Triple-stage Quadrupole Research Articles

Development and validation of a multiplex HPLC-MS/MS assay for the monitoring of JAK inhibitors in patient plasma

Janus kinase inhibitors (JAKi) are oral small molecules used in the treatment of a broad spectrum of autoimmune and myeloproliferative diseases. JAKi exhibit significant intra- and inter-individual pharmacokinetic variabilities, due to fluctuations in compliance with oral treatments and their metabolism essentially driven by cytochrome P450 enzymes.Intrinsically, JAKi have dose–response relationship and narrow therapeutic index: therapeutic drug monitoring (TDM) is expected to optimize and adapt their dosage regimen in order to resolve problems of efficacy and tolerance linked to dose and safety.A sensitive analytical method using multiplex high-performance liquid-chromatography coupled to tandem mass spectrometry (HPLC-MS/MS) was developed and validated for the simultaneous quantification in plasma of the 6 major currently used JAKi, namely abrocitinib, baricitinib, fedratinib, ruxolitinib, tofacitinib, and upadacitinib.Plasma samples are subjected to protein precipitation with MeOH, using stable isotopically labelled internal standards. The separation of JAKi in supernatants diluted 1:1 with ultrapure H2O was performed using a C18 column Xselect HSS T3 2.5 µm, 2.1x150 mm using a mobile phase composed of formic acid (FA) 0.2% and acetonitrile (+FA 0.1%) in gradient mode. The analytical run time for the multiplex assay was 7 min. JAKi drugs were monitored by electrospray ionization in the positive mode followed by triple-stage quadrupole MS/MS analysis. The method was validated according to SFSTP and ICH guidelines over the clinically relevant concentration ranges (0.5–200 ng/mL for abrocitinib, baricitinib and upadacitinib; 1–400 ng/mL for tofacitinib; 0.5–400 ng/mL for ruxolitinib, and 10–800 ng/mL for fedratinib). This multiplex HPLC-MS/MS assay achieved good performances in term of trueness (91.1-113.5%), repeatability (3.0-9.9%), and intermediate precision (4.5-11.3%).We developed and validated a highly sensitive method for the multiplex quantification of the JAKi abrocitinib, baricitinib, fedratinib, ruxolitinib, tofacitinib, and upadacitinib in human plasma. The method will be applied for prospective clinical pharmacokinetic studies to determine whether TDM programs for JAKi based on residual drug concentrations can be recommended using disease-specific therapeutic ranges.

Detection of doping peptides and basic drugs in equine urine using liquid chromatography-mass spectrometry.

The abuse of prohibited agents including peptides and basic small-molecule drugs is an area of great concern in horseracing due to their high potential to act as doping agents. These compound classes include agents such as growth hormone-releasing peptides, peptide analgesics, beta-2-adrenergic receptor agonists, and quaternary ammonium drugs that can be challenging to detect and regulate because of their chemical properties and potential rapid elimination following administration. The use of highly sensitive and selective analytical techniques such as liquid chromatography-mass spectrometry (LC-MS) is necessary to provide coverage of these substances and their potential metabolites. This study describes the development and validation of methodology capable of the detection of over 50 different peptide-based doping agents, related secretagogues, quaternary ammonium drugs, and other challenging small molecules in equine urine following solid-phase extraction using a mixed mode weak cation exchange sorbent. Following sample extraction, the compounds were analyzed using LC-MS with chromatographic separation via a reverse phase gradient and detection via selective reaction monitoring following introduction to a triple-stage quadrupole mass spectrometer using positive mode electrospray ionization. Validation parameters including limits of detection and quantitation, accuracy, precision, linear range, recovery, stability, and matrix effects were determined. Briefly, the limits of detection for most compounds were in the sub-ng/mL ranges with adequate precision and accuracy sufficient for an initial testing procedure. Stability studies indicated that most compounds were sufficiently stable to allow for effective screening using conditions commonly utilized in drug testing laboratories.

Comparative pharmacokinetics of six bioactive components of Shen-Wu-Yi-Shen tablets in normal and chronic renal failure rats based on UPLC-TSQ-MS/MS

Ethnopharmacological relevanceShen-Wu-Yi-Shen tablets (SWYST), a Chinese patent medicine consisting of 12 herbal medicines, was formulated by a famous TCM nephrologist, Zou Yunxiang. It is clinically used to improve the symptoms of nausea, vomiting, poor appetite, dry mouth and throat, and dry stool in patients with chronic renal failure (CRF) accompanied by qi and yin deficiency, dampness, and turbidity. SWYST can reduce urea nitrogen, blood creatinine, and urinary protein loss, and increase the endogenous creatinine clearance rate. However, little is known about its pharmacokinetics. Aim of studyTo compare the pharmacokinetics of six bioactive components after oral administration of SWYST in normal and adenine-induced CRF rats. Materials and methodsA method based on ultra-performance liquid chromatography coupled with a triple-stage quadrupole mass spectrometer (UPLC-TSQ-MS/MS) was developed and validated to determine the six bioactive compounds (albiflorin, paeoniflorin, plantagoguanidinic acid, rhein, aloe-emodin, and emodin) in rat plasma. Rat plasma samples were prepared using protein precipitation. Chromatography was performed on an Agilent Eclipse Plus C18 column (3.0 × 50 mm, 1.8 μm) using gradient elution with a mobile phase composed of acetonitrile and water containing 0.1% (v/v) formic acid, while detection was achieved by electrospray ionization MS under the multiple selective reaction monitoring modes. After SWYST administration, rat plasma was collected at different time points, and the pharmacokinetic parameters of six analytes were calculated and analyzed based on the measured plasma concentrations. ResultsThe UPLC-TSQ-MS/MS method was fully validated for its satisfactory linearity (r ≥ 0.9913), good precisions (RSD <11.5%), and accuracy (RE: -13.4∼13.1%), as well as acceptable limits in the extraction recoveries, matrix effects, and stability (RSD <15%). In normal rats, the six analytes were rapidly absorbed (Tmax ≤ 2 h), and approximately 80% of their total exposure was eliminated within 10 h. Moreover, in normal rats, the AUC0-t and Cmax of albiflorin, plantagoguanidinic acid, and rhein exhibited linear pharmacokinetics within the dose ranges, while that of paeoniflorin is non-linear. However, in CRF rats, the six analytes exhibited reduced elimination and significantly different AUC or Cmax values. These changes may reflect a decreased renal clearance rate or inhibition of drug-metabolizing enzymes and transporters in the liver and gastrointestinal tract caused by CRF. ConclusionsA sensitive UPLC-TSQ-MS/MS method was validated and used to investigate the pharmacokinetics of SWYST in normal and CRF rats. This is the first study to investigate the pharmacokinetics of SWYST, and our findings elucidate the causes of their different pharmacokinetic behaviors in CRF rats. Furthermore, the results provide useful information to guide further research on the pharmacokinetic-pharmacodynamic correlation and clinical application of SWYST.

Potential profound fluctuation in tacrolimus concentration on consumption of pomegranate rind extract: A Pharmacokinetic Experiment.

Background: Presently, varied case reports demonstrated an increase or decrease in blood concentration of diverse conventional drugs, often co-administered with edible fruits, spices, or vegetables. The overarching aim of this research is to elucidate the fluctuations in tacrolimus (TAC) blood concentration on the consumption of pomegranate rind extract (PRE). Methods: A pharmacokinetic (PK) study was conducted with two groups, vis-a-vis PRE + TAC (3mg/kg) and TAC (3mg/kg) alone groups. An experimental study was conducted in three different manners: Single-dose (S) PRE (200mg/kg), 7-day repetitive (7-R) PRE (200mg/kg) dosing, and multiple (M) PRE doses (100, 200, 400, and 800mg/kg). All the blood samples (approximately 300μl) were drawn at different time intervals, i.e., 30min, 1, 2, 4, 8, and 12h after oral administration of TAC (3mg/kg). The estimation of TAC in rat plasma was done using the hyphenated technique LC-MS/MS where the mass spectrometer used was a triple-stage quadrupole in multiple-reaction monitoring (MRM) mode. Results: The findings depict that in comparison with the TAC (3mg/kg) alone group with the 7-day repetitive (7-R) PRE (200mg/kg) dosing, the Cmax was found to be 9.03 ± 1.21ng/ml; AUC from time zero to infinity (AUC0-∞), 61.91 ± 17.37ngh/ml, while the TAC (3mg/kg) + PRE group exhibited an increase in PK parameters of TAC (Cmax 22.48 ± 3.07ng/ml; AUC0-∞ 153.08 ± 13.24ngh/ml). The authors further investigated in what manner the PRE affects the PK of TAC in animals. For this, docking studies with major phytoconstituents present in the PRE with CYP3A4 isoenzyme were carried out. Ellagitannins (dock score, -11.64) and punicalagin (dock score, -10.68) were again used for molecular simulation studies with TAC. To validate our findings, a CYP3A4 inhibitory in vitro assay was conducted. Conclusion: Based on the integrated in vivo and in silico studies, we concluded that pomegranate rind extract interacts strongly with CYP isoenzyme and is therefore responsible for the altered PK profile of TAC.

A fast and simple extraction method for analysing levoglucosan and its isomers in sediments by ion chromatography tandem mass spectrometry.

The ability to trace current and past biomass burning events is important for understanding the links between human activity, fire frequency, and climate. One method of tracing biomass burning is to measure the concentrations of certain monosaccharides anhydrides (MAs), specifically levoglucosan (LEV) and its isomers, mannosan (MAN) and galactosan (GAL), which are products of cellulose and hemicellulose pyrolysis. This work presents a simple extraction method allowing for the rapid, sensitive, and selective determination of MAs in sediments. MAs detection was performed using suppressed ion chromatography with electrospray - triple-stage quadrupole tandem mass spectrometry (IC-TSQ-MS). The extraction method involves ultrasound probe sonication using water as the solvent. Extraction time, amplitude, and sonication mode were optimised. Recoveries higher than 86% for all MAs tested were achieved by applying 70% amplitude in continuous mode for 60 s. Analytical performance of the method included instrumental LODs of 0.10, 0.12 and 0.50 μg L-1 for LEV, MAN and GAL, respectively. No carryover issues, no matrix effect and no co-elution of targeted MAs with other sugars likely present in sediments samples were observed. The developed extraction method was further validated by the analysis of LEV and MAN in NIST® 1649b urban dust reference material and the resulting concentrations were in excellent agreement with previously reported values. MAs quantification in 70 lake sediment samples were carried out with concentrations found to range from 0.009 to 0.390 μg g-1 for LEV and from 0.009 to 0.194 μg g-1 for MAN. Plotting MAs concentrations versus approximate sediment age allowed the reconstruction of recent fire events impacting two locations in the Central Highlands of Tasmania, Australia.

Triple-stage Quadrupole Mass Spectrometer to Determine Ubiquitously Present Per- and Polyfluorinated Alkyl Substances in Drinking Water at Part Per Trillion Levels Using Solid Phase Extraction Approach.

An accurate analytical method was developed to determine selected per- and polyfluorinated alkyl substances (PFAS) at the level of parts per trillion (ppt or ng/L) in drinking water. The method included a concentration step using solid phase extraction (SPE) approach in combination with a liquid chromatography-tandem mass spectrometry system (LC-MS/MS). This method was optimized and validated for the common PFAS contaminants in drinking water. An initial demonstration of capability was established with an acceptable initial calibration, minimum reporting limit (MRL), limit of detection (LOD), initial demonstration of low system background, and initial demonstration of precision (IDP). Isotopically labeled internal standards were used for quantification. Surrogate standards were used to monitor method performance. The current method will help in better understanding of PFAS crisis by providing an efficient measurement of PFAS in water. In this study, the recoveries of four surrogates were between 84 and 113%, and calculated limit of detection (DL) and minimum reporting limits (MRL) were generally 1.0-3.0 and 5-10 ng/L, respectively.

Water coordinated on Cu(I)-based catalysts is the oxygen source in CO2 reduction to CO

Catalytic reduction of CO2 over Cu-based catalysts can produce various carbon-based products such as the critical intermediate CO, yet significant challenges remain in shedding light on the underlying mechanisms. Here, we develop a modified triple-stage quadrupole mass spectrometer to monitor the reduction of CO2 to CO in the gas phase online. Our experimental observations reveal that the coordinated H2O on Cu(I)-based catalysts promotes CO2 adsorption and reduction to CO, and the resulting efficiencies are two orders of magnitude higher than those without H2O. Isotope-labeling studies render compelling evidence that the O atom in produced CO originates from the coordinated H2O on catalysts, rather than CO2 itself. Combining experimental observations and computational calculations with density functional theory, we propose a detailed reaction mechanism of CO2 reduction to CO over Cu(I)-based catalysts with coordinated H2O. This study offers an effective method to reveal the vital roles of H2O in promoting metal catalysts to CO2 reduction.

Internal exposure to perfluoroalkyl substances (PFAS) in vegans and omnivores.

Perfluoroalkyl substances (PFAS) are a complex group of anthropogenic compounds with exceptional properties. Due to their high persistence and mobility, they have caused ubiquitous environmental contamination and in part accumulate in the food chain. In the general population, diet is the main source of PFAS exposure, with the important sources fish and meat. As a vegan diet implies the complete exclusion of any animal products, it might be expected that vegans have lower blood levels of PFAS compared to omnivores. Furthermore, lower levels of cholesterol is one of the well-documented nutritional effects in vegans, but cholesterol levels were also found to be associated with higher PFAS levels in epidemiological studies. To examine the relations of internal PFAS levels and the levels of cholesterol in vegans and omnivores, the cross-sectional "Risks and Benefits of a Vegan Diet" (RBVD) study was used involving 36 vegans and 36 omnivores from Berlin/Germany. Nine perfluoroalkyl substances were quantified in plasma using a triple-stage quadrupole mass spectrometer. Lower median plasma concentrations were found in vegans compared to omnivores for perfluorooctane sulfonic acid (PFOS) (2.31 vs. 3.57ng/ml, respectively; p=0.02) and for perfluorononanoic acid (PFNA) (<0.25 vs. 0.41ng/ml, respectively; p<0.0001). No significant differences of the median concentrations were observed for perfluorooctanoic acid (PFOA) (1.69 vs. 1.44ng/ml, respectively, p=0.26) and perfluorohexane sulfonic acid (PFHxS) (1.96 vs. 1.79ng/ml, respectively; p=0.70). The strongest correlations with food groups, derived from a food frequency questionnaire, were observed between levels of PFOA and water consumption (in case of the total study population, n=72), and between levels of PFOS as well as PFNA and the consumption of 'meat and meat products' (in case of the omnivores, n=36). Levels of Low Density Lipoprotein (LDL) cholesterol were confirmed to be considerably lower in vegans compared to omnivores (86.5 vs. 115.5mg/dl, respectively; p=0.001), but no associations between the four main PFAS and LDL cholesterol were observed (all p>0.05) at the low exposure level of this study. According to the results of our study, a vegan diet may be related to lower PFAS levels in plasma. We highlight the importance of the adjustment of dietary factors like a vegan diet in case of epidemiological studies dealing with the impact of PFAS on the levels of blood lipids.

Ceramide Analysis by Multiple Linked-Scan Mass Spectrometry Using a Tandem Quadrupole Instrument.

Ceramides are a special class of sphingolipids and play a central role in sphingolipid metabolism, and have diverse structures. In this book chapter, tandem quadrupole mass spectrometric approaches applying multiple linked scannings including various constant neutral loss scan (NLS) and precursor ion scan (PIS), the unique applicable feature of a triple-stage quadrupole (TSQ) instrument for analysis of ceramides desorbedas [M-H]- and [M+Li]+ ions are described. Thesemultiple dimensional tandem mass spectrometric approaches are fully adapted to the conventional shotgun lipidomics workflow with minimal or without prior chromatographic separation to profile ceramide molecules, and thus detection of a whole class of ceramide or various specific ceramide subclasses in crude lipid extractcan be achieved. With addition of internal standard(s), semi-quantitation of ceramide in the lipid extract of biological origin is possible. Examples have shown promise in ceramide profiling of several whole lipid extracts from porcine brain, the model Dictyostelium Discoideum cells for cancer study, and skin.

A fast and simple method for the simultaneous analysis of midazolam, 1-hydroxymidazolam, 4-hydroxymidazolam and 1-hydroxymidazolam glucuronide in human serum, plasma and urine

For the quantification of the sedative and anesthetic drug midazolam and its main (active) metabolites 1-hydroxymidazolam, 4-hydroxymidazolam and 1-hydroxymidazolam glucuronide in human serum, human EDTA plasma, human heparin plasma and human urine a single accurate method by ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) has been developed. Protein precipitation as sample preparation, without the need of a time-consuming deglucuronidation step for the quantification of 1-hydroxymidazolam glucuronide, resulted in a simple and rapid assay suitable for clinical practice with a total runtime of only 1.1 min. The four components and the isotope-labeled internal standards were separated on a C18 column and detection was performed with a triple-stage quadrupole mass spectrometer operating in positive ionization mode.The method was validated based on the “Guidance for Industry Bioanalytical Method Validation” (Food and Drug Administration, FDA) and the “Guideline on bioanalytical method validation” of the European Medicines Agency (EMA). Linearity was proven over the ranges of 5–1500 μg/L for midazolam, 1-hydroxymidazolam and 4-hydroxymidazolam and 25–5000 μg/L for 1-hydroxymidazolam glucuronide, using a sample volume of 100 μL. Matrix comparison indicated that the assay is also applicable to other human matrices like EDTA and heparin plasma and urine. Stability experiments showed good results for the stability of midazolam, 1-hydroxymidazolam and 1-hydroxymidazolam glucuronide in serum, EDTA and heparin plasma and urine stored for 7 days under different conditions. At room temperature, 4-hydroxymidazo-lam is stable for 7 days in EDTA plasma, but stable for only 3 days in serum and heparin plasma and less than 24 h in urine. All four compounds were found to be stable in serum, EDTA plasma, heparin plasma and urine for 7 days after sample preparation and for 3 freeze–thaw cycles. The assay has been applied in therapeutic drug monitoring of midazolam for (pediatric) intensive care patients.

Method comparison of HPLC-ninhydrin-photometry and UHPLC-PITC-tandem mass spectrometry for serum amino acid analyses in patients with complex congenital heart disease and controls

IntroductionMetabolomics studies are not routine when quantifying amino acids (AA) in congenital heart disease (CHD).ObjectivesComparative analysis of 24 AA in serum by traditional high-performance liquid chromatography (HPLC) based on ion exchange and ninhydrin derivatisation followed by photometry (PM) with ultra-high-performance liquid chromatography and phenylisothiocyanate derivatisation followed by tandem mass spectrometry (TMS); interpretation of findings in CHD patients and controls.MethodsPM: Sample analysis as above (total run time, ~ 119 min). TMS: Sample analysis by AbsoluteIDQ® p180 kit assay (BIOCRATES Life Sciences AG, Innsbruck, Austria), which employs PITC derivatisation; separation of analytes on a Waters Acquity UHPLC BEH18 C18 reversed-phase column, using water and acetonitrile with 0.1% formic acid as the mobile phases; and quantification on a Triple-Stage Quadrupole tandem mass spectrometer (Thermo Fisher Scientific, Waltham, MA) with electrospray ionisation in the presence of internal standards (total run time, ~ 8 min). Calculation of coefficients of variation (CV) (for precision), intra- and interday accuracies, limits of detection (LOD), limits of quantification (LOQ), and mean concentrations.ResultsBoth methods yielded acceptable results with regard to precision (CV < 10% PM, < 20% TMS), accuracies (< 10% PM, < 34% TMS), LOD, and LOQ. For both Fontan patients and controls AA concentrations differed significantly between methods, but patterns yielded overall were parallel.ConclusionSerum AA concentrations differ with analytical methods but both methods are suitable for AA pattern recognition. TMS is a time-saving alternative to traditional PM under physiological conditions as well as in patients with CHD.Trial registration numberClinicalTrials.gov Identifier NCT03886935, date of registration March 27th, 2019 (retrospectively registered).

Sample extraction and liquid chromatography–tandem mass spectrometry (LC-MS/MS) method development and validation for the quantitative detection of cyanobacterial hepatotoxins and neurotoxins in Singapore's reservoirs

Cyanobacterial blue–green algal toxins are produced by harmful algal blooms (HABs). Most species of phytoplankton are not harmful, but excessive amounts of certain HAB taxa can cause harm to human and animal health, aquatic ecosystems and local economies. To investigate the prevalence of cylindrospermopsin (CYN) and anatoxin-a (ANA) in Singapore’s reservoirs, a hazard analysis was initiated to profile the CYN and ANA levels present. Water samples from 17 reservoirs were monitored monthly over a 12-month period (November 2012–October 2013). Analyses were conducted by liquid chromatography–tandem mass spectrometry (LC-MS/MS) using a triple-stage quadrupole mass spectrometer with a turbo-assisted ion spray source. CYN was more prevalent than ANA. Intracellular CYN concentrations exceeded 0.4μgL–1 in 6 of 17 man-made reservoirs surveyed, and slightly exceeded the provisional CYN drinking water guidelines of 1μgL–1 (National Health and Medical Research Council and National Resource Management Ministerial Council 2011) on one occasion (1.1μgL–1, July 2013) in one reservoir. The dominant cyanobacteria genera during that period were Cylindrospermopsis, Planktolyngbya, Pseudanabaena and Microcystis. For ANA, all 17 reservoirs had concentrations below 0.1μgL–1. Based on random forest analysis, the most important environmental factors affecting CYN concentrations were total nitrogen (most important), nitrate, total phosphorus and Cylindrospermopsis counts (least important). The findings of this study indicate that reducing total nitrogen concentrations may be useful in minimising CYN concentrations in tropical reservoirs.

Preclinical Evaluation of Ropivacaine in 2 Liposomal Modified Systems.

Our research group has recently developed liposomes with ionic gradient and in a combined manner as donor and acceptor vesicles containing ropivacaine (RVC; at 2% or 0.75%). Looking for applications of such novel formulations for postoperative pain control, we evaluated the duration of anesthesia, pharmacokinetics, and tissue reaction evoked by these new RVC formulations. The formulations used in this study were large multivesicular vesicle (LMVV) containing sodium acetate buffer at pH 5.5 or in a combined manner with LMVV as donor and large unilamellar vesicles (LUVs) as acceptor vesicles with an external pH of 7.4. Wistar rats were divided into 6 groups (n = 6) and received sciatic nerve block (0.4 mL) with 6 formulations of RVC (LMVVRVC0.75%, LMVV/LUVRVC0.75%, LMVVRVC2%, LMVV/LUVRVC2%, RVC 0.75%, and RVC 2%). To verify the anesthetic effect, the animals were submitted to the pain pressure test and the motor block was also monitored. Histopathology of the tissues surrounding the sciatic nerve region was also assessed 2 and 7 days after treatment. Rats (n = 6) were submitted to a hind paw incision, and mechanical hypersensitivity was measured via the withdrawal response using von Frey filaments after injection of the 6 formulations. Finally, New Zealand white rabbits (n = 6) received sciatic nerve block (3 mL) with 1 of the 6 formulations of RVC. Blood samples were collected predose (0 minutes) and at 15, 30, 45, 60, 90, 120, 180, 240, 300, 360, 420, 480, and 540 minutes after injection. RVC plasma levels were determined using a triple-stage quadrupole mass spectrometer. Duration and intensity of the sensory block were longer with all liposomal formulations, when compared to the plain RVC solution (P < .05). Histopathological evaluation showed greater toxicity for the positive control (lidocaine 10%), when compared to all formulations (P < .05). After the hind paw incision, all animals presented postincisional hypersensitivity and liposomal formulations showed longer analgesia (P < .05). LMVVRVC0.75% presented higher time to reach maximum concentration and mean residence time than the remaining formulations with RVC 0.75% (P < .05), so LMVV was able to reduce systemic exposure of RVC due to slow release from this liposomal system. All new liposomal formulations containing 0.75% RVC were able to change the pharmacokinetics and enhance anesthesia duration due to slow release of RVC from liposomes without inducing significant toxic effects to local tissues.

Electrochemical oxidation of perfluorooctane sulfonate (PFOS) substitute by modified boron doped diamond (BDD) anodes

BDD/SnO2-F electrode was synthesized by spin-coating and heat treatment, and it was characterized by scanning electron microscope-energy dispersive spectrometer (SEM-EDS), X-ray diffraction (XRD), and linear sweep voltammetry (LSV). Electrochemical oxidation of chlorinated polyfluorinated ether sulfonate (F-53B, ClCF2CF2CF2CF2CF2CF2-O-CF2CF2-SO3K) as PFOS substitute was carried out with the BDD, BDD/SnO2, BDD/PbO2, and BDD/SnO2-F anodes. The effects of F and Sn ratio, current density, initial pH, initial F-53B concentration, the supporting electrolyte, NaCl concentration, and electrode distance on the degradation ratios were investigated. The defluorination and the desulfurization were evaluated by ion chromatography and UV–vis spectra. The intermediate products were analyzed on the liquid chromatography system coupled with a triple-stage quadrupole mass spectrometer (LC-MS/MS). The experimental results showed that BDD/SnO2-F had the most outstanding electrochemical oxidation performance. After introduction of F into SnO2 crystal, the grain size of SnO2-F was reduced, the oxygen evolution potential was increased by 0.13 V, and the degradation ratio of F-53B was increased by 15.07%. Under the optimum operating parameters, F-53B removal ratio of 95.6% was obtained, and the defluorination ratio reached 61.39% with NaCl supporting electrolyte concentration of 1 mol/L at current density of 30 mA/cm2 after electrolysis of 30 min. Finally, the electrochemical degradation pathways and the mechanisms of F-53B were proposed based on the intermediate products and the previous literature reported. This study indicates that the electrochemical performance of BDD anode can be much improved by modification with SnO2-F, and BDD/SnO2-F anode is kind of effective anode to decompose refractory compounds.

Small-Seeded Legumes as a Novel Food Source. Variation of Nutritional, Mineral and Phytochemical Profiles in the Chain: Raw Seeds-Sprouted Seeds-Microgreens.

Growing public concerns about health haves prompted the search for novel food sources. The study is focused on the seeds, sprouted seeds and microgreens of Trifolium pratense, T. medium, Medicago sativa, M. lupulina, Onobrychis viciifolia, Astragalus glycyphyllos and A. cicer species as a potential source of value-added food ingredientsr. The samples were analysed for nutritional (wet chemistry, standard methods) and mineral (atomic absorption spectroscopy, UV-Vis spectrophotometry) profiles, isoflavones (ultra-performance liquid with diode array detector –UPLC-DAD), coumestrol (UPLC-DAD), condensed tannins (CT) (vanillin-H2SO4 assay) and triterpene saponins (UPLC with triple-stage quadrupole MS). In our study, each species displayed high, but species-dependent nutritional, mineral and phytochemical value. All counterparts of legumes were mineral and protein rich. A. glycyphyllos samples, especially seeds, were abundant in iron. Trifolium spp. were found to be important sources of isoflavones, Medicago spp. of coumestrol and saponins, and O. viciifolia of CT. The protein and phytochemical contents increased and total carbohydrates decreased from seeds to microgreens.Our findings proved for the first time that seeds, sprouted seeds, and especially microgreens of small-seeded legumes are promising new sources of ingredients for fortification of staple foods with bioactive compounds, minerals and nutrients.

Quantification of the next-generation oral anti-tumor drugs dabrafenib, trametinib, vemurafenib, cobimetinib, pazopanib, regorafenib and two metabolites in human plasma by liquid chromatography-tandem mass spectrometry

A sensitive and selective method of high performance liquid chromatography (HPLC) coupled to tandem mass spectrometry (MS/MS) has been developed for the simultaneous quantification of six anticancer protein kinase inhibitors (PKIs), dabrafenib, trametinib, vemurafenib, cobimetinib, pazopanib, regorafenib, and two active metabolites (regorafenib-M2 and regorafenib-M5) in human plasma. Plasma protein precipitation with methanol enables the sample extraction of 100 μL aliquot of plasma. Analytes are detected by electrospray triple-stage quadrupole mass spectrometry and quantified using the calibration curves with stable isotope-labeled internal standards. The method was validated based on FDA recommendations, including assessment of extraction yield (74–104%), matrix effects, analytical recovery (94–104%) with low variability (<15%). The method is sensitive (lower limits of quantification within 1 to 200 ng/mL), accurate (intra- and inter-assay bias: −0.3% to +12.7%, and −3.2% to +6.3%, respectively) and precise (intra- and inter-assay CVs within 0.7–7.3% and 2.5–8.0%, respectively) over the clinically relevant concentration range (upper limits of quantification 500 to 100,000 ng/mL). This method is applied in our laboratory for both clinical research programs and routine therapeutic drug monitoring service of PKIs.

A Comparative Pharmacokinetic Study by UHPLC-MS/MS of Main Active Compounds after Oral Administration of Zushima-Gancao Extract in Normal and Adjuvant-Induced Arthritis Rats.

A sensitive and rapid ultra high-performance liquid-chromatography tandem mass spectrometry (UHPLC-MS/MS) method has been applied to investigate the influence of rheumatoid arthritis (RA) on the pharmacokinetics of nine analytes (daphnetin, daphnoretin, 7-hydroxycoumarin, liquiritin, isoliquiritin, liquiritigenin, isoliquiritigenin, glycyrrhizin, and glycyrrhetinic acid), which are major active components in Zushima-Gancao extract. The analytes and internal standard (IS) were separated in a Hypersil Gold C18 column and detected on a triple-stage quadrupole mass spectrometer using the validated method. All analytes exhibited good linearities (R2 > 0.98), and the lower limit of quantification (LLOQs) were sufficient for quantitative analysis. Intra- and inter-batch precision were all within 14.96% while the accuracy of nine analytes ranged from −17.99 to 14.48%, and these results were all within acceptance criteria. The extraction recoveries, matrix effects, and stabilities were all satisfactory. Main pharmacokinetic parameters of each compound were compared, and significant differences were found in parameters of daphnetin, daphnoretin, liquiritin, isoliquiritin, isoliquiritigenin, glycyrrhizin, and glycyrrhetinic acid, especially the last one, between the two groups. Therefore, adjuvant-induced arthritis has different effects on the pharmacokinetics of ingredients in Zushima-Gancao extract. The comparative pharmacokinetic study between normal and adjuvant-induced arthritis rats might provide more comprehensive information to guide the clinical usage of Zushima-Gancao extract for treating RA.

Reliable and Easy-To-Use Liquid Chromatography-Tandem Mass Spectrometry Method for Simultaneous Analysis of Fluconazole, Isavuconazole, Itraconazole, Hydroxy-Itraconazole, Posaconazole, and Voriconazole in Human Plasma and Serum.

A fast and easy-to-use liquid chromatography-tandem mass spectrometry method for the determination and quantification of 6 triazoles [fluconazole (FLZ), isavuconazole (ISZ), itraconazole (ITZ), hydroxy-itraconazole (OH-ITZ), posaconazole (PSZ), and voriconazole (VRZ)] in human plasma and serum was developed and validated for therapeutic drug monitoring. Sample preparation was based on protein precipitation with acetonitrile and subsequent centrifugation. Isotope-labeled analogues for each analyte were used as internal standards. Chromatographic separation was achieved using a 50 × 2.1 mm, 1.9 μm polar Hypersil Gold C18 column and mobile phase consisting of 0.1% formic acid/acetonitrile (45%/55%, vol/vol) at a flow rate of 340 μL/min. The triazoles were simultaneously detected using a triple-stage quadrupole mass spectrometer operated in selected reaction monitoring mode with positive heated electrospray ionization within a single runtime of t = 3.00 minutes. Linearity of all azole concentration ranges was verified by the Mandel test and demonstrated for all azoles. All calibration curves were linear and fitted using least squares regression with a weighting factor of the reciprocal concentration. Limits of detection (μg/L/L) were FLZ, 9.3; ISZ, 0.3; ITZ, 0.6; OH-ITZ, 8.6; PSZ, 3.4; and VRZ, 2.1. The lower limits of quantitation (μg/L/liter) were FLZ, 28.3; ISZ, 1.0; ITZ, 1.7; OH-ITZ, 26.2; PSZ, 10.3; and VRZ, 6.3. Intraday and interday precisions ranged from 0.6% to 6.6% for all azoles. Intraday and interday accuracies (%bias) of all analytes were within 10.5%. In addition, we report on a 29-year-old white woman (94 kg body weight) with a history of acute myeloid leukemia who underwent stem cell transplantation. Because of diagnosis of aspergillus pneumonia, antifungal pharmacotherapy was initiated with different application modes and dosages of ISZ, and plasma concentrations were monitored over a time period of 6 months. A precise and highly sensitive liquid chromatography-tandem mass spectrometry method was developed that enables quantification of triazoles in plasma and serum matrix across therapeutically relevant concentration ranges. It was successfully implemented in our therapeutic drug monitoring routine service and is suitable for routine monitoring of antifungal therapy and in severely ill patients.

ESTIMATION OF CURCUMINOIDS IN CURCUMA KARNATAKENSIS (WHITE TURMERIC) - AN ENDEMIC TAXON

Objective: To estimate the contents of curcuminoids in two samples of Curcuma karnatakensis collected from different localities.Methods: Quantification of curcuminoids was estimated by ultra-high performance liquid chromatography-mass spectrometry/selected reaction monitoring (UHPLC-MS/SRM) method in two samples of C. karnatakensis for the first time. Fine powder of rhizomatous rootstocks of two plant samples collected from different habitats were served as Samples A and B. The MS used for the metabolite analysis is a Vantage TSQ triple stage quadrupole MS equipped with heated electrospray ionization. The MS is coupled with an Agilent 1290 infinity UHPLC system. A stock solution of curcuminoid standard was prepared by dissolving 5 mg of standard in 1 mL of methanol. Seven different concentrations of standard (0.15-10 ng on column) were injected for the UHPLC-MS/SRM analysis. Separations were performed using a C-18 column with a flow rate of 0.2 mL/minute.Results: Contents of curcumin, demethoxycurcumin, and bisdemethoxycurcumin were found to be varied in two samples and lowest than any other species of Curcuma studied. Variation in the contents may be due to their different habitats in which they are growing.Conclusion: The present attempt of analyzing the contents of curcuminoids in this endemic taxon for the first time will provide the basis for further pharmacological analysis to authenticate the efficacy of these active principles as the curcuminoids are known for varied pharmacological activities.

Development and validation of an LC-MS/MS method to quantify lysergic acid diethylamide (LSD), iso-LSD, 2-oxo-3-hydroxy-LSD, and nor-LSD and identify novel metabolites in plasma samples in a controlled clinical trial.

Lysergic acid diethylamide (LSD) is a widely used recreational drug. The aim of this study was to develop and validate a liquid chromatography tandem mass spectrometry (LC-MS/MS) method for the quantification of LSD, iso-LSD, 2-oxo-3-hydroxy LSD (O-H-LSD), and nor-LSD in plasma samples from 24healthy subjects after controlled administration of 100μg LSD in a clinical trial. In addition, metabolites that have been recently described in in vitro studies, including lysergic acid monoethylamide (LAE), lysergic acid ethyl-2-hydroxyethylamide (LEO), 2-oxo-LSD, trioxylated-LSD, and 13/14-hydroxy-LSD, should be identified. Separation of LSD and its metabolites was achieved on a reversed phase chromatography column after turbulent-flow online extraction. For the identification and quantification, a triple-stage quadrupole LC-MS/MS instrument was used. The validation data showed slight matrix effects for LSD, iso-LSD, O-H-LSD, or nor-LSD. Mean intraday and interday accuracy and precision were 105%/4.81% and 105%/4.35% for LSD, 98.7%/5.75% and 99.4%/7.21% for iso-LSD, 106%/4.54% and 99.4%/7.21% for O-H-LSD, and 107%/5.82% and 102%/5.88% for nor-LSD, respectively. The limit of quantification was 0.05ng/mL for LSD, iso-LSD, and nor-LSD and 0.1ng/mL for O-H-LSD. The limit of detection was 0.01ng/mL for all compounds. The method described herein was accurate, precise, and the calibration range within the range of expected plasma concentrations. LSD was quantified in the plasma samples of the 24 subjects of the clinical trial, whereas iso-LSD, O-H-LSD, nor-LSD, LAE, LEO, 13/14-hydroxy-LSD, and 2-oxo-LSD could only sporadically be detected but were too low for quantification.