[Study on anticoagulant material basis and mechanism of Trichosanthis Semen and its shell and kernel based on spectrum-effect relationship integrated molecular docking].

This study explored the anticoagulant material basis and mechanism of Trichosanthis Semen and its shell and kernel based on spectrum-effect relationship-integrated molecular docking. High performance liquid chromatography(HPLC) fingerprints of Trichosanthis Semen and its shell and kernel were established. Prothrombin time(PT) and activated partial thromboplastin time(APTT) in mice in the low-and high-dose(5, 30 g·kg~(-1), respectively) Trichosanthis Semen, the shell, and kernel groups were determined as the coagulation markers. The spectrum-effect relationship and anticoagulant material basis of Trichosanthis Semen and its shell and kernel were analyzed with mean value calculation method of Deng's correlation degree(MATLAB) and the common effective component cluster was obtained. Then the common targets of the component cluster and coagulation were retrieved from TCMSP, Swiss-TargetPrediction, GenCLiP3, GeneCards, and DAVID, followed by Gene Ontology(GO) term enrichment and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment of the targets. The main anticoagulant molecular mechanism of the component cluster was verified by SYBYL-X 2.1.1. The spectrum-effect relationship of Trichosanthis Semen and its shell and kernel was in positive correlation with the dosage. The contribution of each component to anticoagulation was not the same, suggesting that the material basis for anticoagulation was different, but they have common effective components(i.e. common material basis), such as adenine(peak 3), uracil(peak 4), hypoxanthine(peak 6), xanthine(peak 9), and adenosine(peak 11). Network pharmacology showed that these components can act on multiple target proteins such as NOS3, KDR, and PTGS2, and exert anticoagulant effect through multiple pathways such as VEGF signaling pathway. They involved the biological functions such as proteolysis, cell component such as cytosol, and molecular functions. The results of molecular docking showed that the binding free energy of these components with NOS3(PDB ID: 1 D0 C), KDR(PDB ID: 5 AMN), and PTGS2(PDB ID: 4 COX) was ≤-5 kJ·mol~(-1), and the docking conformations were stable. Spectrum-effect relationship-integrated molecular docking can be used for the optimization, virtual screening, and verification of complex chemical and biological information of Chinese medicine. Trichosanthis Semen and its shell and kernel have the common material basis for anticoagulation and they exert the anticoagulant through multiple targets and pathways.

[Mechanism and material basis of Lianhua Qingwen capsule for improving clinical cure rate of COVID-19: a study based on network pharmacology and molecular docking technology].

OBJECTIVE: To explore the potential targets, signal pathways and biological functions that mediate the effect of Lianhua Qingwen capsule in improving clinical cure rate of COVID-19 in light of network pharmacology and molecular docking technology. METHODS: TCMSP, Target, Prediction, CooLGeN, GeneCards, DAVID and other databases were searched for the active components and their target proteins from 13 herbs including Forsythia, Honeysuckle and roasted Ephedra used in Lianhua Qingwen capsule. The common target proteins, signal pathways and biological functions shared by these components and the clinical manifestations of COVID-19 (fever, cough, and fatigue) were identified to construct the network consisting of the component drugs in Lianhua Qingwen capsule, the active ingredients of, their targets of action, and the biological functions involved using Gephi software. RESULTS: A total 160 active components including MOL000522, and MOL003283, MOL003365, MOL003006, MOL003014 in 13 component drugs in Lianhua Qingwen capsule produced therapeutic effects against COVID-19 through 57 target proteins including MAPK1, IL6, HSP90AA1, TNF, and CCL2, involving 35 signaling pathways including NOD-like receptor signaling pathway and Toll-like receptor signaling pathway. The results of molecular docking showed that 83 chemical components had total scores no less than 5.0 for docking with 12 target proteins (including MAPK1, IL6, and HSP90AA1) with high binding activities to form stable conformations. The binding of MOL000522, MOL004989, and MOL003330 with MAPK1; MOL001495 and MOL001494 with NLRP3; MOL004908, MOL004863 and MOL004806 with HSP90AA1; MOL001749 with TLR9; and MOL001495 with AKT1 all had total scores exceeding 9.0. CONCLUSIONS: Lianhua Qingwen capsule contains multiple effective ingredients to improve clinical cure rate of COVID-19, and its therapeutic effect is mediated by multiple protein targets, signal pathways and biological functions.

[Mechanism of Qingfei Paidu decoction for treatment of COVID-19: analysis based on network pharmacology and molecular docking technology].

OBJECTIVE: To explore the target, signaling pathways and their biological functions of Qingfei Paidu Decoction in the treatment of COVID-19 based on network pharmacology and molecular docking technology. METHODS: The active components and target proteins in 21 drugs such as Ephedrae Herba and Pinelliae Rhizoma in Qingfei Paidu decoction were analyzed, and the signaling pathways and biological functions of the target proteins common with COVID-19 were screened by using TCMSP, Swiss Target Prediction, CooLGeN, GeneCards, DAVID and other databases. The network diagram of Qingfei Paidu decoction was constructed using Gephi software. RESULTS: We identified 163 active ingredients, including MOL004798, MOL000519, MOL004824, MOL000554, MOL010428, and MOL013443, from 18 drugs in Qingfei Paidu decoction (such as Ephedrae Herba, Pinelliae Rhizoma, Glycyrrhizae Radix Et Rhiizoma, Farfarae Flos, Asteris Radix Et Rhizoma and Aurantii Fructus Immaturus). These ingredients activate renin-angiotensin system signaling pathway and apoptosis signaling pathway by regulating 10 protein targets (ACE, ACE2, AGTR1, FURIN, TNF, CASP3, CASP6, DPP4, MCL1 and POLD1) to execute 42 biological functions such as renin-angiotensin regulation of blood volume and systemic arterial blood pressure to treat COVID-19. The results of preliminary molecular docking showed that MOL000519 (from Pinelliae Rhizoma), MOL000554 (from Farfarae Flos), MOL004798 (from Ephedrae Herba), MOL004824 (from Glycyrrhizae Radix Et Rhiizoma), MOL010428 (from Asteris Radix Et Rhizoma), and MOL013443 (from Aurantii Fructus Immaturus) had good affinity with SARS-CoV-2 3CL hydrolase to form complexes with stable conformations and high binding activity (binding energy ≤- 5 kJ/mol). CONCLUSIONS: Qingfei Paidu decoction can treat COVID-19 through its multiple medicinal ingredients that have multiple targets and involve multiple signaling pathways for different biological functions. Our finding provides reference for further investigation into the pharmacological mechanism of Qingfei Paidu decoction in treating COVID-19.

[LC-MS/MS analysis of pharmacokinetics of 3, 29-dibenzoyl karounitriol in rats following administration of Trichosanthes sustained-release pellets].

OBJECTIVE: To study the plasma concentration and pharmacokinetics of 3, 29-Dibenzoyl Karounitriol (3, 29-DK) from sustained- release pellets and extracts of Trichosanthes at different time points in rats using high-performance liquid chromatography- tandem mass spectrometry (LC-MS/MS). METHODS: Healthy male SD rats were given a single gavage of Trichosanthes sustained-release pellets or Trichosanthes extract, and orbital blood samples were taken at different time points within 48 h after drug administration in the pellet group and within 5 h in Trichosanthes extract group for determination of the plasma concentrations of 3, 29-DK using LC-MS/MS. The standard curve of 3, 29-DK content was established, and the specificity, minimum detection limit, precision and accuracy, extraction recovery, stability and matrix effect of LC-MS/MS analysis were assessed. The mean plasms levels of 3, 29-DK at different time points after the drug administration were determined and its pharmacokinetic parameters were calculated using Das 2.0 software. RESULTS: LC-MS/MS analysis showed a good linearity of 3, 29-DK concentration within the range of 0.5-32 ng/mL, and the results of methodological validation confirmed the validity of this method for biological sample determination. Trichosanthes sustained-release pellets and Trichosanthes extract showed significant differences in their AUC(0-t), AUC(0-∞), MRT(0-t), MRT(0-∞), t1/2z and Tmax of 3, 29-DK after administration in rats (P < 0.05). CONCLUSIONS: Trichosanthes sustained-release pellets are capable of sustained-release of 3, 29-DK in rats, and thus provides a basis for the study of new dosage forms of Trichosanthes.

[Anti-platelet aggregation and anti-thrombotic mechanism of Trichosanthis Fructus combined with aspirin based on network pharmacology].

To explore the anti-platelet aggregation and anti-thrombotic mechanisms of Trichosanthis Fructus combined with aspirin based on network pharmacology and the validation of arteriovenous by pass model in rats. The databases of Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP),Drug Repositioning and Adverse Drug Reaction Chemical-Protein Interactome(DRAR-CPI),Universal Protein Resource(Uniprot) and the Database for Annotation,Visualization,and Integrated Discovery(DAVID) were used to predict protein targets and analyze biological pathway and signal pathway in the combination of Trichosanthis Fructus with aspirin. The effects of pretreatment with Trichosanthis Fructus pellets,aspirin pellets and their combination on thromboxane B2(TXB2),6-keto prostaglandin F1α(6-keto-PGF1α) and cyclic adenosine monophosphate(c AMP) in rat thrombotic model were studied. Through the study of network pharmacology,12 components of aspirin and Trichosanthis Fructus,including hydroxygenkwanin,quercetin and adenosine,were found to show the anti-platelet aggregation and anti-thrombosis mechanisms through9 common protein targets,such as SRC,RAC1,MAPK14,MAPK1,AKT1,and 14 common signaling pathways,such as VEGF signaling pathway. After the intervention with Trichosanthis Fructus pellets combined with aspirin pellets,the vascular endothslia growth factor(VEGF) signaling pathway can be activated to inhibit platelet aggregation and improve vascular endothelial function,and show the anti-platelet aggregation and anti-thrombosis mechanisms,which verify the results of the network pharmacology,and explain the anti-platelet aggregation and anti-thrombotic mechanisms of the combination of Trichosanthis Fructus pellets with aspirin pellets.

[Research progress on chromatographic fingerprint similarity evaluation method for traditional Chinese medicine in the past 30 years（1988-2017）and its prospect].

Over the past 30 years, the chromatographic fingerprint technology of traditional Chinese medicine (TCM) has been developed from academic discussion to application for the research and development of TCM which has promoted the technological innovation of Chinese medicine industry and the progress of quality standard of TCM. The similarity evaluation method of chromatographic fingerprint of TCM has played a key role in this process. According to the number of literature and research tendency in terms of the chromatographic fingerprint in the last 30 years, the chromatographic fingerprint evaluation could be divided into three stages: the direct comparison stage (1988-1999), similarity evaluation stage (2000-2009) and the similarity evaluation development stage (2010-2017). In this paper, the research progress of chromatographic fingerprints similarity evaluation of TCM in the last 30 years and its prospect were discussed, which may lead to a more mature stage for this method.

[Effect of different data standardization methods on spectrum-effect relationship for anticoagulation of Trichosanthis Fructus dropping pills].

To study the effect of different data standardization methods on the spectrum-effect relationship for anticoagulant effect of Trichosanthis Fructus dropping pills. The spectrum-effect relationship was studied by using grey correlation degree method between three doses of Trichosanthis Fructus dropping pills and prothrombintime (PT) in mice. The effect of 10 data standardization methods, namely minimization method, maximum method, data extreme difference method, standard deviation standardization method, initialization transformation method, mean transformation method, ratio of each chromatographic peak area to the total peak area, ratio of each chromatographic peak area to the common peak area, logarithmic standardization method and tangent normalization method on the spectrum-effect relationship between Trichosanthis Fructus dropping pills and PT in mice was evaluated by using relative correlation degree as the index. The results of spectrum-effect relationship can be expressed by the minimization method, the data extreme difference method, the standard deviation standardization method, the initialization method and the mean transformation method, with highest relative correlation degree by the mean transformation method. As compared with the mean transformation method, there were significant differences between the high dose group and the medium dose group in the minimization method and the data extreme difference method (P<0.01), while the minimization method in the low dose group showed statistical significance (P<0.05). The standard deviation standardization method, initialization method and the mean transformation method can be used to study the spectrum-effect relationship for the anticoagulation of Trichosanthis Fructus dropping pills.

[Spectrum-activity relationship of trichosanthis fructus and trichosanthis fructus strip pieces for rat myocardial ischemia-reperfusion injury].

To investigate the spectrum-activity relationship of Trichosanthis Fructus and Trichosanthis Fructus strip pieces for rat myocardial ischemia-reperfusion injury. HPLC fingerprints of Trichosanthis Fructus and Trichosanthis Fructus strip pieces were established, and the values of creatinekinase-MB (CK-MB), myoglobin (MYO) and cardiac troponin-T (cTNT) in 3 dose groups (2.25, 13.5, 27.0 g·kg⁻¹, equivalent to the crude herb g·kg⁻¹) of Trichosanthis Fructus and Trichosanthis Fructus strip pieces with myocardial ischemia-reperfusion injury in rats were measured, and the grey relational analysis was used to study the spectrum-activity relationship of Trichosanthis Fructus and Trichosanthis Fructus strip pieces for rat myocardial ischemia-reperfusion injury. With the dosage increase from 2.25 g·kg⁻¹ to 27.0 g·kg⁻¹, the correlation degree of spectrum-activity relationship of Trichosanthis Fructus and Trichosanthis Fructus strip pieces was also enhanced, but the change trend was different between these two groups. According to the frequency of the top 10 peaks in the correlation degree, peak 17, 14, 16, 19, 32, 12, 26, 30, 4, 6 and 2 were the basic effective substances group of Trichosanthis Fructus, peak 6,14,12,32,30,4 and 6 were the basic effective substances group of Trichosanthis Fructus strip pieces. Peak 6, 14, 12, 32, 30, 4 and 26 in fingerprints of Trichosanthis Fructus and Trichosanthis Fructus strip pieces were the main common pharmacodynamic substance base, among them, peak 6 was 5-hydroxymethyl furfural, peak 14 was vanillic acid and the peak 28 was rutin, but the correlation degree with the efficacy was different. The effect of Trichosanthis Fructus and Trichosanthis Fructus strip pieces on rat myocardial ischemia-reperfusion injury was due to the synergistic effect of the effective substance groups related to the dosage. The essential pharmacodynamic substance groups of Trichosanthis Fructus and Trichosanthis Fructus strip pieces were different, but they shared a common active ingredient group.

[Pharmacodynamics Study on Gualou Xiebai Dropping Pills and Its Medicinal Ingredients in Prescription].

OBJECTIVE: To study the pharmacodynamics of Gualou Xiebai Dropping Pills and its medicinal ingredients in prescription on anti-myocardial ischemia. METHODS: SPF Rats were divided randomly into eleven groups with ten rats in each group and half male and half female, the rats were respectively given the physiological saline(blank group and model group), Gualou, Xiebai, Gualou Xiebai Baijiutang (all equivalent to the crude herb of 22. 5 g/kg), Gualou Xiebai. Dropping Pills in the doses of 3. 75,11. 25,22. 5,33. 75 and 45 g/kg and Compound Danshen Drop Pills of 0. 085 g/kg by gavage one time a day for seven days. Except blank group, other rats were given by intraperitoneal injection of isoproterenol to establish myocardial ischemia models, changes of ST segments in ECG were observed in all groups, and the levels of SOD, NO, HDL-C, MDA, CAT, LDH and CK in blood plasma were detected, and the pathological changes of myocardial tissues were observed under light microscope by HE staining. RESULTS: Compared with model group, ST segments in ECG dropped markedly at different time point which included 10,11 and 12 (P <0. 05) in Gualou Xiebai Drop Pills groups of 22. 5, 33. 75 and 45 g/kg, time points were more than those of other groups. Gualou Xiebai Dropping Pills groups of 22. 5 and 33. 75 g/kg improved the levels of SOD, MDA, CAT, NO, HDL-C, LDH and CK in blood plasma in model rats significantly (P <0. 01 or P <0. 05). Gualou Xeibai Dropping Pills improved the pathological changes of myocardial tissues at all dosages. CONCLUSION: Gualou Xiebai Drop Pills can effectively restrain the acute myocardial ischemia induced by isoproterenol in rats, compared with Gualou, Xiebai or Gualou Xiebai Baijiutang, Gualou Xiebai Drop Pills obtains a favourable effect.

[Study on HPLC Fingerprint of Trichosanthes Fruit and Its Processed Products].

OBJECTIVE: To study the correlation on chemical fingerprint features between Trichosanthis Fructus and its processed products. METHODS: The chemical fingerprints were established by HPLC for the ethyl acetate extraction and the n-butanol extraction in Trichosanthis Fructus and its processed products,the common pattern was established by the mean and the median, and the similarity degree between Trichosanthis Fructus and its processed products was calculated by the correlation coefficient method and the included angle cosine method. RESULTS: There were 24 common peaks in the fingerprints of ethyl acetate extraction of Trichosanthis Fructus and its processed products,the average similarity degree was calculated separately by the correlation coefficient method and the included angle cosine method:the former as the value was 0. 8497(mean), 0. 8344(median); and the latter as the value was 0. 8429(mean), 0. 8536 (median); there were 6 common peaks in the fingeprints of n-butanol extraction of Trichosanthis Fructus and its processed products, the average similarity degree was calculated by the correlation coefficient method and the included angle cosine method:the former as the value was 0. 9044 (mean), 0. 9076 (median); and the latter as the value was 0. 9075 (mean), 0. 9081 (median). CONCLUSION: Main peaks have good correlation between Trichosanthis Fructus and its processed products. This method can provide theoretical basis for processing and quality control of Trichosanthis Fructus.

[Spectrum-Effect Relationship of GualouXiebai Dropping Pills on Myocardial Ischemia].

OBJECTIVE: To study the relationship between HPLC characteristic spectrum and pharmacodynamics on anti-myocardial ischemia of GualouXiebai dropping pills. METHODS: HPLC characteristic spectrum of GualouXiebai dropping pills was established, dropping pills were divided into five dose groups (3.75, 11.25, 22.5, 33.75 and 45 g/kg, equivalent to the crude herb g/kg), the mice were orally administered dropping pills once daily for 7 d, 90 min after the mice were given by intraperitoneal injection of isoprenaline to establish myocardial ischemia models, the level of CK in blood plasma were detected; Then, the correlation between characteristic spectrum and biochemical index CK was studied by grey relational analysis method. RESULTS: The correlation between each common peak and CK had gradually increased with the dose increased from 3.73 g/kg to 33.75 g/kg, but when the dose reached to 45 g/kg, the correlation between each common peak and CK had decreased. The variation trends of correlation of spectrum-effect relationship for different dose were similar,but the correlation variation trend of the efficacy on the No. 8 peak in 33.75 g/kg group with the other four groups in the opposite, the change trends of the No. 11 peak in 22.5 g/kg group, the No. 24 peak in 33. 75 g/kg group and the No. 37 peak in 45 g/ kg group with 3.75 g/kg group and 11.25 g/kg group on the contrary. The relational orders of spectrum-effect relationship were not consistent, respectively( the first 15 peaks) :11 > 37 > 24 > 30 > 8 > 21 > 2 > 16 > 1 > 3 > 20 > 15 > 12 > 19 > 7;11 > 37 > 30 > 8 > 21 > 24 > 2 > 1 > 16 > 3 > 27 > 12 > 22 > 20 >10; 8 > 30 > 1 > 2 > 21 > 27 > 31 > 22 > 16 > 12 > 3 > 10 > 9 > 20 > 4; 1 > 2 > 27 > 21 > 31 > 22 > 12 > 16 > 9 > 3 > 10 > 4 > 17 > 30 > 20; 8 > 30 > 1 > 2 > 2 > 2 > 7 > 31 > 22 > 16 > 12 > 3 > 9 > 10 > 20 > 17. CONCLUSION: Anti-myocardial ischemia effect of GualouXiebai dropping pills comes from the synergistic or antagonistic effect among various active ingredients related to the dose. With the difference of the dosage, the relational orders of chemical components to play the role is not the same, but the main components to play a pharmacodynamic of five dose groups are consistent,the existence of the component groups lay a foundation for further study of GualouXiebai dropping pills.

[Influence of the Compatibility of Gualou and Xiebai on Pharmacokinetics of Quercetin in Gualou].

OBJECTIVE: To determine the concentration of quercetin in mouse plasma by HPLC-DAD in different time after oral administration with Gualou Xiebai decoction and Gualou decoction, and to explore the influence of the compatibility of Gualou and Xiebai on pharmacokinetics of quercetin in Gualou. METHODS: The plasma sample was simply deproteinized with acetonitrile, extracted three times by aceticether, evaporated to dryness with a gentle N2 stream, dissolved with methanol and determined by HPLC. The pharmacokinetic parameters were separately calculated and compared with data processing software DAS 2. 0 and SPSS 11. 5 software. RESULTS: After oral administration with Gualou Xiebai decoction and Gualou decoction, the pharmacokinetics of quercetin in mouse plasma accorded with the two-compartment model, and t1/2α, t1/2ß, AUC0-t, AUC0-∞, Cmax, tmax, Ka, K10, K12, K21, CL/F and V1/F all had significant differences(P <0. 05 or P <0. 01). CONCLUSION: Xiebai in Gualou Xiebai decoction has a remarkable effect on pharmacokinetic character of quercetin in Gualou, it can promote the absorption and distribution and reduce the elimination of quercetin, in vivo, as well as increase the bioavailability of quercetin.

[Chromatographic fingerprint peak matching of Semen Cassiae by asymptotic window orthoganality projection analytical method].

OBJECTIVE: To investigate the principle and basis about the chromatographic fingerprint peak matching of Semen Cassiae by the asymptotic window orthogonal projection analytical method. METHOD: The samples of Semen Cassiae were hydrolyzed in the 1.5 mol x L(-1) hydrochloride acid and then reflux extracted with chloroform. The chromatographic condition was that the HPLC was run on Agilent 1100 column, Zorbax Eclipse XDB-C18 (4.6 mm x 250 mm, 5 microm). The protected column was Scienhome C18 column (3.0 mm x 20 mm, 5 microm) with acetonitrile-water (0.1% phosphoric acid) as mobile phase in gradient elution at a flow rate of 1.0 mL x min(-1). The detection wavelength and reference wavelength were 278 nm and 550 nm, respectively. RESULT: The characteristic fingerprint collection of illustrative plates of Semen Cassiae were obtained with different HPLC instruments and chromatographic columns. And the asymptotic window orthogonal projection analytical method was used to decide the chromatographic fingerprint peak matching of Semen Cassiae, promptly and prapidly. CONCLUSION: The asymptotic window orthogonal projection analytical method can progress the matching of different fingerprint collection of illustrative plates peak matching well and truly.