Development and validation of stability-indicating assay method and identification of force degradation products of glucagon-like peptide-1 synthetic analog Exenatide using liquid chromatography coupled with Orbitrap mass spectrometer.

Exenatide is a synthetic glucagon-like peptide 1 analog, widely used in the management of type 2 diabetes mellitus. The stability of pharmaceutical products is significantly impacted by various environmental stress conditions. The present study reports the development of a validated reverse-phase high-performance liquid chromatography (RP-HPLC) stability-indicating method for the identification of force degradation products (DPs) of synthetic glucagon-like peptide-1 analog Exenatide using UHPLC-Orbitrap fusionTM mass spectrometer. Force degradation studies were performed by subjecting Exenatide to various stress conditions, such as hydrolytic, oxidative, photolytic and thermal to investigate the stability indicating ability of the method. Significant degradation was observed during acidic, oxidative, photolytic and thermal stress conditions. Exenatide and its major DPs identification and characterization were demonstrated by employing LC-HRMS and MS/MS method. In total, five major stress DPs were characterized, and their fragmentation pathway was proposed using MS/MS studies. Finally, the proposed RP-HPLC method was validated as per ICH guidance.

Exploring the therapeutic potential of Thai medicinal plants: in vitro screening and in silico docking of phytoconstituents for novel anti-SARS-CoV-2 agents.

BACKGROUND: The high virulence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), responsible for coronavirus disease 2019 (COVID-19), has triggered global health and economic concerns. The absence of specific antiviral treatments and the side effects of repurposed drugs present persistent challenges. This study explored a promising antiviral herbal extract against SARS-CoV-2 from selected Thai medicinal plants based on in vitro efficacy and evaluated its antiviral lead compounds by molecular docking. METHODS: Twenty-two different ethanolic-aqueous crude extracts (CEs) were rapidly screened for their potential activity against porcine epidemic diarrhea virus (PEDV) as a surrogate using a plaque reduction assay. Extracts achieving ≥ 70% anti-PEDV efficacy proceeded to the anti-SARS-CoV-2 activity test using a 50% tissue culture infectious dose method in Vero E6 cells. Molnupiravir and extract-free media served as positive and negative controls, respectively. Potent CEs underwent water/ethyl acetate fractionation to enhance antiviral efficacy, and the fractions were tested for anti-SARS-CoV-2 performance. The fraction with the highest antiviral potency was identified using liquid chromatography-high-resolution mass spectrometry (LC-HRMS). Molecular docking analyses of these compounds against the main protease (Mpro) of SARS-CoV-2 (6LU7) were performed to identify antiviral lead molecules. The top three hits were further evaluated for their conformational stability in the docked complex using molecular dynamics (MD) simulation. RESULTS: The water fraction of mulberry (Morus alba Linn.) leaf CE (WF-MLCE) exhibited the most potent anti-SARS-CoV-2 efficacy with low cytotoxicity profile (CC50 of ~ 0.7 mg/mL), achieving 99.92% in pre-entry mode and 99.88% in postinfection treatment mode at 0.25 mg/mL. Flavonoids and conjugates were the predominant compounds identified in WF-MLCE. Molecular docking scores of several flavonoids against SARS-CoV-2 Mpro demonstrated their superior antiviral potency compared to molnupiravir. Remarkably, myricetin-3-O-ß-D-galactopyranoside, maragrol B, and quercetin 3-O-robinobioside exhibited binding energies of ~ - 9 kcal/mol. The stability of each ligand-protein complex of these compounds with the Mpro system showed stability during MD simulation. These three molecules were pronounced as antiviral leads of WF-MLCE. Given the low cytotoxicity and high antiviral potency of WF-MLCE, it holds promise as a candidate for future therapeutic development for COVID-19 treatment, especially considering its economic and pharmacological advantages.

Phenolic Profile, Bioactivities and In Silico Analysis of the Trunk Bark of Acacia Cyanophylla Lindl.

In the current investigation, total phenolics and flavonoids of the methanolic extract obtained from the trunk bark of Acacia cyanophylla Lindl. were quantified by LC-HRMS technique. DPPH and ABTS reagents were employed to assay the antioxidant potential. The anti-tyrosinase and anti-α-amylase potentials were also assayed. The findings revealed that thirteen polyphenolic compounds were detected in the methanolic extract with trans-taxifolin (23.2 g/kg), as the major constituent. A. cyanophylla extract displayed a higher activity with DPPH test (IC50=10.14±1.00 µg/mL) than with ABTS (IC50=15.27±2.09 µg/mL). The same extract also exhibited interesting α-amylase inhibitory action (IC50 value of 4.00±0.17 µg/mL). Moreover, methanolic trunk bark extract exerted strong anti-tyrosinase capacity with an IC50 of 5.12±0.41 µg/mL in comparison to kojic acid (IC50=10.22±0.85 µg/mL) used as positive control. The antioxidant, anti-tyrosinase and anti-α-amylase potentials of the methanolic extract of A. cyanophylla trunk bark were reinforced by in silico molecular docking analyses, which confirmed the results of the in vitro tests.

Chemical profiling of Anthriscus cerefolium (L.) Hoffm., biological potential of the herbal extract, molecular modeling and KEGG pathway analysis.

This study was designed to investigate chemical composition and biological activities of the Anthriscus cerefolium methanolic extract. Chemical characterization of the extracts was performed by LC-HRMS/MS analysis. Antimicrobial activities of the extract were investigated on six bacteria and eight fungi while antioxidant activity was assessed by six different assays. Anti-enzymatic activity of the methanolic extract was tested on five enzymes associated with therapy of neurodegenerative diseases and diabetes mellitus type 2. Cytotoxic properties of the extract were tested on human immortalized keratinocytes (HaCaT) and tumor cell lines (SiHa, MCF7, HepG2). Anti-inflammatory activity of the extract was assessed on bacteria mediated inflammation model using HaCaT cell line. Molecular docking studies of enzymes and KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway analysis were performed. The results showed that the obtained extract was rich in phenolic compounds (a total of seventy-two were identified), with malonyl-1,4-O-dicaffeoylquinic acid and 3,5-O-dicaffeoylquinic acid dominating in the sample. The extract expressed antimicrobial, antioxidant, anti-enzymatic, cytotoxic and anti-inflammatory properties. The identified compounds demonstrated strong binding to the acetylcholinesterase (AChE) and to a lesser extent, to the butyrylcholinesterase (BChE), glucosidase, amylase, and modestly, to tyrosinase. KEGG pathway analysis has shown that the certain phenolic compounds may be related to anti-tumor, anti-inflammatory and anti-microbial activities of the extract. The data obtained suggest that phenolic compounds of the extract and their mixtures should be considered for future research as ingredients in pharmaceutical and nutraceutical formulations.

Characterization of the Polyphenolic Profile in Tomato (Lycopersicon esculentum P. Mill) Peel and Seeds by LC-HRMS/MS.

Peel and seeds are the main byproducts from tomato (Lycopersicon esculentum P. Mill) processing with high concentrations of polyphenols that have been underexploited. Herein, polyphenolic profiles in tomato peel and seeds were elucidated by untargeted liquid chromatography coupled to high-resolution mass spectrometry (LC-HRMS) with an LTQ Orbitrap analyzer. Samples from two Spanish regions─"Murcia" and "Almería"─were analyzed to obtain complementary results. 57 compounds were found, mainly phenolic acids and flavonoids, of which eight were identified for the first time in tomato. Polyphenols were more abundant in byproducts from "Murcia" samples than in those from"Almería" samples, where the abundance of compounds like coutaric, caffeic, neochlorogenic, dicaffeoylquinic and ferulic acids, vanillic acid hexoside, catechin, naringenin, prunin, apigenin-O-hexoside, rutin, and rutin-O-pentoside was even much higher in byproducts than that in whole fruits. These results reveal the wide range of polyphenols found in tomato byproducts, with potential applications in pharmaceutical research, food preservation, and cosmetic development, among others.

Discrepancies between the stated contents and analytical findings for electronic cigarette liquid products: Identification of the new cannabinoid, Δ9-tetrahydrocannabihexol acetate.

A number of synthetic cannabinoids have been appearing in the recreational drug market for more than a decade. Recent additions are so-called semi-synthetic cannabinoids, and they structurally closely resemble the main psychoactive component of cannabis, Δ9-tetrahydrocannabinol. Knowledge of new (semi-)synthetic cannabinoids is essential to help identify them in authentic forensic case samples. Therefore, the aim of the study was to examine two commercially available electronic cigarette liquid products claiming to contain cannabinoids and characterize the structures of the main compounds. The liquid products were analyzed by gas chromatography-mass spectrometry (GC-MS), GC-quadrupole time-of-flight mass spectrometry (GC-QTOF-MS), and liquid chromatography-high-resolution mass spectrometry (LC-HRMS). In product A, typical cannabinoids (cannabidiol, cannabigerol, and cannabinol) and terpenes (α-caryophyllene and ß-caryophyllene) were identified by comparison with reference materials. An unknown peak was isolated by semi-preparative high-performance LC, analyzed by nuclear magnetic resonance (NMR) spectroscopy, and identified to be Δ9-tetrahydrocannabihexol acetate (Δ9-THCH-O). To the authors' knowledge, this is the first report of the identification of Δ9-THCH-O in commercially available products. Another compound estimated as cannabihexol acetate was also detected. In product B, cannabidiol, cannabinol, α-caryophyllene, and ß-caryophyllene were identified, while two unknown peaks were estimated as tetrahydrocannabidiol isomers. Despite products A and B being labeled to contain "60% HHCPM" and "80% 10-OH-HHC," respectively, no such compounds were detected. The findings of this study could help detect Δ9-THCH-O in case samples and highlight the need to keep monitoring commercial products to identify new drugs, while warning that the package labels cannot be trusted.

Comparative metabolomics reveals how the severity of predation by the invasive insect Cydalima perspectalis modulates the metabolism re-orchestration of native Buxus sempervirens.

The recent biological invasion of box tree moth Cydalima perspectalis on Buxus trees has a major impact on European boxwood stands through severe defoliation. This can hinder further regrowth and threaten survival of populations. In a mesocosm approach and controlled larval density over a 2-month period, responses of B. sempervirens essential and specialized metabolites were characterized using metabolomics, combining 1H-NMR and LC-MS/MS approaches. This is the first metabolome depiction of major Buxus responses to boxwood moth invasion. Under severe predation, remaining green leaves accumulate free amino acids (with the noticeable exception of proline). The leaf trans-4-hydroxystachydrine and stachydrine reached 10-13% and 2-3% (DW), while root content was lower but also modulated by predation level. Larval predation promoted triterpenoid and (steroidal) alkaloid synthesis and diversification, while flavonoids did not seem to have a relevant role in Buxus resistance. Our results reveal the concomitant responses of central and specialized metabolism, in relation to severity of predation. They also confirm the potential of metabolic profiling using 1H-NMR and LC-MS to detect re-orchestration of metabolism of native boxwood after severe herbivorous predation by the invasive box-tree moth, and thus their relevance for plant-insect relationships and ecometabolomics.

Detection of the GH analog somatrogon in sports drug testing: Immunological approaches and LC-HRMS/MS.

Due to the presumed lipolytic and anabolic properties, the misuse of human growth hormone (hGH) and its synthetic analogs in sports is prohibited both in- and out-of-competition. Within this research project, the detectability of somatrogon, a recombinant fusion glycoprotein of 22 kDa hGH and the C-terminal peptide (CTP) of the human chorionic gonadotropin (hCG) ß-subunit, with current WADA-approved doping control assays for hGH and hCG was investigated. For that purpose, cross-reactivity tests and a somatrogon administration study were conducted, and only "Kit 2" of the GH isoform differential immunoassays proved applicable to the detection of somatrogon administration in serum. In urine, the immunoassay specific for total hCG yielded presumptively positive findings for several post-administration samples, which can probably be attributed to the presence of an immunoreactive fragment of the hCG ß-subunit. As the detectability of somatrogon with these approaches was found to be limited, a highly specific detection assay (LOD: 10 ng/mL) for the drug in serum samples was developed by using affinity purification with GH receptor (GHR)-conjugated magnetic beads, proteolytic digestion, and liquid chromatography high-resolution tandem mass spectrometry (LC-HRMS/MS). Following optimization, the approach was comprehensively characterized, and authentic post-administration serum samples were successfully analyzed as proof-of-concept, indicating a detection window of at least 96 h. Consequently, the presented method can be employed to confirm the presence of somatrogon in serum samples, where only "Kit 2" of the currently used immunoassay kits yielded an abnormally high Rec/Pit ratio.

LC-MS and GC-MS Analyses on Green Algae Penicillus capitatus: Cytotoxic, Antimicrobial and Anticholinesterase Activity Screening Enhanced by Molecular Docking & Dynamics and ADME Studies.

In this comprehensive screening study, the chemical composition, and cytotoxic, antimicrobial, and anticholinergic activities of the green algae Penicillus capitatus, collected from Antalya-Türkiye, were determined as in vitro and in silico. GC-MS analysis of the hexane extract revealed a high content of fatty acids, with hexadecanoic acid constituting half of the total fatty acid content. LC-HRMS analysis of the DCM:MeOH extract identified ascorbic acid as the most abundant compound, followed by (-)-epigallocatechin and salicylic acid. The DCM:MeOH extract exhibited potent cytotoxicity against MDA-MB-231 and MCF7 breast cancer cell lines, outperforming doxorubicin with lower IC50 values and a higher selectivity index. Additionally, the extract demonstrated significant antimicrobial activity against Staphylococcus aureus, Escherichia coli, and Candida albicans, along with selective inhibition of acetylcholinesterase (hAChE) over butyrylcholinesterase (hBChE). Molecular docking and dynamics studies revealed that apigenin-7-O-glucoside and epigallocatechin form stable interactions with estrogen receptor alpha (ERα) and hAChE, suggesting their potential as inhibitors. In silico ADME studies indicated favorable pharmacokinetic profiles for the detected compounds, supporting their potential as drug candidates. The promising cytotoxic activity of the P. capitatus extracts, coupled with significant antimicrobial properties and selective hAChE inhibition, highlights their therapeutic potential for breast cancer treatment, infection management, and neurodegenerative disease intervention.

In Vitro and In Vivo Human Metabolism of Ostarine, a Selective Androgen Receptor Modulator and Doping Agent.

Ostarine (enobasarm) is a selective androgen receptor modulator with great therapeutic potential. However, it is also used by athletes to promote muscle growth and enhance performances without the typical adverse effects of anabolic steroids. Ostarine popularity increased in recent years, and it is currently the most abused "other anabolic agent" (subclass S1.2. of the "anabolic agents" class S1) from the World Anti-Doping Agency's (WADA) prohibited list. Several cases of liver toxicity were recently reported in regular users. Detecting ostarine or markers of intake in biological matrices is essential to document ostarine use in doping. Therefore, we sought to investigate ostarine metabolism to identify optimal markers of consumption. The substance was incubated with human hepatocytes, and urine samples from six ostarine-positive cases were screened. Analyses were performed via liquid chromatography-high-resolution tandem mass spectrometry (LC-HRMS/MS) and software-assisted data mining, with in silico metabolite predictions. Ten metabolites were identified with hydroxylation, ether cleavage, dealkylation, O-glucuronidation, and/or sulfation. The production of cyanophenol-sulfate might participate in the mechanism of ostarine liver toxicity. We suggest ostarine-glucuronide (C25H22O9N3F3, diagnostic fragments at m/z 118, 185, and 269) and hydroxybenzonitrile-ostarine-glucuronide (C25H22O10N3F3, diagnostic fragments at m/z 134, 185, and 269) in non-hydrolyzed urine and ostarine and hydroxybenzonitrile-ostarine (C19H14O4N3F3, diagnostic fragments at m/z 134, 185, and 269) in hydrolyzed urine as markers to document ostarine intake in doping.

Exploring the cytotoxic potential of biflavones of Araucaria cunninghamii: Precise identification combined by LC-HRMS-metabolomics and database mining, targeted isolation, network pharmacology, in vitro cytotoxicity, and docking studies.

The leaves of Araucaria cunninghamii are known to be nonedible and toxic. Previous studies have identified biflavones in various Araucaria species. This study aimed to investigate the in vitro cytotoxicity of the isolated compounds from Araucaria cunninghamii after metabolomics and network pharmacological analysis. Methanol extract of Araucaria cunninghamii leaves was subjected to bioassay-guided fractionation. The active fraction was analyzed using LC-HRMS, through strategic database mining, by comparing the data to the Dictionary of Natural Products to identify 12 biflavones, along with abietic acid, beta-sitosterol, and phthalate. Eight compounds were screened for network pharmacology study, where in silico ADME analysis, prediction of gene targets, compound-gene-pathway network and hierarchical network analysis, protein-protein interaction, KEGG pathway, and Gene Ontology analyses were done, that showed PI3KR1, EGFR, GSK3B, and ABCB1 as the common targets for all the compounds that may act in the gastric cancer pathway. Simultaneously, four biflavones were isolated via chromatography and identified through NMR as dimeric apigenin with varying methoxy substitutions. Cytotoxicity study against the AGS cell line for gastric cancer showed that AC1 biflavone (IC50 90.58 µM) exhibits the highest cytotoxicity and monomeric apigenin (IC50 174.5 µM) the lowest. Besides, the biflavones were docked to the previously identified targets to analyze their binding affinities, and all the ligands were found to bind with energy ≤-7 Kcal/mol.

Cyanoacetohydrazide as a Novel Derivatization Agent for the Determination of UHPLC-HRMS Steroids in Urine.

The possibility of cyanoacetohydrazide usage as a novel derivatizing agent is demonstrated in the presented article, and a comparison with hydroxylamine as the most commonly used reagent is provided. Optimal conditions for steroid derivatization with cyanoacetohydrazide are provided. According to the collected data, the maximum yield of derivatives was observed at pH 2.8 within 70 min at 40 °C with 5 ng/mL limit of detection for all investigated analytes. It was shown that cyanoacetohydrazide derivatives produces both syn- and anti-forms as well as hydroxylamine, and their ratios were evaluated and shown in presented work. An efficiency enchantment from two to up to five times was achieved with a novel derivatization reagent. Its applicability for qualitative analysis of steroids in urine was presented at real samples. Additionally, the reproducible fragmentation of the derivatizing agent in collision-induced dissociation offers opportunities for simplified non-targeted steroidomic screening. Furthermore, cyanoacetohydrazide increases ionization efficiency in positive mode, which can eliminate the need for redundant high-resolution instrument runs required for both positive and negative mode analyses.

Echinophora tournefortii Jaub. & Spach: Evaluation of the effect on indomethacin-induced gastric ulser in rats and phytochemical analyses by LC-HRMS.

E. tournefortii has wound healing properties in folk medicine and 5% infusions are used for stomach ulcers. It is also used in colds, abdominal pain, digestive problems, as an appetite enhancer and antispasmodic. For this purpose, in the study biochemical and histopathological evaluation of the ulcer protective effect of the extract obtained from the E. tournefortii in the indomethacin-induced gastric ulcer model in rats was aimed to develop new strategies in the treatment of ulcers. The phytochemical profile of the plant was elucidated for the first time by LC-HRMS in this study. The results indicate that, in terms of TNF-α, IL-1ß, IL-8, IL-6, PGE2, NF-κB, VEGF, NO, COX-1 and COX-2 biochemical parameters, E. tournefortii protects the gastric mucosa to the inflammation, and also modulates the PGE2 pathway, and has a similar effect or even a more positive effect than the reference substance lansoprazole. According to LC-HRMS analysis results, chlorogenic acid, genistein and quinic acid were the main constituents of E. tournefortii extract with 1397.081, 1014.177 and 992.527µg/g extract, respectively. Considering the anti-inflammatory and antioxidant effects of these phenolic components, it is thought that the major components are responsible for the anti-ulcer activity of the E. tournefortii extract.

A novel strategy for detoxification of deoxynivalenol via modification of both toxic groups.

Deoxynivalenol (DON) is the most abundant mycotoxin in cereal crops and derived foods and is of great concern in agriculture. Bioremediation strategies have long been sought to minimize the impact of mycotoxin contamination, but few direct and effective enzyme-catalyzed detoxification methods are currently available. In this study, we established a multi-enzymatic cascade reaction and successfully achieved detoxification at double sites: glutathionylation for the C-12,13 epoxide group and epimerization for the C-3 hydroxyl group. This yielded novel derivatives of DON, 3-epi-DON-13-glutathione (3-epi-DON-13-GSH) as well as its by-product, 3-keto-DON-13-GSH, for which precise structures were validated via liquid chromatography-high-resolution tandem mass spectrometry (LC-HRMS) and nuclear magnetic resonance (NMR) spectroscopy. Both cell viability and DNA synthesis assays demonstrated dramatically decreased cytotoxicity of the double-site modified product 3-epi-DON-13-GSH. These findings provide a promising and urgently needed novel method for addressing the problem of DON contamination in agricultural and industrial settings.

Determination and Identification of Antibiotic Residues in Fruits Using Liquid Chromatography-High-Resolution Mass Spectrometry (LC-HRMS).

Antibiotic residues may be present in fruit products from trees that were treated to combat bacterial diseases such as citrus greening or blight. A liquid chromatography-high-resolution mass spectrometry (LC-HRMS) method was developed for the simultaneous determination and identification of streptomycin, kasugamycin, penicillin, and oxytetracycline residues in fruit. Samples were extracted with acidic methanol and separation was optimized for a hydrophilic interaction LC column. A Q-Exactive HRMS instrument was used to obtain product ion spectra for analyte identification. Quantitation was performed with matrix-extracted calibration curves and internal standard correction. The method was tested on many different types of fruit. In general, fortified samples demonstrated acceptable recoveries (82-116%) and reproducibility (<15% RSD). Method detection limits for these analytes were well below the established US EPA tolerance levels. It was also possible to analyze the fruit extracts prepared using this method for additional chemical contaminants using LC-HRMS.

Toxic arsenolipids bioaccumulate in the developing brain of pilot whales.

Arsenic-containing hydrocarbons (AsHC), a subclass of arsenolipids (AsL), have been proven to exert neuro- and cytotoxic effects in in-vitro and in-vivo studies and were shown to pass through biological barriers like the blood-brain barrier. However, there has been no connection as to the environmental relevance of these findings, meaning there is no study based on samples from free living animals that are exposed to these compounds. Here, we report the identification of two AsHC as well as 3 arsenosugar phospholipids (AsPL) in the brains of a pod of stranded long-finned pilot whales (Globicephala melas) as well as the absence of arsenobetaine (AsB) which is often found to be a dominant As species in fish. We show data which suggests that there is an age-dependent accumulation of AsL in the brains of the animals. The results show that, in contrast to other organs, total arsenic as well as arsenolipids accumulate in an asymptotic pattern in the brains of the animals. Total As concentrations were found to range from 87 to 260 µg As/kg wet weight and between 0.6 and 27.6 µg As/kg was present in the form of AsPL958 in the brains of stranded pilot whales which was the most dominant lipophilic species present. The asymptotic relationship between total As, as well as AsPL, concentration in the brain and whale age may suggest that the accumulation of these species takes place prior to the full development of the blood-brain barrier in young whales. Finally, comparison between the organs of local squid, a common source of food for pilot whales, highlighted a comparable AsL profile which indicates a likely bioaccumulation pathway through the food chain.

"Tranq-dope": The first fatal intoxication due to xylazine-adulterated heroin in Italy.

BACKGROUND AND AIMS: The number of xylazine-involved overdose deaths tremendously increased from 2019 onwards in the US. This is due to the "tranq-dope" trend consisting in mixing opioids with the sedative to reduce drug manufacturing costs and enhance their effects. In this study, we report the first fatality involving xylazine-adulterated heroin in the EU. MATERIALS AND METHODS: The subject was a 33-year-old Caucasian male with a documented history of drug abuse who was found dead in a public area with puncture marks at the elbow. Peripheral blood and urine were collected at the autopsy and analyzed by liquid chromatography-high-resolution tandem mass spectrometry (LC-HRMS/MS) after protein precipitation. RESULTS: 6-Monoacetylmorphine, total/free morphine, and codeine blood concentrations of 20.3, 236/105, and 38.3 ng/mL, respectively, indicated recent heroin consumption. Methadone blood concentration was below 10 ng/mL. Alprazolam, nordiazepam, and flurazepam blood concentrations were 23.9, 61.4, and 55.0 ng/mL, respectively. Benzoylecgonine blood concentration was below 5 ng/mL. Xylazine blood and urine concentrations were 105 and 72.6 ng/mL, respectively. CONCLUSION: The combination of central nervous system depressants, i.e., opioids, benzodiazepines, and xylazine, was the principal cause of death by cardiorespiratory failure. The case was promptly reported to the UE Early Warning System on drugs.

Semaglutide and health risk: Development and validation of a LC-HRMS method for testing semaglutide in whole blood and application to real cases.

The use of semaglutide, also known by its trade name Ozempic®, has been increasing worldwide in recent years due to its benefits in treating type II diabetes. Thanks to its effects on appetite regulation, in many countries it is also used to treat obesity. However, due to its promotion by social media and celebrities as a weight-loss treatment, semaglutide is misused by a non-diabetic and non-obese population and by a young public, which is the main target of these media. Following the alert by the ANSM (Agence nationale de sécurité du médicament) in France and the FDA (Food and Drug Administration) in the United States, which imposed the addition of fatal effects to the list of side effects, the misuse of semaglutide seems to be becoming a public health problem. For this reason, it seems important that a toxicology laboratory has the capacity to test for semaglutide in blood. In this study, the authors have developed and validated a method for the identification and quantification of semaglutide in whole blood using a LC-HRMS. After the addition of the internal standard (bovine insulin), the blood was subjected to protein precipitation using a mix of acetonitrile/methanol (70:30,v:v). The validation procedure demonstrated an acceptable linearity between 2 and 500 ng/mL. LOD and LOQ were 1 and 2 ng/mL, respectively. Intra and inter-day precision were below 20 % at three concentrations. The method was successfully applied to the blood samples of 3 diabetic patients under treatment of semaglutide. The samples tested positive with concentrations ranging from 31 to 70 ng/mL which fall within the limits of therapeutic blood concentrations described in the literature.

Study of pesticide transformation processes in different wheat varieties and their effects on plant metabolism.

BACKGROUND: This study aims to obtain systematic understanding of the way by which pesticides are metabolized in plants and the influence of this process on plants' metabolism as this process has a key impact on plant-based food safety and quality. The research was conducted under field conditions, which enabled to capture metabolic processes taking place in plants grown under multihectare cultivation conditions. RESULTS: Research was conducted on three wheat varieties cultivated under field conditions and treated by commercially available preparations (fungicides, herbicides, insecticides, and growth regulator). Plant tissues with distinctions in roots, green parts, and ears were collected periodically during spring-summer vegetation period, harvested grains were also investigated. Sample extracts were examined by chromatographic techniques coupled with tandem mass spectrometry for: dissipation kinetics study, identification of pesticide metabolites, and fingerprint-based assessment of metabolic changes. CONCLUSION: Tissue type and wheat varieties influenced pesticide dissipation kinetics and resulting metabolites. Metabolic changes of plants were influenced by type of applied pesticide and its concentration in plants tissues. Despite differences in plant metabolic response to pesticide stress during cultivation, grain metabolomes of all investigated wheat varieties were statistically similar. 4-[cyclopropyl(hydroxy)methylidene]-3,5-dioxocyclo-hexanecarboxylic acid and trans-chrysantemic acid - metabolites of crop-applied trinexapac-ethyl and lambda-cyhalothrin, respectively, were identified in cereal grains. These compounds were not considered to be present in cereal grains up to now. The research was conducted under field conditions, enabling the measurement of metabolic processes taking place in plants grown under large-scale management conditions. © 2024 Society of Chemical Industry.

Drug-Leachable Interaction Product Evaluation in Prefilled Syringe of Ganirelix Acetate Injection.

The concept of extractables and leachable has introduced a new era for identifying potential impurities in drug products. Pharmaceutical packaging materials encompass a variety of polymers due to their appealing properties for storing the drug product. However, numerous chemical species may leach into the drug product from these polymers, posing significant health hazards in the public domain. Identifying such leachable is crucial for assessing safety and addressing toxicological concerns. Acrylic acids are commonly used materials for adhering needles to the barrel in pre-filled syringes. In this study, we identified acrylic acid leachable impurities in ganirelix drug products available in the market using mass spectrometry as an analytical technique. These impurities leached into the drug product during storage conditions. Characterization of the impurities was carried out through data obtained from HRMS and MS/MS analysis, revealing them as polyacrylic acid-ganirelix adduct impurities. This study offers valuable insights into identifying leachable and susceptible sites, providing a foundation for potential modifications in similar classes of drug products, thereby enhancing their safety and efficacy.