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This study explored the biosynthesis of bufadienolides(BDs) in Bufo bufo gargarizans to solve the dilemma of the decreasing resources of B. bufo gargarizans and provide a theoretical basis for the sustainable utilization of the resources. Ultra-high performance liquid chromatography-Orbitrap-mass spectrometry(UHPLC-Orbitrap-MS) was employed to detect the synthesis sites of BDs in B. bufo gargarizans, and the results were verified by desorption electrospray ionization-mass spectrometry imaging(DESI-MSI) and homogenate incubation experiments. BDs in B. bufo gargarizans had the highest content in the liver and the highest concentration in the gallbladder, in addition to the parotid gland and skin, which suggested that the liver could synthesize BDs. The results of DESI-MSI also showed that BDs were mainly enriched in the liver rather than the immature parotid gland. The incubation experiment of liver homogenates demonstrated the liver of B. bufo gargarizans had the ability to synthesize BDs. This study showed that the liver was a major organ for the synthesis of BDs in B. bufo gargarizans during metamorphosis, development, and growth, which provided strong theoretical support for the biosynthesis of BDs and the sustainable utilization of B. bufo gargarizans resources.
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Bufanólidos , Animales , Bufo bufo , Distribución Tisular , Bufonidae , Espectrometría de Masa por Ionización de ElectrosprayRESUMEN
BACKGROUND: Diabetic nephropathy (DN) was one of the most popular and most significant microvascular complications of diabetes mellitus. Qingxin Lianzi Yin Decoction (QXLZY) was a traditional Chinese classical formula, suitable for chronic urinary system diseases. QXLZY had good clinical efficacy in early DN, but the underlying molecular mechanism remained unrevealed. PURPOSE: This study aimed to establish the content determination method of QXLZY index components and explore the mechanism of QXLZY on DN by network pharmacology and metabolomics studies. METHODS: Firstly, the content determination methods of QXLZY were established with calycosin-7-O-ß-d-glucoside, acteoside, baicalin and glycyrrhizic acid as index components. Secondly, pharmacological experiments of QXLZY were evaluated using db/db mice. UHPLC-LTQ-Orbitrap MS was used to carry out untargeted urine metabolomics, serum metabolomics, and kidney metabolomics studies. Thirdly, employing network pharmacology, key components and targets were analyzed. Finally, targeted metabolomics studies were performed on the endogenous constituents in biological samples for validation based on untargeted metabolomics results. RESULTS: A method for the simultaneous determination of multiple index components in QXLZY was established, which passed the comprehensive methodological verification. It was simple, feasible, and scientific. The QXLZY treatment alleviated kidney injury of db/db mice, included the degree of histopathological damage and the level of urinary microalbumin/creatinine ratio. Untargeted metabolomics studies had identified metabolic dysfunction in pathways associated with amino acid metabolism in db/db mice. Treatment with QXLZY could reverse metabolite abnormalities and influence the pathways related to energy metabolism and amino acid metabolism. It had been found that pathways with a high degree were involved in signal transduction, prominently on amino acids metabolism and lipid metabolism, analyzed by network pharmacology. Disorders of amino acid metabolism did occur in db/db mice. QXLZY could revert the levels of metabolites, such as quinolinic acid, arginine, and asparagine. CONCLUSION: This study was the first time to demonstrate that QXLZY alleviated diabetes-induced pathological changes in the kidneys of db/db mice by correcting disturbances in amino acid metabolism. This work could provide a new experimental basis and theoretical guidance for the rational application of QXLZY on DN, exploring the new pharmacological effect of traditional Chinese medicine, and promoting in-depth research and development.
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Nefropatías Diabéticas , Medicamentos Herbarios Chinos , Ratones , Animales , Medicamentos Herbarios Chinos/farmacología , Medicamentos Herbarios Chinos/química , Farmacología en Red , Metabolómica/métodos , Medicina Tradicional China/métodos , Nefropatías Diabéticas/tratamiento farmacológico , AminoácidosRESUMEN
With increasing stress in daily life and work, subhealth conditions induced by "Shi-Re Shanghuo" syndrome was gradually universal. "Huanglian Jiedu Wan" (HLJDW) was the first new syndrome Chinese medicine approved for the treatment of "Shi-Re Shanghuo" with promising clinical efficacy. Preliminary small-sample clinical studies have identified some notable biomarkers (succinate, 4-hydroxynonenal, etc.). However, the correlation and underlying mechanism between these biomarkers of HLJDW intervention on "Shi-Re Shanghuo" syndrome remained ambiguous. Therefore, this study was designed as a randomized, double-blind, multicenter, placebo-controlled Phase II clinical trial, employing integrated analysis techniques such as non-targeted and targeted metabolomics, salivary microbiota, proteomics, parallel peaction monitoring, molecular docking and surface plasmon resonance (SPR). The results of the correlation analysis indicated that HLJDW could mediate the balance between inflammation and immunity through succinate produced via host and microbial source to intervene "Shi-Re Shanghuo" syndrome. Further through the HIF1α/MMP9 pathway, succinate regulated downstream arachidonic acid metabolism, particularly the lipid peroxidation product 4-hydroxynonenal. Finally, an animal model of recurrent oral ulcers induced by "Shi-Re Shang Huo" was established and HLJDW was used for intervention, key essential indicators (succinate, glutamine, 4-hydroxynonenal, arachidonic acid metabolism) essential in the potential pathway HIF1α/MMP9 discovered in clinical practice were validated. The results were found to be consistent with our clinical findings. Taken together, succinate was observed as an important signal that triggered immune responses, which might serve as a key regulatory metabolic switch or marker of "Shi-Re Shanghuo" syndrome treated with HLJDW.
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Medicamentos Herbarios Chinos , Metaloproteinasa 9 de la Matriz , Animales , Ácido Araquidónico , Biomarcadores , Simulación del Acoplamiento Molecular , Succinatos/uso terapéutico , Ácido Succínico , HumanosRESUMEN
This study aims to establish the ultra-performance liquid chromatography(UPLC) fingerprint and multi-indicator quantitative analysis method for Schisandrae Sphenantherae Fructus(SSF) and to screen out the potential quality markers(Q-markers) of hepatoprotection based on network pharmacology. The similarity analysis was performed using the Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System, which showed that the similarity of the fingerprints of 15 samples from different regions ranged from 0.981 to 0.998. Eighteen common components were identified, from which 3 differential components were selected by cluster analysis and principal component analysis. The "component-target-pathway" network was built to predict the core components related to the hepatoprotective effects. Fourteen core components were screened by network pharmacology. They acted on the targets such as AKT1, CCND1, CYP1A1, CYP3A4, MAPK1, MAPK3, NOS2, NQO1, and PTGS2 to regulate the signaling pathways of lipid metabolism and atherosclerosis, hepatitis B, interleukin-17, and tumor necrosis factor. Considering the chemical measurability, characteristics, and validity, schisantherin A, anwulignan, and schisandrin A were identified as the Q-markers. The content of schisantherin A, anwulignan, and schisandrin A in the test samples were 0.20%-0.57%, 0.13%-0.33%, and 0.42%-0.70%, respectively. Combining the fingerprint, network pharmacology, and content determination, this study predicted that schisantherin A, anwulignan, and schisandrin A were the Q-markers for the hepatoprotective effect of SSF. The results can provide reference for improving the quality evaluation standard and exploring the hepatoprotective mechanism of SSF.
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Enfermedad Hepática Inducida por Sustancias y Drogas , Medicamentos Herbarios Chinos , Schisandra , Schisandra/química , Farmacología en Red , Medicamentos Herbarios Chinos/farmacología , Medicamentos Herbarios Chinos/química , Enfermedad Hepática Inducida por Sustancias y Drogas/tratamiento farmacológicoRESUMEN
This study aims to reveal the endogenous metabolic characteristics of acteoside in the young rat model of purinomycin aminonucleoside nephropathy(PAN) by non-targeted urine metabolomics and decipher the potential mechanism of action. Biochemical indicators in the urine of rats from each group were determined by an automatic biochemical analyzer. The potential biomarkers and related core metabolic pathways were identified by ultra-high performance liquid chromatography coupled with linear ion trap-Orbitrap mass spectrometry(UHPLC-LTQ-Orbitrap MS) combined with principal component analysis(PCA) and orthogonal partial least squares-discriminant analysis(OPLS-DA). MetaboAnalyst 5.0 was used to establish the receiver operating characteristic(ROC) curve for evaluating the clinical diagnostic performance of core metabolites. The results showed that acteoside significantly decreased urinary protein-to-creatinine ratio in PAN young rats. A total of 17 differential metabolites were screened out by non-targeted urine metabolomics in PAN young rats and they were involved in phenylalanine metabolism and phenylalanine, tyrosine and tryptophan biosynthesis. Thirtten differential metabolites were screened by acteoside intervention in PAN young rats, and they were involved in phenylalanine metabolism and arginine and proline metabolism. Among them, leucylproline and acetophenone were the differential metabolites that were significantly recovered after acteoside treatment. These pathways suggest that acteoside treats PAN in young rats by regulating amino acid metabolism. The area under the curve of two core biomarkers, leucylproline and acetophenone, were both greater than 0.9. In summary, acteoside may restore amino acid metabolism by regulating endogenous differential metabolites in PAN young rats, which will help to clarify the mechanism of acteoside in treating chronic glomerulonephritis in children. The characteristic biomarkers screened out have a high diagnostic value for evaluating the treatment of chronic glomerulonephritis in children with acteoside.
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Glomerulonefritis , Puromicina Aminonucleósido , Humanos , Niño , Ratas , Animales , Metabolómica/métodos , Biomarcadores/orina , Cromatografía Líquida de Alta Presión/métodos , Acetofenonas , Fenilalanina , AminoácidosRESUMEN
This study comprehensively analyzed the active components of Sanhan Huashi Formula using qualitative and quantitative mass spectrometry techniques, laying the foundation for understanding its pharmacological substance basis. UHPLC-LTQ-Orbitrap-MS and GC-MS technologies were used to analyze and identify the volatile and non-volatile components in Sanhan Huashi Formula. UHPLC-QQQ-MS/MS technology was used to simultaneously determine the content of 27 major active components in the formula. The results showed that 308 major chemical components were identified in Sanhan Huashi Formula, among which 60 compounds were identified by comparing with reference standards, mainly including alkaloids, flavonoids, coumarins, triterpenoid saponins, amino acids, and nucleosides. GC-MS technology preliminarily identified 52 volatile compounds, with γ-eudesmol and ß-eudesmol as the main components. The quantitative results demonstrated good linearity(r>0.99) for the 27 active components, indicating the stability, simplicity, and reliability of the established method. Among them, amygdalin, nodakenin, arecoline, ephedrine, and pseudoephedrine had relatively high content and were presumably the main pharmacologically active substances. In conclusion, this study systematically and comprehensively characterized the major chemical components and patterns in Sanhan Huashi Formula, providing a basis for understanding its pharmacological mechanisms and clinical applications.
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Medicamentos Herbarios Chinos , Espectrometría de Masas en Tándem , Cromatografía Líquida de Alta Presión , Cromatografía de Gases y Espectrometría de Masas , Reproducibilidad de los Resultados , Medicamentos Herbarios Chinos/químicaRESUMEN
A randomized, double-blind, placebo-controlled, multi-center phase â ¡ clinical trial design was used in this study to recruit subjects who were in line with the syndrome of excess heat and fire toxin, and were diagnosed as recurrent oral ulcers, gingivitis, and acute pharyngitis. A total of 240 cases were included and randomly divided into a placebo group and a Huanglian Jiedu Pills group. The clinical efficacy of Huanglian Jiedu Pills in treating the syndrome of excess heat and fire toxin was evaluated by using the traditional Chinese medicine(TCM) syndrome scale. Enzyme-linked immunosorbent assay(ELISA) was used to determine and evaluate the levels of adenosine triphosphate(ATP), 4-hydroxynonenal(4-HNE), and adrenocorticotropic hormone(ACTH) in plasma of the two groups before and after administration and to predict their application value as clinical biomarkers. The results showed that the disappearance rate of main symptoms in the Huanglian Jiedu Pills group was 69.17%, and that in the placebo group was 50.83%. The comparison between the Huanglian Jiedu Pills group and the placebo group showed that 4-HNE before and after administration was statistically significant(P<0.05). The content of 4-HNE in the Huanglian Jiedu Pills group decreased significantly after administration(P<0.05), but that in the placebo group had no statistical significance and showed an upward trend. After administration, the content of ATP in both Huanglian Jiedu Pills group and placebo group decreased significantly(P<0.05), indicating that the energy metabolism disorder was significantly improved after administration of Huanglian Jiedu Pills and the body's self-healing ability also alleviated the increase in ATP level caused by the syndrome of excess heat and fire toxin to a certain extent. ACTH in both Huanglian Jiedu Pills group and placebo group decreased significantly after administration(P<0.05). It is concluded that Huanglian Jiedu Pills has a significant clinical effect, and can significantly improve the abnormal levels of ATP and 4-HNE in plasma caused by the syndrome of excess heat and fire toxin, which are speculated to be the effective clinical biomarkers for Huanglian Jiedu Pills to treat the syndrome of excess heat and fire toxin.
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Hormona Adrenocorticotrópica , Calor , Humanos , Medicina Tradicional China , Adenosina TrifosfatoRESUMEN
BACKGROUND: Loropetalum chinensis (R.Br) Oliv (Bhjm), a Chinese folk herbal medicine, was traditionally used in the treatment of wound bleeding and skin ulcers. A new drug named JIMUSAN granules used for gastrosia was developed by our group, and clinical trials have been approved. However, as the principal herb, the material basis and underlying mechanisms of Bhjm in attenuating gastrointestinal mucosa damage (GMD) remain to be systemically illuminated. PURPOSE: An integrated strategy was used to explore the therapeutic effects and mechanisms of Bhjm and ellagic acid (EA) on GMD zebrafish, using network pharmacology, transcriptomics, lipidomics, and real-time quantitative PCR (RT-qPCR) verification. METHODS: First, network pharmacological analysis was used to infer the major effective constituents and targets of Bhjm. Ultra high performance liquid chromatography-linear ion trap/orbitrap high resolution mass spectrometry (UHPLC-LTQ-Orbitrap HRMS) and ultra-high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) were employed to identify the chemical constituents and quantify the different types of constituents. Second, zebrafish model of GMD was established by using 2,4,6-trinitrobenzenesulfonic acid (TNBS) to evaluate the efficacy of Bhjm and EA. The potential mechanism was examined by integrated transcriptomics and lipidomics analysis. Finally, validation tests were implemented using RT-qPCR. RESULTS: In this study, targets indentified by network pharmacology were related to inflammation and mucosal damage. Ten representative components that interacted with these targets were simultaneously determined by UHPLC-MS/MS. Sixty four compounds were identified or tentatively characterized, most of which were flavonoids and polyphenols. Bhjm and EA alleviated mucosal damage and reduced inflammation in a TNBS-induced zebrafish GMD model, indicating that EA was the main active compounds. Eight common differentially expressed genes were downregulated by Bhjm and EA, as determined by transcriptomics analysis. Lipidomics analysis confirmed 12 differential lipids, including phosphatidylcholine (PC) and triglyceride (TG). Further network enrichment analysis demonstrated that differential lipid metabolism was regulated by klf4 and hist1h2ba, and were validated by RT-qPCR. CONCLUSION: In our study, the chemical profile of Bhjm was clarified. Moreover, the GMD repair effect and the mechanism of Bhjm and EA was comprehensively analyzed for the first time, involving inflammation and lipid metabolism. Collectively, these findings will be significantly helpful for deeply exploring the clinical application value of Bhjm.
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Medicamentos Herbarios Chinos , Espectrometría de Masas en Tándem , Animales , Espectrometría de Masas en Tándem/métodos , Pez Cebra , Lipidómica , Transcriptoma , Medicamentos Herbarios Chinos/química , Cromatografía Líquida de Alta Presión/métodos , Membrana Mucosa/químicaRESUMEN
BACKGROUND: Panax notoginseng (PN) was an edible Chinese herbal medicine. PN's current quality control standard cannot precisely match the traditional grading experience. PURPOSE: In this study, under the guidance of the traditional grading experience, the combined metabolomics and biological effect evaluation were used to reveal the distinct chemical quality of PN. METHODS: The quality of PN was evaluated by traditional experience and characterized by the electronic tongue. A zebrafish myocardial ischemia model was developed to verify the grading experience. The untargeted metabolomics method was used to identify and validate the grading markers of PN. RESULTS: The taste was the critical indicator for classifying the quality. Based on the experience sensory scores (ranged from 47.0 to 87.8), PNs could be divided into two grades. The experience scores were significantly associated with umami and richness of the electronic tongue(p<0.01). Besides, superior PN showed substantially stronger anti-myocardial ischemia activity(p<0.001). Thirty-nine differential components were found using UHPLC-LTQ-Orbitrap MS, of which 22 were identified. A new kind of grading quality markers alkynols in PN-associated efficacy was identified, which revealed stronger anti-myocardial ischemia activities than saponin. CONCLUSION: This study evaluated PN through untargeted metabolomics and anti-myocardial ischemia evaluation of zebrafish and proposed the critical role of alkynols in PN's quality classification.
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Isquemia Miocárdica , Panax notoginseng , Saponinas , Animales , Panax notoginseng/química , Pez Cebra , Isquemia Miocárdica/tratamiento farmacológico , Metabolómica , IsquemiaRESUMEN
BACKGROUND: The novel coronavirus pneumonia (COVID-19) has spread rapidly around the world. As a member against the epidemic, Qingfei Paidu Decoction (QFPDD) has been approved for the treatment of COVID-19 in China. However, its antiviral mechanism was still largely unclear. PURPOSE: An integrated strategy was used to explore the antiviral mechanisms of QFPDD in cold and damp environment, including pharmacokinetic (PK), network pharmacology, metabolomics and protein verification. METHODS: Firstly, the pharmacokinetic study of the prototype absorbed ingredients were analyzed by UHPLC-QqQ-MS. Secondly, the metabolomics analysis of the endogenous constituents was carried out. Based on the aforementioned results, an integrated network was constructed to identify the curative components, crucial endogenous differential metabolites and related pathways. Finally, the validation tests were implemented by molecular docking and western blotting (WB). RESULTS: According to the pharmacokinetic behaviors analysis of 31 components in vivo, the flavonoids presented more longer residence time and higher exposure compared with the other compounds. The efficacy and antiviral mechanism of QFPDD were verified by the poly-pharmacology, metabolomics, molecular docking and WB. For the occurrence of metabolic disorder, the change of amino acid transporters should not be neglected. Afterward, 8 curative compounds, 6 key genes and corresponding metabolic pathways were filtered by compound-reaction-enzyme-gene network. The molecular docking verified that the active ingredients bound to the relevant targets well. CONCLUSION: In the present study, an in vivo comprehensive pharmacokinetic behaviors of QFPDD was analyzed for the first time. The results illustrated that QFPDD could exhibit immune regulation, anti-infection, anti-inflammation and metabolic disorder to perform a corresponding therapeutic effect. Moreover, our findings highlighted the roles of amino acid transporters in the coronavirus infection situation.
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Tratamiento Farmacológico de COVID-19 , Coronavirus Humano 229E , Medicamentos Herbarios Chinos , Humanos , Simulación del Acoplamiento Molecular , Medicamentos Herbarios Chinos/química , Metabolómica , Antivirales/farmacología , Antivirales/uso terapéutico , TecnologíaRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: Acteoside (ACT) is the main ingredient derived from the leaves of Rehmannia glutinosa (Dihuangye). Dihuangye has the function of clearing heat, replenishing qi and activating blood, nourishing yin and tonifying kidney in traditional Chinese medicine. Recent studies have demonstrated that Dihuangye can be used to treat nephritis and ACT is a promising antinephritic agent. AIM OF THE STUDY: To clarify the metabolites of ACT in biological samples and investigate the renoprotective effect and mechanism of ACT in rats with chronic glomerulonephritis (CGN). MATERIALS AND METHODS: In this study, the biotransformation of ACT in rat biological samples was clarified by quadrupole time-of-flight tandem mass spectrometry. The metabolites were validated by urine samples in nephropathy model rats. The effect of ACT and its metabolites was evaluated by glomerular podocyte injury due to high glucose. Based on an analysis of the ingredients in vivo, the potential therapeutic targets in the treatment of CGN were investigated by using network pharmacological analysis and molecular docking. Then, the renoprotective effect and mechanism of ACT were determined in rats in a passive Heymann nephritis (PHN) model. RESULTS: A total of 49 metabolites of ACT were detected and identified. Meanwhile, 21 metabolites were detected in nephropathy model rats. ACT was absorbed rapidly and transferred from the kidney, and the metabolites were eliminated via urine. The whole process lasted approximately 8 h. ACT had a significant protective effect on glomerular podocytes damaged by high glucose and 3,4-dihydroxyphenylacetic acid might be the main metabolite of ACT underlying its functions in vivo. The network pharmacology and molecular docking results showed 84 ACT-CGN targets, among which MAPK1, HRAS, AKT1, EGFR, and others were a highly correlated. In the PHN rat model, ACT significantly reduced the 24-h urine protein and serum creatinine concentrations, suppressed the leukocyte CD18 expression levels, decreased the serum tumor necrosis factor α (TNF-α) levels and tended to reduce serum interleukin 6 (IL-6) levels. ACT significantly reduced the platelet aggregation rate and inhibited the proliferative activity of splenic lymphocytes in response to the mitogen concanavalin A. Meanwhile, ACT inhibited transforming growth factor-ß and fibronectin expression in renal tissues and dose-dependently inhibited TNF-α and IL-6 production in RAW264.7 mouse macrophages at doses ranging from 1.8 to 1330 µg/mL. CONCLUSIONS: ACT had therapeutic effects on PHN rats, and its mechanism might be related to the inhibition of intercellular or intercellular-matrix adhesion, suppression of inflammatory response, regulation of immune function, improvement of tissue hemodynamics and hemorheology, and relief of fibrotic lesions.
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Medicamentos Herbarios Chinos , Glomerulonefritis , Ratones , Ratas , Animales , Factor de Necrosis Tumoral alfa , Simulación del Acoplamiento Molecular , Interleucina-6 , Glomerulonefritis/tratamiento farmacológico , Glomerulonefritis/patología , Medicamentos Herbarios Chinos/farmacología , Medicamentos Herbarios Chinos/uso terapéutico , Enfermedad Crónica , GlucosaRESUMEN
Quality evaluation of Chinese medicinal decoction pieces is vital for the development of the downstream industries, and is an important channel for implementing the strategy of "higher quality, higher price, and priority for the high quality" for traditional Chinese medicine. At the moment, the quality of Chinese medicinal decoction pieces is mainly evaluated based on chemical component examination. Considering the weak preliminary research foundation and poor research conditions, traditional experience-based evaluation is undervalued in the quality rating of Chinese medicinal decoction pieces. However, traditional experience is a summary of the quality of Chinese medicinal materials based on clinical experience, which thus can be a potential basis for the quality evaluation of the decoction pieces. It is a challenge in the evaluation of Chinese medicinal decoction pieces to objectify the traditional experience-based evaluation from multiple aspects such as chemistry, effect, and characterization via modern techniques. Therefore, this study developed the "experience-ingredients-activity-electronic sensing" evaluation system for Chinese medicinal decoction pieces on the basis of experience-based assessment, chemical ingredients that can truly reflect the traditional experience, biological effect assessment, and electronic sensory evaluation, which is expected to quantify the traditional experience of quality evaluation of Chinese medicinal decoction pieces via chemistry, biology, and sensory simulation. The evaluation system can serve as a reference for clinical experience-based quality evaluation of Chinese medicinal decoction pieces.
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Medicamentos Herbarios Chinos , China , Electrónica , Medicina Tradicional China , Restricción FísicaRESUMEN
To establish a method for the simultaneous determination of ellagic acid, quercetin, gallic acid, kaempferol, myricetin, tiliroside, salidroside, isoquercetin, chlorogenic acid, and quinic acid in the leaves, flowers, fruits, and roots of Loropetalum chinensis by ultra-performance liquid chromatography-tandem mass spectrometry, and provide references for the development and utilization of L. chinensis resources. The analysis was performed on the chromatographic column ACQUITY UPLC HSS T3(2.1 mm×100 mm, 1.8 µm) with a gradient mobile phase of acetonitrile-0.2% formic solution at the flow rate of 0.3 mL·min~(-1). Column temperature was 30 â and injection volume was 2 µL. Multiple reactive ion monitoring mode(MRM) was used in the negative ion ionization mode of electrospray ion source. The 10 active components had a good linear relationship, and the established method was stable, simple, and accurate. The 10 active components existed in different parts of L. chinensis, with significant different content. The main components in different parts of L. chinensis were polyphenols, with the highest content, followed by flavonoids. The content of 10 active components was generally high in flowers. Among them, the content of quinic acid was the highest, reaching 22.539 1 mg·g~(-1). This study elucidates the differences of active components in the same part and the different parts of L. chinensis, thereby providing basis for the research on the pharmacodynamic substances of L. chinensis and references for the comprehensive development and utilization of L. chinensis resources.
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Medicamentos Herbarios Chinos , Espectrometría de Masas en Tándem , Cromatografía Líquida de Alta Presión/métodos , Espectrometría de Masas en Tándem/métodos , Ácido Quínico , Cromatografía Liquida , Medicamentos Herbarios Chinos/químicaRESUMEN
Indolealkylamines(IAAs) are the main hydrophilic substances in toad skin, mainly including free N-methyl-5-hydroxytryptamine, bufotenine, bufotenidine, dehydrobufotenine, and binding bufothionine. In this study, the LPS-activated neutrophils were used to investigate the structure-activity relationship and anti-inflammatory mechanism of the above-mentioned five monomers from the toad skin in vitro. The neutrophils were divided into the control group, model group(1 µg·mL~(-1) LPS), positive drug group(100 µg·mL~(-1) indometacin), as well as the low-(50 µg·mL~(-1)), medium-(100 µg·mL~(-1)) and high-dose(200 µg·mL~(-1)) free N-methyl-5-hydroxytryptamine, bufotenine, bufotenidine, dehydrobufotenine, and binding bufothionine groups. The levels of IL-6, TNF-α and IL-1ß in the neutrophil supernatant of each group was measured by enzyme-linked immunosorbent assay(ELISA) after LPS stimulation, followed by the detection of apoptosis in each group after Annexin V/PI staining. The protein expression levels of caspase-3, Bax, Bcl-2, beclin1, LC3-I, and LC3-â ¡ were assayed by Western blot. The results showed that IAAs reduced the excessive secretion of inflammatory cytokines caused by LPS compared with the model group. Besides, the activity of each free IAAs(N-methyl-5-hydroxytryptamine, bufotenine, bufotenidine and dehydrobufotenine), especially bufotenine, was stronger than that of the binding bufothionine. As revealed by Annexin V/PI staining, LPS delayed the early apoptosis of neutrophils compared with the control group, while bufotenine promoted the apoptosis of neutrophils in a dose-dependent manner, which might be related to the elevated expression of apoptosis-related protein Bax/Bcl-2. In addition, LPS activated the autophagy pathways in neutrophils. This study confirmed the efficacy of IAAs in reducing the secretion of inflammatory cytokines in neutrophils induced by LPS for the first time. For instance, bufotenine exerts the anti-inflammatory effect possibly by inducing the apoptosis of neutrophils.
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Lipopolisacáridos , Neutrófilos , Animales , Antiinflamatorios/farmacología , Apoptosis , Bufonidae , Lipopolisacáridos/toxicidad , PielRESUMEN
Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disease. Bu-Shen-Tong-Du prescription (BSP) has traditionally been used in to treat RA but its underlying mechanisms remain unclear. In this study, we explored the potential mechanisms of BSP in collagen-induced arthritis (CIA) rats, a classic animal model of RA. We employed an integrated pharmacology approach in combination with network pharmacology, 1H-nuclear magnetic resonance (NMR) metabolomics, and biochemical analyses to determine the mechanisms of BSP for treating RA. We found that BSP can regulate immunity and inflammation by decreasing the spleen index; inhibiting hyperplasia of the white pulp; reducing the levels of IL-1ß, IL-6, IL-17A, and IFN-γ; and increasing the levels of IL-10 in the serum. Network pharmacology was utilized to predict related signal transduction pathways of BSP in RA treatment. 1H NMR metabolomics of the serum confirmed that BSP regulated energy metabolism and amino acid metabolism. Finally, we validated the Toll-like receptor 4 (TLR4)/nuclear factor (NF)-κB signaling pathway using immunohistochemical methods, which demonstrated that BSP controlled RA-induced inflammation by inhibiting the TLR4/NF-κB signaling pathway. These results confirm the therapeutic effect of BSP in a CIA rat model, which is exerted via the inhibition of the inflammation and the improvement of the immune function, balancing energy metabolism and amino acid metabolism, and inhibiting the TLR4/NF-κB signaling pathway. This study provides an experimental basis for using BSP as a combinatorial drug to inhibit inflammation and regulate immunity in the treatment of RA.
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Antiinflamatorios/farmacología , Antirreumáticos/farmacología , Artritis Experimental/tratamiento farmacológico , Medicamentos Herbarios Chinos/farmacología , Articulaciones/efectos de los fármacos , Farmacología en Red , Animales , Artritis Experimental/inducido químicamente , Artritis Experimental/inmunología , Artritis Experimental/metabolismo , Colágeno Tipo II , Citocinas/metabolismo , Metabolismo Energético/efectos de los fármacos , Articulaciones/inmunología , Articulaciones/metabolismo , Articulaciones/patología , Masculino , Medicina Tradicional China , FN-kappa B/metabolismo , Ratas Sprague-Dawley , Transducción de Señal , Receptor Toll-Like 4/metabolismoRESUMEN
"Shanghuo" ("excessive internal heat") is caused by exuberant endogenous fire, which does not have a comprehensive and systematic traditional Chinese medicine theory. In previous study, we had evaluated the therapeutic effect of Huanglian Jiedu Decoction (HLJDD) (granule) on patients with "Shanghuo", however, the specific mechanism was not clear, which need further exploration. To explain its intervention mechanism, we select 57 patients with oral diseases caused by "Shanghuo" and 20 health volunteers to divide into oral disease group, HLJDD intervention group and healthy control group. Firstly, biochemical indicators before and after HLJDD intervention are detected, such as inflammatory factors, oxidative stress factors and energy metabolism factors. The results exhibit that HLJDD significantly decreases indicators succinic acid (p < 0.001); tumor necrosis factor-alpha, adenosine triphosphate, citric acid (p < 0.01); interleukin-8 (IL-8), 4-hydroxynonenal, pyruvic acid, lactate dehydrogenase (p < 0.05). The levels of glucocorticoid, adrenocorticotropic hormone (p < 0.01); lactic acid, IL-4, IL-10 (p < 0.05) significantly increase after HLJDD intervention. In addition, we adopt multi-omics analysis approach to investigate the potential biomarkers. Nontargeted metabolomics demonstrate that the levels of 7 differential metabolites approach that in the healthy control group after HLJDD intervention, which are correlated with histidine metabolism, beta-alanine metabolism and sphingolipid metabolism through metabolic pathway analysis. Targeted lipidomics results and receiver operating characteristic curve analysis show that 13 differential lipids are identified in the three groups mainly focuse on lysophosphatidylcholines, lysophosphatidylethanolamines. Finally, the network associations of those differential biomarkers reveal the regulation of adenosine triphosphate and tricarboxylic acid cycle play essential role in the therapeutic effect mechanism of HLJDD in "Shanghuo". The study has laid the foundation for further revealing the mechanism and finding clinical biomarkers related to "Shanghuo".
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BACKGROUND: In recent years, excellent results have suggested an association between the "brain-gut" axis and Alzheimer's disease (AD) progression, yet the role of the "brain-gut" axis in AD pathogenesis still remains obscure. Herein, we provided a potential link between the central and peripheral neuroinflammatory disorders in AD progression. METHODS: The Morris water maze (MWM) test, immunohistochemistry, ELISA, ProcartaPlex Multiplex immunoassay, multiple LC-MS/MS methods, and the V3-V4 regions of 16S rRNA genes were applied to explore potential biomarkers. RESULTS: In Tg-APP/PS1 mice, gut dysbiosis and lipid metabolism were highly associated with AD-like neuroinflammation. The combination of inflammatory factors (IL-6 and INF-γ), phosphatidylcholines (PCs) and SCFA-producing bacteria were expected to be early diagnostic biomarkers for AD. Huanglian Jiedu decoction (HLJDD) suppressed gut dysbiosis and the associated Aß accumulation, harnessed neuroinflammation and reversed cognitive impairment. CONCLUSION: Together, our findings highlighted the roles of neuroinflammation induced by gut dysbiosis and lipid metabolism disorder in AD progression. This integrated metabolomics approach showed its potential to understand the complex mechanisms of HLJDD in the treatment of AD.
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Enfermedad de Alzheimer , Enfermedad de Alzheimer/tratamiento farmacológico , Enfermedad de Alzheimer/genética , Animales , Encéfalo , Cromatografía Liquida , Progresión de la Enfermedad , Medicamentos Herbarios Chinos , Ratones , ARN Ribosómico 16S , Espectrometría de Masas en TándemRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: Qingxin Lianzi Yin Decoction (QXLZY), a Chinese classical formula, has been widely used in the treatment of various chronic kidney diseases over 1,000 years. However, the current studies on QXLZY were mostly focused on its clinical efficacy, lacking systematic material basis research on constituents. AIM OF THE STUDY: This work aims to elucidate and quantify the chemical constituents, clarify the blood-absorbed components and excretion pathways, predict major bioactive constituents and discover potential therapeutic targets. MATERIALS AND METHODS: UHPLC-LTQ-Orbitrap HRMS was employed to clarify the chemical constituents and metabolites of QXLZY. The extraction of diagnostic ion and neutral loss fragment was aimed for searching specific type of constituents. The plasma, urine, bile and feces samples of rats after oral administration of QXLZY were systematically studied. UHPLC-QQQ-MS/MS was employed to simultaneously detect different types of constitutes. Based on the analysis of ingredients in vivo, the bioactive constituents and potential therapeutic targets in the treatment of diabetic nephropathy (DN) was investigated by using network pharmacological analysis. RESULTS: Totally, 220 compounds were identified or tentatively characterized by UHPLC-LTQ-Orbitrap HRMS. Among them, 59 compounds were confirmed by reference standards. Meanwhile, 21 representative components were simultaneously determined within 15 min by UHPLC-QQQ-MS/MS. 123 components (74 prototypes as well as 49 metabolites) were identified or tentatively characterized. By using network pharmacological analysis, baicalein, liquiritigenin, succinic acid, formononetin, wogonin might be the major effective constituents in QXLZY during the treatment of DN. CONCLUSIONS: Flavonoids, saponins and organic acids were the major chemical ingredients of QXLZY. Flavonoids were the main components absorbed into blood, followed by organic acids. Phase II conjugation reaction was the major metabolic type. The pathways that QXLZY in the treatment of DN were probably related to glucose and lipid metabolism, oxidative stress and inflammation.
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Medicamentos Herbarios Chinos/química , Medicamentos Herbarios Chinos/farmacología , Fitoquímicos/química , Fitoquímicos/farmacología , Ácidos/análisis , Alcaloides/análisis , Animales , Cromatografía Líquida de Alta Presión , Nefropatías Diabéticas/tratamiento farmacológico , Nefropatías Diabéticas/metabolismo , Medicamentos Herbarios Chinos/administración & dosificación , Medicamentos Herbarios Chinos/análisis , Flavonoides/análisis , Masculino , Metaboloma , Fitoquímicos/administración & dosificación , Fitoquímicos/análisis , Mapas de Interacción de Proteínas , Ratas Sprague-Dawley , Estándares de Referencia , Reproducibilidad de los Resultados , Saponinas/análisis , Espectrometría de Masas en TándemRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: Arenobufagin (ArBu) is an important anti-tumor ingredient of Chan'su which has long been used as traditional Chinese medicine in clinic for tumor therapy in China. AIM OF THE STUDY: The purpose of our study is to investigate the lipid homeostasis regulation effects of ArBu on zebrafish model of liver cancer and hepatoma cells, and to provide a reference for further clarifying its active mechanisms. MATERIALS AND METHODS: The zebrafish xenograft model was established by injecting HepG2 cells stained with CM-Dil red fluorescent dye. Both the xenograft model and HepG2 cells were used to evaluate the anti-hepatoma activity of ArBu. High performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) was the main method to study lipidomics, proteomics and the semiquantification of endogenous metabolites. Bioinformatics was used as an assistant tool to further explore the antitumor mechanism of ArBu. RESULTS: The lipidomics analysis revealed that ArBu caused differential lipids changes in a dose-dependent manner, including PCs, PEs, TGs, SMs, DGs, Cer and PA. PCs, PEs, SMs and TGs were markedly altered in both two models. The influence of glycerophospholipid metabolism was the major and commonly affected pathway. Notably, DGs and Cer were significantly changed only in HepG2 cells. Furthermore, the proteomics research in HepG2 cells fished the target proteins related to lipid homeostasis abnormalities and tumor suppression. ArBu reduced the expression of 65 differential proteins associated with the lipid metabolism, apoptosis and autophagy, such as LCLAT1, STAT3, TSPO and RPS27. Meanwhile, 7 amino acids of 29 determined metabolites were significantly changed, including tyrosine, glutamate, glutamine, leucine, threonine, arginine and isoleucine. CONCLUSION: ArBu has a significant anti-hepatoma effect in vitro and a therapeutic effect on zebrafish xenograft model. It regulated the lipid homeostasis. Activated SM synthase and arginine deiminase, inhibited sphingomyelinase, amino acid supply and JAK-STAT3 signaling pathway, and the affected glycerophospholipid metabolism might explain these results.
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Antineoplásicos/farmacología , Bufanólidos/farmacología , Carcinoma Hepatocelular/tratamiento farmacológico , Metabolismo de los Lípidos/efectos de los fármacos , Lipidómica , Neoplasias Hepáticas/tratamiento farmacológico , Proteómica , Animales , Apoptosis/efectos de los fármacos , Proteínas Reguladoras de la Apoptosis/metabolismo , Autofagia/efectos de los fármacos , Proteínas Relacionadas con la Autofagia/metabolismo , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patología , Regulación Neoplásica de la Expresión Génica , Células Hep G2 , Humanos , Metabolismo de los Lípidos/genética , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , Mapas de Interacción de Proteínas , Transducción de Señal , Ensayos Antitumor por Modelo de Xenoinjerto , Pez CebraRESUMEN
Huang-Lian-Jie-Du Decoction (HLJDD), traditional Chinese medicine (TCM), is proven to have ameliorative effects on learning and memory deficits of Alzheimer's disease (AD). The current study aims to reveal the underlying mechanism of HLJDD in the treatment of AD by simultaneous determination on the regulation of HLJDD on oxidative stress, neurotransmitters, and AMPK-SIRT1 pathway in AD. AD model rat was successfully established by injection of D-galactose and Aß 25-35-ibotenic acid. Morris Water Maze (MWM) test was used to evaluate the success of AD modelling. On this basis, an advanced technique with UPLC-QqQ MS/MS was built up and applied to determine the levels of 8 neurotransmitters in rat plasma. Significant alternation in methionine, glutamine, and tryptophan was observed in AD rats' plasma after the administration of HLJDD, relative to the model group. Meanwhile, HLJDD could upregulate the levels of SOD, GSH-Px, AMPK, and SIRT1 and downregulate the content of MDA in the peripheral system of the AD rats. The underlying therapeutic mechanism of HLJDD for the treatment of AD was associated with alleviating oxidation stress, inflammation, neurotransmitters, and energy metabolism. These data provide solid foundation for the potential use of HLJDD to treat AD.