhance oil yields while a range of moisture content was observed to favour higher oil yields. When samples were first subjected to high pressures up to 700 MPa before pressing at 38 MPa there was no increase in the oil yields. Ground samples incubated with a mixture of α- Amylase, Alcalase, and Viscozyme (a mixture of cell wall degrading enzyme) as a pre-treatment, increased oil yield by pressing and 90% of oil was recovered as a result. When aqueous enzymatic extraction was carried out on ground samples, the use of α- Amylase, Alcalase, and Celluclast independently improved extraction oil yields compared to oil extraction without enzymes by 34.5, 23.4 and 14.7% respectively. A mixture of the three enzymes further augmented the oil yield and different operational factors were individually studied for their effects on the process. These include time, total mixed enzyme concentration, linear agitation speed, and solid-liquid ratio. The largest oil yields were obtained with a solid-liquid ratio of 1:6, mixed enzyme concentration of 1% (w/w) and 6 h incubation time although the longer time allowed for the formation of an emulsion. Using stationary samples during incubation surprisingly gave the highest oil yields, and this was observed to be as a result of gravity separation occurring during agitation. Furthermore, the use of high pressure processing up to 300 MPa as a pre-treatment enhanced oil yields but additional pressure increments had a detrimental effect. The quality of oils recovered from both mechanical and aqueous enzymatic extraction based on the percentage free fatty acid (% FFA) and peroxide values (PV) all reflected the good stabilities of the oils with the highest % FFA of 1.8 and PV of 1.7. The fatty acid profiles of all oils also remained unchanged. The level of tocopherols in oils were enhanced with both enzyme aided pressing (EAP) and high pressure processing before AEE. Analysis on the residual meals revealed DP 3 and DP 4 oligosaccharides present in EAP samples but these would require further assessment on their identity and quality.

Characterisation of synoptic conditions and cyclones associated with top ranking potential wind loss events over Iberia

Intense extra-tropical cyclones are often associated with strong winds, heavy precipitation and socio-economic impacts. Over southwestern Europe, such storms occur less often, but still cause high economic losses. We characterise the largescale atmospheric conditions and cyclone tracks during the top-100 potential losses over Iberia associated with wind events. Based on 65 years of reanalysis data,events are classified into four groups: (i) cyclone tracks crossing over Iberia on the event day (“Iberia”), (ii) cyclones crossing further north, typically southwest of the British Isles (“North”), (iii) cyclones crossing southwest to northeast near the northwest tip of Iberia (“West”), and (iv) so called “Hybrids”, characterised by a strong pressure gradient over Iberia due to the juxtaposition of low and high pressure centres. Generally, “Iberia” events are the most frequent (31% to 45% for top-100 vs.top-20), while “West” events are rare (10% to 12%). 70% of the events were primarily associated with a cyclone. Multi-decadal variability in the number of events is identified. While the peak in recent years is quite prominent, other comparably stormy periods occurred in the 1960s and 1980s. This study documents that damaging wind storms over Iberia are not rare events, and their frequency of occurrence undergoes strong multi-decadal variability.