Ultra-performance liquid chromatography mass spectrometry and sensitive bioassay methods for quantification of posaconazole plasma concentrations after oral dosing.

Posaconazole (POS) is a new antifungal agent for prevention and therapy of mycoses in immunocompromised patients. Variable POS pharmacokinetics after oral dosing may influence efficacy: a trough threshold of 0.5 ?g/ml has been recently proposed. Measurement of POS plasma concentrations by complex chromatographic techniques may thus contribute to optimize prevention and management of life-threatening infections. No microbiological analytical method is available. The objective of this study was to develop and validate a new simplified ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method and a sensitive bioassay for quantification of POS over the clinical plasma concentration range. The UPLC-MS/MS equipment consisted of a triple quadrupole mass spectrometer, an electrospray ionization (ESI) source, and a C(18) analytical column. The Candida albicans POS-hypersusceptible mutant (MIC of 0.002 ?g/ml) ?cdr1 ?cdr2 ?flu ?mdr1 ?can constructed by targeted deletion of multidrug efflux transporters and calcineurin genes was used for the bioassay. POS was extracted from plasma by protein precipitation with acetonitrile-methanol (75%/25%, vol/vol). Reproducible standard curves were obtained over the range 0.014 to 12 (UPLC-MS/MS) and 0.028 to 12 ?g/ml (bioassay). Intra- and interrun accuracy levels were 106% ± 2% and 103% ± 4% for UPLC-MS/MS and 102% ± 8% and 104% ± 1% for bioassay, respectively. The intra- and interrun coefficients of variation were 7% ± 4% and 7% ± 3% for UPLC-MS/MS and 5% ± 3% and 4% ± 2% for bioassay, respectively. An excellent correlation between POS plasma concentrations measured by UPLC-MS/MS and bioassay was found (concordance, 0.96). In 26 hemato-oncological patients receiving oral POS, 27/69 (39%) trough plasma concentrations were lower than 0.5 ?g/ml. The UPLC-MS/MS method and sensitive bioassay offer alternative tools for accurate and precise quantification of the plasma concentrations in patients receiving oral posaconazole.

Therapeutic concepts in adult and paediatric eosinophilic oesophagitis.

Eosinophilic oesophagitis (EoE) was first described in the early 1990s. Although initially reported to be a rare entity, EoE has rapidly become a regularly diagnosed disease with a prevalence of approximately 1 in 2,000 individuals in the USA and Europe. The disease is characterized by a combination of oesophageal dysfunction and predominant eosinophilic infiltration of the oesophageal tissue. At diagnosis, other diseases that can be associated with oesophageal eosinophilic infiltration must be ruled out. Children with EoE present with a wide variety of symptoms, whereas adults mostly present with dysphagia for solid food and chest pain. Histologic features of EoE resemble those of T-helper type 2 inflammation. Endoscopy should be carried out to establish the diagnosis, but endoscopic abnormalities are not pathognomonic for EoE and the examination might not show histologic abnormality. Treatment modalities for EoE include drugs (corticosteroids, PPIs, antiallergic and biologic agents), hypoallergenic diets and oesophageal dilatation for strictures that are unresponsive to medical therapy. Unresolved eosinophilic inflammation leads to the formation of oesophageal strictures, which probably increase the risk of food bolus impactions. To date, long-term strategies for the therapeutic management of this chronic inflammatory disease remain poorly defined.

Analytical methods for the quantitative determination of selective serotonin reuptake inhibitors for therapeutic drug monitoring purposes in patients.

Five selective serotonin reuptake inhibitors (SSRIs) have been introduced recently: citalopram, fluoxetine, fluvoxamine, paroxetine and sertraline. Although no therapeutic window has been defined for SSRIs, in contrast to tricyclic antidepressants, analytical methods for therapeutic drug monitoring of SSRIs are useful in several instances. SSRIs differ widely in their chemical structure and in their metabolism. The fact that some of them have N-demethylated metabolites, which are also SSRIs, requires that methods be available which allow therapeutic drug monitoring of the parent compounds and of these active metabolites. most procedures are based on prepurification of the SSRIs by liquid-liquid extraction before they are submitted to separation by chromatographic procedures (high-performance liquid chromatography, gas chromatography, thin layer chromatography) and detection by various detectors (UV, fluorescence, electrochemical detector, nitrogen-phosphorus detector, mass spectrometry). This literature review shows that most methods allow quantitative determination of SSRIs in plasma, in the lower ng/ml range, and that they are, therefore, suitable for therapeutic drug monitoring purposes of this category of drugs.

Fast gas chromatography and negative-ion chemical ionization tandem mass spectrometry for forensic analysis of cannabinoids in whole blood.

The present work describes a fast gas chromatography/negative-ion chemical ionization tandem mass spectrometric assay (Fast GC/NICI-MS/MS) for analysis of tetrahydrocannabinol (THC), 11-hydroxy-tetrahydrocannabinol (THC-OH) and 11-nor-9-carboxy-tetrahydrocannabinol (THC-COOH) in whole blood. The cannabinoids were extracted from 500 microL of whole blood by a simple liquid-liquid extraction (LLE) and then derivatized by using trifluoroacetic anhydride (TFAA) and hexafluoro-2-propanol (HFIP) as fluorinated agents. Mass spectrometric detection of the analytes was performed in the selected reaction-monitoring mode on a triple quadrupole instrument after negative-ion chemical ionization. The assay was found to be linear in the concentration range of 0.5-20 ng/mL for THC and THC-OH, and of 2.5-100 ng/mL for THC-COOH. Repeatability and intermediate precision were found less than 12% for all concentrations tested. Under standard chromatographic conditions, the run cycle time would have been 15 min. By using fast conditions of separation, the assay analysis time has been reduced to 5 min, without compromising the chromatographic resolution. Finally, a simple approach for estimating the uncertainty measurement is presented.

The Effect of Adding CO2 to Hypoxic Inspired Gas on Cerebral Blood Flow Velocity and Breathing during Incremental Exercise

Hypoxia increases the ventilatory response to exercise, which leads to hyperventilation-induced hypocapnia and subsequent reduction in cerebral blood flow (CBF). We studied the effects of adding CO2 to a hypoxic inspired gas on CBF during heavy exercise in an altitude naïve population. We hypothesized that augmented inspired CO2 and hypoxia would exert synergistic effects on increasing CBF during exercise, which would improve exercise capacity compared to hypocapnic hypoxia. We also examined the responsiveness of CO2 and O2 chemoreception on the regulation ventilation (E) during incremental exercise. We measured middle cerebral artery velocity (MCAv; index of CBF), E, end-tidal PCO2, respiratory compensation threshold (RC) and ventilatory response to exercise (E slope) in ten healthy men during incremental cycling to exhaustion in normoxia and hypoxia (FIO2 = 0.10) with and without augmenting the fraction of inspired CO2 (FICO2). During exercise in normoxia, augmenting FICO2 elevated MCAv throughout exercise and lowered both RC onset andE slope below RC (P<0.05). In hypoxia, MCAv and E slope below RC during exercise were elevated, while the onset of RC occurred at lower exercise intensity (P<0.05). Augmenting FICO2 in hypoxia increased E at RC (P<0.05) but no difference was observed in RC onset, MCAv, or E slope below RC (P>0.05). The E slope above RC was unchanged with either hypoxia or augmented FICO2 (P>0.05). We found augmenting FICO2 increased CBF during sub-maximal exercise in normoxia, but not in hypoxia, indicating that the 'normal' cerebrovascular response to hypercapnia is blunted during exercise in hypoxia, possibly due to an exhaustion of cerebral vasodilatory reserve. This finding may explain the lack of improvement of exercise capacity in hypoxia with augmented CO2. Our data further indicate that, during exercise below RC, chemoreception is responsive, while above RC the ventilatory response to CO2 is blunted.

Contribution to the Development of the LHCb Vertex Locator Readout Electronics

L'expérience LHCb sera installée sur le futur accélérateur LHC du CERN. LHCb est un spectromètre à un bras consacré aux mesures de précision de la violation CP et à l'étude des désintégrations rares des particules qui contiennent un quark b. Actuellement LHCb se trouve dans la phase finale de recherche et développement et de conception. La construction a déjà commencé pour l'aimant et les calorimètres. Dans le Modèle Standard, la violation CP est causée par une phase complexe dans la matrice 3x3 CKM (Cabibbo-Kobayashi-Maskawa) de mélange des quarks. L'expérience LHCb compte utiliser les mesons B pour tester l'unitarité de cette matrice, en mesurant de diverses manières indépendantes tous les angles et côtés du "triangle d'unitarité". Cela permettra de surdéterminer le modèle et, peut-être, de mettre en évidence des incohérences qui seraient le signal de l'existence d'une physique au-delà du Modèle Standard. La reconstruction du vertex de désintégration des particules est une condition fondamentale pour l'expérience LHCb. La présence d'un vertex secondaire déplacé est une signature de la désintégration de particules avec un quark b. Cette signature est utilisée dans le trigger topologique du LHCb. Le Vertex Locator (VeLo) doit fournir des mesures précises de coordonnées de passage des traces près de la région d'interaction. Ces points sont ensuite utilisés pour reconstruire les trajectoires des particules et l'identification des vertices secondaires et la mesure des temps de vie des hadrons avec quark b. L'électronique du VeLo est une partie essentielle du système d'acquisition de données et doit se conformer aux spécifications de l'électronique de LHCb. La conception des circuits doit maximiser le rapport signal/bruit pour obtenir la meilleure performance de reconstruction des traces dans le détecteur. L'électronique, conçue en parallèle avec le développement du détecteur de silicium, a parcouru plusieurs phases de "prototyping" décrites dans cette thèse.<br/><br/>The LHCb experiment is being built at the future LHC accelerator at CERN. It is a forward single-arm spectrometer dedicated to precision measurements of CP violation and rare decays in the b quark sector. Presently it is finishing its R&D and final design stage. The construction already started for the magnet and calorimeters. In the Standard Model, CP violation arises via the complex phase of the 3 x 3 CKM (Cabibbo-Kobayashi-Maskawa) quark mixing matrix. The LHCb experiment will test the unitarity of this matrix by measuring in several theoretically unrelated ways all angles and sides of the so-called "unitary triangle". This will allow to over-constrain the model and - hopefully - to exhibit inconsistencies which will be a signal of physics beyond the Standard Model. The Vertex reconstruction is a fundamental requirement for the LHCb experiment. Displaced secondary vertices are a distinctive feature of b-hadron decays. This signature is used in the LHCb topology trigger. The Vertex Locator (VeLo) has to provide precise measurements of track coordinates close to the interaction region. These are used to reconstruct production and decay vertices of beauty-hadrons and to provide accurate measurements of their decay lifetimes. The Vertex Locator electronics is an essential part of the data acquisition system and must conform to the overall LHCb electronics specification. The design of the electronics must maximise the signal to noise ratio in order to achieve the best tracking reconstruction performance in the detector. The electronics is being designed in parallel with the silicon detector development and went trough several prototyping phases, which are described in this thesis.

Solid and liquid biofuels markets in Finland - a study on international biofuels trade

This study considered the current situation of solid and liquid biofuels markets and international biofuels trade in Finland and identified the challenges ofthe emerging international biofuels markets for Finland. The fact that industryconsumes more than half of the total primary energy, widely applied combined heat and power production (CHP) and a high share of biofuels in the total energy consumption are specific to the Finnish energy system. One third of the electricity is generated in CHP plants. As much as 27% of the total energy consumption ismet by using wood and peat, which makes Finland the leading country in the use of biofuels. Finland has made a commitment to maintain greenhouse gas emissions at the 1990 level at the highest during the period 2008-2012. The Finnish energypolicy aims to achieve the target, and a variety of measures are taken to promote the use of renewable energy sources and especially wood fuels. In this study, the wooden raw material streams of the forest industry were included the international biofuels trade in addition to biomass streams that are traded for energy production. In 2004, as much as 45% of the raw wood importedinto Finland ended up in energy production. The total international trading of biofuels was evaluated at 72 PJ, of which the majority, 58 PJ, was raw wood. About 22% of wood based energy in Finland originated from imported raw wood. Tall oil and wood pellets composed the largest export streams of biofuels. The annual turnover of international biofuels trade was estimated at about ¤ 90 million fordirect trade and at about ¤ 190 million for indirect trade. The forest industryas the biggest user of wood, and the producer and user of wood fuels has a central position in biomass and biofuels markets in Finland. Lately, the international aspects of Finnish biofuels markets have been emphasised as the import of rawwood and the export of wood pellets have increased. Expanding the use of biofuels in the road transportation sector would increase the international streams ofbiofuels in Finland. In coming years, the international trading of biomass for energy purposes can be expected to continue growing.

Diffusion-weighted MR imaging in musculoskeletal diseases: Current concepts.

MR imaging is currently regarded as a pivotal technique for the assessment of a variety of musculoskeletal conditions. Diffusion-weighted MR imaging (DWI) is a relatively recent sequence that provides information on the degree of cellularity of lesions. Apparent diffusion coefficient (ADC) value provides information on the movement of water molecules outside the cells. The literature contains many studies that have evaluated the role of DWI in musculoskeletal diseases. However, to date they yielded conflicting results on the use and the diagnostic capabilities of DWI in the area of musculoskeletal diseases. However, many of them have showed that DWI is a useful technique for the evaluation of the extent of the disease in a subset of musculoskeletal cancers. In terms of tissue characterization, DWI may be an adjunct to the more conventional MR imaging techniques but should be interpreted along with the signal of the lesion as observed on conventional sequences, especially in musculoskeletal cancers. Regarding the monitoring of response to therapy in cancer or inflammatory disease, the use of ADC value may represent a more reliable additional tool but must be compared to the initial ADC value of the lesions along with the knowledge of the actual therapy.

AltitudeOmics : impaired pulmonary gas exchange efficiency and blunted ventilatory acclimatization in humans with patent foramen ovale after 16 days at 5,260 m.

A patent foramen ovale (PFO), present in ∼40% of the general population, is a potential source of right-to-left shunt that can impair pulmonary gas exchange efficiency [i.e., increase the alveolar-to-arterial Po2 difference (A-aDO2)]. Prior studies investigating human acclimatization to high-altitude with A-aDO2 as a key parameter have not investigated differences between subjects with (PFO+) or without a PFO (PFO-). We hypothesized that in PFO+ subjects A-aDO2 would not improve (i.e., decrease) after acclimatization to high altitude compared with PFO- subjects. Twenty-one (11 PFO+) healthy sea-level residents were studied at rest and during cycle ergometer exercise at the highest iso-workload achieved at sea level (SL), after acute transport to 5,260 m (ALT1), and again at 5,260 m after 16 days of high-altitude acclimatization (ALT16). In contrast to PFO- subjects, PFO+ subjects had 1) no improvement in A-aDO2 at rest and during exercise at ALT16 compared with ALT1, 2) no significant increase in resting alveolar ventilation, or alveolar Po2, at ALT16 compared with ALT1, and consequently had 3) an increased arterial Pco2 and decreased arterial Po2 and arterial O2 saturation at rest at ALT16. Furthermore, PFO+ subjects had an increased incidence of acute mountain sickness (AMS) at ALT1 concomitant with significantly lower peripheral O2 saturation (SpO2). These data suggest that PFO+ subjects have increased susceptibility to AMS when not taking prophylactic treatments, that right-to-left shunt through a PFO impairs pulmonary gas exchange efficiency even after acclimatization to high altitude, and that PFO+ subjects have blunted ventilatory acclimatization after 16 days at altitude compared with PFO- subjects.

Use of the dried blood spot sampling process coupled with fast gas chromatography and negative-ion chemical ionization tandem mass spectrometry: application to fluoxetine, norfluoxetine, reboxetine, and paroxetine analysis.

The objective of this work was to combine the advantages of the dried blood spot (DBS) sampling process with the highly sensitive and selective negative-ion chemical ionization tandem mass spectrometry (NICI-MS-MS) to analyze for recent antidepressants including fluoxetine, norfluoxetine, reboxetine, and paroxetine from micro whole blood samples (i.e., 10 microL). Before analysis, DBS samples were punched out, and antidepressants were simultaneously extracted and derivatized in a single step by use of pentafluoropropionic acid anhydride and 0.02% triethylamine in butyl chloride for 30 min at 60 degrees C under ultrasonication. Derivatives were then separated on a gas chromatograph coupled with a triple-quadrupole mass spectrometer operating in negative selected reaction monitoring mode for a total run time of 5 min. To establish the validity of the method, trueness, precision, and selectivity were determined on the basis of the guidelines of the "Société Française des Sciences et des Techniques Pharmaceutiques" (SFSTP). The assay was found to be linear in the concentration ranges 1 to 500 ng mL(-1) for fluoxetine and norfluoxetine and 20 to 500 ng mL(-1) for reboxetine and paroxetine. Despite the small sampling volume, the limit of detection was estimated at 20 pg mL(-1) for all the analytes. The stability of DBS was also evaluated at -20 degrees C, 4 degrees C, 25 degrees C, and 40 degrees C for up to 30 days. Furthermore, the method was successfully applied to a pharmacokinetic investigation performed on a healthy volunteer after oral administration of a single 40-mg dose of fluoxetine. Thus, this validated DBS method combines an extractive-derivative single step with a fast and sensitive GC-NICI-MS-MS technique. Using microliter blood samples, this procedure offers a patient-friendly tool in many biomedical fields such as checking treatment adherence, therapeutic drug monitoring, toxicological analyses, or pharmacokinetic studies.

Blood monitoring of perfluorocarbon compounds (F-tert-butylcyclohexane, perfluoromethyldecalin and perfluorodecalin) by headspace-gas chromatography-tandem mass spectrometry.

A headspace-gas chromatography-tandem mass spectrometry (HS-GC-MS/MS) method for the trace measurement of perfluorocarbon compounds (PFCs) in blood was developed. Due to oxygen carrying capabilities of PFCs, application to doping and sports misuse is speculated. This study was therefore extended to perform validation methods for F-tert-butylcyclohexane (Oxycyte(®)), perfluoro(methyldecalin) (PFMD) and perfluorodecalin (PFD). The limit of detection of these compounds was established and found to be 1.2µg/mL blood for F-tert-butylcyclohexane, 4.9µg/mL blood for PFMD and 9.6µg/mL blood for PFD. The limit of quantification was assumed to be 12µg/mL blood (F-tert-butylcyclohexane), 48µg/mL blood (PFMD) and 96µg/mL blood (PFD). HS-GC-MS/MS technique allows detection from 1000 to 10,000 times lower than the estimated required dose to ensure a biological effect for the investigated PFCs. Thus, this technique could be used to identify a PFC misuse several hours, maybe days, after the injection or the sporting event. Clinical trials with those compounds are still required to evaluate the validation parameters with the calculated estimations.

CT attenuation values of blood and myocardium: rationale for accurate coronary artery calcifications detection with multi-detector CT.

OBJECTIVES: To determine inter-session and intra/inter-individual variations of the attenuations of aortic blood/myocardium with MDCT in the context of calcium scoring. To evaluate whether these variations are dependent on patients' characteristics. METHODS: Fifty-four volunteers were evaluated with calcium scoring non-enhanced CT. We measured attenuations (inter-individual variation) and standard deviations (SD, intra-individual variation) of the blood in the ascending aorta and of the myocardium of left ventricle. Every volunteer was examined twice to study the inter-session variation. The fat pad thickness at the sternum and noise (SD of air) were measured too. These values were correlated with the measured aortic/ventricular attenuations and their SDs (Pearson). Historically fixed thresholds (90 and 130 HU) were tested against different models based on attenuations of blood/ventricle. RESULTS: The mean attenuation was 46 HU (range, 17-84 HU) with mean SD 23 HU for the blood, and 39 HU (10-82 HU) with mean SD 18 HU for the myocardium. The attenuation/SD of the blood were significantly higher than those of the myocardium (p < 0.01). The inter-session variation was not significant. There was a poor correlation between SD of aortic blood/ventricle with fat thickness/noise. Based on existing models, 90 HU threshold offers a confidence interval of approximately 95% and 130 HU more than 99%. CONCLUSIONS: Historical thresholds offer high confidence intervals for exclusion of aortic blood/myocardium and by the way for detecting calcifications. Nevertheless, considering the large variations of blood/myocardium CT values and the influence of patient's characteristics, a better approach might be an adaptive threshold.

Analytical Strategies for Doping Control Purposes: Needs, Challenges, and Perspectives.

The fight against doping in sports has been governed since 1999 by the World Anti-Doping Agency (WADA), an independent institution behind the implementation of the World Anti-Doping Code (Code). The intent of the Code is to protect clean athletes through the harmonization of anti-doping programs at the international level with special attention to detection, deterrence and prevention of doping.1 A new version of the Code came into force on January 1st 2015, introducing, among other improvements, longer periods of sanctioning for athletes (up to four years) and measures to strengthen the role of anti-doping investigations and intelligence. To ensure optimal harmonization, five International Standards covering different technical aspects of the Code are also currently in force: the List of Prohibited Substances and Methods (List), Testing and Investigations, Laboratories, Therapeutic Use Exemptions (TUE) and Protection of Privacy and Personal Information. Adherence to these standards is mandatory for all anti-doping stakeholders to be compliant with the Code. Among these documents, the eighth version of International Standard for Laboratories (ISL), which also came into effect on January 1st 2015, includes regulations for WADA and ISO/IEC 17025 accreditations and their application for urine and blood sample analysis by anti-doping laboratories.2 Specific requirements are also described in several Technical Documents or Guidelines in which various topics are highlighted such as the identification criteria for gas chromatography (GC) and liquid chromatography (LC) coupled to mass spectrometry (MS) techniques (IDCR), measurements and reporting of endogenous androgenic anabolic agents (EAAS) and analytical requirements for the Athlete Biological Passport (ABP).