Synthesis of [11C]BIIB104, an α-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic-Acid-Positive Allosteric Modulator, and Evaluation of the Bio-Distribution in Non-Human Primate Brains Using Positron Emission Tomography.

The aim of this study was to measure the brain penetrance and kinetics of BIIB104, a first-in-class AMPA receptor potentiator developed for cognitive impairment associated with schizophrenia. It was recently halted in phase 2 clinical development, and there are a lack of tools to directly measure AMPA receptor engagement. To achieve this, the drug candidate was radiolabeled with carbon-11, and its brain penetrance and kinetics were measured in non-human primates via dynamic PET scans. Radiolabeling was achieved through a three-step nucleophilic [11C]cyanation reaction in one pot, resulting in the high radioactivity and radiochemical purity (>99%) of [11C]BIIB104. The study found that [11C]BIIB104 entered the non-human primate brains at 4-5% ID at peak, with a homogeneous distribution. However, a mild regional heterogeneity was observed in the thalamus. The lack of conclusive evidence for a change in regional values after BIIB104 dosing suggests that any specific binding component of BIIB104 is negligible compared to the free and non-specific components in the living brain. Overall, the study demonstrated high brain uptake with minor variability in [11C]BIIB104 distribution across various brain regions, its kinetics were consistent with those of passive diffusion, and the dominating components were the free concentration and non-specific binding. This information is valuable for understanding the potential effects and mechanisms of BIIB104 in the brain.

Development of 11C-Labeled ASEM Analogues for the Detection of Neuronal Nicotinic Acetylcholine Receptors (α7-nAChR).

The homo-pentameric alpha 7 receptor is one of the major types of neuronal nicotinic acetylcholine receptors (α7-nAChRs) related to cognition, memory formation, and attention processing. The mapping of α7-nAChRs by PET pulls a lot of attention to realize the mechanism and development of CNS diseases such as AD, PD, and schizophrenia. Several PET radioligands have been explored for the detection of the α7-nAChR. 18F-ASEM is the most functional for in vivo quantification of α7-nAChRs in the human brain. The first aim of this study was to initially use results from in silico and machine learning techniques to prescreen and predict the binding energy and other properties of ASEM analogues and to interpret these properties in terms of atomic structures using 18F-ASEM as a lead structure, and second, to label some selected candidates with carbon-11/hydrogen-3 (11C/3H) and to evaluate the binding properties in vitro and in vivo using the labeled candidates. In silico predictions are obtained from perturbation free-energy calculations preceded by molecular docking, molecular dynamics, and metadynamics simulations. Machine learning techniques have been applied for the BBB and P-gp-binding properties. Six analogues of ASEM were labeled with 11C, and three of them were additionally labeled with 3H. Binding properties were further evaluated using autoradiography (ARG) and PET measurements in non-human primates (NHPs). Radiometabolites were measured in NHP plasma. All six compounds were successfully synthesized. Evaluation with ARG showed that 11C-Kln83 was preferably binding to the α7-nAChR. Competition studies showed that 80% of the total binding was displaced. Further ARG studies using 3H-KIn-83 replicated the preliminary results. In the NHP PET study, the distribution pattern of 11C-KIn-83 was similar to other α7 nAChR PET tracers. The brain uptake was relatively low and increased by the administration of tariquidar, indicating a substrate of P-gp. The ASEM blocking study showed that 11C-KIn-83 specifically binds to α7 nAChRs. Preliminary in vitro evaluation of KIn-83 by ARG with both 11C and 3H and in vivo evaluation in NHP showed favorable properties for selectively imaging α7-nAChRs, despite a relatively low brain uptake.

Clinically Applicable Cyclotron-Produced Gallium-68 Gives High-Yield Radiolabeling of DOTA-Based Tracers.

By using solid targets in medical cyclotrons, it is possible to produce large amounts of 68GaCl3. Purification of Ga3+ from metal ion impurities is a critical step, as these metals compete with Ga3+ in the complexation with different chelators, which negatively affects the radiolabeling yields. In this work, we significantly lowered the level of iron (Fe) impurities by adding ascorbate in the purification, and the resulting 68GaCl3could be utilized for high-yield radiolabeling of clinically relevant DOTA-based tracers. 68GaCl3 was cyclotron-produced and purified with ascorbate added in the wash solutions through the UTEVA resins. The 68Ga eluate was analyzed for radionuclidic purity (RNP) by gamma spectroscopy, metal content by ICP-MS, and by titrations with the chelators DOTA, NOTA, and HBED. The 68GaCl3eluate was utilized for GMP-radiolabeling of the DOTA-based tracers DOTATOC and FAPI-46 using an automated synthesis module. DOTA chelator titrations gave an apparent molar activity (AMA) of 491 ± 204 GBq/µmol. GMP-compliant syntheses yielded up to 7 GBq/batch [68Ga]Ga-DOTATOC and [68Ga]Ga-FAPI-46 (radiochemical yield, RCY ~ 60%, corresponding to ten times higher compared to generator-based productions). Full quality control (QC) of 68Ga-labelled tracers showed radiochemically pure and stable products at least four hours from end-of-synthesis.

Fully automated production of the fibroblast activation protein radiotracer [18 F]FAPI-74.

We report herein an efficient and fully automated protocol for the radiosynthesis of [18 F]FAPI-74, a new positron emission tomography (PET) radiopharmaceutical for in vivo detection of the fibroblast activation protein. [18 F]FAPI-74 was synthesized via a rapid [18 F]aluminum fluoride coordination reaction, which was first developed on the flexible GE TRACERLab FX2N (FXN) platform and later translated to the cassette-based module Trasis AllInOne (AIO). The results obtained with both modules were comparable in terms of yield and reproducibility. Automation of [18 F]FAPI-74 radiosynthesis on the FXN was carried out in 35 min with a radiochemical yield (RCY) of 18.5 ± 2.5% (n = 5, relative to starting [18 F]fluoride). Method transfer to the AIO platform following minor optimizations allowed for the production of [18 F]FAPI-74 in an isolated RCY of 20 ± 2.5% [n = 3] with an overall synthesis time of 40 min. The radiochemical purity was greater than 95% for [18 F]FAPI-74, obtained from both modules. Overall, the protocol reliably provides a sterile and pyrogen-free good manufacturing practice (GMP) compliant product of [18 F]FAPI-74 suitable for clinical PET imaging.

Advancements of chiral molecularly imprinted polymers in separation and sensor fields: A review of the last decade.

Since chiral recognition mechanism based on molecularly imprinted polymers immerged, it has assisted countless chemical and electrochemical analytical sample preparation techniques. It has done this by enhancing the enatioseparation abilities of these techniques. The preparation and optimization of chiral molecularly imprinted polymers (CMIPs) are two favored methods in the separation and sensor fields. This review aims to present an overview of advances in the preparation and application of CMIPs in analytical approaches in different available formats (eg. column, monolithic column, cartridge, membrane, nanomaterials, pipette tip and stir bar sorptive) over the last decade. In addition, progress in the preparation and development of CMIPs-based sensor fields have been also discussed. Finally, the main application challenges of CMIPs are also summarily explained, as well as upcoming prospects in the field.

Preclinical Comparison of the Blood-brain barrier Permeability of Osimertinib with Other EGFR TKIs.

PURPOSE: Osimertinib is a potent and selective EGFR tyrosine kinase inhibitor (EGFR-TKI) of both sensitizing and T790M resistance mutations. To treat metastatic brain disease, blood-brain barrier (BBB) permeability is considered desirable for increasing clinical efficacy. EXPERIMENTAL DESIGN: We examined the level of brain penetration for 16 irreversible and reversible EGFR-TKIs using multiple in vitro and in vivo BBB preclinical models. RESULTS: In vitro osimertinib was the weakest substrate for human BBB efflux transporters (efflux ratio 3.2). In vivo rat free brain to free plasma ratios (Kpuu) show osimertinib has the most BBB penetrance (0.21), compared with the other TKIs (Kpuu ≤ 0.12). PET imaging in Cynomolgus macaques demonstrated osimertinib was the only TKI among those tested to achieve significant brain penetrance (C max %ID 1.5, brain/blood Kp 2.6). Desorption electrospray ionization mass spectroscopy images of brains from mouse PC9 macrometastases models showed osimertinib readily distributes across both healthy brain and tumor tissue. Comparison of osimertinib with the poorly BBB penetrant afatinib in a mouse PC9 model of subclinical brain metastases showed only osimertinib has a significant effect on rate of brain tumor growth. CONCLUSIONS: These preclinical studies indicate that osimertinib can achieve significant exposure in the brain compared with the other EGFR-TKIs tested and supports the ongoing clinical evaluation of osimertinib for the treatment of EGFR-mutant brain metastasis. This work also demonstrates the link between low in vitro transporter efflux ratios and increased brain penetrance in vivo supporting the use of in vitro transporter assays as an early screen in drug discovery.

Brain exposure of the ATM inhibitor AZD1390 in humans-a positron emission tomography study.

BACKGROUND: The protein kinase ataxia telangiectasia mutated (ATM) mediates cellular response to DNA damage induced by radiation. ATM inhibition decreases DNA damage repair in tumor cells and affects tumor growth. AZD1390 is a novel, highly potent, selective ATM inhibitor designed to cross the blood-brain barrier (BBB) and currently evaluated with radiotherapy in a phase I study in patients with brain malignancies. In the present study, PET was used to measure brain exposure of 11C-labeled AZD1390 after intravenous (i.v.) bolus administration in healthy subjects with an intact BBB. METHODS: AZD1390 was radiolabeled with carbon-11 and a microdose (mean injected mass 1.21 µg) was injected in 8 male subjects (21-65 y). The radioactivity concentration of [11C]AZD1390 in brain was measured using a high-resolution PET system. Radioactivity in arterial blood was measured to obtain a metabolite corrected arterial input function for quantitative image analysis. Participants were monitored by laboratory examinations, vital signs, electrocardiogram, adverse events. RESULTS: The brain radioactivity concentration of [11C]AZD1390 was 0.64 SUV (standard uptake value) and reached maximum 1.00% of injected dose at Tmax[brain] of 21 min (time of maximum brain radioactivity concentration) after i.v. injection. The whole brain total distribution volume was 5.20 mL*cm-3. No adverse events related to [11C]AZD1390 were reported. CONCLUSIONS: This study demonstrates that [11C]AZD1390 crosses the intact BBB and supports development of AZD1390 for the treatment of glioblastoma multiforme or other brain malignancies. Moreover, it illustrates the potential of PET microdosing in predicting and guiding dose range and schedule for subsequent clinical studies.

Application of chiral chromatography in radiopharmaceutical fields: A review.

Chiral column chromatography (CCC) is a revolutionary analytical methodology for the enantioseparation of novel positron emission tomography (PET) tracers in the primary stages of drug development. Due to the different behaviors of tracer enantiomers (e.g. toxicity, metabolism and side effects) in administrated subjects, their separation and purification is a challenging endeavor. Over the last three decades, different commercial chiral columns have been applied for the enantioseparation of PET-radioligand (PET-RL) or radiotracers (PET-RT), using high-performance liquid chromatography (HPLC). The categorization and reviewing of them is a vital topic. This review presents a brief overview of advances, applications, and future prospectives of CCC in radiopharmaceutical approaches. In addition, the effective chromatographic parameters and degravitation trends to enhance enantioseparation resolution are addressed. Moreover, the application and potential of chiral super fluidical chromatography (CSFC) as an alternative for enantioseparation in the field of radiopharmaceutical is discussed. Finally, the crucial application challenges of CCC are explained and imminent tasks are suggested.

Sample preparation techniques for protein binding measurement in radiopharmaceutical approaches: A short review.

Plasma protein binding (PPB) measurement is a key step in radiopharmaceutical studies for the development of positron emission tomography (PET) radioligands. PPB refers to the binding degree of a radioligand, radiotracer, or drug to blood plasma proteins or tissues after administration into the body. Several techniques have been successfully developed and applied for PPB measurement of PET radioligands. However, there is room for progress among these techniques in relation to duration time, adaptability with nonpolar radioligands, in vivo measurement, specificity, and selectivity. This mini review gives a brief overview of advances, limitations, and prospective applications of commercially-available PPB methods.

Quantification and reliability of [11C]VC - 002 binding to muscarinic acetylcholine receptors in the human lung - a test-retest PET study in control subjects.

BACKGROUND: The radioligand [11C]VC-002 was introduced in a small initial study long ago for imaging of muscarinic acetylcholine receptors (mAChRs) in human lungs using positron emission tomography (PET). The objectives of the present study in control subjects were to advance the methodology for quantification of [11C]VC-002 binding in lung and to examine the reliability using a test-retest paradigm. This work constituted a self-standing preparatory step in a larger clinical trial aiming at estimating mAChR occupancy in the human lungs following inhalation of mAChR antagonists. METHODS: PET measurements using [11C]VC-002 and the GE Discovery 710 PET/CT system were performed in seven control subjects at two separate occasions, 2-19 days apart. One subject discontinued the study after the first measurement. Radioligand binding to mAChRs in lung was quantified using an image-derived arterial input function. The total distribution volume (VT) values were obtained on a regional and voxel-by-voxel basis. Kinetic one-tissue and two-tissue compartment models (1TCM, 2TCM), analysis based on linearization of the compartment models (multilinear Logan) and image analysis by data-driven estimation of parametric images based on compartmental theory (DEPICT) were applied. The test-retest repeatability of VT estimates was evaluated by absolute variability (VAR) and intraclass correlation coefficients (ICCs). RESULTS: The 1TCM was the statistically preferred model for description of [11C]VC-002 binding in the lungs. Low VAR (< 10%) across analysis methods indicated good reliability of the PET measurements. The VT estimates were stable after 60 min. CONCLUSIONS: The kinetic behaviour and good repeatability of [11C]VC-002 as well as the novel lung image analysis methodology support its application in applied studies on drug-induced mAChR receptor occupancy and the pathophysiology of pulmonary disorders. TRIAL REGISTRATION: ClinicalTrials.gov identifier: NCT03097380, registered: 31 March 2017.

Microextraction approaches for bioanalytical applications: An overview.

Biological samples are usually complex matrices due to the presence of proteins, salts and a variety of organic compounds with chemical properties similar to those of the target analytes. Therefore, sample preparation is often mandatory in order to isolate the analytes from troublesome matrices before instrumental analysis. Because the number of samples in drug development, doping analysis, forensic science, toxicological analysis, and preclinical and clinical assays is steadily increasing, novel high throughput sample preparation approaches are calling for. The key factors in this development are the miniaturization and the automation of the sample preparation approaches so as to cope with most of the twelve principles of green chemistry. In this review, recent trends in sample preparation and novel strategies will be discussed in detail with particular focus on sorptive and liquid-phase microextraction in bioanalysis. The actual applicability of selective sorbents is also considered. Additionally, the role of 3D printing in microextraction for bioanalytical methods will be pinpointed.

New approach in radiometabolite analysis of positron emission tomography (PET) radioligands; lead-shielded microextraction by packed sorbent as a tool for in vivo radiometabolite analysis of [11C]SMW139 in rat plasma.

The blood plasma radiometabolite analysis (RMA) is crucial procedure for quantitative analysis of positron emission tomography (PET) radioligands with no original reference region in the brain. Blood sampling and separating plasma for RMA has been a challenge especially with small animal PET research. This work aimed to overcome this problem by presenting a semi-automated lead shielded microextraction by packed sorbents (SLS-MEPS). The capability of SLS-MEPS was evaluated for RMA of [11C]SMW139 as a PET radioligand in rat plasma samples. [11C]SMW139 is a potential radioligand with high receptor binding for P2X7R in vivo. The validation method was individually performed for [11C]SMW139 and [12C]SMW139 with reversed phase high performance liquid chromatography (HPLC) utilizing radio- and UV-detector, respectively. The limits of detection (LOD) and quantification (LOQ) [11C]SMW139 in rat plasma were 3 and 10 Becquerel (Bq), respectively. The standard calibration curve was obtained for [11C]SMW139 within the concentration range 10-15000 Bq with over 0.995 coefficients for all assays. In addition, the capability of SLS-MEPS in combination with HPLC-UV for extraction of [12C]SMW139 in rat plasma was also challenged and the obtained results showed a proper linearity (20-2000 µg mL-1), accuracy and repeatability.

The pro-psychotic metabotropic glutamate receptor compounds fenobam and AZD9272 share binding sites with monoamine oxidase-B inhibitors in humans.

The metabotropic glutamate receptor 5 (mGluR5) ligands fenobam and AZD9272 have been reported to induce psychosis-like adverse events and to bind at unknown, non-GluR5-related, sites. Based on similarities of the regional binding patterns for [11C]AZD9272 and the monoamine oxidase-B (MAO-B) radioligand [11C]L-deprenyl-D2 in PET studies of the human brain we tested the hypothesis that the unique binding of fenobam and AZD9272 may represent specific binding to the MAO-B. PET data previously acquired for subjects examined using [11C]AZD9272 or [11C]L-deprenyl-D2 were re-evaluated to assess the correlations between radioligand binding parameters in human brain. In addition, the pharmacology of AZD9272 binding sites was characterized using competition binding studies carried out in vivo in non-human primates (NHPs) and in vitro using autoradiography in selected human brain regions. The regional binding of [11C]AZD9272 in human brain was closely correlated with that of [11C]L-deprenyl-D2. In PET studies of NHP brain administration of the MAO-B ligand L-deprenyl inhibited binding of radiolabeled AZD9272 and administration of fenobam inhibited binding of [11C]L-deprenyl-D2. Binding of radiolabeled AZD9272 in vitro was potently inhibited by fenobam or MAO-B compounds, and [11C]L-deprenyl-D2 binding was inhibited by fenobam or AZD9272. The findings are consistent with the hypothesis that both fenobam and AZD9272 bind to the MAO-B, which may be of relevance for understanding the mechanism of the psychosis-like adverse events reported for these compounds. Such understanding may serve as a lead to generate new models for the pathophysiology of psychosis.

Development of a 18F-labeled PET radioligand for imaging 5-HT1B receptors: [18F]AZ10419096.

INTRODUCTION: In the last decade PET has been useful in studying and understanding the 5-HT1B receptor. [11C]AZ10419369 and [11C]P943 have been applied as radioligands in these studies. Both use carbon-11 (t1/2 = 20.4 min) as radionuclide, which limits the application to PET centres that have an on-site cyclotron and radiochemistry facilities. In this paper we report the synthesis and initial evaluation of the first fluorine-18 PET radioligand to image 5-HT1B receptors in brain, [18F]AZ10419096. MATERIALS AND METHODS: A boronate-precursor for [18F]AZ10419096 was synthesized from an intermediate provided by AstraZeneca and was labeled with fluorine 18 using Cu-mediated radio-fluorination. [18F]AZ10419096 was used in PET baseline and pretreatment measurements in nonhuman primates. PET data were analyzed using SRTM using the cerebellum as reference region. Blood samples for radio-metabolite analysis were collected during PET measurements. RESULTS: Radio-fluorination gave [18F]AZ10419096 in sufficient amounts and molar activity and with high radiochemical purity to be applied in PET measurements. In a baseline PET measurement [18F]AZ10419096 showed a high brain uptake and regional distribution consistent with reported 5-HT1B receptor densities. In a pretreatment PET measurement, AR-A000002 (2.0 mg/kg) blocked the binding of [18F]AZ10419096 to 5-HT1B receptors in occipital cortex by 80%, thereby demonstrating high specific binding. CONCLUSION: [18F]AZ10419096 is the first fluorine-18 PET radioligand for imaging 5-HT1B receptors in vivo with high specific binding and binding potential. [18F]AZ10419096 is a candidate for further development for use in clinical PET studies.

Graphene Oxide/Polyethylene Glycol-Stick for Thin Film Microextraction of ß-Blockers from Human Oral Fluid by Liquid Chromatography-Tandem Mass Spectrometry.

A wooden stick coated with a novel graphene-based nanocomposite (Graphene oxide/polyethylene glycol (GO/PEG)) is introduced and investigated for its efficacy in solid phase microextraction techniques. The GO/PEG-stick was prepared and subsequently applied for the extraction of ß-blockers, acebutolol, and metoprolol in human oral fluid samples, which were subsequently detected by liquid chromatography tandem mass spectrometry (LC-MS/MS). Experimental parameters affecting the extraction protocol including sample pH, extraction time, desorption time, appropriate desorption solvent, and salt addition were optimized. Method validation for the detection from oral fluid samples was performed following FDA (Food and Drug Administration) guidelines on bioanalytical method validation. Calibration curves ranging from 5.0 to 2000 nmol L-1 for acebutolol and 25.0 to 2000 nmol L-1 for metoprolol were used. The values for the coefficient of determination (R2) were found to be 0.998 and 0.996 (n = 3) for acebutolol and metoprolol, respectively. The recovery of analytes during extraction was 80.0% for acebutolol and 62.0% for metoprolol, respectively. The limit of detections (LODs) were 1.25, 8.00 nmol L-1 for acebutolol and metoprolol and the lower limit of quantifications (LLOQ) were 5.00 nmol L-1 for acebutolol and 25.0 nmol L-1 for metoprolol. Validation experiments conducted with quality control (QC) samples demonstrated method accuracy between 80.0% to 97.0% for acebutolol and from 95.0% to 109.0% for metoprolol. The inter-day precision for QC samples ranged from 3.6% to 12.9% for acebutolol and 9.5% to 11.3% for metoprolol. Additionally, the GO/PEG-stick was demonstrated to be reusable, with the same stick observed to be viable for more than 10 extractions from oral fluid samples.

Recent Applications of Molecularly Imprinted Sol-Gel Methodology in Sample Preparation.

Due to their selectivity and chemical stability, molecularly imprinted polymers have attracted great interest in sample preparation. Imprinted polymers have been applied for the extraction and the enrichment of different sorts of trace analytes in biological and environmental samples before their analysis. Additionally, MIPs are utilized in various sample preparation techniques such as SPE, SPME, SBSE and MEPS. Nevertheless, molecularly imprinted polymers suffer from thermal (stable only up to 150 °C) and mechanical stability issues, improper porosity and poor capacity. The sol-gel methodology as a promising alternative to address these limitations allowing the production of sorbents with controlled porosity and higher surface area. Thus the combination of molecularly imprinted technology and sol-gel technology can create influential materials with high selectivity, high capacity and high thermal stability. This work aims to present an overview of molecularly imprinted sol-gel polymerization methods and their applications in analytical and bioanalytical fields.

Polyacrylonitrile / graphene oxide nanofibers for packed sorbent microextraction of drugs and their metabolites from human plasma samples.

In the present study, a new graphene based nanofibers material (Polyacrylonitrile/Graphene Oxide (PAN/GO)) was used for microextraction by packed sorbent (MEPS). The PAN/GO nanofiber was synthesized using the electrospinning technique. MEPS online with liquid chromatography-tandem mass spectrometry (LC-MS/MS) was utilized for the extraction and determination of two local anesthetic drugs (lidocaine, prilocaine) and their major metabolites (2,6-xylidine, o-toluidine) in human plasma samples. Parameters affecting the extraction efficiency were investigated and optimized (including sample pH, washing solution and elution solution). The validation of the method was based on FDA (Food and Drug Administration) guidelines for bioanalytical methods. The calibration curve ranged from 2.00 to 2000 nmol/L for lidocaine and prilocaine, and from 10.0 to 2000 nmol/L for 2,6-xylidine and o-toluidine. The coefficient of determination (R2) values were 0.996, 0.995, 0.995, 0.996 (n = 3) for lidocaine, prilocaine, 2,6-xylidine and o-toluidine, respectively. The extraction recovery was 93.0% for lidocaine, 96.0% for prilocaine, 68.0% for 2,6-xylidine and 69.0% for o-toluidine. The limits of detection (LODs) were 0.25, 0.50, 2.50, 1.25 nmol/L for lidocaine, prilocaine, 2,6-xylidine and o-toluidine, respectively. The lower limits of quantification (LLOQs) were 2.0 nmol/L for lidocaine and prilocaine, and 10 nmol/L for 2,6-xylidine and o-toluidine, respectively. The accuracy values for the quality control (QC) samples were in the range of 91.0-111% for lidocaine, 92.0-118% for prilocaine, 84.0-98.0% for 2,6-xylidine and 82.0-90.0% for o-toluidine. The inter-day precisions for QC samples ranged from 7.0% to 11.8% for lidocaine, from 8.6% to 11.7% for prilocaine, from 8.0% to 10.0% for 2,6-xylidine and from 8.0% to 9.0% for o-toluidine. The matrix effect values were in the range of -2.3% to -8.6% for lidocaine, -2.7% to -10.2% for prilocaine, 4.8%-5.2% for 2, 6-xylidine and -8.2% to 9.4% for o-toluidine.

Graphene Oxide Tablets for Sample Preparation of Drugs in Biological Fluids: Determination of Omeprazole in Human Saliva for Liquid Chromatography Tandem Mass Spectrometry.

In this study, a novel sort of sample preparation sorbent was developed, by preparing thin layer graphene oxide tablets (GO-Tabs) utilizing a mixture of graphene oxide and polyethylene glycol on a polyethylene substrate. The GO-Tabs were used for extraction and concentration of omeprazole (OME) in human saliva samples. The determination of OME was carried out using liquid chromatography-tandem mass spectrometry (LC⁻MS/MS) under gradient LC conditions and in the positive ion mode (ESI+) with mass transitions of m/z 346.3→198.0 for OME and m/z 369.98→252.0 for the internal standard. Standard calibration for the saliva samples was in the range of 2.0⁻2000 nmol L-1. Limits of detection and quantification were 0.05 and 2.0 nmol L-1, respectively. Method validation showed good method accuracy and precision; the inter-day precision values ranged from 5.7 to 8.3 (%RSD), and the accuracy of determinations varied from -11.8% to 13.3% (% deviation from nominal values). The extraction recovery was 60%, and GO-Tabs could be re-used for more than ten extractions without deterioration in recovery. In this study, the determination of OME in real human saliva samples using GO-Tab extraction was validated.

Reduced graphene oxide as an efficient sorbent in microextraction by packed sorbent: Determination of local anesthetics in human plasma and saliva samples utilizing liquid chromatography-tandem mass spectrometry.

Herein, reduced graphene oxide (RGO) has been utilized as an efficient sorbent in microextraction by packed sorbent (MEPS). The combination of MEPS and liquid chromatography-tandem mass spectrometry has been used to develop a method for the extraction and determination of three local anesthetics (i.e. lidocaine, prilocaine, and ropivacaine) in human plasma and saliva samples. The results showed that the utilization of RGO in MEPS could minimize the matrix effect so that no interfering peaks at the retention times of the analytes or internal standard was observed. The high extraction efficiency of this method was approved by mean recoveries of 97.26-106.83% and 95.21-105.83% for the studied analytes in plasma and saliva samples, respectively. Intra- and inter-day accuracies and precisions for all analytes were in good accordance with the international regulations. The accuracy values (as percentage deviation from the nominal value) of the quality control samples were between -2.1 to 13.9 for lidocaine, -4.2 to 11.0 for prilocaine and between -4.5 to -2.4 for ropivacaine in plasma samples while the values were ranged from -4.6 to 1.6 for lidocaine, from -4.2 to 15.5 for prilocaine and from -3.3 to -2.3 for ropivacaine in human saliva samples. Lower and upper limit of quantification (LLOQ, ULOQ) were set at 5 and 2000 nmol L-1 for all of the studied drugs. The correlation coefficients values were ≥0.995. The limit of detection values were obtained 4 nmol L-1 for lidocaine and prilocaine, and 2 nmol L-1 for ropivacaine.

Development of [ Carbonyl-11C]AZ13198083, a Novel Histamine Type-3 Receptor Radioligand with Favorable Kinetics.

The histamine subtype-3 receptor (H3R) is implicated in a range of central nervous system disorders, and several radioligands have been developed for H3R positron emission tomography imaging. However, a limitation of currently used PET radioligands for H3R is the slow binding kinetics in high density brain regions. To address this, we herein report the development of three novel candidate H3R radioligands, namely, [ carbonyl-11C]AZ13153556 ([ carbonyl-11C]4), [ carbonyl-11C]AZD5213([ carbonyl-11C]5), and [ carbonyl-11C]AZ13198083 ([ carbonyl-11C]6), and their subsequent preclinical evaluation in nonhuman primates (NHP). Radioligands [ carbonyl-11C]4-6 were produced and isolated in high radioactivity (>1000 MBq), radiochemical purity (>99%), and moderate molar activity (19-28 GBq/µmol at time of injection) using a palladium-mediated 11C-aminocarbonylation protocol. All three radioligands showed high brain permeability as well as a regional brain radioactivity distribution in accordance with H3R expression (striatum > cortex > cerebellum). [ Carbonyl-11C]6 displayed the most favorable in vivo kinetics and brain uptake, with an early peak in the striatal time-activity curve followed by a progressive washout from the brain. The specificity and on-target kinetics of [ carbonyl-11C]6 were next investigated in pretreatment and displacement studies. After pretreatment or displacement with 5 (0.1 mg/kg), a uniformly low distribution of radioactivity across the NHP brain was observed. Collectively, this work demonstrates that [ carbonyl-11C]6 is a promising candidate for H3R imaging in human subjects.