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@#Cinnabaris(α-HgS) is a mineral traditional Chinese material medica, as a tranquilizer and sedative, which is widely used in combination with herbs for the treatment of children high fever and convulsion.However, a large amount of mercury in Cinnabaris poses a potential risk to the immature central nervous system of children and probably causes severe memory disorders.Inthisstudy,three groups of juvenile rats were given low, medium, and high doses of Cinnabaris by oral gavage once a day for 14 continuous weeks, respectively.The blood mercury concentrations of the rats at different growth phases were monitored by atomic fluorescence spectrometry.The brain structural and functional changes related to the memory functions were investigated through HE staining and Morris water-maze test. Correlation analysis was conducted to clarify the dose- mercury exposure-toxic effect relationship of Cinnabaris and memory disorders.It was found thatthe blood mercury levels increased in both time- and dose-dependent manner.After the 14-week continuous administration of Cinnabaris, the pathological lesions in hippocampal neurons of rats in the high dose group were observed including pyknosis and disordered cell arrangement.In the Morris water-maze test, compared with the control group, rats in the high dose group exhibited the significantly prolonged latency to find the platform and the target quadrant, and the time spent in the target quadrant was obviously shortened. Thus, the significant correlations were established between Cinnabaris dose and mercury exposure,mercury exposure and memory disorders, respectively. In conclusion, the long-term and overdose administration of Cinnabaris in juvenile rats can increase the in-vivo mercury level, destroy the normal hippocampal morphological structure, and lead to memory disorders. This study provided scientific references for the potential mercury poisoning risks pharmacovigilance of Cinnabaris-containing paediatric formulations.
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Vascular endothelial growth factor (VEGF) is the main regulator of physiological angiogenesis during embryonic development, bone growth, and reproductive function, and it also participates in a series of pathological changes. Traditional Chinese medicine (TCM), with a history of more than 2000 years, has been widely used in clinical practice, while the exploration of its mechanisms has only begun. This review summarizes the research of recent years on the influence of TCM on VEGF. It is found that many Chinese medicines and recipes have a regulatory effect on VEGF, indicating that Chinese medicine has broad prospects as a complementary and alternative therapy, providing new treatment ideas for clinical applications and the theoretical basis for research on the mechanisms of TCM.
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Drugs, Chinese Herbal/therapeutic use , Medicine, Chinese Traditional , Vascular Endothelial Growth Factor AABSTRACT
With the recent upsurge of studies in the field of microbiology, we have learned more about the complexity of the gastrointestinal microecosystem. More than 30 genera and 1000 species of gastrointestinal microflora have been found. The structure of the normal microflora is relatively stable, and is in an interdependent and restricted dynamic equilibrium with the body. In recent years, studies have shown that there is a potential relationship between gastrointestinal microflora imbalance and gastric cancer (GC) and precancerous lesions. So, restoring the balance of gastrointestinal microflora is of great significance. Moreover, intervention in gastric premalignant condition (GPC), also known as precancerous lesion of gastric cancer (PLGC), has been the focus of current clinical studies. The holistic view of traditional Chinese medicine (TCM) is consistent with the microecology concept, and oral TCM can play a two-way regulatory role directly with the microflora in the digestive tract, restoring the homeostasis of gastrointestinal microflora to prevent canceration. However, large gaps in knowledge remain to be addressed. This review aims to provide new ideas and a reference for clinical practice.
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Humans , Drugs, Chinese Herbal/therapeutic use , Gastrointestinal Microbiome , Medicine, Chinese Traditional , Precancerous Conditions/pathology , Stomach Neoplasms/pathologyABSTRACT
@#This study aimed to identify the related substances of lorazepam tablets by liquid chromatography mass spectrometry (LC-MS). To separate the related substances of lorazepam tablets, gradient elution was performed using acetonitrile and 0.1% acetic acid -20 mmol/L of ammonium acetate as mobile phase on Inert Sustain C18 (250 mm × 4.6 mm, 5 μm).The accurate mass and elemental composition of the parent ions and their product ions of related substances were determined by electrospray-ionization quadrupole time-of-flight high resolution mass spectrometry (ESI-Q-TOF/MS).The structures of the related substances were identified by spectral analysis. Under the established analytical condition, lorazepam and its related substances were adequately separated, and 22 major related substances with content greater than 0.1% were detected and identified by hyphenated techniques in lorazepam tablets and their stressed samples.Among them, 5 were the impurities listed in the USP or EP, and the others were unknown related substances identified for the first time in this paper.The LC-MS technique can effectively separate and identify the related substances of lorazepam tablets, which provides some reference for quality control.
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@#To establish a method for the determination of formaldehyde and glyoxal simultaneously in varenicline tartrate active pharmaceutical ingredient (API) and its intermediate, formaldehyde and glyoxal were derivatized by 2, 4-dinitrophenylhydrazine (2,4-DNPH) to improve the HPLC retention and UV detection sensitivity. Separation was performed on a C8 (150 mm × 4.6 mm, 5 μm) column by linear gradient elution using acetonitrile and water as the mobile phase; the detective wavelength was set at 380 nm.Formaldehyde and glyoxal were quantitatively determined by an external reference method.Linear calibration was established for both formaldehyde and glyoxal in the range from 0.094 to 1.88 μg/mL.The detection and the quantification limits were 0.047 μg/mL (19 μg/g) and 0.094 μg/mL (38 μg/g), respectively.The recoveries were (95.0±1.1)% and (99.4 ± 2.6)% for formaldehyde and glyoxal, respectively.This method has been fully validated to be applicable to quantitative analysis of trace amount of formaldehyde and glyoxal in varenicline tartrate API and its intermediate.Test results demonstrated that the contents of both formaldehyde and glyoxal met the permitted daily exposure (PDE) limits for the finished products of varenicline tartrate API as well as its intermediate, though the glyoxal contents in the crude intermediates were likely to exceed the limit.The established method is valuable for the manufacturing process and quality control of varenicline tartrate.
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Objective:To analyze the differences and similarities of pre-treatment and post-treatment lung microbiome of acute respiratory distress syndrome (ARDS) and find out the change rules of the lung microbiome in the progression of ARDS according to different prognosis.Methods:A retrospective study was conducted. Patients with ARDS caused by severe pneumonia admitted to intensive care unit (ICU) of Jiangmen Central Hospital from February 2019 to January 2020 were enrolled as the study subjects. The patients were divided into pre-treatment (ARDS-preT) group (24 cases), post-treatment survival (ARDS-poT-Survival) group (17 cases), and post-treatment death (ARDS-poT-Dead) group (7 cases). ICU patients with mild pulmonary infection and non-ARDS admitted to ICU during the same period were enrolled as control group (25 cases). The similarities and differences of lung microbiome in four groups were analyzed and compared, and the possible pathogenic bacteria (potential risk factors for death) and probiotics (potential survival and protective factors) related to death caused by ARDS were screened.Results:In terms of pathogenic microorganisms, the positive rates of Escherichia coli and Candida albicans in the ARDS-poT-Dead group were significantly higher than those in the ARDS-poT-Survival group [57.1% (4/7) vs. 5.9% (1/17) and 57.1% (4/7) vs. 0% (0/7), both P < 0.05]. In the screening of background bacteria, the decrease of bacteria in the ARDS-preT group compared with the ARDS-poT-Survival group, the ARDS-poT-Dead group compared with the ARDS-poT-Survival group, the ARDS-poT-Dead group compared with the control group, the reduced bacteria might be pulmonary probiotics (potential protective factor for ARDS). The screening result was Hydrobacter [ARDS-preT group vs. ARDS-poT-Survival group: 62.5% (15/24) vs. 94.1% (16/17); ARDS-poT-Dead group vs. ARDS-poT-Survival group: 14.3% (1/7) vs. 94.1% (16/17); ARDS-poT-Dead vs. control: 14.3% (1/7) vs. 96.0% (24/25), all P < 0.05]. In the screening of background bacteria, the increase of bacteria in the ARDS-poT-Dead group compared with the ARDS-preT group, the ARDS-poT-Dead group compared with the ARDS-poT-Survival group, the ARDS-poT-Dead group compared with the control group, and the increased bacteria might be potential pulmonary pathogen (potential risk factor for death of ARDS), which belonged to Enterobacteria: Edwardsiella, Enterobacteriaceae, Escherichia, Klebsiella, Kluyvera, Lelliottia, Pantoea, Raoultella. Conclusions:The results revealed the increase of Escherichia coli or Candida albicans in pulmonary pathogenic microorganisms, or the increase of Enterobacteria in background bacteria may be the risk factors for the death of ARDS. Additionally, background bacteria Hydrobacter probably is a protective factor for the survival of ARDS. Whether it can be used as a novel treatment for ARDS is worth further investigation.
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@#This study aimed to identify the related substances of midazolam hydrochloride by liquid chromatography-mass spectrometry (LC-MS). To separate the related substances of midazolam hydrochloride, gradient elution was performed using acetonitrile and 25 mmol/L of ammonium acetate (pH was adjusted to 5.5 with acetic acid) as mobile phase on Thermo BDS Hypersil C18 column (100 mm × 4.6 mm, 2.4 μm). The accurate mass and elemental composition of the parent ions and their product ions of related substances were determined by electrospray-ionization quadrupole time-of-flight mass spectrometry (ESI-Q-TOF/MS). The structures of the related substances were identified by spectral analysis and process synthesis, and some of them were further confirmed by reference substances. Under the established HPLC condition, midazolam hydrochloride and its related substances were adequately separated, and 22 major related substances were detected and identified by hyphenated techniques in midazolam hydrochloride and its stressed samples, of which 8 were recorded as impurities in the United States Pharmacopeia. The LC-MS techniques can effectively separate and identify the related substances of midazolam hydrochloride and provide reference for the establishment of storage condition, optimization of synthetic processes and quality control.
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Lao-Xiang-Huang (LXH) of Chaozhou is the processed product of Fructus Citri sarcodactylis.LXH from different producing areas were used as research objects in the study for the establishment of HPLC fingerprints of LXH.Contents of hesperidin and 5,7-dimethoxycoumarin were also determined to provide scientific basis for the establishment of quality standard of LXH.Samples were separated by an Agilent Eclipse XDB C18 column (4.6 mm × 250 mm,5 μm) using acetonitrile-0.1％ phosphoric acid with water gradient system as a mobile phase.The flow rate was 1.0 mL· min-1.The injection volume was 20 μL.The detection wavelength was at 283 nm.The results showed that HPLC fingerprint of LXH was established with good separation and repeatability.The similarity evaluation on 27 batches samples of LXH showed that there was a certain similarity on the HPLC fingerprints of LXH.However,there was a certain difference as a whole.Contents of hesperidin and 5,7-dimethoxycoumarin in LXH were simultaneously determined.It was concluded that the established HPLC fingerprint of LXH and content determination of hesperidin and 5,7-dimethoxycoumarin method were accurate,sensitive and repeatable.It provided scientific evidence for the quality control standard of LXH of Chaozhou.
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A sensitive and selective method was developed for the separation and characterization of related substances (RSs) in EVT-401 by hyphenated LC–MS techniques. Complete separation of the RSs was achieved with an Inertsil ODS-SP column (250 mm×4.6 mm, 5 μm) by linear gradient elution using a mobile phase consisting of 0.2% formic acid solution, methanol and acetonitrile. EVT-401 was found to be susceptible to acid, alkaline and oxidative stresses, while relatively stable under photolytic and thermal dry stress conditions. Fourteen RSs including six process-related substances and eight degradation products were detected and identified in EVT-401 with positive ESI high-resolution TOF-MS analysis of their parent ions and the corresponding product mass spectra elucidation, and some of them were further verified by chemical synthesis and NMR spectroscopy. The specific LC–MS method developed for separation, identification and characterization of RSs is valuable for EVT-401 manufacturing process optimization and quality control.
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Objective To screen and confirm cell fusion by DNA technology of parentage identification based on detecting of short tandem repeats.Methods With 20％ polyethylene glycol (PEG)-6000,human myeloma cell lines and health individual peripheral blood mononuclear cell were fused.Then selected by hypoxantin,aminopterin,thymidin (HAT) medium,and fusion cell were sub-cloned.Morphology of fusion cells was checked by regular microscope.Concentration of DNA was compared to parental cells.Allele genes,identified by short tandem repeats,of fusion cell line were sequenced and compared with each other.Results The fused cells from myeloma cell line and peripheral blood mononuclear cell (PBMC) were slightly larger than primary cells,and the proliferation cycle was not changed significantly.DNA concentration of the fused cell DNA was increased by two times.Sequences of short tandem repeats (STR) showed that the fused cell included all original genetic materials of parent cells.Conclusions DNA technology of parentage identification is a convenient and reliable method to screen and confirm fused cell.
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AIM: To investigate the differences in the distribution of SRY-related HMG box 5 (SOX5) gene single nucleotide polymorphisms (SNPs) among stable chronic obstructive pulmonary disease (COPD) patients, COPD with pulmonary hypertension (PH) patients and healthy controls, and to explore the association of the SOX5 SNPs in COPD-related PH.METHODS: From April 2013 to April 2015, 250 patients with stable COPD were enrolled continuous-ly in Ningxia People’s Hospital according to COPD treatment guidelines (2013 edition).All the patients received echocar-diography, and were divided into COPD with PH group [pulmonary artery systolic pressure (PASP)≥50 mmHg, n =103] and COPD without PH group (PASP loci was performed using MassARRAY genotyping system ( Sequenom).Genotype frequencies were calculated.RE-SULTS: Age, sex and smoking index showed no significantly difference between control group and COPD group, neither between COPD with PH group and COPD without PH group.Genotype frequencies of SOX5 gene rs10842262 and rs11046966 loci between control group and COPD group was of significant difference (P<0.05).Genotype frequencies of SOX5 gene rs10842262 and rs11046966 loci showed no significant difference between COPD with PH group and COPD without PH group.CONCLUSION: SOX5 gene rs10842262 and rs11046966 loci may play an important role in COPD, but not in COPD-related PH.
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Objective To construct HaCaT cell lines stably expressing the wild type human GJB6 gene or its mutant by using a Tet-On lentiviral vector, and to lay an experimental foundation for studies on pathogenesis of hidrotic ectodermal dysplasia. Methods The wild-type human GJB6 gene and its mutant (A88V)were amplified by PCR, and then inserted into the Tet-on lentivirus plasmid to construct recombinant lentivirus vectors. The recombinants were identified by gene sequencing and enzymatic digestion. Cultured HaCaT cells were classified into three groups to be transfected with a negative control lentiviral vector (NC group), the lentivirus vector expressing the wild-type human GJB6 gene (WT group), or the lentivirus vector expressing the mutant human GJB6 gene (MU group). Puromycin was used to select HaCaT cell clones stably expressing the GJB6 gene which encodes the connexin 30 (Cx30)protein. The selected HaCaT cell clones were cultured with or without tetracycline for 48 hours, thereafter, real-time PCR(RT-PCR) was performed to detect GJB6 gene mRNA expression, Western-blot analysis to measure expressions of Cx30 and FLAG-tag proteins, and cell counting kit 8 (CCK8)assay to evaluate cellular proliferative activity. Results Enzymatic digestion and gene sequencing showed that recombinant lentivirus plasmids were successfully constructed. RT-PCR showed evidently increased mRNA expression of the GJB6 gene in stably transfected HaCaT cells. Moreover, the expression abundance of the GJB6 gene was 112.369 times higher in the WT group induced by tetracycline than in that without tetracycline treatment (P 0.05). Conclusion HaCaT cell lines which stably express the wild-type GJB6 gene or its mutant(A88V)are successfully constructed.
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Reproductive toxicity research takes an important place in traditional Chinese medicine pre-clinical safety evaluation. Modern reproductive toxicity experiment includes drug-related miscarriage, fetal death, teratism, and adverse effects on fertility, genital system, embryonic development and fetus, which is different from contraindicated in pregnancy in traditional Chinese medicine theory. Now the three-phases reproductive toxicity study is the method mainly applied in traditional Chinese medicine reproductive toxicity evaluation. Besides that, alternative methods of whole embryos culture and embryonic stem cell test are also used in traditional Chinese medicine embryo toxicity evaluation. This article reviews research progress and pre-clinical evaluation on reproductive toxicity of traditional Chinese medicine.
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The purpose of this study is to evaluate the embryotoxicity of alkaloids in Senecionis Scandentis Hebra on in vitro cultured mouse embryos. Mouse whole embryo culture (WEC) was applied in this study. Post-implantation (8.5 d) mouse embryos were isolated from their mothers, and cultured in medium of immediately centrifuged serum (ICS) with different concentrations of seneciphylline (target concentrations were 100, 50, 25 and 12.5 μg x mL(-1)) or senkirkine (target concentrations were 50, 25 and 12.5 μg x mL(-1)) for 48 h. After culturing completed, the development and organic morphodifferentiation of the cultured embryos were evaluated microscopically. Treatment with seneciphylline and senkirkine had adverse effects on the development and organic morphodifferentiation of embryos. The effect also had clear dose-response. Alkaloidals in Senecionis Scandentis Hebra had embryotoxicity on cultured embryos, which indicated that pregnant people exposed to Senecionis Scandentis Hebra may get potential risk on fetus.