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OBJECTIVE To study the mechanism of Compound zaoren granule in improving insomnia. METHODS Forty-nine mice were divided into blank group， model group， positive control group 1 （Estazolam tablets 0.5 mg/kg），control group 2 （Shumian capsule 0.6 g/kg）， Compound zaoren granule low-dose， medium-dose and high-dose groups （2.5， 5， 10 g/kg）， with 7 mice in each group. The insomnia model was established by chronic unpredictable mild stress combined with 4-chloro-DL- phenylacetic acid. The behavioral changes of mice were investigated through open field test and pentobarbital sodium synergistic hypnosis experiment， as well as the pathomorphology of mice hypothalamus tissue was observed by HE staining. The metabonomics analysis and multivariate statistical analysis of serum in mice were performed by UHPLC-Q-TOF-MS/MS， and the differential metabolites were screened out； the metabolic pathway analysis was conducted based on MetaboAnalyst 5.0 database. RESULTS Compared with blank group， the total travelling distance， the number of entering the central region and the moving distance in the central region of the model group were significantly reduced （P＜0.05）， the proportion of total rest time was significantly increased （P＜0.05）， the sleep duration of mice was significantly shortened （P＜0.05）， and hypothalamic nerve cells damaged and severely vacuolated. Compared with model group， the total travelling distance of Compound zaoren granule low-dose and medium-dose groups were increased significantly and the proportions of total rest time of those groups were decreased significantly （P＜0.05）， and the sleep duration of mice in Compound zaoren granule high-dose group was prolonged significantly （P＜0.05）； the hypothalamic nerve cells of mice in each administration group recovered to varying degrees， and the hypothalamus histiocytes of mice in the Compound zaoren granules high-dose group were closer to those in the blank group. A total of 18 differential metabolites （such as phenylalanine， taurine， norvaline， methionine） and 4 important amino acid metabolic pathways （L-phenylalanine， tyrosine and tryptophan biosynthesis； taurine and hypotaurine metabolism； L-phenylalanine metabolism； cysteine and methionine metabolism） were identified through metabolomics analysis. CONCLUSIONS Compound zaoren granules can normalize the disordered metabolism in vivo by regulating differential metabolites such as phenylalanine， taurine， and four amino acid metabolic pathways， so as to improve insomnia.
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ObjectiveTo perform a predictive analysis of the quality marker（Q-Marker） for the anticoagulant activity of Kunning granules. MethodThe chemical components of Kunning granules were analyzed by ultra high performance liquid chromatography-quadrupole-time of flight tandem mass spectrometry（UHPLC-Q-TOF-MS/MS） on a Waters ACQUITY UPLC HSS T3 column（2.1 mm×100 mm， 1.8 μm） with the mobile phase of acetonitrile（A）-25 mmol∙L-1 ammonium acetate aqueous solution（B） for gradient elution （0-5 min， 5%-22%A； 5-10 min， 22%-30%A； 10-15 min， 30%-95%A； 15-20 min， 95%-5%A； 20-30 min， 5%A）， flow rate of 0.2 mL∙min-1， column temperature at 30 ℃， injection volume of 1 μL， electrospray ionization（ESI）， positive and negative ion detection modes. Interaction analysis between the targets of chemical components and the targets of abnormal uterine bleeding（AUB） was performed by network pharmacology， and the key components were screened through network topology analysis. The fingerprints of 10 batches of Kunning granules were established by high performance liquid chromatography（HPLC）， the anticoagulant activity of the granules was determined by blood coagulation method and fibrinogen plate method， and the spectrum-effective relationship was established. The components co-occurring in the topological analysis and spectrum-effective relationship were selected as Q-Markers， and their anticoagulant activities were verified and confirmed. ResultA total of 475 chemical components were identified from Kunning Granule， of which 22 key components such as salvianolic acid B， paeoniflorin， naringin and neohesperidin， were the potential material basis for the treatment of AUB. The spectrum-effective analysis showed that peaks 7（paeoniflorin）， 9（naringin）， 10（neohesperidin） and 11（salvianolic acid B） were the optimal principal components， and in vitro activity test showed that these four components could better characterize their anticoagulant activity. ConclusionSalvianolic acid B， paeoniflorin， neohesperidin and naringin may be Q-Markers for the anticoagulant activity of Kunning granules.
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OBJECTIVE：To identi fy chemical components of Actinidia chinensis root rapidly ，and to provide reference for further material basis and quality control study of the crude medicine. METHODS ：UHPLC-Q-TOF-MS/MS technique was used to detect chemical components of A. chinensis root. The separation was performed on Waters XSelect HSS T 3 column with mobile phase consisted of 0.1％ formic acid acetonitrile solution- 0.1％ formic acid water solution （gradient elution ）at the flow rate of 0.3 mL/min. The column temperature was set at 40 ℃，and sample size was 3 μL. Electrospray ion source was adopted，the data was collected under negative ion mode ；the scanning range was m/z 50-1 500；the drying gas temperature was 350 ℃，the atomizing air pressure was 45 psi，the capillary voltage was 3 500 V，and sheath gas temperature was 350 ℃. According to the information of excimer ion and secondary fragment ion ，the chemical components were identified by combining with the relevant literature ，the retention time of the reference substance and the law of mass spectrometry cracking. RESULTS & CONCLUSIONS ：Totally 58 chemical components was identified ，which included 16 pentacyclic triterpenes （such as hydroxyasiatic acid ，asiatic acid ，maslinic acid，corosolic acid ，oleanic acid ，ursolic acid ，etc.），12 flavonoids（such as rutin ，quercitrin，cynaroside，astragalin，etc.），17 organic acids （such as cryptochlorogenic acid ，chlorogenic acid ，isochlorogenic acid A ，isochlorogenicacid C ，etc.）. There were 9 components（such as procydanidin B 1，B2 and luteolin ，etc.）identified for the first time in A. chinensis root. UHPLC-Q-TOF-MS/ MS technique can be used for the rapid identification of chemical components in A. chinensis root.
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Objective: To identify the in vivo metabolites of icaritin and speculate its metabolic profiling in rats. Methods: The plasma, bile, urine, and feces of rats were collected after orally administration of icaritin at a dose of 100 mg/kg and detected by an ultra-high performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC/Q-TOF-MS/MS)in both positive and negative modes. The data of treated and control groups were compared and analyzed with the aid of Metabolynx XS software. Results: A total of 25 metabolites were identified in the biosamples, and 14 of them were reported for the first time to our knowledge. Conclusion: The main metabolite types of icaritin in rats were glucuronide conjugation, methylation, hydroxylation, reduction, and acetylation.
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The root of Aster tataricus L. f. (RA) has been widely used in the clinic for moistening lung, dispelling phlegm and relieving cough because of its significant therapeutic effects on respiratory diseases. In this study, a systematic data acquisition and mining strategy was established aimed at solving the complexity of the traditional Chinese medicine using ultra high performance liquid chromatography coupled with quadruple time of flight mass spectrometry (UHPLC-Q-TOF-MS). A total of 132 chemical constituents, including 43 terpenes, 31 flavonoids, 22 organic acids, 18 peptides, 9 coumarins, 3 steroids, 3 anthraquinones and 3 aldehydes were identified or tentatively characterized, among which 59 components were confirmed by comparison with the standard references. Meanwhile, the accurate mass measurements of the identified components were all with ±5 ppm error. Therefore, this work provided not only reliable data supports for the comprehensive analysis of the chemical constituents in RA, but also provided an efficient data acquisition and mining strategy to profile the chemical constituents for other traditional Chinese medicine complex system.
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The Chinese herbal Sophora alopecuroides is widely used to clean intestine and eliminate dampness, and it has good therapeutic effects on treating bacillary dysentery and inflammatory bowel disease, etc. in clinics. However, the mechanism of treatment is not yet well understood. The present study was aimed to explore the mechanism of Sophora alopecuroides treatment of large intestine dampness-heat syndrome (LIDHS). The LIDHS model was performed by the comprehensive factors, including high temperature and humidity environment, high-sugar and high-fat diet, and intraperitoneal injection of Escherichia coli. The blood routine, serum proinflammatory cytokine levels and histopathological changes of intestine were detected and observed. Meanwhile, the serum metabolomic approach was conducted using the method of ultra performance liquid chromatography coupled to quadrupole time-of-flight mass/mass spectrometry (UHPLC-Q/TOF-MS/MS). The results showed that Sophora alopecuroides has good therapeutic effects on the LIDHS rat models. After treatment with Sophora alopecuroides, the abnormality of blood routine indexes as well as proinflammatory cytokines, including IL-1β, IL-2, IL-6 and TNF-α in vivo, tended to be normal, and the histopathological changes of intestine were improved. Through metabolic profiling and protocol analysis, 9 potential metabolic markers may be closely related with the treatment mechanism of Sophora alopecuroides on this disease, including taurine, L-tryptophan, LysoPE, LysoPC, LPA, DG, chenodeoxycholic acid disulfate, traumatic acid and 7-ketodeoxycholic acid, which were involved in taurine and hypotaurine metabolism, glycerophospholipid metabolism, glycerolipid metabolism, tryptophan metabolism and primary bile acid biosynthesis etc. The serum metabolomic approach can be applied to clarify the therapeutic mechanism of Sophora alopecuroides on LIDHS, and provide the theoretical basis for Sophora alopecuroides in clinical practice.
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Objective To investigate the prototype components and metabolites of Mentha haplocalyx in rats plasma. Methods UHPLC-Q-TOF-MS/MS method was developed and applied to identify the components and metabolites of rat plasma. The analysis was carried out on an Eclipse Plus C18 column (100 mm × 4.6 mm, 3.5 μm, Agilent) with the mobile phase of 0.1% acetic acid solution (A)-acetonitrile (B) at a flowing rate of 0.3 mL/min, and the injection volume was 5 μL. Results The developed method was applicable to the analysis and identification of metabolites of M. haplocalyx after oral administration. A total of 67 compounds, including 28 prototype components and 39 metabolites (one of which was a new metabolite of luteolin unreported), were identified by comparison of their retention time, ion fragmentation information with that of the blank biological samples, herb extract, and reference compounds. Conclusion The metabolic pathway of M. haplocalyx in rats were consisted of oxidation, reduction, methylation, sulfation, and glucuronidation, and the main metabolic pathway was phase II metabolic pathway among of them. This experimental method is simple and reliable, which could provide theoretical basis for elucidating the bioactive components of M. haplocalyx.
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In this study, we developed a qualitative analytical method based on liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry (UHPLC-Q-TOF-MS/MS) for identification of multi-constituents of raw Fructus Arctii (RFA) and processed Fructus Arctii (PFA). We established a UHPLC-UV analytical method for simultaneously determining 6 major compounds in Fructus Arctii. UHPLC- Q-TOF-MS/MS qualitative analysis was performed under negative and positive ion modes and a total of 23 chemical compounds were identified. The analysis data were subjected to a principle component analysis with a t-test. Ten peaks were found to be the main difference (P<0.05) between RFA and PFA. HPLC-UV quantitative method result showed the contents of 6 constituents were different between RFA and PFA. The results indicated that there was less arctiin, chlorogenic acid, isochlorogenic acid A in PFA than in RFA. However, there were higher levels of arctigenin, isochlorogenic acid B, isochlorogenic acid C in the PFA than RFA, which may be the main reason for different clinical efficacy of RFA and PFA.
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OBJECTIVE: To identify the metabolites of Flos scabiosae extract in rats after oral administration for exploring its metabolism mode in vivo. METHODS: Welch Ultimate UHPLC C18 column (2.1 mm×100 mm, 1.7 μm) was used for the analysis, and the column temperature was maintained at 25℃. Gradient elution was conducted with mobile phase consisting of 0.1% acetic acid water (A) and acetonitrile (B) at a flowing rate of 0.35 mL·min-1. The injection volume was 6 μX. Urine, plasma, and feces samples were collected during 0-24 h or 0-12 h after oral administration of Flos scabiosae extract to rats. The samples and control samples were analyzed by UHPLC-Q-T MS/MS in the scanning mode to acquire the full-scan chromatograms. The data acquisition and processing analysis were conducted using compound retention time, precise molecular mass, elemental composition, isotopic abundance, and mass spectra with Peak View v1.2 data processing software. Metabolites were identified by comparison of chromatograms of the urine, feces and plasma samples after administraion of Flos scabiosae extract with those of the blank biological samples and the herb extract. RESULTS: The developed method is applicable to the analysis and identification of metabolites of Flos scabiosae extract in biological matrices after oral administration. Based on the investigation of Flos scabiosae extract in rats, 28 parent compounds with 31 metabolites were detected in rat urine, feces, and plasma. CONCLUSION: In this study, the developed method is simple and efficient. The main metabolism pathways of Flos scabiosae extract in rats may include oxidation, demethylation, glucuronidation, and sulfation. This study provides a novel pattern and illumination for the research of material basis of Flos scabiosae.
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OBJECTIVE: To qualitatively analyze the chemical constituents of Folium hibisci Mutabilis by high performance liquid chromatography-electrospray ionization-quadrupole-time of flight-mass spectrometry (UHPLC-Q-TOF-MS/MS). METHODS: The separation was performed on a UHPLC Welch C18 column (2.1 mm×100 mm, 1.7 μm), with acetonitrile-0.1% formic acid as the mobile phase for gradient elution; ESI ion source was used; the data was collected in a negative ion mode. The chemical components of Folium hibisci Mutabilis were identified by analyzing the retention time, exact relative molecular mass, and cleavage fragments of MS/MS. RESULTS: A total of 73 compounds were identified in Folium hibisci Mutabilis, including 42 flavonoids, 15 triterpenoids, 11 organic acids, 3 coumarins, and 2 other kinds of compounds by UPLC-Q-TOF-MS. CONCLUSION: LC-MS/MS can identify the chemical components of Folium hibisci Mutabilis in a simple, which would provide a foundation for further exploration of the effective substances of Folium hibisci Mutabilis.