RESUMEN
Centipeda minima (CMX) has been widely investigated using network pharmacology and clinical studies for its effects on hair growth via the JAK/STAT signaling pathway. Human hair follicle papilla cells exhibit hair regrowth through the expression of Wnt signaling-related proteins. However, the mechanism of action of CMX in animals has not been elucidated fully. This study examined the effect of induced hair loss and its side-effects on the skin, and observed the mechanism of action of an alcoholic extract of CMX (DN106212) on C57BL/6 mice. Our results showed that DN106212 was more effective in promoting hair growth than dimethyl sulfoxide in the negative control and tofacitinib (TF) in the positive control when mice were treated with DN106212 for 16 days. We confirmed that DN106212 promotes the formation of mature hair follicles through hematoxylin and eosin staining. We also found that the expression of vascular endothelial growth factor (Vegfa), insulin-like growth factor 1 (Igf1), and transforming growth factor beta 1 (Tgfb1) is related to hair growth using PCR. DN106212-treated mice had significantly higher expression of Vegfa and Igf1 than TF-treated ones, and inhibiting the expression of Tgfb1 had similar effects as TF treatment. In conclusion, we propose that DN106212 increases the expression of hair growth factors, promotes the development of hair follicles, and promotes hair growth. Although additional experiments are needed, DN106212 may serve as an experimental basis for research on natural hair growth-promoting agents.
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Chronic neurodegenerative diseases are typically associated with oxidative stress conditions leading to neuronal cell death. We aimed to investigate the neuroprotective effect of three pyranocoumarins (decursin, decursinol angelate, and decursinol) targeting oxidative stress factors. Decursin (also known as dehydro-8-prenylnaringenin) is a prenylated coumarin compound consisting of a coumarin ring system with a prenyl group attached to one of the carbons in the ring. As a secondary metabolite of plants, pyranocoumarin decursin from Angelica gigas Nakai presented protective effects against glutamate-induced oxidative stress in HT22, a murine hippocampal neuronal cell line. Decursinol (DOH) is a metabolite of decursin, sharing same coumarin ring system but a slightly different chemical structure with the prenyl group replaced by a hydroxyl group (-OH). In our findings, DOH was ineffective while decursin was, suggesting that this prenyl structure may be important for compound absorption and neuroprotection. By diminishing the accumulation of intracellular reactive oxygen species as well as stimulating the expression of HO-1, decursin triggers the self-protection system in neuronal cells. Additionally, decursin also revealed an anti-apoptotic effect by inhibiting chromatin condensation and reducing the forming of annexin-V-positive cells.
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In this study, an automated online micro-solid-phase extraction (µSPE)-liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for the detection of metabolites of cannabidiol (CBD), Δ8-tetrahydrocannabinol (Δ8-THC), and Δ9-tetrahydrocannabinol (Δ9-THC), particularly 7-carboxy- cannabidiol (7-COOH-CBD), 11-nor-9-carboxy-Δ8-tetrahydrocannabinol (Δ8-THCCOOH), 11-nor-9-carboxy-Δ9-tetrahydrocannabinol (Δ9-THCCOOH), and 11-nor-9-carboxy-Δ9- tetrahydrocannabinol-glucuronide (Δ9-THCCOOH-glu) in urine. An instrument top sample preparation (ITSP) cartridge was introduced to increase the sensitivity toward analytes and decrease the matrix effect of the urine. LC-MS/MS analysis was performed in the multiple-reaction monitoring mode, and the analytes were separated using an Acquity UPLC HSS T3 (2.1 × 100 mm, 1.8 µm) column and gradient elution with water containing 0.05 % acetic acid and methanol as the mobile phase. The calibration range was 0.5-200 ng/mL for all the analytes, with a correlation coefficient (r) of ≥0.996 and a weighting factor of 1/x2. The limits of detection for 7-COOH-CBD, Δ8-THCCOOH, Δ9-THCCOOH, and Δ9-THCCOOH-glu were 0.06, 0.02, 0.03, and 0.1 ng/mL, respectively. The intra- and inter-day accuracy ranged from -8.0 to 6.2 % and -7.3 to 7.8 % with a precision of ≤7.2 % and ≤6.2 %, respectively. The method was also validated for selectivity, recovery, matrix effect, stability, and dilution integrity. The developed method was successfully applied to the analysis of 78 urine samples, and 7-COOH-CBD, Δ8-THCCOOH, Δ9-THCCOOH, and Δ9-THCCOOH-glu were detected in 54 urine samples at normalized concentrations of 1.1, 0.6-939.1, 0.9-2595.0, and 1.3-527.6 ng/mg creatinine, respectively.
Asunto(s)
Cannabidiol , Cannabinoides , Cannabidiol/análisis , Dronabinol/análisis , Cromatografía Liquida/métodos , Cannabinoides/análisis , Espectrometría de Masas en Tándem/métodos , Límite de Detección , Extracción en Fase SólidaRESUMEN
Etomidate, with efficacy similar to that of propofol, has been used as a propofol substitute because propofol is a designated narcotic drug, and an increase in the frequency of illegal distribution and misuse has been reported in Korea. Previous analytical studies on etomidate used blood and urine. For long-term use and timing estimation, a method for etomidate analysis using hair should be developed. Therefore, in this study, an analytical method using LC-MS/MS was developed to determine etomidate and its major metabolite in hair. Human hair samples were segmented after washing to eliminate possible contaminants on the hair and stirred with methanol. The LC-MS/MS conditions were optimized, and the chromatographic separation time was 10 min. Selectivity, linearity, limit of detection, limit of quantification, precision, accuracy, recovery, process efficiency, matrix effect, and stability were evaluated to validate the analytical method. The calibration curves ranged from 0.25 to 50 pg/mg for etomidate and 2-250 pg/mg for etomidate acid; the coefficients of determination were higher than 0.997. The intra- and inter-assay precision results for all the compounds were <15% and satisfied at recovery, process efficiency, matrix effect, and stability. In addition, this method was applied to the hair of 4 rats which are administered with etomidate to evaluate. The etomidate concentrations in the rat hair ranged from 2.60 to 8.50 pg/mg, and the etomidate acid concentrations were 2.06-7.13 pg/mg. Thus, this method can be used as basic data for monitoring etomidate in hair.
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Etomidato , Propofol , Humanos , Ratas , Animales , Cromatografía Liquida/métodos , Espectrometría de Masas en Tándem/métodos , Propofol/análisis , Propofol/química , Cabello/química , Detección de Abuso de Sustancias/métodosRESUMEN
With an increase in the obese population, the indiscriminate demand for anti-obesity drugs for rapid weight loss or maintenance has grown. As a result, illegal substances that could induce unexpected negative health effects or fatal side effects are being produced and mixed into consumer products. In the present study, the metabolites of five major illegal anti-obesity drugs are analyzed for the first time. Our data can be utilized to identify related compounds and predict their toxicological effects. Didesmethylsibutramine, desmethylsibutramine, homosibutramine, chlorosibutramine, and benzylsibutramine were metabolized in in vitro and in vivo models, and the metabolites were identified using liquid chromatography quadrupole-time of flight mass spectrometry (LC-Q-TOF-MS) and tandem mass spectrometry (LC-Q-TOF-MS/MS). The in vivo metabolite analysis was carried out using urine and feces samples from rats, and the in vitro metabolite analysis was performed by incubating the analogues with human liver microsomes. We found that each sibutramine analogue was metabolized into several constituents: 2 (M1-2), 5 (M1-5), 11 (M1-11), 7 (N1-7), and 5 (O1-5). In conclusion, our metabolic study could be used for toxicological detection of illegal obesity treatments and metabolite identification in forensic cases.
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Fármacos Antiobesidad , Cromatografía Liquida/métodos , Ciclobutanos , Drogas Ilícitas , Espectrometría de Masas en Tándem/métodos , Animales , Fármacos Antiobesidad/análisis , Fármacos Antiobesidad/metabolismo , Ciclobutanos/análisis , Ciclobutanos/metabolismo , Humanos , Drogas Ilícitas/análisis , Drogas Ilícitas/metabolismo , Masculino , Microsomas Hepáticos/metabolismo , Ratas , Ratas Sprague-DawleyRESUMEN
Centipeda minima (L.) A. Braun & Asch is a well-studied plant in Chinese medicine that is used for the treatment of several diseases. A recent study has revealed the effects of extract of Cetipeda minima (CMX) standardized by brevilin A in inducing hair growth. However, the mechanism of action of CMX in human hair follicle dermal papilla cells (HFDPCs) has not yet been identified. We aimed to investigate the molecular basis underlying the effect of CMX on hair growth in HFDPCs. CMX induced the proliferation of HFDPCs, and the transcript-level expression of Wnt family member 5a (Wnt5a), frizzled receptor (FZDR), and vascular endothelial growth factor (VEGF) was upregulated. These results correlated with an increase in the expression of growth-related factors, such as VEGF and IGF-1. Immunoblotting and immunocytochemistry further revealed that the phosphorylation of ERK and JNK was enhanced by CMX in HFDPCs, and ß-catenin accumulated significantly in a dose-dependent manner. Therefore, CMX substantially induced the expression of Wnt signaling-related proteins, such as GSK phosphorylation and ß-catenin. This study supports the hypothesis that CMX promotes hair growth and secretion of growth factors via the Wnt/ß-catenin, ERK, and JNK signaling pathways. In addition, computational predictions of drug-likeness, together with ADME property predictions, revealed the satisfactory bioavailability score of CMX compounds, exhibiting high gastrointestinal absorption. We suggest that CMX could be used as a promising treatment for hair regeneration and minimization of hair loss.
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Asteraceae/química , Regulación de la Expresión Génica/efectos de los fármacos , Folículo Piloso/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Fitoquímicos , Extractos Vegetales , Alopecia/tratamiento farmacológico , Alopecia/metabolismo , Línea Celular , Humanos , Fitoquímicos/química , Fitoquímicos/farmacología , Extractos Vegetales/química , Extractos Vegetales/farmacologíaRESUMEN
Vardenafil, a remedy for erectile dysfunction, is easily modified, facilitating the creation of analogues that have been illegally added to functional foods and counterfeit medications. However, the medical profile of these analogues, including their safety, efficacy, safe drug combinations, metabolism and excretion, has not been completely evaluated, which could cause serious health problems. In this study, two representative vardenafil analogues, pseudovardenafil and hydroxyvardenafil, were metabolized with in-vitro model (human liver microsome) and in-vivo model (rats). The metabolized samples were extracted and characterized, using liquid chromatography quadrupole-time of flight mass spectrometry (LC-Q-TOF-MS). Some imprecise interpretations were evaluated with tandem mass spectrometry (LC-Q-TOF-MS/MS) for mass fragmentation analysis. A total of 11 metabolites of pseudovardenafil and 13 metabolites of hydroxyvardenafil that were identified have never been reported. These new metabolites could be usefully applied to forensic science and other metabolic fields. Furthermore, they could serve as principal references for the toxicity, danger, and side effects of unlawful vardenafil counterfeits.
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Cromatografía Líquida de Alta Presión/métodos , Espectrometría de Masas en Tándem/métodos , Diclorhidrato de Vardenafil , Animales , Humanos , Masculino , Microsomas Hepáticos/metabolismo , Ratas , Ratas Sprague-Dawley , Diclorhidrato de Vardenafil/análogos & derivados , Diclorhidrato de Vardenafil/análisis , Diclorhidrato de Vardenafil/metabolismoRESUMEN
Pyrethroids, organic compounds similar to natural pyrethrums, constitute the majority of insecticides. Pyrethroids are widely used around the world owing to their excellent selective toxicity to certain insects. In addition, they are easily found in daily life, accounting for most household pesticides. Owing to the easy access to pyrethroid insecticides, pyrethroid-related accidents and suicides have occurred yearly. For the first time, nine pyrethroids commonly used in South Korea and their seven major metabolites were simultaneously analyzed and validated in human plasma using liquid chromatography triple quadrupole mass spectrometry. Plasmas spiked with these pyrethroids and their metabolites were prepared and deproteinized via the addition of acetonitrile. This deproteinized supernatant was filtered and directly injected to ascertain the liquid chromatography-tandem mass spectrometry. For a sensitive and reproducible analysis, all the pyrethroid and metabolite analysis conditions for the multiple reaction monitoring mode were optimized in advance and employed. The validation parameters of the method, including the specificity, linearity, limit of detection, limit of quantification, accuracy, precision, recovery, matrix effect, and stability were also evaluated. The R2 value of linearity was greater than 0.997 for all the analytes, the accuracy ranged from 81.8% to 112.3%, the precision from 0% to 10.1%, and the recovery from 90.9% to 112.4%, depending on the analyte. The stability was 97.0% to 107.0% in fresh plasma and 97.6% to 107.7% in corrupt plasma. The results were satisfactory for all the validation parameters. Furthermore, authentic pyrethroid-poisoned samples were analyzed using this validation method, to determine the suitability; deltamethrin and its metabolites, cis-DBCA and 3-PBA, were successfully analyzed.
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Toxicología Forense/métodos , Insecticidas/sangre , Piretrinas/sangre , Cromatografía Liquida , Estabilidad de Medicamentos , Humanos , Límite de Detección , Manejo de Especímenes , Espectrometría de Masas en TándemRESUMEN
One hundred and twenty six seized methamphetamine (MA) samples were analyzed using GC-MS. All the peaks that appeared in the chromatograms were investigated and 61 impurities including n-octacosane (internal standard) were identified. Among them, 37 impurities were already known or newly identified by comparing with commercial library entries and 18 impurities were detected for the first time. To estimate the synthetic routes of MA samples, route specific impurities had to be selected for each method. Two naphthalenes, 1,3-dimethyl-2-phenylnaphthalene and 1-benzyl-3-methylnaphthalene were selected as Nagai route specific impurities and three diasteromers, UK-19.62(58_165_178) I, UK-19.95(58_165_178) II, UK-20.49(58_165_178) III were also selected not only for their high frequency detection only in Nagai samples but also for the high principal component analysis (PCA) correlation values. For the Emde route, N,N-dimethyl-3,4-diphenylhexane-2,5-diamine and N-methyl-1-{4-[2-(methylamino)propyl]phenyl}-1-phenylpropan-2-amine were selected as route specific impurities, and N,N-di(ß-phenylisopropyl)amine I (DPIA I), N,N-di(ß-phenylisopropyl)amine II (DPIA II), N,N-di(ß-phenylisopropyl)methylamine I (DPIMA I) and N,N-di(ß-phenylisopropyl)methylamine II (DPIMA II) were selected for the Leuckart route. With these route specific impurities, synthetic routes could be identified for 78 of the 126 samples. The 61 impurities were registered in AMDIS target component library and the GC-MS data were deconvoluted. After AMDIS deconvolution, a matrix file was composed and then multivariate analyses were performed to estimate the synthetic route for unknown samples. The unsupervised methods, hierarchical clustering analysis (HCA) and PCA clustered the samples according to the closeness between samples. Two classification functions were obtained from discriminant analysis (DA) and the synthetic routes of the unknown samples were predicted using these two functions.
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Cromatografía de Gases y Espectrometría de Masas , Drogas Ilícitas/análisis , Metanfetamina/análisis , Análisis por Conglomerados , Análisis de Componente PrincipalRESUMEN
Dried Blood Spot (DBS) has been used a blood extraction method for inherited metabolic disorder screening since 1960s. With introduction of LC-MS/MS, not only DBS could be used to analysis drugs in small blood volume, but in various fields, such as toxicology, drug therapeutic monitoring, drug diagnostic screening, and illicit drugs. In toxicology field, many drugs (e.g. benzodiazepines, acetaminophen, small molecule drugs) have been tested with DBS. Compared with earlier blood extraction methods (SPE and LLE), DBS has lots of advantages; lower blood volume (less than 50µL), shorter analysis time caused by a more concise analysis procedure and lower cost. We optimized the DBS procedure and LC-MS/MS conditions for 18 benzodiazepines, seven benzodiazepine metabolites, and one z-drug (zolpidem) analysis in blood. 30µL of whole blood was spotted on FTA DMPK card C and dried for 2h in a desiccator. A 6-mm disk was punched and vortexed for 1min in a centrifuge tube with 300µL methanol/acetonitrile mixture (1:1, v/v). After evaporation, redissolved in 100µL mobile phase of LC-MS/MS and 5µL was injected. In the analysis for 26 target compounds in blood, all of the method validation parameters - LLOD, LLOQ, accuracy (intra- and inter-assay), and precision (intra- and inter-assay) - were satisfied with method validation criteria, within 15%. The results of matrix effect, recovery, and process efficiency were good. We developed a fast and reliable sample preparation method using DBS for 26 benzodiazepines, benzodiazepine metabolites, and z-drug (zolpidem).
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Benzodiazepinas/sangre , Pruebas con Sangre Seca , Cromatografía Liquida , Toxicología Forense/métodos , Humanos , Hipnóticos y Sedantes/sangre , Espectrometría de Masas , Piridinas/sangre , ZolpidemRESUMEN
Nereistoxin(NTX) was originated from a marine annelid worm Lumbriconereis heteropoda and its analogue pesticides including cartap, bensultap, thiocyclam and thiobensultap have been commonly used in agriculture, because of their low toxicity and high insecticidal activity. However, NTX has been reported about its inhibitory neuro toxicity in human and animal body, by blocking nicotinic acetylcholine receptor and it cause significant neuromuscular toxicity, resulting in respiratory failure. We developed a new method to determine NTX in biological fluid. The method involves mixed-mode cationic exchange based solid phase extraction and gas chromatography/mass spectrometry for final identification and quantitative analysis. The limit of detection and recovery were substantially better than those of other methods using liquid-liquid extraction or headspace solid phase microextraction. The good recoveries (97±14%) in blood samples were obtained and calibration curves over the range 0.05-20 mg/L have R2 values greater than 0.99. The developed method was applied to a fatal case of cartap intoxication of 74 years old woman who ingested cartap hydrochloride for suicide. Cartap and NTX were detected from postmortem specimens and the cause of the death was ruled to be nereistoxin intoxication. The concentrations of NTX were 2.58 mg/L, 3.36 mg/L and 1479.7 mg/L in heart, femoral blood and stomach liquid content, respectively. The heart blood/femoral blood ratio of NTX was 0.76.
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Toxinas Marinas/análisis , Anciano , Femenino , Toxicología Forense , Cromatografía de Gases y Espectrometría de Masas , Contenido Digestivo/química , Humanos , Límite de Detección , Plaguicidas/análisis , Reproducibilidad de los Resultados , Extracción en Fase Sólida , Suicidio , Tiocarbamatos/análisisRESUMEN
The information about the sources of supply, trafficking routes, distribution patterns and conspiracy links can be obtained from methamphetamine profiling. The precursor and synthetic method for the clandestine manufacture can be estimated from the analysis of minor impurities contained in methamphetamine. Also, the similarity between samples can be evaluated using the peaks that appear in chromatograms. In South Korea, methamphetamine was the most popular drug but the total seized amount of methamphetamine whole through the country was very small. Therefore, it would be more important to find the links between samples than the other uses of methamphetamine profiling. Many Asian countries including Japan and South Korea have been using the method developed by National Research Institute of Police Science of Japan. The method used gas chromatography-flame ionization detector (GC-FID), DB-5 column and four internal standards. It was developed to increase the amount of impurities and minimize the amount of methamphetamine. After GC-FID analysis, the raw data have to be processed. The data processing steps are very complex and require a lot of time and effort. In this study, Microsoft Visual Basic Application (VBA) modules were developed to handle these data processing steps. This module collected the results from the data into an Excel file and then corrected the retention time shift and response deviation generated from the sample preparation and instruments analysis. The developed modules were tested for their performance using 10 samples from 5 different cases. The processed results were analyzed with Pearson correlation coefficient for similarity assessment and the correlation coefficient of the two samples from the same case was more than 0.99. When the modules were applied to 131 seized methamphetamine samples, four samples from two different cases were found to have the common origin and the chromatograms of the four samples were appeared visually identical. The developed VBA modules could process raw data of GC-FID very quickly and easily. Also, they could assess the similarity between samples by peak pattern recognition using whole peaks without spectral identification of each peak that appeared in the chromatogram. The results collectively suggest that the modules would be useful tools to augment similarity assessment between seized methamphetamine samples.