Isorhynchophylline improves lipid metabolism disorder by mediating a circadian rhythm gene Bmal1 in spontaneously hypertensive rat.

Hypertension is a progressive metabolic disease characterized by circadian regulation of lipid metabolism disorder. Identifying specific lipid components and maintaining circadian homeostasis of lipid metabolism might be a promising therapeutic strategy for hypertension. Isorhynchophylline (IRP) can regulate lipid metabolism; however, the underlying mechanism of IRP in improving lipid metabolism rhythm disorder is still unclear. The lipid circadian biomarkers and abnormal metabolic pathways intervened by IRP were investigated using diurnal lipidomic research methods. The 24-h circadian changes in mRNA and protein expression levels of circadian genes, including Bmal1, Clock, Cry1, Cry2, Per1, and Per2, and lipid metabolism-related factors (PPARα and LPL) were determined using RT-PCR and western blot analyses, respectively. The underlying mechanisms were intensively investigated by inhibiting Bmal1. Molecular docking and drug affinity responsive target stability analyses were performed to assess the binding affinity of IRP and Bmal1. IRP treatment could effectively improve 24-h blood pressure, ameliorate the lipid metabolic rhythm disorder, reverse the expression levels of circadian rhythm genes, and regulate lipid metabolism-related genes (PPARα and LPL) by mediating Bmal1. This study highlighted the potential effects of IRP in maintaining the circadian homeostasis of lipid metabolism and the treatment of hypertension.

The activation of histone deacetylases 4 prevented endothelial dysfunction: A crucial mechanism of HuangqiGuizhiWuwu Decoction in improving microcirculation dysfunction in diabetes.

ETHNOPHARMACOLOGICAL RELEVANCE: The regulation of epigenetic factors is considered a crucial target for solving complex chronic diseases such as cardio-cerebrovascular diseases. HuangqiGuizhiWuwu Decoction (HGWWD), a classic Chinese prescription, is mainly used to treat various vascular diseases. Although our previous studies reported that HGWWD could effectively prevent vascular dysfunction in diabetic rodent models, the precise mechanism is still elusive. AIM OF THE STUDY: In this study, we investigated the epigenetic mechanisms of modulating the damage of vascular endothelial cells in diabetes by HGWWD. METHODS: We first analyzed common active components of HGWWD by using HPLC-Q-TOF-MS/MS analysis, and predicted the isoforms of histone deacetylase (HDAC) that can potentially combine the above active components by systems pharmacology. Next, we screened the involvement of specific HDAC isoforms in the protective effect of HGWWD on vascular injury by using pharmacological blockade combined with the evaluation of vascular function in vivo and in vitro. RESULTS: Firstly, HDAC1, HDAC2, HDAC3, HDAC4, HDAC6, HDAC7, SIRT2, and SIRT3 have been implicated with the possibility of binding to the thirty-one common active components in HGWWD. Furthermore, the protective effect of HGWWD is reversed by both TSA (HDAC inhibitor) and MC1568 (class II HDAC inhibitor) on vascular impairment accompanied by reduced aortic HDAC activity in STZ mice. Finally, inhibition of HDAC4 blocked the protective effect of HGWWD on microvascular and endothelial dysfunction in diabetic mice. CONCLUSIONS: These results prove the key role of HDAC4 in diabetes-induced microvascular dysfunction and underlying epigenetic mechanisms for the protective effect of HGWWD in diabetes.

Isorhynchophylline Regulates the Circadian Rhythm of the Hypothalamus in Spontaneously Hypertensive Rats to Treat Hypertension.

BACKGROUND: The neurotransmitter metabolism in spontaneously hypertensive rats (SHR) is disordered, and these disturbances in neurotransmitter levels can further exacerbate the development of hypertension. Neurotransmitters can affect the expression of circadian clock genes. OBJECTIVE: To clarify the time-dependent internal mechanism of the imbalance of the target neurotransmitter metabolic rhythm of spontaneously hypertensive rats, the circadian research was carried out by the method of targeted metabolomics and molecular biology technology. METHODS: We have explored the mechanism of isorhynchophylline regulating the circadian rhythm through the ERK signaling pathway and thus treating hypertension by detecting the changes of central hypothalamic biological clock rhythm genes after isorhynchophylline intervention, from hypothalamic neurotransmitter rhythmicity. RESULTS: The expression of rhythm genes in normal rats showed a certain rhythm at 6 time points, while the expression of rhythm genes in model rats decreased, and the gene rhythm returned to normal after isorhynchophylline treatment. Cosine analysis of 12 neurotransmitters in hypothalamus showed that there were 6 rhythmic neurotransmitters in the normal group, while in the model group, 4 of the 6 neurotransmitters lost their rhythmicity, and the rhythmicity returned to normal after isorhynchophylline intervention. Compared with the normal group, the expression of ERK protein in the model group increased significantly and decreased after isorhynchophylline treatment. CONCLUSION: The mechanism of isorhynchophylline treating hypertension is not only the regulation of serum neurotransmitters rhythm, but also acting on rhythm genes in the feedback loop of the central biological clock.

Antioxidant Effects of Roasted Licorice in a Zebrafish Model and Its Mechanisms.

Licorice (Gan-Cao, licorice) is a natural antioxidant and roasted licorice is the most common processing specification used in traditional Chinese medicine prescriptions. Traditional Chinese medicine theory deems that the honey-roasting process can promote the efficacy of licorice, including tonifying the spleen and augmenting "Qi" (energy). The antioxidant activity and mechanisms underlying roasted licorice have not yet been reported. In this study, we found that roasted licorice could relieve the oxidative stress injury induced by metronidazole (MTZ) and could restrain the production of excessive reactive oxygen species (ROS) induced by 2,2'-azobis (2-methylpropionamidine) dihydrochloride (AAPH) in a zebrafish model. It was further found that roasted licorice could exert its oxidative activity by upregulating the expression of key genes such as heme oxygenase 1 (HO-1), NAD(P)H quinone dehydrogenase 1 (NQO1), glutamate-cysteine ligase modifier subunit (GCLM), and glutamate-cysteine ligase catalytic subunit (GCLC) in the nuclear factor erythroid 2-related factor 2 (NRF2) signaling pathway both in vivo and in vitro. Furthermore, consistent results were obtained showing that rat serum containing roasted licorice was estimated to reduce cell apoptosis induced by H2O2. Then, the UHPLC-Q-Exactive Orbitrap MS analysis results elucidated the chemical composition of rat plasma containing roasted licorice extracts, including ten prototype chemical components and five metabolic components. Among them, six compounds were found to have binding activity with Kelch-like ECH-associated protein 1 (KEAP1), which plays a crucial role in the transcriptional activity of NRF2, using a molecular docking simulation. The results also showed that liquiritigenin had the strongest binding ability with KEAP1. Immunofluorescence further confirmed that liquiritigenin could induce the nuclear translocation of NRF2. In summary, this study provides a better understanding of the antioxidant effect and mechanisms of roasted licorice, and lays a theoretical foundation for the development of a potential antioxidant for use in clinical practice.

Deciphering the pharmacological mechanisms of Chaenomeles Fructus against rheumatoid arthritis by integrating network pharmacology and experimental validation.

Chaenomeles Fructus is a plant that can be used for both food and medicine. Modern studies have shown that Chaenomeles Fructus has anti-inflammatory and immunosuppressive effects on arthritis. However, the mechanism of action of Chaenomeles Fructus on rheumatoid arthritis (RA) and its main active ingredients are still unclear. This study was aimed at devising an integrated strategy for investigating the bioactivity constituents and possible pharmacological mechanisms of Chaenomeles Fructus against RA. The components of Chaenomeles Fructus were analyzed using UPLC-Q-Exactive orbitrap MS techniques and applied to screen the active components of Chaenomeles Fructus according to their oral bioavailability and drug-likeness index. Then, we speculated on the potential molecular mechanisms of Chaenomeles Fructus against RA through a network pharmacology analysis. Finally, the potential molecular mechanisms of Chaenomeles Fructus against RA were validated in a complete Freund's adjuvant (CFA)-induced RA rat model. We identified 48 components in Chaenomeles Fructus and screened seven bioactive ingredients. The results of the network pharmacology prediction and the experimental verification results were analyzed by Venn analysis, and the experimental results concluded that Chaenomeles Fructus mainly interferes with the inflammation of RA by inhibiting arachidonic acid metabolism and the MAPK signaling pathway. This study identified the ingredients of Chaenomeles Fructus by UPLC-Q-Exactive orbitrap MS and explained the possible mechanisms of Chaenomeles Fructus against RA by integrating network pharmacology and experimental validation.

Discovery of Biomarkers and Potential Mechanisms of Agarwood Incense Smoke Intervention by Untargeted Metabolomics and Network Pharmacology.

BACKGROUND: Agarwood, as a traditional Chinese medicine, has great potential value for the treatment of tranquilization. However, its potential mechanisms and biomarkers are still unclear. METHODS: In this study, ultra-high performance liquid chromatography-quadrupole-Exactive Orbitrap mass spectrometry (UHPLC-Q-Exactive Orbitrap-MS)-based metabonomics was adopted to discover the potential biomarkers in mice after agarwood incense smoke (AIS) intervention. Furthermore, the chemical components in agarwood were identified based on UHPLC-Q-Exactive Orbitrap-MS. The global view of potential compound-target-pathway (C-T-B) network was constructed through network pharmacology to understand the potentially material basis of biomarkers. RESULTS: Metabolic profiling indicated that the metabolic changed significantly in mice serum after AIS intervention. A total of 18 potential biomarkers closely related to insomnia and emotional disease were identified, mainly involving in tryptophan metabolism, arginine and proline metabolism, cysteine and methionine metabolism and steroid hormone biosynthesis pathways. A total of 138 components in agarwood were identified based on UHPLC-Q-Exactive Orbitrap-MS. The results showed that mainly compounds such as flidersia type 2-(2-phenylethyl) chromones (FTPECs) and sesquiterpenes exerted good docking abilities with key target proteins, which were involved in multiple diseases including depression and hypnosis. CONCLUSION: In conclusion, this study enhanced current understanding of the change of metabolic markers after AIS intervention. Meanwhile, it also confirmed the feasibility of combining metabolomics and network pharmacology to identify active components and elucidate the material basis of biomarkers and mechanisms.

Toward the Development of Personalized Syndrome Discriminant Systems: A Discriminant System for Hypertension with Liver Yang Hyperactivity Syndrome.

Traditional Chinese medicine has shown promising results in treating the symptoms of hypertension, a major global health concern not yet fully managed by modern medicine. It is, therefore, of high priority to clarify the altered pathophysiology of hypertension in individuals with liver Yang hyperactivity syndrome (HLYH) in response to effective treatments to better understand this disorder. The primary aim of this study was to construct a personalized syndrome discriminant system based on data capable of informing management strategies prior to the initiation of antihypertensive therapy or the implementation of screening strategies in at-risk HLYH. Based on the successful replication of HLYH rat models, we extracted the core discriminant factors of the disorder through the integration of physical signs, biochemical indicators, and metabolic markers. Macro and micro information was correlated to construct a syndrome discriminant system. At the macroscopic level, HLYH rat models characterized by elevated blood pressure were found to be associated with significant changes in water intake, pain threshold, retention time on a rotating platform, and body surface temperature. A total of 27 potential biomarkers and 14 metabolic pathways appeared to reflect the primary metabolic characteristics. Through the integration of these data, we successfully constructed a combined macro-micro personalized syndrome discriminant system, which provides a foundation for research regarding the risk loci of HLYH. Our findings also broaden our understanding of the biological pathways involved in HLYH.

Separation of chemical constituents in Bidens pilosa Linn. var. radiata Sch. Bip. by elution-extrusion counter-current chromatography using two new three-phase solvent systems.

Two new three-phase solvent systems combined with elution-extrusion counter-current chromatography mode were used to study the chemical constituents in Bidens pilosa Linn. var. radiata Sch. Bip. The first novel solvent system consisted of n-hexane, acetonitrile, chloroform, and water in a ratio of 5:5:1:5, which was selected for elution-extrusion counter-current chromatography to separate the n-hexane extraction part. A total of six constituents were obtained from this part in the up phase as the stationary phase and the middle phase as the mobile phase. The second novel solvent system, composed of n-hexane-butyl acetate-acetonitrile-water (3:1:4:3, v/v/v/v), was used for separating ethyl acetate extract of Bidens pilosa Linn. var. radiata Sch. Bip. Eight compounds were successfully isolated using elution-extrusion counter-current chromatography elution-extrusion mode. Fourteen chemical constituents were identified as 2-ß-D-glucopyranosytoxy-1-hydroxy-5(E)-tridecene-7,9,11-triyne (Y1), 3-ß-D-glucopyranosyloxy -1-hydroxy-6(E)-tetradecene-8,10,12-triyne (Y2), 1, 2-dihydroxy-5(E)-tridecene-7,9, 11-triyne (Y3), isorhamnetin (Y4), kaempferol (Y5), icthyothereolacetate (Y6), quercetin-3-O-ß-D- galactopyranosyl-7-O-ß-D-glucopyranoside (W1), quercetin 3-O-ß-L-rhamnopyranoside (W2), neosperidin dihydrochalcone (W3), quercetin (W4), quercetagetin-3,6,4' -trimethoxyl- 7-O-ß-D-glucopyranoside (W5), taxifolin (W6), luteolin (W7), and apigenin (W8) by spectra of 1 H-NMR and 13 C-NMR data. Among them, compounds Y1, Y2, Y3, and Y6 belong to polyacetylene compounds, and the rest were flavonoids. In addition, counter-current chromatography has been used to separate polyacetylene compounds for the first time. All compounds in this method were isolated from Bidens pilosa Linn. var. radiata Sch. Bip. for the first time.

Screening for Anti-Influenza Actives of Prefractionated Traditional Chinese Medicines.

Traditional Chinese medicines (TCMs) have proven to possess advantages in counteracting virus infections according to clinical practices. It's therefore of great value to discover novel antivirals from TCMs. In this paper, One hundred medicinal plants which have been included in TCM prescriptions for antiviral treatment were selected and prefractionated into 5 fractions each by sequentially using cyclohexane, dichloromethane, ethyl acetate, n-butanol, and water. 500 TCM-simplified extracts were then subjected to a phenotypic screening using a recombinant IAV expressing Gaussia luciferase. Ten TCM fractions were identified to possess antiviral activities against influenza virus. The IC50's of the hit fractions range from 1.08 to 6.45 µg/mL, while the SIs, from 7.52 to 98.40. Furthermore, all the ten hit fractions inhibited the propagation of progeny influenza virus significantly at 20 µg/mL. The hit TCM fractions deserve further isolation for responsible constituents leading towards anti-influenza drugs. Moreover, a library consisting of 500 simplified TCM extracts was established, facilitating antiviral screening in quick response to emerging and re-emerging viruses such as Ebola virus and current SARS-CoV-2 pandemic.

A novel time-dimension and circadian rhythm-dependent strategy for pharmacodynamic evaluation of Uncaria in the regulation of neurotransmitter circadian metabolic homeostasis in spontaneously hypertensive rats.

In the present study, we aimed to use metabolomics platforms to examine circadian-regulated neurotransmitters across a 24-h day and the effects of Uncaria administration on daily rhythmicity in order to establish a strategy for evaluating the spatiotemporal efficacy evaluation strategy of Uncaria. By using targeted ultrahigh performance liquid chromatography-mass spectrometry metabolomics, we quantified 32 neurotransmitter metabolites every 4 h over 24 h. To assess 24-h metabolite rhythmicity, we performed cosinor analysis. The expression of hypothalamic rhythm genes was detected by reverse transcription polymerase chain reaction (RT-PCR). Data revealed circadian loss of many neurotransmitters during the entire circadian cycle in the serum of group M, indicating that hypertension causes circadian rhythm disorders. Cosinor analysis of the rhythmic oscillations of the levels of 32 neurotransmitters in the three groups showed that the metabolite rhythms peaked at approximately the same time of day (ZT4 and ZT16). Moreover, the amplitudes of the metabolite rhythms were altered. After treatment with Uncaria, the amplitudes of 13 neurotransmitters reverted to normal. The change trends in the hypothalamic rhythm genes confirmed the rhythm changes in neurotransmitters. Collectively, a novel pharmacodynamic evaluation strategy was established to dynamically observe the holistic effects of Uncaria on 32 circulating neurotransmitters within 24 h from the perspective of time dimension.

An integrated data filtering and identification strategy for rapid profiling of chemical constituents, with Arnebiae Radix as an example.

Profiling the chemical components of complicated herbal extracts using traditional analytical methods is time-consuming and laborious. In this study, an integrated data filtering and identification strategy was developed to efficiently identify the chemical constituents in Arnebiae Radix. The post-acquisition data processing steps with this strategy were as follows: (1) data acquisition by ultra-high performance liquid chromatography-quadrupole-Orbitrap high-resolution mass spectrometry (UPLC-Q-Orbitrap-MS); (2) background subtraction on the basis of the total ion chromatogram (TIC) to obtain the background-subtracted ion chromatogram; (3) construction of a diagnostic ion database based on the measured MS/MS fragment ions of reference standards and auxiliary diagnostic information according to literatures; (4) mass defect filtering (MDF) to filter the background-subtracted ion chromatogram; and (5) rapid structural identification in the MDF-processed ion chromatogram on the basis of the diagnostic ion database and further structural confirmation by analysing the retention time, fragment behaviour, and online databases (Chemspider, PubChem, and SciFinder). In this study, the herbal medicine Arnebiae Radix was used to illustrate this strategy. A total of 96 compounds were efficiently exposed and characterized from Arnebiae Radix samples obtained from 20 sources, and 13 of these compounds were confirmed by comparison with the reference standards. Thirty components with a low abundance, that remained undetected in the TIC, were identified in the MDF-processed ion chromatogram. Nine of these compounds had not been identified from Arnebiae Radix previously, and were tentatively screened as unknowns. The chemical components in traditional Chinese medicine preparations are considered to be the material basis for the effectiveness of this medical system, and are closely related to the pharmacological activities of the drugs. The pharmacodynamics of these drugs are known to be influenced by the synergistic effects of various components. Therefore, comprehensive profiling of the chemical compositions of herbal extracts is essential for systematic elucidation of the pharmacodynamics of these medicines.

A rapid and practical prediction method for the Arizona solvent system family used in high speed countercurrent chromatography.

Selecting the appropriate solvent system is the key to the successful separation of samples by using countercurrent chromatography. Although high-speed countercurrent chromatography has been widely used in the separation and preparation of natural products, the selection of a solvent system has always been a stumbling block to the application of high-speed countercurrent chromatography. In order to explore a rapid and practical prediction method to select countercurrent chromatography solvent system, five linear prediction models of the Arizona solvent system family (HEMW) was established by using fourteen compounds with different structures and five HPLC columns of different brands. And two different solvent system selection methods (The partition coefficient K of the target compound in the solvent system was in the range of 0.25 < K < 2.5) were proposed for targeted separation of compounds and multi-component separation in a complex sample respectively. The appropriate HSCCC solvent system of five known compounds was determined by a HPLC analysis and a shake flask test and the appropriate HSCCC solvent system of two Chinese herbal extracts was determined by a HPLC analysis to verify the prediction method. In this study, solid-liquid partition chromatography (HPLC) and liquid-liquid partition chromatography (HSCCC) were linked by polarity to simplify the screening process of solvent system. This method reduced the difficulty and workload of solvent system selection, which provided methods and ideas for more solvent system prediction models.

[Review of chemical constituents, pharmacological effects and clinical applications of Jiaotai Pills and predictive analysis of its quality marker(Q-marker)].

Jiaotai Pills is a traditional medical prescription to treat the incompatibility of heart and kidney. It has the distinctive functions of heart and kidney communication, sedation and hypnosis, anti-anxiety and depression, as well as the improvement of insulin resistance. However, this pill is broadly used to cure insomnia, anxiety, depression, and diabetes in the contemporary clinical trials. Based on the article, it illustrates the research progress of the chemical ingredients, pharmacological actions, and clinical applications of Jiaotai Pills. With respect to the "five principles" of Q-marker in Chinese medicine, the Q-marker of Jiaotai Pills is comprehensively predicted and analyzed, noting that berberine, epiberberine, coptisine chloride, palmatine chloride, berberine chloride, berberrubine chloride, ferulic acid, cinnamic acid, cinnamaldehyde, proanthocyanidin B2 can be treated as the Q-marker of Jiaotai Pills. In addition, these components of Q-marker have been selected as indicators to provide a significant reference for the quality control and surveillance research of Jiaotai Pills.

Traditional uses, phytochemistry and pharmacological activities of the genus Cinnamomum (Lauraceae): A review.

Species of Cinnamomum exhibit excellent economic and medicinal value, and have found use in traditional medicine, are consumed as a spice, as well as being cultivated as landscape plants. Investigations into the pharmacological activities of the genus Cinnamomum revealed that it manifested a wide range of pharmacological properties including antimicrobial, antioxidant, anti-inflammatory and analgesic, antitumor, anti-diabetic and anti-obesity, immunoregulation, insecticidal and acaricidal, cardiovascular protective, cytoprotective, as well as neuroprotective properties both in vivo and in vitro. In the past five years, approximately 306 chemical constituents have been separated and identified from the genus Cinnamomum, covering 111 terpenes, 44 phenylpropanoids, 51 lignans, 17 flavonoids, 53 aromatic compounds, 17 aliphatic compounds, four coumarins, two steroids. This article highlights the traditional uses, phytochemistry and pharmacological properties of the few studied taxa of Cinnamomum through searching for the pieces of literature both at home and abroad, which would provide a reference for the pharmaceutical research and clinical application of this genus.

Deciphering the Mechanism of the Anti-Hypertensive Effect of Isorhynchophylline by Targeting Neurotransmitters Metabolism of Hypothalamus in Spontaneously Hypertensive Rats.

Essential hypertension is a major risk factor for cardiovascular disease that can lead eventually to structural and functional alterations in the brain. Accumulating evidence has suggested that the increased activities in renin-angiotensin system and sympathetic nerve participated in the pathogenesis of hypertension that is related to the imbalance between neurotransmitters. The potential role in essential hypertension arising from alterations of neurotransmitters in the central nervous system remains understudied. Isorhynchophylline is a major oxindole alkaloid extracted from Uncaria rhynchophylla, which has been widely used for treating hypertension and neurodegenerative diseases. Whether isorhynchophylline acts on neurotransmitters to lower blood pressure has been hypothesized but rarely demonstrated unequivocally. Here, we studied the metabolic neurotransmitter profiles in the hypothalamus using a targeted metabolomic approach in spontaneously hypertensive rats after isorhynchophylline intervention. Our study demonstrated that isorhynchophylline exhibited a strong anti-hypertensive effect in spontaneously hypertensive rats by improving the neurotransmitter imbalance in the hypothalamus and inhibiting the overactivation of the renin-angiotensin system and sympathetic nerve system. Overall, this study played an essential role in enhancing our understanding of the mechanism of isorhynchophylline in essential hypertension and in providing theoretical evidence for future research and clinical application.

[Establishment of rat heart-kidney insomnia model consistent with traditional Chinese medicine syndrome and its serum metabolomics].

Enzyme-linked immunosorbent assay(ELISA) and metabolomics were used to analyze and compare two animal models of heart-kidney insomnia, in order to explore a more ideal animal model and preliminarily explore the essence of heart-kidney insomnia. Based on the clinical symptoms and disease characteristics of heart-kidney insomnia, the animal model of heart-kidney insomnia was reproduced through intraperitoneal injection with p-chlorophenylalanine(PCPA) and multi-factor interaction. The animal model of disease-syndrome combination was evaluated by behavioral observation, ELISA and metabolomics. Wistar rats were randomly divided into normal group, PCPA group and compound model group(FH). The rats' behavior, body weight, adrenal index and spleen index were recorded. The levels of corticotropin releasing hormone(CRH) and adrenocorticotropin(ACTH) in serum were detected by ELISA, and the differential metabolites in serum were detected by UPLC-QE-MS. The body weight and adrenal index in FH group were significantly lower than those in PCPA group(P<0.05); whereas ACTH and CRH in FH group were significantly higher than those in PCPA group by ELISA; nine potential biomarkers were identified by serum sample statistics. There were four main metabolic pathways in cardiorenal insomnia: pentose phosphate metabolism, alanine, aspartic acid and glutamic acid metabolism, histidine metabolism, and taurine and subtaurine metabolism. PCPA and multi-factor interaction method can successfully replicate the insomnia model, but multi-factor modeling method is more similar to clinical traditional Chinese medicine syndrome. Animal behavior, ELISA and metabolomics were used to evaluate the rat model of cardiorenal insomnia from in vitro to in vivo, from macro to micro, and from individual to the whole.

Qualitative and Quantitative Analysis for the Chemical Constituents of Tetrastigma hemsleyanum Diels et Gilg Using Ultra-High Performance Liquid Chromatography/Hybrid Quadrupole-Orbitrap Mass Spectrometry and Preliminary Screening for Anti-Influenza Virus Components.

Tetrastigma hemsleyanum Diels et Gilg (T. hemsleyanums) is a kind of traditional folk medicinal plant which has been used widely in China for its antivirus, antitumor, and other clinical effects. In this study, ultra-high performance liquid chromatography coupled with hybrid quadrupole-orbitrap mass spectrometry (UPLC-Q-Exactive/MS) was utilized to analyze the chemical constituents of T. hemsleyanums. Fifty-one constituents were clarified, including flavonoids, anthraquinones, esters, fatty acids, phenols, and catechins. In the subsequent quantitative analysis, the contents of ten compounds of rutin, kaempferol, astragalin, quercitrin, quercetin, vitexin-rhamnoside, isorhamnetin, vitexin, emodin-8-O-ß-D-glucoside, and isoquercetin in 18 batches of T. hemsleyanums collected from different places of cultivation were determined. Meanwhile, anti-influenza virus bioactivity in vitro of the above samples was detected with Gaussia Luciferase viral titer assay. It was found that the antiviral bioactivity varied from batches to batches in accordance with content difference of the chemical constituents in T. hemsleyanums. Correlation analysis was performed with SPSS software for the association between LC-MS chemometrics and bioactivity of influenza virus inhibition, and 8 constituents of flavonoids showed positive correlation coefficient, which may provide a valuable clue for searching potential antiviral components in T. hemsleyanums.

Simultaneous quantification of fifteen compounds in rat plasma by LC-MS/MS and its application to a pharmacokinetic study of Chaihu-Guizhi decoction.

A simple, sensitive and selective high-performance liquid chromatography electrospray ionization tandem mass spectrometry (HPLC-ESI-MS/MS) method was developed and validated for simultaneous determination and pharmacokinetic study of 15 active compounds (Saikosaponin A, Baicalin, Wogonin, Glycyrrhizic acid, Glycyrrhetinic acid, Albiflorin, Paeoniflorin, Liquiritin, Isoliquiritin, Liquiritigenin, Isoliquiritigenin, Cinnamic acid, Gallic acid, Wogonoside and Oroxylin A) in rat plasma. After a feasible protein precipitation using methanol for sample preparation, chromatographic separation was carried out with a Halo® C18 column (2.1 × 100 mm, 2.7 µm) at 35 °C, water containing 0.1% formic acid and acetonitrile were used as the mobile phase with a flow rate of 0.3 mL/min. Multiple reaction monitoring (MRM) with positive and negative ion switching mode was performed for the quantification of the standards and internal standard in plasma. All the calibration curves showed good linear regression within the linear range (r2 > 0.9923). In particular, the results of the method validation including specificity, linearity, accuracy, precision, extraction recovery, matrix effect, and stability of compounds in bio-samples were all within the current acceptance criteria. The established method was successfully applied to the pharmacokinetic study of 15 compounds in rats after oral administration of CGD and laid the foundation for studying the active components and mechanism of CGD in vivo.

Study on the Intervention Effects of Pinggan Prescription () on Spontaneously Hypertensive Rats Based on Metabonomic and Pharmacodynamic Methods.

OBJECTIVE: To investigate the effects of Pinggan Prescription (, PGP) on hypertension by the associated methods of metabonomic and pharmacodynamic. METHODS: A total of 32 male spontaneously hypertensive rats (SHRs) were randomly divided into two groups by using the random number table method: a treatment group (n=18) and a model group (n=14). The Wistar rats (n=14) were used as the normal group. Different prescription were used to intervene three groups: the treatment group in which PGP extract was administered orally at a dose of 18.336 g/kg (PGP/body weight), and the model group in which physiological saline was administered at the equivalent dose. The same treatment was applied to the normal group as the model group. The blood pressure was measured by tail-cuff method, and pharmacodynamic indexes including cyclic adenosine monophosphate (cAMP) and angiotensin II (Ang II) were tested by enzyme-linked immunosorbent assay. The plasma samples from three groups were detected by gas chromatography-mass spectrometry (GC-MS). RESULTS: Compared with the model group, blood pressure of treatment group was obviously reduced after continuous curing with PGP (P<0.01). The pharmacodynamic results illustrated that the content of Ang II increased with the raised blood pressure and the cAMP expressed the converse trend. After curing with PGP, the content of Ang II decreased, the difference between model group and treatment group was significant (P<0.01), and the cAMP expressed the converse trend. Five potential biomarkers were identified, including arachidonic acid, hexadecanoic acid, elaidic acid, octadecanedioic acid and 9,12-octadecadienoic acid. These metabolites had shown significantly changes as followed: arachidonic acid, hexadecanoic acid and elaidic acid were significantly higher and octadecanedioic acid and 9,12-octadecadienoic acid were lowered in the model group than those in the normal group. After the treatment of PGP, the metabolites had the trends of returning to normal along with the reduced blood pressure. CONCLUSIONS: PGP intervention for hypertension played a major role in the metabolism of arachidonic acid and linoleic acid. Metabonomic with pharmacodynamic methods could be potentially powerful tools to investigate the mechanism of Chinese medicine.

Metabolomics Research Reveals the Mechanism of Action of Astragalus Polysaccharide in Rats with Digestive System Disorders.

With the diversity of modern dietary lifestyles, digestive system disorders (DSD) have become a frequently occurring disease in recent years. Astragalus polysaccharide (APS) is a homogeneous polysaccharide extracted from Astragalus, which might ameliorate the digestive and absorptive functions. However, the treatment mechanisms remain unclear. In this study, rats with DSD were fed a high-fat⁻low-protein diet and subjected to weight-bearing swimming until exhaustion. When body weight and autonomous activities of the rats decreased, they were administered APS. After two weeks, serum metabolomics analysis based on LC-MS was performed to validate the therapeutic effect of APS and explore its mechanism. APS pharmacodynamics was determined in this study, and serum metabolomics analysis discovered and identified 16 significant, differentially produced metabolites involved in energy, amino acid, and lipid metabolism, including citric acid, lactic acid, alanine, phosphatidylcholine, phenylalanine. After treatment with APS, the levels of the above small-molecule metabolites were reversed. Our results show the efficacy of APS in DSD treatment through the regulation of perturbed metabolic pathways related to energy, amino acid, and lipid metabolism.