High-performance Liquid Chromatography-mass Spectrometry Research Articles

Detection of Antagonistic Compounds Synthesized by Bacillus velezensis against Xanthomonas citri subsp. citri by Metabolome and RNA Sequencing.

Biological control of plant diseases has gained attraction for controlling various bacterial diseases at a field trial stage. An isolated endophytic bacterium, Bacillus velezensis 25 (Bv-25), from Citrus species had strong antagonistic activity against Xanthomonas citri subsp. citri (Xcc), which causes citrus canker disease. When Bv-25 was incubated in Landy broth or yeast nutrient broth (YNB), the ethyl acetate extract of Landy broth exhibited higher levels of antagonistic activity against Xcc compared to that of YNB. Therefore, the antimicrobial compounds in the two ethyl acetate extracts were detected by high performance liquid chromatography-mass spectrometry. This comparison revealed an increase in production of several antimicrobial compounds, including difficidin, surfactin, fengycin, and Iturin-A or bacillomycin-D by incubation in Landy broth. RNA sequencing for the Bv-25 grown in Landy broth were performed, and the differential expressions were detected for the genes encoding the enzymes for the synthesis of antimicrobial compounds, such as bacilysin, plipastatin or fengycin, surfactin, and mycosubtilin. Combination of metabolomics analysis and RNA sequencing strongly suggests that several antagonistic compounds, especially bacilysin produced by B. velezensis, exhibit an antagonistic effect against Xcc.

4-Hydroxybenzyl alcohol from Sinapis alba mustard seed meal extract for control of liverwort (Marchantia polymorpha)

Sinapis alba mustard seed meal extract is a by-product of mustard oil extraction, that is used for biodiesel and renewable jet fuel production. Recently, S. alba mustard seed meal has been shown to have herbicidal activity for a range of problematic weeds. The objectives of the present research were 1) to optimize the preparation of S. alba mustard seed meal extract; 2) to explore the applicability of S. alba mustard seed meal extract for control of liverwort, a common pest in ornamental plants greenhouses; and 3) to identify the major herbicidal compound(s) in these extracts. For extraction, methanol content in aqueous solution was optimized between 0% and 100%. Extracts prepared with 0–30% aqueous methanol provided 98–100% control of liverwort after single application and prevented liverwort plants from recovering. Extracts prepared using sequential extraction with 60% methanol provided an additional benefit of low particulate solutions. The efficiency of the produced S. alba mustard seed meal extract was comparable to two commercially available herbicides labeled for liverwort control under greenhouse conditions. Composition of S. alba mustard seed meal extracts was analyzed by high-performance liquid chromatography mass spectrometry and ion chromatography. 4-Hydroxybenzyl alcohol was identified as a main herbicidal compound based on the analysis and using commercially available reference solution. To the best of our knowledge, this is the first report of 4-hydroxybenzyl alcohol herbicidal activity toward nonvascular plants. The high concentrations of 4-hydroxybenzyl alcohol naturally produced from S. alba mustard seed meal, makes it a potential herbicide feedstock material.

Validation of Fourier Transform Infrared Microspectroscopy for the Evaluation of Enzymatic Cross-Linking of Bone Collagen.

Enzymatic cross-linking of the bone collagen is important to resist to crack growth and to increased flexural strength. In the present study, we proposed a new method for assessment of enzymatic cross-link based on Fourier transform infrared (FTIR) microspectroscopy that takes into account secondary structure of type I collagen. Briefly, femurs were collected from sham or ovariectomized mice and subjected either to high-performance liquid chromatography-mass spectrometry or embedded in polymethylmethacrylate, cut and analyzed by FTIR microspectroscopy. FTIR acquisition was recorded before and after ultraviolet (UV) exposure or acid treatment. In addition, femurs from a second animal study were used to compare gene expression of Plod2 and Lox enzymes and enzymatic cross-links determined by FTIR microspectroscopy. We evidenced here that intensities and areas of subbands located at ~1660, ~1680, and ~1690cm-1 were positively and significantly associated with the concentration of pyridinoline (PYD), deoxypyridinoline, or immature dihydroxylysinonorleucine/hydroxylysinonorleucine cross-links. Seventy-two hours exposure to UV light significantly reduced by ~86% and ~89% the intensity and area of the ~1660cm-1 subband. Similarly, 24h of acid treatment significantly reduced by 78% and 76% the intensity and area of the ~1690cm-1 subband. Plod2 and Lox expression were also positively associated to the signal of the ~1660 and ~1690cm-1 subbands. In conclusion, our study provided a new method for decomposing the amide I envelope of bone section that positively correlates with PYD and immature collagen cross-links. This method allows for investigation of tissue distribution of enzymatic cross-links in bone section.

Development and validation of a peptide-mapping procedure for a novel C1 esterase inhibitor

Peptide mapping is a key method for studying the primary structure of proteins. With its sensitivity to the slightest changes in the covalent structure of a protein, this method is applicable both to medicinal product identification at the control stage and to production process stability monitoring.The aim of the study was to develop and validate a peptide-mapping procedure for the identification of a novel highly glycosylated recombinant C1 esterase inhibitor.Materials and methods. The authors studied recombinant human C1 esterase inhibitor products and trypsin. The study involved peptide mapping using reverse-phase high-performance liquid chromatography and high-resolution mass spectrometry. The following statistics were calculated to evaluate the results: mean, standard deviation, and coefficient of variation. The validation parameters included specificity, precision, and robustness.Results. The authors tested several variants of sample preparation for tryptic digests, including additional N-glycanase treatment and complete deglycosylation, and established the optimal conditions for sample preparation and chromatographic separation of C1 esterase inhibitor peptides to obtain consistent chromatographic profiles (peptide maps). The authors identified characteristic peaks and the corresponding relative retention time and area ranges. The absolute retention time of the second (characteristic) peak was approximately 16.5–16.9 minutes. The relative retention times were 2.14–2.21 for peak 9, 2.55–2.64 for peak 12, 2.97–3.14 for peak 14, 3.11–3.29 for peak 15, and 6.20–6.63 for peak 28.Conclusions. The authors developed a peptide-mapping procedure for C1 esterase inhibitors and optimised the conditions to achieve an over 18-hour reduction in sample preparation time. This procedure met the established acceptance criteria for specificity, precision, and robustness.

Development of biomimetic hexapeptide functionalized monolithic material for capillary microextraction of microcystins in environmental waters

In this work, a histidine-tagged hexapeptide functionalized monolithic material is described for the efficient capillary microextraction (CME) of microcystins (MCs) from environment waters. 1-Vinylimidazole (VIM) monomer was utilized to synthesize the matrix polymer monolith by the copolymerization with divinylbenzene (DVB) crosslinker in the presence of radical initiator azobisisobutyronitrile (AIBN). The affinity sorbent was then prepared by a metal ion chelation-based multi-step approach, in which the histidine tag peptide Trp-His-Trp-Leu-Gln-Trp (WHWLQW) was functionalized on a Cu2+ modified polymer monolith via a chelation reaction between the imidazole group of histidine and Cu2+. The resultant WHWLQW functionalized monolithic material had high affinity, hierarchical porosity and good mechanical stability. Especially, it exhibited favorable enrichment capacity for MCs compared to other affinity materials as previously reported. With the optimized CME protocol, MCs at ultra-trace level can be analyzed by CME coupled to high performance liquid chromatography-mass spectrometry (HPLC-MS) in different aqueous environmental matrices. The average recoveries were ranged from 89.3% to 104.5% with relative standard deviations (RSD%) of 2.1–8.4, demonstrating high extraction efficiency and repeatability. The limits of detection (LODs) of MCs were determined to be 0.28–0.45 ng L-1. This novel biomimetic sorbent shows promising potential for the enrichment of ultra-trace pollutants in environment waters.

Effect of Chemical Refining on the Reduction of β-Carboline Content in Sesame Seed Oil.

β-carbolines (harman and norharman) are potentially mutagenic and have been reported in some vegetable oils. Sesame seed oil is obtained from roasted sesame seeds. During sesame oil processing, roasting is the key procedure to aroma enhancement, in which β-carbolines are produced. Pressed sesame seed oils cover most market share, while leaching solvents are used to extract oils from the pressed sesame cake to improve the utilization of the raw materials. β-carbolines are nonpolar heterocyclic aromatic amines with good solubility in leaching solvents (n-hexane); therefore, the β-carbolines in sesame cake migrated to the leaching sesame seed oil. The refining procedures are indispensable for leaching sesame seed oil, in which some small molecules can be reduced. Thus, the critical aim is to evaluate the changes in β-carboline content during the refining of leaching sesame seed oil and the key process steps for the removal of β-carbolines. In this work, the levels of β-carbolines (harman and norharman) in sesame seed oil during chemical refining processes (degumming, deacidification, bleaching and deodorization) have been determined using solid phase extraction and high performance liquid chromatography-mass spectrometry (LC-MS). The results indicated that in the entire refining process, the levels of total β-carbolines greatly decreased, and the adsorption decolorization was the most effective process in reducing β-carbolines, which might be related to the adsorbent used in the decolorization process. In addition, the effects of adsorbent type, adsorbent dosage and blended adsorbent on β-carbolines in sesame seed oil during the decolorization process were investigated. It was concluded that oil refining can not only improve the quality of sesame seed oil, but also reduce most of the harmful β-carbolines.

HIGH SERUM TRIMETHYLAMINE N-OXIDE LEVEL IS ASSOCIATED WITH ENDOTHELIAL DYSFUNCTION IN PATIENTS WITH HYPERTENSION

Objective: Trimethylamine N-oxide (TMAO) is a metabolite derived from gut microbial metabolism and dietary intake and plays important roles in cardiovascular disease pathogenesis. Endothelial dysfunction is associated with risk factors of cardiovascular disease in patients with hypertension. The present study aimed to determine the correlation between serum TMAO and endothelial function, measured by vascular reactivity index (VRI) in patients with hypertension. Design and method: The present study collected fasting blood samples from 110 patients with hypertension. Serum TMAO levels were determination of by high-performance liquid chromatography–mass spectrometry. The endothelial function and VRI values were measured using digital thermal monitoring test, and VRI values of >2.0, 1.0–1.9 and <1.0 indicated good, intermediate and poor vascular reactivity, respectively. Results: Ten hypertensive patients (9.1%) were categorized as poor vascular reactivity (VRI < 1.0), 57 hypertensive patients (51.8%) were categorized as intermediate vascular reactivity (1.0 < VRI < 2.0), and 43 hypertensive patients had good vascular reactivity. Older age (P = 0.010), higher levels of serum total cholesterol (TCH, P = 0.007), low-density lipoprotein cholesterol (LDL-C, P = 0.009), and TMAO levels (P < 0.001) were associated with poor vascular reactivity. Furthermore, after multivariable forward stepwise linear regression analysis noted that logarithmically transformed serum level of TMAO (log-TMAO, adjusted R2 change = 0.274, P < 0.001) was significantly and independently associated with VRI values in hypertensive patients. Compared with good vascular reactivity index, patients with vascular reactivity dysfunction (intermediate vascular reactivity and poor vascular reactivity; odds ratio (OR) = 1.100; 95% confidence interval (CI) = 1.047–1.155; P < 0.001) or poor vascular reactivity (OR = 1.580, 95% CI = 1.002–2.492, P = 0.049) also noted TMAO was positively associated with poor vascular reactivity after multivariate logistic regression analysis. Conclusions: Serum TMAO levels were negatively associated with VRI and associated with endothelial dysfunction in patients with hypertension.

Microbial diversity, transformation and toxicity of azo dye biodegradation using thermo-alkaliphilic microbial consortia

In this research, the transformation and toxicity of Reactive Red 141 and 239 biodegraded under anaerobic-aerobic conditions as well as metagenomic analysis of Reactive Red 239 degrading microbial consortia collected from Shala Hot spring were investigated. Toxicity of dyes before treatment and after treatment on three plants, fish and microorganisms were done. A halotolerant and thermo-alkaliphilic bacterial consortia decolorizing azo dyes (>98% RR 141 and > 96% RR 239 in 7 h) under optimum conditions of salt concentration (0.5%), temperature (55 °C) and pH (9), were used. Toxicity effect of untreated dyes and treated dyes in Tomato > Beetroot > Cabbage plants, while the effect was Leuconostoc mesenteroides > Lactobacillus plantarum > Escherichia coli in microorganisms. Among fishes, the toxicity effect was highest in Oreochromis niloticus followed by Cyprinus carpio and Clarias gariepinus. The three most dominant phyla that could be in charge of decolorizing RR 239 under anaerobic-aerobic systems were Bacteroidota (22.6–29.0%), Proteobacteria (13.5–29.0%), and Chloroflexi (8.8–23.5%). At class level microbial community structure determination, Bacteroidia (18.9–27.2%), Gammaproteobacteria (11.0–15.8%), Alphaproteobacteria (2.5–5.0%) and Anaerolineae (17.0–21.9%) were dominant classes. The transformation of RR 141 and RR 239 into amine compounds were proposed via high performance liquid chromatography-mass spectroscopy (HPLC/MS) and fourier transform infrared spectroscopy (FT-IR). Overall, dye containing wastewaters treated under anaerobic-aerobic systems using thermo-alkaliphilic microbial consortia were found to be safe to agricultural (fishes and vegetables) purposes.

Plasma Metabolomics Predicts Chemotherapy Response in Advanced Pancreatic Cancer.

Pancreatic cancer (PC) is one of the deadliest cancers. Developing biomarkers for chemotherapeutic response prediction is crucial for improving the dismal prognosis of advanced-PC patients (pts). To evaluate the potential of plasma metabolites as predictors of the response to chemotherapy for PC patients, we analyzed plasma metabolites using high-performance liquid chromatography-mass spectrometry from 31 cachectic, advanced-PC subjects enrolled into the PANCAX-1 (NCT02400398) prospective trial to receive a jejunal tube peptide-based diet for 12 weeks and who were planned for palliative chemotherapy. Overall, there were statistically significant differences in the levels of intermediates of multiple metabolic pathways in pts with a partial response (PR)/stable disease (SD) vs. progressive disease (PD) to chemotherapy. When stratified by the chemotherapy regimen, PD after 5-fluorouracil-based chemotherapy (e.g., FOLFIRINOX) was associated with decreased levels of amino acids (AAs). For gemcitabine-based chemotherapy (e.g., gemcitabine/nab-paclitaxel), PD was associated with increased levels of intermediates of glycolysis, the TCA cycle, nucleoside synthesis, and bile acid metabolism. These results demonstrate the feasibility of plasma metabolomics in a prospective cohort of advanced-PC patients for assessing the effect of enteral feeding as their primary source of nutrition. Metabolic signatures unique to FOLFIRINOX or gemcitabine/nab-paclitaxel may be predictive of a patient's response and warrant further study.

Determination of human milk oligosaccharides by porous graphite carbon adsorption coupled with high-performance liquid chromatography-mass spectrometry

Determination of human milk oligosaccharides by porous graphite carbon adsorption coupled with high-performance liquid chromatography-mass spectrometry

Simultaneous quantitation of ketamine, norketamine and dehydronorketamine in human milk using a novel ultra high-performance liquid chromatography-mass spectrometry (UPLC-MS/MS) assay

There is a paucity of data on the transfer of ketamine from maternal blood into human milk. Quantification of ketamine in human milk provides information about the potential exposure of the infant to ketamine and its metabolites from the mother during lactation. A highly specific, reproducible, and sensitive UPLC-MS/MS based analytical method was developed and validated for the quantitation of ketamine and its metabolites (norketamine and dehydronorketamine) in human milk. Samples were subjected to a simple protein precipitation and ketamine-d4 and norketamine-d4 were used as internal standards. Separation of the analytes was achieved using an Acquity UPLC equipped with BEH RP18 1.7 µm, 2.1 × 100 mm column. Mass spectrometric analysis of the analyte ions was carried out using electrospray with positive ionization and multiple reaction monitoring mode. The assay was linear over a concentration range of 1–100 ng/mL for ketamine and norketamine, and 0.1–10 ng/mL for dehydronorketamine. Acceptable intra-day and inter-day accuracy and precision were observed for all the analytes. High recovery of the analytes and minimal matrix effect were observed. Stability of analytes was confirmed at the tested conditions. This assay was successfully used to measure analytes in human milk samples collected from lactating women enrolled in a clinical research study. This is the first validated method that simultaneously quantified ketamine and its metabolites in human milk.

Efficient Electrochemical Oxidation of Chloramphenicol by Novel Reduced TiO2 Nanotube Array Anodes: Kinetics, Reaction Parameters, Degradation Pathway and Biotoxicity Forecast.

The key component of electrochemical advanced oxidation technology are high-efficiency anodes, and highly efficient and simple-to-prepare materials have generated a lot of interest. In this study, novel self-supported Ti3+-doped titanium dioxide nanotube arrays (R-TNTs) anodes were successfully prepared by a two-step anodic oxidation and straightforward electrochemical reduction technique. The electrochemical reduction self-doping treatment produced more Ti3+ sites with stronger absorption in the UV-vis region, a band gap reduction from 2.86 to 2.48 ev, and a significant increase in electron transport rate. The electrochemical degradation effect of R-TNTs electrode on chloramphenicol (CAP) simulated wastewater was investigated. At pH = 5, current density of 8 mA cm-2, electrolyte concentration of 0.1 M sodium sulfate (Na2SO4), initial CAP concentration of 10 mg L-1, CAP degradation efficiency exceeded 95% after 40 min. In addition, molecular probe experiments and electron paramagnetic resonance (EPR) tests revealed that the active species were mainly •OH and SO4-, among which •OH played a major role. The CAP degradation intermediates were discovered using high-performance liquid chromatography-mass spectrometry (HPLC-MS), and three possible degradation mechanisms were postulated. In cycling experiments, the R-TNTs anode demonstrated good stability. The R-TNTs prepared in this paper were an anode electrocatalytic material with high catalytic activity and stability, which could provide a new approach for the preparation of electrochemical anode materials for difficult-to-degrade organic compounds.

Transcriptome Analysis Reveals the Genes Involved in Oxidative Stress Responses of Scallop to PST-Producing Algae and a Candidate Biomarker for PST Monitoring.

Paralytic shellfish toxins (PST) could be accumulated in bivalves and cause safety problems. To protect public health, bivalves are examined for PST contamination before entering the market, usually by high-performance liquid chromatography (HPLC) or LC-tandem mass spectrometry (LC-MS/MS) in the lab, which needs PST standards not all available and is time-consuming for large sample sizes. To detect PST toxicity in bivalves rapidly and sensitively, a biomarker gene is highly demanded, but the related study is very limited. In this study, we fed a commercially important bivalve, Patinopecten yessoensis, with the PST-producing dinoflagellate Alexandrium catenella. After 1, 3, and 5 days of exposure, both PST concentrations and toxicity levels in the digestive gland continuously increased. Transcriptome analysis revealed that the differentially expressed genes were significantly enriched in oxidation-reduction process, which included the cytochrome P450 genes (CYPs), type I iodothyronine deiodinase (IOD1s), peroxidasin (PXDN), and acyl-Coenzyme A oxidase 1 (ACOX1) at day 1 and a superoxide dismutase (SOD) at day 5, highlighting the crucial roles of these genes in response to oxidative stress induced by PST. Among the 33 continuously upregulated genes, five showed a significant correlation between gene expression and PST concentration, with the highest correlation present in PyC1QL4-1, the gene encoding Complement C1Q-like protein 4, C1QL4. In addition, the correlation between PyC1QL4-1 expression and PST toxicity was also the highest. Further analysis in another aquaculture scallop (Chlamys farreri) indicated that the expression of CfC1QL4-1, the homolog of PyC1QL4-1, also exhibited significant correlations with both PST toxicity and concentration. Our results reveal the gene expression responses of scallop digestive glands to PST-producing algae and indicate that the C1QL4-1 gene might be a potential biomarker for PST monitoring in scallops, which may provide a convenient way for the early warning and sensitive detection of PST contamination in the bivalves.

ELABORATION OF CHEMICAL SENSOR SENSITIVE TO BORON BENZILATE

Over the last decades, the development of simple and selective methods for the determination of organic analytes has been getting attention from analytical chemists. Particularly, the same applies to biologically active substances of natural and artificial origin, precursors, and metabolites of pharmaceuticals[1]. An example of such substance is benzylic acid (2, 2-diphenyl-2-hydroxyacetic acid) (BA), gravimetric reagent for zirconium [2], a product of hydrolysis and a metabolite of the antispasmodic and sedative drugs Pipoxolan, Benactyzine, or Metacin [3]. The complexes of BA have been proposed as corrosion inhibitors for some steels and aluminum alloys [4]. Its derivatives are being studied as new analgesics [5], anticancer drugs [6], and also as deep eutectic solvents [7]. The BA was used in the synthesis of the psychotropic combatant substance 3-quinuclidinyl benzilate (BZ), which causes mental slowing, and hydrolyzes to form BC [8-11]. For BA determination high-performance liquid chromatography and chromatography-mass spectrometry [12-15] are mostly used, hence the development of simple methods for determining benzylic acid is pertinent and relevant.
 Boric acid is known to form compounds with diols, α-hydroxy acids, dicarboxylic acids, and α-diketones [16] and that was used in developing potentiometric sensors for the determination of mandelic acid and vitamin C [17, 18]. As BA also forms complexes with boric acid and various metals [19, 20] the H3BO3 has been used in the present study as a reagent for obtaining boric acid benzilate ester - the analytical form of BA, as well as the ionic associate of its anion with tetraoctylammonium(TOA) as the active substance of a potentiometric PVC plasticized sensor sensitive to the mentioned above analytical form. The complex formation has been confirmed using IR spectroscopy. On the IR spectrum of BA, the absorption bands in the region of 3393. 69 cm-1 and 1715. 77 cm-1, which correspond to the vibrations of –ОН and >С=О groups, respectively have been detected [23]. At the same time, in the IR spectrum of the complex, the absorption band of –ОН groups in the region of 3300 - 3400 cm-1 was absent, that indicates their participation in the formation of ether bonds, additionally, the intensive bands of >C=O in the region of 1723. 41 cm-1 and 1742. 05 cm-1 has appeared. The ion pair of tetraoctylammonium cation with synthesized complex ion [B(Benz)2]- has been used as an active substance of the potentiometric sensor. The metrological and operational characteristics of the sensor and the optimal conditions of the analytical form obtaining have been systematically studied. The optimized composition of the sensitive PVC membrane was 0. 1% of IP, 33. 0% of PVC and 66. 9% of o-NFOE. For high yield [B(Benz)(ОН)2]--analytical form preparation the boiling of BA for 15 minutes in the medium of a buffer solution that consists of H3BO3 (sat.) - 0. 1 M NaOH - 0. 1 M H3PO4 with a pH of 5. 5 is needed. In optimal conditions, the slope of the sensor developed is close to the theoretical value of 55. 2 mV/pС, has a detection limit pCmin= 4. 9, and the linearity range 10-2 – 10-4mol/l. The response time of a sensor ranges from 5 to 30 seconds depending on the added quantity. High selectivity of the sensor to metal cations, anions of organic and inorganic acids, as well as polyatomic alcohols that are able to form complex compounds with BA or H3BO3 has been shown. The sensor developed was used for benzylic acid analysis in "Difenin", which can be formed during the synthesis of this drug [27].
 It has been shown that the use of potentiometry in combination with the conversion of the analyte into an active form using a cheap reagent - boric acid allows to develop simple methods of the organic analytes determination for which the use of potentiometry is not common. Preparation of the complex boronate ester of benzylic acid in the sample provides the selectivity of the technique to the target analyte.
 Keywords: benzylic acid; boron benzilate; potentiometric sensor; ion pair.

An interference-free SERS-based aptasensor for chlorpyrifos detection

In this study, we propose an interference-free SERS-based aptasensor for trace detection of chlorpyrifos (CPF) in real samples. In the aptasensor, gold nanoparticles coated with Prussian blue (Au@PB NPs) were employed as SERS tags to provide a sole and intense Raman emission at 2160 cm−1, which could avoid overlapping with the Raman spectrum of the real samples in 600–1800 cm−1 to improve the anti-matrix effect ability of the aptasensor. Under the optimum conditions, this aptasensor displayed a linear response for CPF detection in the range of 0.1–316 ng mL−1 with a low detection limit of 0.066 ng mL−1. In addition, the prepared aptasensor shows excellent application to determine CPF in cucumber, pear and river water samples. The recovery rates were highly correlated with high-performance liquid chromatography‒mass spectrometry (HPLC‒MS/MS). This aptasensor shows interference-free, specific and sensitive detection for CPF and offers an effective strategy for other pesticide residue detection.

Identification of human in vitro metabolites of the haemoglobin S polymerization inhibitor voxelotor for doping control purposes.

Voxelotor (GBT440) is a haemoglobin S polymerization inhibitor used to treat anaemia in sickle cell disease. Due to an increase of arterial oxygen saturation as well as serum erythropoietin and haemoglobin, the World Anti-Doping Agency included voxelotor in the list of prohibited substances and methods in 2023. The objective of the present study was to identify and characterize metabolites of voxelotor to detect a potential misuse by athletes. The biotransformation was studied in vitro using the human hepatocellular cell line HepG2 and pooled human liver microsomes. The metabolites were analysed using high-performance liquid chromatography (high-resolution) mass spectrometry. In total, three phase I metabolites and six phase II metabolites (resulting from glucuro-conjugation and O-methylation) were formed by the HepG2 cells in a time-dependent manner, and two phase I metabolites were generated by the liver microsomes, among them one also found in the HepG2 incubations. A reduced metabolite and the glucuro-conjugate of a reduced metabolite were the most abundant formed by HepG2 cells. In addition, metabolites resulting from mono-hydroxylation, reduction and O-methylation in different combinations were identified. Voxelotor was also found as glucuro-conjugate with a low abundance. With the spectrometric behaviour of voxelotor and its in vitro metabolites described herein, an implementation in doping control screening and, consequently, a detection of an abuse in an athlete urine sample might be possible.

Benzoin Resin: An Overview on Its Production Process, Phytochemistry, Traditional Use and Quality Control.

Benzoin is a pathologic exudation produced by plants of the family Styrax. It is secreted by traumatic resin ducts after injury, which are derived from parenchymal cells in secondary xylem by schizolysigeny. Some 63 chemical constituents have been isolated and identified from this resin, including balsamic acid esters, lignans and terpenoids. It has a long history of applications, including as incense along with olibanum, a flavor enhancer in the food industry, materials in the daily chemistry industry as well as therapeutic uses. Up to now, high-performance liquid chromatography (HPLC) and gas chromatography mass spectrometry (GC-MS) have been widely used in qualitative and quantitative analysis of benzoin. Other technologies, including near-infrared reflectance spectroscopy (NIR), proton transfer reaction time-of-flight mass spectrometry (PTR-ToF-MS) and Fourier-transform infrared spectroscopy (FT-IR), have also been used to distinguish different resins. Herein, this paper provides a comprehensive overview of the production process, phytochemistry, traditional uses and quality control of benzoin and looks to the future for promoting its further research and applications.

Enhanced degradation of quinoline in three-dimensional electro-Fenton system through NiCo2S4/g-C3N4 particles

The stability and toxicity of quinolines are difficult to degrade by conventional physicochemical and biological methods, posing a threat to human health and the environment. In this study, we prepared NiCo2S4/g-C3N4 particles and applied them in an electrochemical reactor to form a three-dimensional catalytic particle electro-Fenton system (3D-EF), which can efficiently remove quinoline from wastewater. The NiCo2S4/g-C3N4 catalytic particles were characterized by XRD, SEM, TEM, XPS. The optimum conditions for 3D-EF were 30 min reaction time, 60 g/L NiCo2S4/g-C3N4 particles dosage, pH value of 3, 67.6 mmol/L H2O2 concentration, 12.1 ms/cm conductivity and 5 A current. Under the optimum conditions, a chemical oxygen demand (COD) removal rate of 95.6% was achieved. NiCo2S4/g-C3N4 catalytic particles can be easily recovered by filtration and can be reused. Kinetic analysis showed that the COD degradation of quinoline solutions by the 3D-EF followed a first-order kinetic model. To determine the important role of hydroxyl radicals in the electrochemical process, electron paramagnetic reaction (EPR) and radical scavenging experiments were performed. Finally, in order to elucidate the degradation mechanism, the intermediates were identified by high performance liquid chromatography-mass spectrometry (HPLC-MS) and two possible degradation pathways were proposed. Meanwhile, the biochemical analysis of quinoline wastewater was also performed.

Circulating retinol and 25(OH)D contents and their association with symptoms in children with chronic tic disorders

The present study measured serum levels of vitamin A (VA) and vitamin D (VD) in children with chronic tic disorders (CTD) and investigated their potential association with CTD and comorbidity of attention deficit hyperactivity disorder (ADHD) and the association of their co-insufficiencies or deficiencies with CTD symptoms. A total of 176 children (131 boys and 45 girls, median age of 9 years) with CTD were recruited as the CTD group. During the same period, 154 healthy children were selected as the healthy control (HC) cohort. Circulating retinol and 25-hydroxyvitamin D (25[OH]D) levels were measured for all participants using high-performance liquid chromatography (HPLC) and tandem mass spectrometry. The Yale Global Tic Severity Scale (YGTSS) was employed for the assessment of tic status and CTD impairment. The Swanson, Nolan, and Pelham Rating Scale (SNAP-IV) and the Children’s Yale-Brown Obsessive–Compulsive Scale (CY-BOCS) were used to evaluate comorbidity symptoms. CTD pediatric participants exhibited markedly diminished circulating retinol and 25(OH)D levels compared to HCs. Moreover, VA and VD deficiencies and their co-insufficiencies/deficiencies were more prevalent in CTD participants than HCs. Circulating 25(OH)D levels were inversely proportional to the YGTSS motor tic scores. YGTSS scores in CTD children with only VA or VD insufficiency or deficiency or with VA and VD co-insufficiency/deficiency did not differ from those in CTD children with normal VA and VD. CTD children with comorbid ADHD displayed reduced circulating retinol and 25(OH)D concentrations and elevated prevalence of VD deficiency compared to CTD participants without comorbid ADHD. Lower serum retinol content was intricately linked to the presence of elevated CTD and comorbid ADHD. VA and VD deficiencies and their co-insufficiencies/deficiencies were markedly enhanced in CTD pediatric participants compared to HCs. Lower VA concentration was linked to the presence of enhanced CTD and comorbid ADHD. Therefore, children with CTD, especially with comorbid ADHD, may be at a higher risk of VA or VD deficiency, which may prompt the clinicians to consider whether blood tests for VA and VD in CTD children would be helpful for clinical care.

Natural Clearance of Chlamydia trachomatis Infection Is Associated With Distinct Differences in Cervicovaginal Metabolites.

Natural clearance of Chlamydia trachomatis in women occurs in the interval between screening and treatment. In vitro, interferon-γ (IFN-γ)-mediated tryptophan depletion results in C. trachomatis clearance, but whether this mechanism occurs in vivo remains unclear. We previously found that women who naturally cleared C. trachomatis had lower cervicovaginal levels of tryptophan and IFN-γ compared to women with persisting infection, suggesting IFN-γ-independent pathways may promote C. trachomatis clearance. Cervicovaginal lavages from 34 women who did (n = 17) or did not (n = 17) naturally clear C. trachomatis were subjected to untargeted high-performance liquid chromatography mass-spectrometry to identify metabolites and metabolic pathways associated with natural clearance. In total, 375 positively charged metabolites and 149 negatively charged metabolites were annotated. Compared to women with persisting infection, C. trachomatis natural clearance was associated with increased levels of oligosaccharides trehalose, sucrose, melezitose, and maltotriose, and lower levels of indoline and various amino acids. Metabolites were associated with valine, leucine, and isoleucine biosynthesis pathways. The cervicovaginal metabolome in women who did or did not naturally clear C. trachomatis is distinct. In women who cleared C. trachomatis, depletion of various amino acids, especially valine, leucine, and isoleucine, suggests that amino acids other than tryptophan impact C. trachomatis survival in vivo.