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Objectives: Maternal self-reported ethnicity is recognised as a risk factor for pre-eclampsia in clinical screening tools and models. This study investigated whether ethnicity is acting as a proxy for genetic variants in this context., Study Design: A total of 436 women from multi-ethnic backgrounds recruited to two UK observational pregnancy hypertension cohort studies were genotyped. Genetically-computed individual ancestry estimates were calculated for each individual through comparison to the multi-ethnic 1000 Genomes reference panel genotypes. Regression models for pre-eclampsia using clinical risk factors including self-reported ethnicity with and without ancestry estimates were built and compared using Likelihood Ratio Tests (LRT)., Main Outcome Measures: Pre-eclampsia (early- and late-onset)., Results: In these multi-ethnic cohorts (mean age 34.9 years; 41.3 % White, 34.2 % Black, 13.1 % Asian ethnic backgrounds; 82.6 % chronic hypertension), discrepancies between self-reported ethnicity and genetically-computed individual ancestry estimates were present in all ethnic groups, particularly minority groups. Genetically-computed pan-African ancestry percentage was associated with early-onset (< 34 weeks) pre-eclampsia in adjusted models (aOR 100 % vs 0 % African ancestry: 3.81, 95 % CI 1.04-14.14, p-value 0.044) independently of self-reported ethnicity and established clinical risk factors. Addition of genetically-computed African ancestry to a clinical risk factor model including self-reported ethnicity, improved model fit (Likelihood ratio test p-value 0.023)., Conclusions: Self-reported maternal ethnicity is an imperfect proxy for genetically-computed individual ancestry estimates, particularly in ethnic minority groups. Genetically-computed African ancestry percentage was associated with early-onset pre-eclampsia independently of self-reported maternal ethnicity. Well-powered studies in multi-ethnic cohorts are required to delineate the genetic contribution to pre-eclampsia., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Obesity, diabetes, hypertension and dyslipidemia are well-established risk factors for cardiovascular diseases (CVDs), and have been associated with exposure to persistent organic pollutants. However, studies have been lacking as regards effects of non-persistent pesticides on CVD risk factors. Here, we investigated whether background chronic exposure to polychlorinated biphenyls (PCBs) and multiclass pesticides were associated with the prevalence of these CVD risk factors in 502 Belgian and 487 Luxembourgish adults aged 18-69 years from the Nutrition, environment and cardiovascular health (NESCAV) study 2007-2013. We used hair analysis to evaluate the chronic internal exposure to three PCBs, seven organochlorine pesticides (OCs) and 18 non-persistent pesticides. We found positive associations of obesity with hexachlorobenzene (HCB), β-hexachlorocyclohexane (β-HCH) and chlorpyrifos, diabetes with pentachlorophenol (PCP), fipronil and fipronil sulfone, hypertension with PCB180 and chlorpyrifos, and dyslipidemia with diflufenican and oxadiazon, among others. However, we also found some inverse associations, such as obesity with PCP, diabetes with γ-HCH, hypertension with diflufenican, and dyslipidemia with chlorpyrifos. These results add to the existing evidence that OC exposure may contribute to the development of CVDs. Additionally, the present study revealed associations between CVD risk factors and chronic environmental exposure to currently used pesticides such as organophosphorus and pyrethroid pesticides., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Rare genetic diseases affect millions, and identifying causal DNA variants is essential for patient care. Therefore, it is imperative to estimate the effect of each independent variant and improve their pathogenicity classification. Our study of 140 214 unrelated UK Biobank (UKB) participants found that each of them carries a median of 7 variants previously reported as pathogenic or likely pathogenic. We focused on 967 diagnostic-grade gene (DGG) variants for rare bleeding, thrombotic, and platelet disorders (BTPDs) observed in 12 367 UKB participants. By association analysis, for a subset of these variants, we estimated effect sizes for platelet count and volume, and odds ratios for bleeding and thrombosis. Variants causal of some autosomal recessive platelet disorders revealed phenotypic consequences in carriers. Loss-of-function variants in MPL, which cause chronic amegakaryocytic thrombocytopenia if biallelic, were unexpectedly associated with increased platelet counts in carriers. We also demonstrated that common variants identified by genome-wide association studies (GWAS) for platelet count or thrombosis risk may influence the penetrance of rare variants in BTPD DGGs on their associated hemostasis disorders. Network-propagation analysis applied to an interactome of 18 410 nodes and 571 917 edges showed that GWAS variants with large effect sizes are enriched in DGGs and their first-order interactors. Finally, we illustrate the modifying effect of polygenic scores for platelet count and thrombosis risk on disease severity in participants carrying rare variants in TUBB1 or PROC and PROS1, respectively. Our findings demonstrate the power of association analyses using large population datasets in improving pathogenicity classifications of rare variants., (© 2023 by The American Society of Hematology. Licensed under Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0), permitting only noncommercial, nonderivative use with attribution. All other rights reserved.)

Background: Bariatric surgery leads to profound changes in gut microbiota and dietary patterns, both of which may interact to impact gut-brain communication. Though cognitive function improves postsurgery, there is a large variability in outcomes. How bariatric surgery-induced modifications in the gut microbiota and dietary patterns influence the variability in cognitive function is still unclear., Objectives: To elucidate the associations between bariatric surgery-induced changes in dietary and gut microbiota patterns with cognition and brain structure., Setting: University hospital., Methods: A total of 120 adult patients (≥30 years) scheduled to undergo a primary bariatric surgery along with 60 age-, sex-, and body mass index-matched patients on the surgery waitlist will undergo assessments 3-months presurgery and 6- and 12-month postsurgery (or an equivalent time for the waitlist group). Additionally, 60 age-and sex-matched nonbariatric surgery eligible individuals will complete the presurgical assessments only. Evaluations will include sociodemographic and health behavior questionnaires, physiological assessments (anthropometrics, blood-, urine-, and fecal-based measures), neuropsychological cognitive tests, and structural magnetic resonance imaging. Cluster analyses of the dietary and gut microbiota changes will define the various dietary patterns and microbiota profiles, then using repeated measures mixed models, their associations with global cognitive and structural brain alterations will be explored., Results: The coordinating study site (Centre intégré universitaire de santé et de services sociaux du Nord-de-l'Île-de-Montréal, QC, Canada), provided the primary ethical approval (Research Ethics Board#: MP-32-2022-2412)., Conclusions: The insights generated from this study can be used to develop individually-targeted neurodegenerative disease prevention strategies, as well as providing critical mechanistic information., (Copyright © 2023 American Society for Metabolic and Bariatric Surgery. Published by Elsevier Inc. All rights reserved.)

Background: Blood platelets are mediators of atherothrombotic disease and are regulated by complex sets of genes. Association studies in European ancestry populations have already detected informative platelet regulatory loci. Studies in other ancestries can potentially reveal new associations because of different allele frequencies, linkage structures, and variant effects., Objectives: To reveal new regulatory genes for platelet count (PLT)., Methods: Genome-wide association studies (GWAS) were performed in 20,218 Bangladeshi and 9198 Pakistani individuals from the Genes & Health study. Loci significantly associated with PLT underwent fine-mapping to identify candidate genes., Results: Of 1588 significantly associated variants ( P  < 5 × 10 -8 ) at 20 loci in the Bangladeshi analysis, most replicated findings in prior transancestry GWAS and in the Pakistani analysis. However, the Bangladeshi locus defined by rs946528 (chr1:46019890) did not associate with PLT in the Pakistani analysis but was in the same linkage disequilibrium block ( r 2 ≥ 0.5) as PLT-associated variants in prior East Asian GWAS. The single independent association signal was refined to a 95% credible set of 343 variants spanning 8 coding genes. Functional annotation, mapping to megakaryocyte regulatory regions, and colocalization with blood expression quantitative trait loci identified the likely mediator of the PLT phenotype to be PIK3R3 encoding a regulator of phosphoinositol 3-kinase (PI3K)., Conclusion: Abnormal PI3K activity in the vessel wall is already implicated in the pathogenesis of atherothrombosis. Our identification of a new association between PIK3R3 and PLT provides further mechanistic insights into the contribution of the PI3K pathway to platelet biology., (© 2023 The Authors.)

This work presents an evaluation of solid-phase microextraction (SPME) SPME in combination with liquid chromatography-high resolution mass spectrometry (LC-HRMS) as an analytical approach for untargeted brain analysis. The study included a characterization of the metabolite coverage provided by C18, mixed-mode (MM, with benzene sulfonic acid and C18 functionalities), and hydrophilic lipophilic balanced (HLB) particles as sorbents in SPME coatings after extraction from cow brain homogenate at static conditions. The effects of desorption solvent, extraction time, and chromatographic modes on the metabolite features detected were investigated. Method precision and absolute matrix effects were also assessed. Among the main findings of this work, it was observed that all three tested coating chemistries were able to provide comparable brain tissue information. HLB provided higher responses for polar metabolites; however, as these fibers were prepared in-house, higher inter-fiber relative standard deviations were also observed. C18 and HLB coatings offered similar responses with respect to lipid-related features, whereas MM and C18 provided the best results in terms of method precision. Our results also showed that the use of methanol is essential for effective desorption of non-polar metabolites. Using a reversed-phase chromatographic method, an average of 800 and 1200 brain metabolite features detected in positive and negative modes, respectively, met inter-fibre RSD values below 30% (n=4) after removal of fibre and solvent artefacts from the associated datasets. For features detected using a lipidomics method, a total of 900 and 1800 features detected using C18 fibers in positive and negative mode, respectively, met the same criteria. In terms of absolute matrix effects, the majority of the model metabolites tested showed values between 80 and 120%, which are within the acceptable range. Overall, the findings of this work lay the foundation for further optimization of parameters for SPME-LC-HRMS methods suitable for in vivo and ex vivo brain (and other tissue) untargeted studies, and support the applicability of this approach for non-destructive tissue metabolomics., Competing Interests: Declaration of Competing Interest The authors have no competing interests to declare., (Copyright © 2021. Published by Elsevier B.V.)

To increase metabolome coverage in global LC-MS metabolomics, often both reversed-phase liquid chromatography (RPLC) and hydrophilic-interaction liquid chromatography (HILIC) are implemented in parallel. However, there is a lack of consensus in the literature on the best HILIC stationary phase to employ for global metabolomics of human biological fluids. The objective of this study was to compare in detail the performance of two commonly employed HILIC phases: zwitterionic sulfobetaine ZIC-HILIC stationary phase and an underivatized silica HILIC stationary phase. During method development, the effect of salt concentration in the mobile phase was also investigated, and 5 mM ammonium acetate was selected. The stationary phases were evaluated using a mixture of 37 polar standards covering a range of logP values (-10 to 3.73), molecular weights (59-776 Da), charges (15 anions, 11 cations, and 11 neutral) as well as 17 lipid standards to understand phospholipid behaviour on the two stationary phases. The criteria used for the comparison included the quality of the chromatographic peak shape, adequate analyte retention, peak separation capability, and metabolite coverage. The zwitterionic ZIC-HILIC column provided better chromatographic performance over the silica stationary phase with 14 standards achieving good quality peaks compared to the 7 with the silica column. Only 2 standards were undetected with the ZIC-HILIC column compared to the 14 undetected with the silica column. In human plasma, 1966 and 1650 metabolites were observed on the ZIC-HILIC column in positive and negative electrospray ionization (ESI) respectively. On the silica HILIC column, 1773 and 2028 metabolites were observed in positive and negative ESI respectively, showing comparable performance of the two phases. Next, the effect of adding 10 mM ammonium phosphate to the samples to improve the analyte peak shape and metabolite coverage was investigated for both ZIC-HILIC and silica HILIC. In contrast with recently reported results for pZIC-HILIC, there was no clear evidence that ammonium phosphate addition was beneficial for human plasma samples. In conclusion, ZIC-HILIC provided better chromatographic performance for polar plasma metabolomics than underivatized silica in terms of chromatographic peak shape and chromatographic resolution, while maintaining comparable metabolite coverage. The addition of ammonium phosphate to human plasma was not beneficial for either of the two stationary phases., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Mycotoxins are secondary metabolites produced by filamentous fungi. Primary route of human exposure to mycotoxins is the intake of the contaminated food. Minimizing mycotoxin exposure is important for population health, as their chronic toxic effects have been associated with kidney and liver diseases, some types of cancer and immunosuppression. The objective of this work was to develop and validate a multi-class mycotoxin method suitable for exposure monitoring of mycotoxins in human plasma. A sensitive liquid chromatography - mass spectrometry method was developed for 17 mycotoxins: nivalenol (NIV), deoxynivalenol, fusarenon X, 3-acetyldeoxynivalenol, 15-acetyldeoxynivalenol, T-2 toxin, HT-2 toxin, aflatoxin B1, aflatoxin B2, aflatoxin G1, aflatoxin G2, zearalenone, α-zearalenol (α-ZOL), β-zearalenol, zearalanone, α-zeranol and, and β-zeranol. The method relies on three-step liquid-liquid extraction with ethyl acetate to eliminate the need for immunoaffinity extraction and minimize ionization matrix effects. Chromatographic separation of mycotoxins, including all isomers, was achieved with pentafluorophenyl column and water/methanol mobile phase. Mycotoxin detection and quantitation were performed using high-resolution mass spectrometry on LTQ Velos Orbitrap, in both positive and negative electrospray ionization (ESI(+) and (ESI(-)). The use of 0.02% acetic acid as mobile phase additive for ESI(-) resulted in significant increase in ionization efficiency ranging from 1.7 to 26 times for mycotoxins that ionize better in ESI(-). The optimized method was validated according to FDA guidance procedures. LOQs of all mycotoxins ranged from 0.1 to 0.5 ng/ml, except NIV which resulted in LOQ of 3 ng/ml because of low extraction recovery of this highly polar mycotoxin. Mean intra-day accuracy ranged from 85.8% to 116.4%, and intra-day precision (n = 6) ranged from 1.6% to 12.5% RSD for all mycotoxins except α-ZOL where mean accuracy ranged from 72.9% to 97.2%. Inter-day accuracy and precision were 85.6%-111.5% and 2.7-15.6% RSD respectively, showing good analytical performance of the method for biomonitoring., (Copyright © 2018 Elsevier B.V. All rights reserved.)

As the lipidomics field continues to advance, self-evaluation within the community is critical. Here, we performed an interlaboratory comparison exercise for lipidomics using Standard Reference Material (SRM) 1950-Metabolites in Frozen Human Plasma, a commercially available reference material. The interlaboratory study comprised 31 diverse laboratories, with each laboratory using a different lipidomics workflow. A total of 1,527 unique lipids were measured across all laboratories and consensus location estimates and associated uncertainties were determined for 339 of these lipids measured at the sum composition level by five or more participating laboratories. These evaluated lipids detected in SRM 1950 serve as community-wide benchmarks for intra- and interlaboratory quality control and method validation. These analyses were performed using nonstandardized laboratory-independent workflows. The consensus locations were also compared with a previous examination of SRM 1950 by the LIPID MAPS consortium. While the central theme of the interlaboratory study was to provide values to help harmonize lipids, lipid mediators, and precursor measurements across the community, it was also initiated to stimulate a discussion regarding areas in need of improvement., (Copyright © 2017 by the American Society for Biochemistry and Molecular Biology, Inc.)

Solid phase microextraction (SPME) has experienced rapid development and growth in number of application areas since its inception over 20 years ago. It has had a major impact on sampling and sample preparation practices in chemical analysis, bioanalysis, food and environmental sciences. A significant impact is expected in clinical analysis as well as pharmaceutical and medical sciences in the near future. In this review, recent developments of SPME and related technologies are discussed including an in-vial standard gas system for calibration of SPME in high throughput mode; a thin film geometry with high extraction efficiency SPME for gas chromatography (GC) and liquid chromatography (LC) analyses; and couplings of SPME with portable instruments permitting on-site measurements. Also, the latest advances in the preparation of sorbents applicable for direct extraction from complex biological matrices as well as applications of these extraction phases in food analysis and biomedical studies such as therapeutic drug monitoring and pharmacokinetics are described. Finally, recent trends in metabolomics analysis and examples of clinical monitoring of biomarkers with SPME are reviewed., (Copyright © 2012 Elsevier B.V. All rights reserved.)

In vivo solid-phase microextraction (SPME) can be used to sample the circulating blood of animals without the need to withdraw a representative blood sample. In this study, in vivo SPME in combination with liquid-chromatography tandem mass spectrometry (LC-MS/MS) was used to determine the pharmacokinetics of two drug analytes, R,R-fenoterol and R,R-methoxyfenoterol, administered as 5 mg kg(-1) i.v. bolus doses to groups of 5 rats. This research illustrates, for the first time, the feasibility of the diffusion-based calibration interface model for in vivo SPME studies. To provide a constant sampling rate as required for the diffusion-based interface model, partial automation of the SPME sampling of the analytes from the circulating blood was accomplished using an automated blood sampling system. The use of the blood sampling system allowed automation of all SPME sampling steps in vivo, except for the insertion and removal of the SPME probe from the sampling interface. The results from in vivo SPME were compared to the conventional method based on blood withdrawal and sample clean up by plasma protein precipitation. Both whole blood and plasma concentrations were determined by the conventional method. The concentrations of methoxyfenoterol and fenoterol obtained by SPME generally concur with the whole blood concentrations determined by the conventional method indicating the utility of the proposed method. The proposed diffusion-based interface model has several advantages over other kinetic calibration models for in vivo SPME sampling including (i) it does not require the addition of a standard into the sample matrix during in vivo studies, (ii) it is simple and rapid and eliminates the need to pre-load appropriate standard onto the SPME extraction phase and (iii) the calibration constant for SPME can be calculated based on the diffusion coefficient, extraction time, fiber length and radius, and size of the boundary layer. In the current study, the experimental calibration constants of 338.9±30 mm(-3) and 298.5±25 mm(-3) are in excellent agreement with the theoretical calibration constants of 307.9 mm(-3) and 316.0 mm(-3) for fenoterol and methoxyfenoterol respectively., (Copyright © 2012 Elsevier B.V. All rights reserved.)

A solid phase microextraction (SPME) method followed by LC-MS/MS analysis was developed to determine the concentration of tranexamic acid (TA) in plasma. The use of a new biocompatible C18 coating allowed the direct extraction from complex biological samples without prior sample preparation; no matrix effect was observed. The results revealed that SPME was suitable for the analysis of polar drugs such as TA; such an application was previously inaccessible because of the limited availability of SPME coatings that can extract polar molecules. The proposed method was validated according to the bioanalytical method validation guidelines. LOD and LLOQ were 0.5 and 1.5 μg/ml, respectively. The recovery for the method was 0.19%, and the accuracy and precision of the method were <9 and <11%, respectively, with the exception of LLOQ, where the values were <16 and <13%, respectively. The performance of the proposed method was also compared against that of the standard techniques of protein precipitation and ultrafiltration. A statistical analysis indicated a clinically significant agreement among all assays. Another advantage of SPME over conventional techniques was the easy automation and feasibility of in vivo analysis; this advantage makes it possible to use the proposed method for an on-site analysis in clinical practice., (Copyright © 2011 Elsevier B.V. All rights reserved.)

The use of solid-phase microextraction (SPME) for in vivo sampling of drugs and metabolites in the bloodstream of freely moving animals eliminates the need for blood withdrawal in order to generate pharmacokinetics (PK) profiles in support of pharmaceutical drug discovery studies. In this study, SPME was applied for in vivo sampling in mice for the first time and enables the use of a single animal to construct the entire PK profile. In vivo SPME sampling procedure used commercial prototype single-use in vivo SPME probes with a biocompatible extractive coating and a polyurethane sampling interface designed to facilitate repeated sampling from the same animal. Pre-equilibrium in vivo SPME sampling, kinetic on-fibre standardization calibration and liquid chromatography-tandem mass spectrometry analysis (LC-MS/MS) were used to determine unbound and total circulating concentrations of carbamazepine (CBZ) and its active metabolite carbamazepine-10,11-epoxide (CBZEP) in mice (n=7) after 2mg/kg intravenous dosing. The method was linear in the range of 1-2000ng/mL CBZ in whole blood with acceptable accuracy (93-97%) and precision (<17% RSD). The single dose PK results obtained using in vivo SPME sampling compare well to results obtained by serial automated blood sampling as well as by the more conventional method of terminal blood collection from multiple animals/time point. In vivo SPME offers the advantages of serial and repeated sampling from the same animal, speed, improved sample clean-up, decreased animal use and the ability to obtain both free and total drug concentrations from the same experiment., (Copyright © 2010 Elsevier B.V. All rights reserved.)

The primary objective of this review is to discuss recent technological developments in the field of solid-phase microextraction that have enhanced the utility of this sample preparation technique in the field of bioanalysis. These developments include introduction of various new biocompatible coating phases suitable for bioanalysis, such as commercial prototype in vivo SPME devices, as well as the development of sampling interfaces that extend the use of this methodology to small animals such as mice. These new devices permit application of in vivo SPME to a variety of analyses, including pharmacokinetics, bioaccumulation and metabolomics studies, with good temporal and spatial resolution. New calibration approaches have also been introduced to facilitate in vivo studies and provide fast and quantitative results without the need to achieve equilibrium. In combination with the drastic improvement in the analytical sensitivity of modern liquid chromatography-tandem mass spectrometry instrumentation, full potential of in vivo SPME as a sample preparation tool in life sciences can finally be explored. From the instrumentation perspective, SPME was successfully automated in 96-well format for the first time. This opens up new opportunities for high-throughput applications (>1000 samples/day) such as for the determination of unbound and total drug concentrations in complex matrices such as whole blood with no need for sample pretreatment, studies of distribution of drugs in various compartments and/or determination of plasma protein binding and other ligand-receptor binding studies, and this review will summarize the progress in this research area to date., (Copyright (c) 2010 Elsevier B.V. All rights reserved.)

The success of in vivo solid phase microextraction (SPME) depends significantly on the selection of calibration method. Three kinetic in vivo SPME calibration methods are evaluated in this paper: (1) on-fibre standardization (OFS), (2) dominant pre-equilibrium desorption (DPED), and (3) the diffusion-based interface (DBI) model. These are compared in terms of precision, accuracy, and ease of experimental use by employing a flow device simulating an animal circulatory system. In addition, the kinetic calibration methods were validated against established SPME equilibrium extraction (EE) external calibration and a conventional sample preparation method involving protein precipitation. The comparison was performed using a hydrophilic drug fenoterol as the analyte of interest. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used for the determinations. All three kinetic methods compared well with both EE extraction and the conventional method in terms of accuracy (93-119%). In terms of precision, the DBI model had the best precision in whole blood and buffered phosphate saline solution with %RSD similar to the standard techniques (9-15%). DPED had the poorest precision of %RSD (20-30%) possibly due to errors associated with uncertainty in the amount of standard loaded on-fibre and remaining on the fibre after desorption. In addition, incurred errors could result due to the greater number of fibres used in comparison to the other two calibration methods. The precision of the OFS procedure was better than for DPED primarily because the use of multiple fibres is eliminated. In terms of the ease of use for calibration, the DBI model was the simplest and most convenient as it did not require standards once it had been calibrated or the uptake constant was calculated. This research suggests the potential use of DBI model as the best kinetic calibration method for future in-vein blood SPME investigations., (Copyright 2010 Elsevier B.V. All rights reserved.)

An in situ application of solid-phase microextraction (SPME) as a sampling and sample preparation method coupled to HPLC-MS/MS for direct monitoring of ochratoxin A (OTA) distribution at different locations in a single cheese piece is proposed. To be suited to the acidic analyte, the extraction phase (carbon-tape SPME fiber) was acidified with aqueous solution of HCl at pH 2, instead of the traditional sample pre-treatment with acids before SPME sampling. For calibration, kinetic on-fiber-standardization was used, which allowed the use of short sampling time (20 min) and accurate quantification of the OTA in the semi-solid cheese sample. In addition, the traditional kinetic calibration that used deuterated compounds as standards was extended to use a non-deuterated analogue ochratoxin B (OTB) as the standard of the analyte OTA, which was supported by both theoretical discussion and experimental verification. Finally, the miniaturized SPME fiber was adopted so that the concentration distribution of OTA in a small-sized cheese piece could be directly probed. The detection limit of the resulting SPME method in semi-solid gel was 1.5 ng/mL and the linear range was 3.5-500 ng/mL. The SPME-LC-MS/MS method showed good precision (RSD: approximately 10%) and accuracy (relative recovery: 93%) in the gel model. The direct cheese analysis showed comparable accuracy and precision to the established liquid extraction. As a result, the developed in situ SPME-LC-MS/MS method was sensitive, simple, accurate and applicable for the analysis of complicated lipid-rich samples such as cheese.

A new line of solid-phase microextraction (SPME) coatings suitable for use with liquid chromatography applications was recently developed to address the limitations of the currently available coatings. The proposed coatings were immobilized on the metal fiber core and consisted of a mixture of proprietary biocompatible binder and various types of coated silica (octadecyl, polar embedded and cyano) particles. The aim of this research was to perform in vitro assessment of these new SPME fibers in order to evaluate their suitability for drug analysis and in vivo SPME applications. The main parameters examined were extraction efficiency, solvent resistance, preconditioning, dependence of extraction kinetics on coating thickness, carryover, linear range and inter-fiber reproducibility. The performance of the proposed coatings was compared against commercial Carbowax-TPR (CW-TPR) coating, when applicable. The fibers were evaluated for the extraction of drugs of different classes (carbamazepine, propranolol, pseudoephedrine, ranitidine and diazepam) from plasma and urine. The analyses were performed using liquid chromatography-tandem mass spectrometry. The results show that the fibers perform very well for the extraction of biological fluids with no sample pre-treatment required and can also be used for in vivo sampling applications of flowing blood. A coating thickness of 45 microm was found to be a good compromise between extraction capacity and extraction kinetics. Due to the high extraction efficiency of these coatings, pre-equilibrium SPME with very short extraction times (2 min) can be employed to increase sample throughput. Inter-fiber reproducibility was < or = 11% R.S.D. (n=10) for model drugs examined in plasma, which is a significant improvement over polypyrrole coatings reported in literature, and permits single fiber use for in vivo applications.

A new automated, high-throughput method for the determination of ochratoxin A (OTA) in human urine samples has been optimized and validated using solid-phase microextraction coupled to liquid chromatography-tandem mass spectrometry (SPME-LC-MS/MS). High-throughput was achieved by simultaneous preparation of up to 96 samples using multi-fiber SPME device and multi-well plates. A carbon-tape coating was chosen for the first time as the best extracting phase for this contaminant. The proposed method required only minimal sample pre-treatment to adjust sample pH to 3.0 using a dilution (1:1) with 0.5M phosphate-buffered saline. A simple gradient guaranteed a good chromatographic separation from matrix interferences in only 8min. Relative recovery (%), precision and linearity validation results met Food and Drug Administration acceptance criteria at three concentration levels (1, 10, and 50ng/mL), indicating excellent performance of the proposed method. Limits of detection and quantitation were 0.3 and 0.7ng/mL in urine, respectively. OTA determination in urine is a good marker for human exposure to this mycotoxin. It is also less invasive than blood analysis. This method is fully automated and the SPME technique is simpler, less time-consuming and cheaper compared with most widely adopted clean-up procedures for OTA extraction from urine.