Discrimination of Diptera order insects based on their saturated cuticular hydrocarbon content using a new microextraction procedure and chromatographic analysis.

The nature and proportions of hydrocarbons in the cuticle of insects are characteristic of the species and age. Chemical analysis of cuticular hydrocarbons allows species discrimination, which is of great interest in the forensic field, where insects play a crucial role in estimating the minimum post-mortem interval. The objective of this work was the differentiation of Diptera order insects through their saturated cuticular hydrocarbon compositions (SCHCs). For this, specimens fixed in 70 : 30 ethanol : water, as recommended by the European Association for Forensic Entomology, were submitted to solid-liquid extraction followed by dispersive liquid-liquid microextraction, providing preconcentration factors up to 76 for the SCHCs. The final organic extract was analysed by gas chromatography coupled with flame ionization detection (GC-FID), and GC coupled with mass spectrometry was applied to confirm the identity of the SCHCs. The analysed samples contained linear alkanes with the number of carbon atoms in the C9-C15 and C18-C36 ranges with concentrations between 0.1 and 125 ng g-1. Chrysomya albiceps (in its larval stage) showed the highest number of analytes detected, with 21 compounds, while Lucilia sericata and Calliphora vicina the lowest, with only 3 alkanes. Non-supervised principal component analysis and supervised orthogonal partial least squares discriminant analysis were performed and an optimal model to differentiate specimens according to their species was obtained. In addition, statistically significant differences were observed in the concentrations of certain SCHCs within the same species depending on the stage of development or the growth pattern of the insect.

Dosage Individualization of Linezolid: Precision Dosing of Linezolid To Optimize Efficacy and Minimize Toxicity.

The high interindividual variability in the pharmacokinetics (PK) of linezolid has been described, which results in an unacceptably high proportion of patients with either suboptimal or potentially toxic concentrations following the administration of a fixed regimen. The aim of this study was to develop a population pharmacokinetic model of linezolid and use this to build and validate alogorithms for individualized dosing. A retrospective pharmacokinetic analysis was performed using data from 338 hospitalized patients (65.4% male, 65.5 [±14.6] years) who underwent routine therapeutic drug monitoring for linezolid. Linezolid concentrations were analyzed by using high-performance liquid chromatography. Population pharmacokinetic modeling was performed using a nonparametric methodology with Pmetrics, and Monte Carlo simulations were employed to calculate the 100% time >MIC after the administration of a fixed regimen of 600 mg administered every 12 h (q12h) intravenously (i.v.). The dose of linezolid needed to achieve a PTA ≥ 90% for all susceptible isolates classified according to EUCAST was estimated to be as high as 2,400 mg q12h, which is 4 times higher than the maximum licensed linezolid dose. The final PK model was then used to construct software for dosage individualization, and the performance of the software was assessed using 10 new patients not used to construct the original population PK model. A three-compartment model with an absorptive compartment with zero-order i.v. input and first-order clearance from the central compartment best described the data. The dose optimization software tracked patients with a high degree of accuracy. The software may be a clinically useful tool to adjust linezolid dosages in real time to achieve prespecified drug exposure targets. A further prospective study is needed to examine the potential clinical utility of individualized therapy.

Population Pharmacokinetics of Anidulafungin in Critically Ill Patients.

A two-compartment pharmacokinetic (PK) population model of anidulafungin was fitted to PK data from 23 critically ill patients (age, 65 years [range, 28 to 81 years]; total body weight [TBW], 75 kg [range, 54 to 168 kg]). TBW was associated with clearance and incorporated into a final population PK model. Simulations suggested that patients with higher TBWs had less-extensive MIC coverage. Dosage escalation may be warranted in patients with high TBWs to ensure optimal drug exposures for treatment of Candida albicans and Candida glabrata infections.

A simple device for headspace sorptive extraction prior to gas chromatography-mass spectrometry analysis.

A device for headspace sorptive extraction (HSSE) combined with gas chromatography separation and mass spectrometry detection (GC-MS) is presented. The gadget is based on a simple magnetic disk that permits the stir bar to be placed at the top of the sample vial. The complete surface of the coated stir bar is exposed to the headspace atmosphere, and the movement of the bar caused by the magnetic stirrer in the liquid sample facilitates the sorption process. The final consequence is increased sensitivity and repeatability when compared with a glass-made comercial device for the same purpose. Successful determination of ten chlorobenzenes in waters testifies the good performance of the new device.

In situ ionic liquid dispersive liquid-liquid microextraction coupled to gas chromatography-mass spectrometry for the determination of organophosphorus pesticides.

Nine organophosphorus pesticides (OPPs) were determined in environmental waters from different origins using in situ ionic liquid dispersive liquid microextraction (IL-DLLME). This preconcentration technique was coupled to gas chromatography-mass spectrometry (GC-MS) using microvial insert thermal desorption, an approach that uses a thermal desorption injector as sample introduction system. The parameters affecting both the microextraction and sample injection steps were optimized. The proposed method showed good precision, with RSD values ranging from 4.1 to 9.7%, accuracy with recoveries in the 85-118% range, and sensitivity with DLs ranging from 5 to 16 ng L-1.

Glyoxal and methylglyoxal as urinary markers of diabetes. Determination using a dispersive liquid-liquid microextraction procedure combined with gas chromatography-mass spectrometry.

Glyoxal (GO) and methylglyoxal (MGO) are α-oxoaldehydes that can be used as urinary diabetes markers. In this study, their levels were measured using a sample preparation procedure based on salting-out assisted liquid-liquid extraction (SALLE) and dispersive liquid-liquid microextraction (DLLME) combined with gas chromatography-mass spectrometry (GC-MS). The effect of the derivatization reaction with 2,3-diaminonaphthalene, the addition of acetonitrile and sodium chloride to urine, and the DLLME step using the acetonitrile extract as dispersant solvent and carbon tetrachloride as extractant solvent were carefully optimized. Quantification was performed by the internal standard method, using 5-bromo-2-chloroanisole. The intraday and interday precisions were lower than 6%. Limits of detection were 0.12 and 0.06ngmL-1, and enrichment factors 140 and 130 for GO and MGO, respectively. The concentrations of these α-oxoaldehydes in urine were between 0.9 and 35.8ngg-1 levels (creatinine adjusted). A statistical comparison of the analyte contents of urine samples from non-diabetic and diabetic patients pointed to significant differences (P=0.046, 24 subjects investigated), particularly regarding MGO, which was higher in diabetic patients. The novelty of this study compared with previous procedures lies in the treatment of the urine sample by SALLE based on the addition of acetonitrile and sodium chloride to the urine. The DLLME procedure is performed with a sedimented drop of the extractant solvent, without a surfactant reagent, and using acetonitrile as dispersant solvent. Separation of the analytes was performed using GC-MS detection, being the analytes unequivocal identified. The proposed procedure is the first microextraction method applied to the analysis of urine samples from diabetic and non-diabetic patients that allows a clear differentiation between both groups using a simple analysis.

Direct sample introduction-gas chromatography-mass spectrometry for the determination of haloanisole compounds in cork stoppers.

A solventless analytical method is proposed for analyzing the compounds responsible for cork taint in cork stoppers. Direct sample introduction (DSI) is evaluated as a sample introduction system for the gas chromatography-mass spectrometry (GC-MS) determination of four haloanisoles (HAs) in cork samples. Several parameters affecting the DSI step, including desorption temperature and time, gas flow rate and other focusing parameters, were optimized using univariate and multivariate approaches. The proposed method shows high sensitivity and minimises sample handling, with detection limits of 1.6-2.6ngg-1, depending on the compound. The suitability of the optimized procedure as a screening method was evaluated by obtaining decision limits (CCα) and detection capabilities (CCß) for each analyte, which were found to be in 6.9-11.8 and 8.7-14.8ngg-1, respectively, depending on the compound. Twenty-four cork samples were analysed, and 2,4,6-trichloroanisole was found in four of them at levels between 12.6 and 53ngg-1.

Cloud point extraction and gas chromatography with direct microvial insert thermal desorption for the determination of haloanisoles in alcoholic beverages.

A sensitive analytical procedure for the determination of four haloanisoles (2,4,6 trichloroanisole, 2,4,6-tribromoanisole, 2,3,4,6-tetrachloroanisole and pentachloroanisole) related with cork taint defects in wines, in different types of alcoholic beverages has been developed. The analytes were extracted from the matrix samples by cloud point extraction (CPE) using Triton X-114 heated to 75°C, and the surfactant rich phase was separated by centrifugation. By means of direct microvial insert thermal desorption, 20µL of the CPE obtained extract was submitted to gas chromatography-mass spectrometry (GC-MS) analysis. The parameters affecting the CPE and microvial insert thermal desorption were optimized. Quantification was carried by matrix-matched calibration using an internal standard. Detection limits ranged between 12.9 and 20.8ngL(-1), depending on the compound, for beer and wine samples, whereas for whiskies values in the 46.3-48ngL(-1) range were obtained, since these samples were diluted for analysis. Recoveries for alcoholic beverages were in the 89-111% range, depending on the analyte and the sample.

Urinary micafungin levels are sufficient to treat urinary tract infections caused by Candida spp.

Six cases of patients diagnosed with urinary tract infection (UTI) successfully treated with micafungin are reported. Four were infected with fluconazole-resistant Candida spp. and two (with hepatic injury) were infected with fluconazole-sensitive Candida spp. Traditionally, echinocandins have not been considered for the treatment of UTIs. However, despite its low urinary excretion rate, therapeutic drug monitoring of micafungin urinary levels could be helpful in order to achieve optimal pharmacokinetic/pharmacodynamic (PK/PD) indices for treating UTIs caused by Candida spp. resistant to fluconazole.

Dispersive liquid-liquid microextraction for the determination of nitrophenols in soils by microvial insert large volume injection-gas chromatography-mass spectrometry.

A rapid and sensitive procedure for the determination of six NPs in soils by gas chromatography and mass spectrometry (GC-MS) is proposed. Ultrasound assisted extraction (UAE) is used for NP extraction from soil matrices to an organic solvent, while the environmentally friendly technique dispersive liquid-liquid microextraction (DLLME) is used for the preconcentration of the resulting UAE extracts. NPs were derivatized by applying an "in-situ" acetylation procedure, before being injected into the GC-MS system using microvial insert large volume injection (LVI). Several parameters affecting UAE, DLLME, derivatization and injection steps were investigated. The optimized procedure provided recoveries of 86-111% from spiked samples. Precision values of the procedure (expressed as relative standard deviation, RSD) lower than 12%, and limits of quantification ranging from 1.3 to 2.6ngg(-1), depending on the compound, were obtained. Twenty soil samples, obtained from military, industrial and agricultural areas, were analyzed by the proposed method. Two of the analytes were quantified in two of the samples obtained from industrial areas, at concentrations in the 4.8-9.6ngg(-1) range.

Plasma and peritoneal fluid population pharmacokinetics of micafungin in post-surgical patients with severe peritonitis.

OBJECTIVES: Limited information about the pharmacokinetics of micafungin in the peritoneal cavity is available. The aim of this study was to explore the pharmacokinetics/pharmacodynamics of micafungin in plasma and peritoneal fluid in post-surgical critically ill patients with proven or suspected intra-abdominal fungal infection. METHODS: Patients were administered 100 mg/day micafungin. Serial blood and peritoneal fluid samples were collected on day 1 and day 3 (steady-state) of treatment. Concentrations were determined by validated chromatography and were subject to a population pharmacokinetic analysis with Pmetrics(®). Monte Carlo simulations were performed for AUC0-24/MIC ratios in plasma. The PTA was calculated using AUC0-24/MIC cut-offs: 285 for Candida parapsilosis and 3000 for non-parapsilosis Candida spp. RESULTS: Ten patients were included; six were male. The median (range) age, APACHE II score and Mannheim peritonitis index were 72 (43-85) years, 15 (11-36) and 26 (8-37), respectively. On day 1, median (SD) penetration of micafungin into the peritoneal cavity was 30% (30%-40%). A three-compartment model adequately described the data. The mean (SD) estimates for clearance and volume of distribution of the central compartment were 1.27 (0.75) L/h and 9.26 (1.11) L, respectively. In most patients, the PTA in plasma was ≥ 90% for MICs of 0.008-0.016 mg/L for Candida spp. and 0.125-0.25 mg/L for C. parapsilosis. CONCLUSIONS: After the first dose, micafungin at 100 mg/day achieves pharmacokinetic/pharmacodynamic targets in plasma for Candida spp. and C. parapsilosis MICs of 0.008-0.016 and 0.125-0.25 mg/L, respectively.

Evaluation of three headspace sorptive extraction coatings for the determination of volatile terpenes in honey using gas chromatography-mass spectrometry.

Headspace sorptive extraction (HSSE) was used to preconcentrate seven monoterpenes (eucalyptol, linalool, menthol, geraniol, carvacrol, thymol and eugenol) for separation by gas chromatography and mass spectrometry (GC-MS). Three commercially available coatings for the stir bars, namely Polydimethylsiloxane (PDMS), polyacrilate (PA) and Ethylene glycol-silicone (EG-Silicone), were tested, and the influential parameters both in the adsorption and the thermal desorption steps were optimized. PDMS provided the best sensitivity for linalool, geraniol, menthol and eucalyptol, whereas EG-Silicone was best for extracting the phenolic monoterpenes studied. Considering the average obtained slopes from all compounds, PDMS pointed as the best option, and the analytical characteristics for the HSSE-TD-GC-MS method using this coating were obtained. Quantification of the samples was carried out by matrix-matched calibration using a synthetic honey. Detection limits ranged between 0.007 and 0.032 ng g(-1), depending on the compound. Twelve honey samples of different floral origins were analyzed using the HSSE-GC-MS method, the analytes being detected at concentrations up to 64 ng g(-1).

Direct sample introduction gas chromatography and mass spectrometry for the determination of phthalate esters in cleaning products.

A method using direct sample introduction (DSI) coupled to gas chromatography-mass spectrometry (GC-MS) is developed for the determination of six phthalate esters (dimethyl, diethyl, dibutyl, butylbenzyl, diethylhexyl and dioctyl phthalate) in cleaning products. The different variables involved in the DSI step, including venting time and temperature, vaporisation time and temperature, injector heating temperature and gas flow rate and pressure, were evaluated and optimised using Taguchi orthogonal arrays. The proposed method, using calibration against methanolic standards, showed good linearity in the 0.05-15 µg g(-1) range and good repeatability, with RSD values ranging from 3.5% to 5.7%. Quantification limits between 0.010 and 0.041 µg g(-1), depending on the compound, were attained, while recovery assays provided values from 83% to 115%. Twenty-seven cleaning products were analysed using the DSI-GC-MS method, being four phthalates (dimethyl, diethyl, dibutyl and diethylhexyl phthalate) found in fourteen of them at concentration levels in the 0.1-21 µg g(-1) range. Compared with the most common GC injection technique, which uses the split/splitless injector, the proposed DSI procedure provided larger peak areas and lower detection limits, as result of the greater injected volume and reduction in noise.

Plasma and cerebrospinal fluid concentrations of linezolid in neurosurgical critically ill patients with proven or suspected central nervous system infections.

Linezolid is a valuable treatment option for central nervous system (CNS) infections caused by multidrug-resistant Gram-positive micro-organisms. Data regarding its penetration into the CNS have shown wide variability. The aim of this study was to describe the population pharmacokinetics of linezolid in plasma and cerebrospinal fluid (CSF) in critically ill patients with external CSF drainage and proven or suspected CNS infections. This was an observational pharmacokinetic (PK) study in 11 critically ill patients with proven or suspected CNS infection receiving linezolid. Serial blood and CSF samples were taken and were subject to population PK analysis. The median (interquartile range) of AUC(0-12h) was 47.6 (17.9-58.6) mgh/L in plasma and 21.1 (18.8-30.4) mgh/L in CSF, with a median CSF/plasma ratio of 0.77. At pre-dose at steady state, a strong positive correlation was observed between linezolid concentrations in CSF and plasma (Spearman's rho=0.758; P=0.011). For a minimum inhibitory concentration (MIC) of 2 mg/L, the median AUC(0-24h)/MIC values in plasma and CSF were <80 in all patients. A three-compartment linear model was found to be most appropriate. The mean value for linezolid clearance was 16.6L/h and mean volume of distribution was 101.3 L. No covariate relationships could be supported on any of the parameters. Linezolid demonstrated good penetration into the CNS but high interindividual PK variability. Administration of higher than standard doses of linezolid and therapeutic drug monitoring should therefore be considered as options to optimise linezolid dosing in critically ill patients with CNS infections.

Use of headspace sorptive extraction coupled to gas chromatography-mass spectrometry for the analysis of volatile polycyclic aromatic hydrocarbons in herbal infusions.

A solvent-free method is described for the determination of 10 volatile polycyclic aromatic hydrocarbons (PAHs), considered as priority pollutants by the EU, in different herbal infusions using headspace sorptive extraction (HSSE) and gas chromatography-mass spectrometry (GC-MS). The parameters affecting both the extraction and thermal desorption steps in the HSSE were optimized by means of Plackett-Burman designs. Ten millilitres of the herbal infusion was submitted to the HSSE preconcentration in the presence of salt for 4h at 88 °C. The use of d(10)-phenanthrene as internal standard not only improved the repeatability of the method but allowed quantification of the samples against external aqueous standards. Detection limits ranged between 11 and 26 ng L(-1).

Headspace sorptive extraction for the detection of combustion accelerants in fire debris.

A novel method for separation and identification of ignitable liquid residues in fire debris by gas chromatography and mass spectrometry is presented. Preconcentration of the analytes was carried out using the simple headspace sorptive extraction (HSSE) technique. Polydimethylsiloxane stir bars were used as the enrichment phase, and parameters affecting both the adsorption and desorption stages were carefully optimized. Extraction was carried out at 50°C for 1h. Stir bars were desorbed thermally in the GC injection port, thus avoiding the use of organic solvents. The results for five ignitable liquids, including gasoline and diesel fuel, using HSSE were compared with those obtained with a solid-phase microextraction method, with HSSE appearing as a more sensitive alternative.

Dispersive liquid-liquid microextraction for the determination of three cytokinin compounds in fruits and vegetables by liquid chromatography with time-of-flight mass spectrometry.

The paper presents a novel approach for the determination of three cytokinin compounds, thidiazuron (TDZ), 1,3-diphenylurea (1,3-DPU) and forchlorfenuron (CPPU), in fruit and vegetables samples using liquid chromatography with electrospray ionization and time-of-flight mass spectrometry (LC-ESI-TOFMS). Analytes were extracted from the sample matrix with ethanol, and the extract, after dilution with water, was submitted to dispersive liquid-liquid microextraction (DLLME). Once acetonitrile and 1,2-dichloroethane had been selected as extraction and disperser solvents, respectively, the influence of the following experimental parameters was studied using a Plackett-Burman design: volume of extraction and disperser solvents, sample mass and time and speed of centrifugation. The best analytical conditions were 250 µL 1,2-dichloroethane, 1.5 mL acetonitrile, 5 g sample mass, and centrifugation at 3000 rpm for 3 min. The optimized method provided DLs in the range 0.02-0.05 ng g(-1), depending on the compound. Satisfactory recovery values between 89 and 106% were obtained for spiked samples (kiwifruit, watermelon, grape and tomato) in the 0.2-1.0 ng g(-1) concentration range, depending on the compound. None of the target analytes was detected in any of the samples analyzed.

Stir bar sorptive extraction with gas chromatography-mass spectrometry for the determination of resveratrol, piceatannol and oxyresveratrol isomers in wines.

A simple and highly sensitive procedure based on stir bar sorptive extraction coupled to gas chromatography-mass spectrometry by means of a thermal desorption unit (SBSE-TD-GC-MS) has been optimized for the determination of cis/trans isomers of resveratrol, piceatannol and oxyresveratrol in wine samples. Quantification of the cis-isomers was carried out by generating the standards from the corresponding trans-species once they had been preconcentrated on the SBSE extracting phase. The optimization of the acetylation derivatization, SBSE extraction and thermal desorption steps was investigated using Plackett-Burman designs, taking into account the high number of variables to be considered. The use of bisphenol F as internal standard allowed quantification of the samples against aqueous standards. Repeatability, expressed as relative standard deviation of 10 successive analyses was between 5% and 9%, confirming the high precision attained under the optimized conditions. Satisfactory recovery values of between 79% and 109% were obtained for spiked samples in the 0.2-1.0 µgL(-1) concentration range, depending on the compound. The main compound determined in the analyzed samples was trans-resveratrol, with concentrations in the range of 3-230 µgL(-1), depending on the type of wine.

Headspace sorptive extraction for the analysis of organotin compounds using thermal desorption and gas chromatography with mass spectrometry.

A method based on headspace sorptive extraction (HSSE) in combination with thermal desorption-gas chromatography-mass spectrometry (TD-GC-MS) has been developed for the simultaneous determination of six organotin compounds (OTCs), corresponding to mono- and di-substituted methyltin, butyltin and octyltin species. Several parameters affecting both the headspace extraction and thermal desorption steps were carefully optimized using multivariate designs. Analytes were derivatized by in situ ethylation with sodium tetraethylborate. The optimized method was applied to the analysis of water samples of different origins, as well as to checking the migration of the studied compounds from commercially available plastic containers to the adequate liquid simulant. Quantification was carried out against aqueous calibration curves using diphenyltin as internal standard, providing detection limits of between 1.7 and 7.0 ng(Sn) L(-1), depending on the compound, and repeatabilities lower than 10% in terms of relative standard deviation. The applicability of the method was assessed by means of recovery studies and satisfactory values for all compounds were attained. The release of OTCs from the tested packages to the liquid simulant was confirmed, concentrations as high as 2.4 µg(Sn) L(-1) being found for dioctyltin. Even though the proposed method was developed for organotin halides, its application to an organotin ester shows its suitability for determining these compounds in migration assays.

The value of selected in vitro and in silico methods to predict acute oral toxicity in a regulatory context: results from the European Project ACuteTox.

ACuteTox is a project within the 6th European Framework Programme which had as one of its goals to develop, optimise and prevalidate a non-animal testing strategy for predicting human acute oral toxicity. In its last 6 months, a challenging exercise was conducted to assess the predictive capacity of the developed testing strategies and final identification of the most promising ones. Thirty-two chemicals were tested blind in the battery of in vitro and in silico methods selected during the first phase of the project. This paper describes the classification approaches studied: single step procedures and two step tiered testing strategies. In summary, four in vitro testing strategies were proposed as best performing in terms of predictive capacity with respect to the European acute oral toxicity classification. In addition, a heuristic testing strategy is suggested that combines the prediction results gained from the neutral red uptake assay performed in 3T3 cells, with information on neurotoxicity alerts identified by the primary rat brain aggregates test method. Octanol-water partition coefficients and in silico prediction of intestinal absorption and blood-brain barrier passage are also considered. This approach allows to reduce the number of chemicals wrongly predicted as not classified (LD50>2000 mg/kg b.w.).