Search

Oxysterols play a key role in many (patho)physiological processes and they are potential biomarkers for oxidative stress in several diseases. Here we developed a rapid gas chromatographic-mass spectrometry-based method for the separation and quantification of 11 biologically relevant oxysterols bearing hydroxy, epoxy, and dihydroxy groups. Efficient chromatographic separation (resolution ≥ 1.9) was achieved using a medium polarity 35%-diphenyl/65%-dimethyl polysiloxane stationary phase material (30 m × 0.25 mm inner diameter and 0.25 μm film thickness). Based on thorough analysis of the fragmentation during electron ionization we developed a strategy to deduce structural information of the oxysterols. Optimized sample preparation includes (i) extraction with a mixture of n-hexane/iso-propanol, (ii) removal of cholesterol by solid phase extraction with unmodified silica, and (iii) trimethylsilylation. The method was successfully applied on the analysis of brain samples, showing consistent results with previous studies and a good intra- and interday precision of ≤20%. Finally, we used the method for the investigation of oxysterol formation during oxidative stress in HepG2 cells. Incubation with tert-butyl hydroperoxide led to a massive increase in free radical formed oxysterols (7-keto-chol 7β-OH-chol 7α-OH-chol), while 24 h incubation with the glutathione peroxidase 4 inhibitor RSL3 showed no increase in oxidative stress based on the oxysterol pattern. Overall, the new method described here enables the robust analysis of a biologically meaningful pattern of oxysterols with high sensitivity and precision allowing us to gain new insights in the biological formation and role of oxysterols.

Enzymatic and nonenzymatic oxidation of linoleic (LA) and α-linolenic acid (ALA) during pressing and storage of plant oils leads to a variety of oxylipins. We pressed oils from flaxseeds, rapeseeds, and sunflower seeds and analyzed the oxylipin pattern in freshly pressed oils. 9-/13-Hydro(pero)xy-LA/-ALA occurred in high concentration resulting probably from lipoxygenase-catalyzed reactions as well as autoxidation and photooxidation. However, in flaxseed and rapeseed oil, the highest concentrations were found for the terminal epoxy-ALA (15(16)-EpODE) and the hardly known 15-hydroxy-LA (15-HODE, 80 mg/100 g in flaxseed oil). Oils were stored for 6 months and the peroxide value (PV) as well as oxylipin and secondary volatile aldehyde concentrations were determined. While lipid peroxidation in flaxseed oil was surprisingly low, the oxylipin concentration and PV massively increased in rapeseed oil dependent on oxygen availability. Oxylipin concentrations correlated well with the PV, while secondary volatile aldehydes did not reflect the changes of oxylipins and PVs. The comprehensive analysis of hydroxy-, epoxy-, and dihydroxy-LA/-ALA reveals new and unique insights into the composition of plant oils and ongoing oxidation processes.

Analysis of fatty acids (FA) in food and biological samples such as blood is indispensable in modern life sciences. We developed a rapid, sensitive and comprehensive method for the quantification of 41 saturated and unsaturated fatty acids by means of LC-MS. Optimized chromatographic separation of isobaric analytes was carried out on a C8 reversed phase analytical column (100 × 2.1 mm, 2.6 μm core–shell particle) with a total run time of 15 min with back pressure lower than 300 bar. On an old triple quadrupole instrument (3200, AB Sciex), pseudo selected reaction monitoring mode was used for quantification of the poorly fragmenting FA, yielding limits of detection of 5–100 nM. Sample preparation was carried out by removal of phospholipids and triglycerides by solid-phase extraction (non-esterified fatty acids in oils) or saponification in iso-propanol (fatty acyls). This is not only a rapid strategy for quantification of fatty acyls, but allows the direct combination with the LC-MS-based analysis of fatty acid oxidation products (eicosanoids and other oxylipins) from the same sample. The concentrations of fatty acyls determined by means of LC-MS were consistent with those from GC-FID analysis demonstrating the accuracy of the developed method. Moreover, the method shows high precisions with a low intra-day (≤ 10% for almost all fatty acids in plasma and ≤ 15% in oils) and inter-day as well as inter-operator variability (< 20%). The method was successfully applied on human plasma and edible oils. The possibility to quantify non-esterified fatty acids in samples containing an excess of triacylglycerols and phospholipids is a major strength of the described approach allowing to gain new insights in the composition of biological samples. Graphical abstract

This in vivo study aimed to test if a diet enriched with 6% walnuts alone or in combination with physical activity supports healthy ageing by changing the oxylipin profile in brain and liver, improving motor function, cognition, and cerebral mitochondrial function. Female NMRI mice were fed a 6% walnut diet starting at an age of 12 months for 24 weeks. One group was additionally maintained in an enriched environment, one group without intervention served as control. After three months, one additional control group of young mice (3 weeks old) was introduced. Motor and cognitive functions were measured using Open Field, Y-Maze, Rotarod and Passive Avoidance tests. Lipid metabolite profiles were determined using RP-LC-ESI(-)-MS/MS in brain and liver tissues of mice. Cerebral mitochondrial function was characterized by the determination of ATP levels, mitochondrial membrane potential and mitochondrial respiration. Expression of genes involved with mito- and neurogenesis, inflammation, and synaptic plasticity were determined using qRT-PCR. A 6% walnut-enriched diet alone improved spatial memory in a Y-Maze alternation test (p p* p** p*** p

Supplementing long-chain omega-3 polyunsaturated fatty acids (n3-PUFA) improves health. We characterized the pattern of total and non-esterified oxylipins and fatty acids in n3 supplements made of fish, krill, or micro-algae oil by LC-MS. All supplements contained the declared amount of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA); however, their content per capsule and the concentration of other fatty acids varied strongly. Krill oil contained the highest total n3 oxylipin concentration (6000 nmol/g) and the highest degree of oxidation (EPA 0.7%; DHA 1.3%), while micro-algae oil (

Fetal bovine serum (FBS) has been used as a universal supplement in cell culture for more than six decades. This includes the investigation of lipid and lipid mediator formation and biology. Little is known about the (polyunsaturated) fatty acid composition and their oxidation products in FBS. Therefore, we analyzed six different FBS purchased from three different companies regarding their fatty acid and oxylipin concentrations. We found pronounced differences in the fatty acid concentrations. Even two batches of "standardized" FBS batches from one company showed drastic differences (e.g., for eicosapentaenoic acid 5 ± 1 μM vs. 11 ± 1 μM). Oxylipin concentrations also markedly differ between the FBS lots. The highest differences were found for 12-lipoxygenase products (e.g., 12-hydroxyeicosatetraenoic acid free 21-87 nM and total 58-108 nM), probably due to inconsistent serum generation procedures. Our results indicate that for cell culture studies dealing with lipid metabolism, researchers should carefully characterize their used FBS to ensure reliability and reproducibility of study outcomes.

OBJECTIVES: Neuroinflammation contributes to brain-aging which may be mitigated by anti-inflammatory oxylipins. Based on our previous findings that a 6% walnut-enriched diet alone, and additional physical activity (PA), enhanced cognition in 18 months old NMRI, we now investigated the effects of this diet on oxylipin- and inflammatory marker levels in liver and brain. METHODS: 18 months and 3 months old female NMRI mice were fed with a 6% walnut-enriched diet. Oxylipins were determined in brain and liver sections using LC-MS. Expression of IL1β gene was determined by qRT-PCR. RESULTS: The walnut diet compensates for the age related increase in IL1β gene expression in the liver of mice, whereas expression in the brain was not affected. Basal levels of oxylipins in brain and liver samples isolated from young mice were generally lower compared to aged mice. The walnut diet further increased oxylipin levels of walnut specific fatty acids in liver and brain of aged mice. Enrichment of linoleic acid (LA) and α-linolenic acid (ALA) derived oxylipin levels were quantitatively higher in the liver compared to the brain (P

International audience; Oxidized unsaturated fatty acids - i.e. eicosanoids and other oxylipins - are lipid mediators involved in the regulation of numerous physiological functions such as inflammation, blood coagulation, vascular tone and endothelial permeability. They have raised strong interest in clinical lipidomics in order to understand their role in health and diseases and their use as biomarkers. However, before the clinical translation, it is crucial to validate the analytical reliability of oxylipins. This notably requires to assess the putative artificial formation or degradation of oxylipins by (unsuitable) blood handling during plasma generation, storage and sample preparation. Using a liquid chromatography-mass spectrometry method covering 133 oxylipins we comprehensively analyzed the total (free + esterified) oxylipin profile in plasma and investigated the influence of i) addition of additives during sample preparation, ii) different storage times and temperatures during the transitory stage of plasma generation and iii) long-term storage of plasma samples at -80 degrees C. Addition of radical scavenger butylated hydroxytoluene reduced the apparent concentrations of hydroxy-PUFA and thus should be added to the samples at the beginning of sample preparation. The concentrations of all oxylipin classes remained stable (within analytical variance of 20%) during the transitory stage of plasma generation up to 24 h at 4 degrees C or 4 h at 20 degrees C before centrifugation of EDTA-whole blood and up to 5 days at - 20 degrees C after plasma separation. The variations in oxylipin concentrations did not correlate with storage time, storage temperature or stage of plasma generation. A significant increase of potentially lipoxygenase derived hydroxy-PUFA compared to immediate processing was only detected when samples were stored for longer times before centrifugation, plasma separation as well as freezing of plasma revealing residual enzymatic activity. Autoxidative rather than enzymatic processes led to a slightly increased concentration of 9-HETE when plasma samples were stored at - 80 degrees C for 15 months. Overall, we demonstrate that total plasma oxylipins are robust regarding delays during plasma generation and long-term storage at -80 degrees C supporting the application of oxylipin profiling in clinical research.

Objectives:We sought to assess whether the GlideScope Ranger video laryngoscope may be a reliable alternative to direct laryngoscopy in the prehospital setting.Design:Multicenter, prospective, randomized, control trial with patient recruitment over 18 months.Setting:Four study centers operating phys

A major part of oxygenated metabolites of polyunsaturated fatty acids - i.e. eicosanoids and other oxylipins - in biological samples is found in the esterified form. Yet, their biological role is only poorly understood. For quantification of esterified oxylipins in biological samples current protocols mostly apply alkaline hydrolysis with or without prior lipid extraction to release oxylipins into their free form which can be subsequently quantified via liquid chromatography-mass spectrometry. Herein, a detailed protocol for precise and reproducible quantification of esterified oxylipins in plasma is presented comprising i) extraction of lipids and removal of proteins with iso-propanol, ii) base hydrolysis with potassium hydroxide to saponify lipids and iii) solid phase extraction of the liberated oxylipins on C8/anion exchange mixed mode material. Unequal extraction of internal standards and lipid classes during lipid extraction before hydrolysis led to distorted concentrations, emphasizing that the choice of solvent used in this step is important to minimize discrimination. Regarding the hydrolysis conditions, at least 30 min incubation at 60 °C is required with 0.1 M KOH in sample. Drying of the SPE cartridges is a critical parameter since autoxidation processes of PUFA, which are present in high concentrations after cleavage, lead to artificial formation of epoxy fatty acids. With the developed protocol, inter-day, intra-day and inter-operator variance was21% for most oxylipins including hydroxy-, dihydroxy-, and epoxy-PUFA. The applicability of the developed methodology is demonstrated by investigating the changes in the oxylipin pattern following omega-3 fatty acid feeding to rats.

Die Rechtsordnungen in Europa entscheiden sich bei der Bestimmung der Mutterschaft gegen die Genetik und fur die Geburt. Traditionell ordnen sie das Kind abstammungsrechtlich der Frau zu, die es geboren hat. Auch wenn das Kind unter Verwendung einer fremden Eizelle entstanden ist und genetisch nicht von ihr abstammt, gilt, wie das deutsche Recht formuliert, der Satz: Mutter eines Kindes ist die Frau, die es geboren hat. Mit der Anknupfung an die Geburt bleibt es also bei der aus dem romischen Recht bekannten zweifelsfreien Statuszuordnung: mater semper certa est (Paul. D. 2.4.5). Die einmal erfolgte abstammungsrechtliche Zuordnung zur Mutter ist nach den europaischen Rechtsordnungen dann auch stabil. Anders als die Vaterschaft kann die Mutterschaft nicht infrage gestellt und, etwa auf eine Anfechtung hin, wieder aufgehoben werden.

The Pan European Phenology (PEP) project is a European infrastructure to promote and facilitate phenological research, education, and environmental monitoring. The main objective is to maintain and develop a Pan European Phenological database (PEP725) with an open, unrestricted data access for science and education. PEP725 is the successor of the database developed through the COST action 725 “Establishing a European phenological data platform for climatological applications” working as a single access point for European-wide plant phenological data. So far, 32 European meteorological services and project partners from across Europe have joined and supplied data collected by volunteers from 1868 to the present for the PEP725 database. Most of the partners actively provide data on a regular basis. The database presently holds almost 12 million records, about 46 growing stages and 265 plant species (including cultivars), and can be accessed via http://www.pep725.eu/ . Users of the PEP725 database have studied a diversity of topics ranging from climate change impact, plant physiological question, phenological modeling, and remote sensing of vegetation to ecosystem productivity.

AIMS OF THE STUDY: Nonmelanoma skin cancer (NMSC) is the most common cancer in Switzerland and Europe. The main causative factor is exposure to ultraviolet radiation, which puts outdoor workers in general at a higher risk of developing NMSC than indoor workers. However, few studies have clinically examined the risk of developing NMSC to outdoor workers, especially mountain guides. We aimed to investigate the prevalence of NMSC and corresponding precancerous lesions, and the associated risk behaviour of mountain and ski guides in order to develop future prevention programmes.METHODS: We conducted a cross-sectional study including mountain and ski guides from southern Germany, who underwent a full-body skin check-up by a dermatologist. We assessed their NMSC awareness and risk behaviour using a paper-based questionnaire.RESULTS: Of the 62 state-certified mountain and ski guides (55 men, 7 women; mean age 52.9 ± 13.4 years) included in this study, 27 (43.5%) were diagnosed with NMSC or its premalignant stages. In addition, 59.7% of the participants expressed the opinion that their protection from ultraviolet radiation exposure needs to be improved; 83.6% requested further information on NMSC, and 48.5% had never undergone a skin check-up or consulted a dermatologist before.CONCLUSIONS: Mountain and ski guides are at a high risk for developing NMSC. Their unmet medical needs indicate an underestimation of NMSC prevalence, which is usually based on reports by insurance companies, and offer the chance for developing evidence-based awareness and prevention tools that can be promoted to individuals with other outdoor jobs.

The article concerns the situation of refugees and immigrants from countries outside the European Union /European Economic Area with regard to social cash benefits in Denmark. At present these immigrants are treated different than inhabitants of Danish origin since their social cash benefits are reduced to a considerable extent. The conditions of these immigrants are discussed from an international human rights perspective and the article applies two approaches: a poverty threshold approach and a non-discrimination approach. The author concludes that the reduced social cash benefits are in violation of Denmark’s human rights obligation under socio-economic as well as civil-political treaties. In this way the article confirms that human rights are indivisible, interrelated and interdependent. The circumstances in Denmark during the last 12–14 years with regard to immigrants’ rights to social cash benefits has made it natural and even necessary to consider the situation from a legal as well as a political perspective. Thus it seems that the problems in Denmark for immigrants with regard to social cash benefits will be solved by politicians in Parliament in a foreseeable future whereas the human rights machinery because of its sluggishness has only to a limited extent been able to demonstrate its effectiveness.

To urism and recreation are important economic factors which are directly connected to weather and climate of a specific destination. Based on the observation network of the Central Institute of Meteorology and Geodynamics of Austria (ZAMG), data of 37 stations has been collected and analysed for tourism and recreation purposes. The analysis was based on long term data sets which were processed in relevant ways for tourism and recreation, resulting in frequency diagrams of Physiologically Equivalent Temperature (PET) and precipitation. Additionally, we prepared the results according to the demands of tourism and recreation authorities and industry using the Climate-Tourism/Transfer-Information-Scheme (CTIS). Applying data from the regional climate models REMO and CLM we can provide information on future climate conditions in Austria’s recreation areas. We chose two different time slices (2021–2050, 2071–2100) and IPCC emission scenarios (A1B, B1). The data was processed based on the threshold factors which are included in the CTIS (e.g. thermal comfort, heat stress, cold stress, sunshine, etc.). For the time slice 2021–2050 only moderate changes can be expected. But for 2071–2100 one can observe a distinct decrease of cold stress and the skiing potential. On the other hand, moderate increases of thermal comfort, heat stress, sultriness and sunshine are expected. No tendencies can be seen in precipitation and wind conditions.

The relationship between heat stress and mortality in the federal state of Vienna (Austria) was analyzed from 1970 to 2007. Long-term trends of mortality data and short-term adaptation to heat stress were considered by two complex approaches. The evaluation is based on the human biometeorological parameter, physiologically equivalent temperature. The results revealed a significant impact of heat stress on the human health, with a significantly higher sensitivity on women compared to men. Additionally, higher risks of deaths due to cardiovascular and respiratory diseases were found. During the long period of 38 years, some significant decreases of the sensitivity were found, especially in the medium heat stress levels. This could indicate active processes of long-term adaptation to the increasing heat stress.

The potential development of heat-related mortality in the 21th century for Vienna (Austria) was assessed by the use of two regional climate models based on the IPCC emissions scenarios A1B and B1. Heat stress was described with the human-biometeorological index PET (Physiologically Equivalent Temperature). Based on the relation between heat stress and mortality in 1970-2007, we developed two approaches to estimate the increases with and without long-term adaptation. Until 2011-2040 no significant changes will take place compared to 1970-2000, but in the following decades heat-related mortality could increase up to 129% until the end of the century, if no adaptation takes place. The strongest increase occurred due to extreme heat stress (PET >or= 41 degrees C). With long-term adaptation the increase is less pronounced, but still notable. This encourages the requirement for additional adaptation measurements.

Several daily measurements of air temperature, relative humidity, wind velocity and mean cloud cover are required for the calculation of the Physiologically Equivalent Temperature (PET). Using data obtained from daily measurements at 7, 14 and 19 CET over the period from 1991 to 2000 of 201 Austrian climate stations, we analysed the thermal bioclimate in Austria. The calculated PET-values were compared with the results of another computation using synoptic data not only from Austria but also from neighbouring countries. The mean radiant temperature is rather complex, as it covers all relevant short and long-wave radiation fluxes and plays an important role in the energy balance of humans. Based on the individual current conditions, this parameter was calculated using the well-established ‘RayMan’ model, which was development based on guidelines of the German Engineering Society, i.e. VDI 3789 ( VDI, 1994 ). The mean radiant temperature is determined based on the mean cloud cover and the maximum possible global radiation at a certain time and location. Statistical and GIS methods were used to convert the point-data consisting of PET-values for individual climate stations into a continuous surface. PET was spatially interpolated through a multiple linear regression analysis on a given grid. The results reveal fundamental information which is of particular relevance to the tourism and recreation authorities. The results are also important for spas and wellness resorts.

We present a quantitative and qualitative analysis of the influence of altitude on plant phenology over an area, delimited by 46 to 49°N latitude and 5 to 15°E longitude, that includes major parts of the Alpine region for the period 1971-2000. Our attention is focused on flowering of widely distributed plants in Europe, including some important allergenic species. We calculated the dependence of phenological mean dates on altitude for different phenophases as a linear regres- sion. Results show a statistically significant linear dependence (p < 0.0001) for each phenophase, with regression coefficients that range from 0.92 ± 0.21 d (100 m) -1 altitude for beginning of flower- ing of Norway spruce to 4.56 ± 0.21 d (100 m) -1 for beginning of flowering of common hazel. Pheno- logical temporal trends and thermal temporal trends show a less strong dependence on altitude, predominantly non-significant. The most significant regression coefficients are negative for pheno- logical trends (beginning of flowering of common alder, -0.065 ± 0.028 d yr -1 (100 m) -1 , p = 0.0187, and Norway spruce, -0.049 ± 0.020 d yr -1 (100 m) -1 , p = 0.0167) and positive for temperature trends (February, 0.0017°C yr -1 (100 m) -1 , p < 0.0001). The significant linear dependence of February tem- perature trends on altitude induces an inverse significant linear dependence for phenological trends of flowering of common alder (occurring in March). The prevalence of negative mean pheno- logical trends suggests a stronger advance of flowering phases at higher altitude. Finally, a regional analysis suggests a tendency towards a stronger altitudinal response in the northern than in the southern Alps.

In a considerable number of cases the European Court of Human Rights has held that civil and political rights encompass (social) fulfilment elements, which are otherwise considered non-justiciable under the conventions on economic, social and cultural rights. This integrated approach has frequently been described in legal literature. However, a satisfactory normative explanation to the phenomenon has not been given, and we have not developed clear legal principles of guidance to treaty bodies when having to decide when and to what extent social facts can be considered legally relevant under the conventions on civil and political rights. The article applies a hermeneutic perspective in search of a better understanding of the integrated approach, the legal implications of the notion of the indivisibility of human rights and the justiciability of economic, social and cultural rights.

Global climate change impacts can already be tracked in many physical and biological systems; in particular, terrestrial ecosystems provide a consistent picture of observed changes. One of the preferred indicators is phenology, the science of natural recurring events, as their recorded dates provide a high-temporal resolution of ongoing changes. Thus, numerous analyses have demonstrated an earlier onset of spring events for mid and higher latitudes and a lengthening of the growing season. However, published single-site or single-species studies are particularly open to suspicion of being biased towards predominantly reporting climate change-induced impacts. No comprehensive study or meta-analysis has so far examined the possible lack of evidence for changes or shifts at sites where no temperature change is observed. We used an enormous systematic phenological network data set of more than 125 000 observational series of 542 plant and 19 animal species in 21 European countries (1971–2000). Our results showed that 78% of all leafing, flowering and fruiting records advanced (30% significantly) and only 3% were significantly delayed, whereas the signal of leaf colouring/fall is ambiguous. We conclude that previously published results of phenological changes were not biased by reporting or publication predisposition: the average advance of spring/summer was 2.5 days decade � 1 in Europe. Our analysis of 254 mean national time series undoubtedly demonstrates that species’ phenology is responsive to temperature of the preceding

The year to year variability and trends of animal phenological phases (honey bee, cockchafer, 3 butterfly species, swallow and cuckoo) of the Austrian phenological observational network were related to each other and to mean monthly temperatures over the time period 1951-1998. Insect phases were well correlated with each other (r 2 = 0.4 to 0.6) and with temperature (r 2 = 0.25 to 0.55), whereas both bird phases were only well correlated with each other (r 2 = 0.57), but showed low common variance values with temperature and with other animal phases. The slope of the temperature-pheno regression, also termed as temperature sensitivity of the phenological phase, was high in the case of the insect phases (-3 to -5 days/°C), but low in the cases of both bird phases (about -1 days/°C). All animal phenological time series showed a trend towards later occurrence dates. The trends of the bird phases were even significant (p