Method for analyzing association between bletilla ochracea and diseases based on UHPLC-Q-Exactive Plus Orbitrap-HRMS in combination with network pharmacology

Abstract

The invention relates to the technical field of traditional Chinese medicine medicinal materials, and discloses a method for analyzing correlation between bletilla ochracea and diseases on the basis of UHPLC-Q-Exactive Plus Orbitrap-HRMS (Ultra High Performance Liquid Chromatography-Q-Exactive Plus Orbitrap-High Resolution Mass Spectrometry) and network pharmacology, and the method comprises the following steps: S1, analyzing and identifying medicinal components in bletilla ochracea on the basis of UHPLC-Q-Exactive Plus Orbitrap-HRMS; s2, constructing an active component data set by using the bletilla ochracea and the active component targets; s3, finding out common targets between the bletilla ochracea and the active component-disease by taking the intersection of the two; s4, determining core target screening conditions and performing screening analysis; and S5, constructing a visual graph of'active ingredient-target spot-pathway-disease '. According to the method disclosed by the invention, the medicinal components of the bletilla ochracea are analyzed through UHPLC-Q-Exactive Plus Orbitrap-HRMS (Ultra High Performance Liquid Chromatography-Q-Exactive Plus Orbitrap-High Resolution Mass Spectrometry), and common target analysis of the medicinal components and the diseases is carried out in combination with network pharmacology, so that the association rule between the bletilla ochracea and the diseases is obtained.

Description

Technical Field

[0001] The present invention relates to the technical field of traditional Chinese medicine materials, and specifically relates to a method for analyzing the association between Bletilla ochracea and diseases based on UHPLC-Q-Exactive Plus Orbitrap-HRMS combined with network pharmacology. Background Technique

[0002] Bletilla ochracea is the dried tuber of the orchid plant Bletilla ochracea Schltr., which has the effects of moistening the lungs and relieving cough, arresting bleeding with astringents, and promoting tissue regeneration and subsiding swelling. It is widely used in the clinical treatment of gastric ulcers. Pharmacological studies have found that the water extract of Bletilla ochracea can reduce ethanol-induced gastric damage. At the inflammatory level, the extract of Bletilla ochracea can reduce the release of TNF-α, IL-6, IL-1β and ET-1 and the protein expression of COX-2 and NF-κB p65, and at the same time has immunomodulatory and anti-tumor activities. However, there is currently no research on the analysis of the active ingredients of Bletilla ochracea and the analysis of the common targets of the active ingredients and diseases in combination with network pharmacology to clarify the association law between Bletilla ochracea and diseases.

[0003] Therefore, there is an urgent need to provide a method for analyzing the association between Bletilla ochracea and diseases based on UHPLC-Q-Exactive Plus Orbitrap-HRMS combined with network pharmacology, and then analyze the association law among the active ingredients-disease targets-gastric ulcer of Bletilla ochracea, so as to provide a reference for clarifying the mechanism of action of this medicine on gastric ulcer, clinical rational drug use and new drug research and development. Summary of the Invention

[0004] The present invention aims to provide a method for analyzing the association between Bletilla ochracea and diseases based on UHPLC-Q-Exactive Plus Orbitrap-HRMS combined with network pharmacology. By analyzing and identifying the active ingredients of Bletilla ochracea by UHPLC-Q-Exactive Plus Orbitrap-HRMS and combining network pharmacology to analyze the common targets of the active ingredients and diseases, the association law between Bletilla ochracea and diseases can be obtained.

[0005] To achieve the above object, the present invention adopts the following technical scheme: A method for analyzing the association between Bletilla ochracea and diseases based on UHPLC-Q-Exactive Plus Orbitrap-HRMS combined with network pharmacology, including the following steps:

[0006] Step S1: Analyze and identify the active ingredients in Bletilla ochracea based on UHPLC-Q-Exactive Plus Orbitrap-HRMS, and identify 14 active ingredients in Bletilla ochracea;

[0007] Step S2: Import the 14 pharmacodynamic components of Bletilla ochracea in Step S1 into TCMIP, TCMSP and PharmMapper respectively. Meanwhile, in combination with relevant documents, collect the corresponding targets of the active components of Bletilla ochracea, delete the duplicate targets of the active components of Bletilla ochracea, and construct an active component dataset with the targets of the active components of Bletilla ochracea;

[0008] Step S3: In the Genecard database, retrieve the target information related to GU by using "Gastric ulcer" as the keyword, and then perform intersection mapping with the targets of the active components of Bletilla ochracea obtained in Step S2 and the GU targets, and take the intersection of the two to find the common potential targets between the active components of Bletilla ochracea and the disease;

[0009] Step S4: Import the common potential targets between the active components of Bletilla ochracea and the disease in Step S3 into the String database to construct a PPI network, use Cytoscape software to analyze the topological structure characteristic parameters of the network nodes, determine the core target screening conditions and conduct subsequent screening and analysis;

[0010] Step S5: Import the core targets obtained from the PPI network analysis in Step S4 into the David database, select "Homo sapiens" for the species, conduct GO analysis and KEGG enrichment analysis on the core targets, output the corresponding charts, and construct a visualization graph of "active component - target - pathway - disease".

[0011] Furthermore, in Step S1, the analysis and identification of the pharmacodynamic components in Bletilla ochracea include the preparation of the test solution, mass spectrometry analysis, and data processing. Among them, the preparation of the test solution includes the following steps:

[0012] (1) Take 10 g of Bletilla ochracea, add 100 ml of distilled water, soak for 30 min, and obtain the soaked Bletilla ochracea;

[0013] (2) Boil the soaked Bletilla ochracea over high heat and then simmer for 30 min, take out the decoction of Bletilla ochracea and set it aside;

[0014] (3) Take the residue of Bletilla ochracea, boil it over high heat and then simmer for 15 min, combine the decoctions, take 20 mL, evaporate to dryness in an evaporating dish, add 20 mL of methanol, stir to dissolve, shake well, filter, precisely absorb 1 mL of the filtrate into a 10 mL volumetric flask, add methanol to volume to the scale, filter with a 0.22 μm microporous filter membrane, and take the subsequent filtrate.

[0015] Furthermore, the chromatographic conditions for the chromatographic analysis are as follows: Chromatographic column: Hypersil GOLD C 18(100mm×2.1mm, 1.9um); mobile phase A was 0.1% formic acid in acetonitrile, and mobile phase B was 0.1% formic acid in water. The elution gradient was: 0-1min, 3% A; 1-3min, 3%→8% A; 3-4min, 8%→20% A; 4-7min, 20%→35% A; 7-10min, 35%→45% A; 10-12min, 45%→75% A; 12-13min, 75%→97% A; 13-15min, 97%→97% A; 15-16min, 97%→3% A; 16-18min, 3% A; column temperature 45°C; flow rate 0.30mL / min; injection volume 1μL.

[0016] Furthermore, the mass spectrometry conditions are as follows: HESI ion source was used to collect data in positive and negative ion modes, with positive and negative ion voltages of 2.5 kV and 3.5 kV, respectively; collision energies of 20, 40, and 60 eV; sheath gas flow rate of 30 Arb; auxiliary gas flow rate of 10 Arb; capillary temperature of 320°C; scan range of m / z 100-1500; and scan mode of Full MS / ddMS. 2 Full MS primary mass spectrometry resolution: 70,000; secondary mass spectrometry resolution: 17,500; probe heater temperature: 350°C.

[0017] Furthermore, the data processing is as follows: the mass spectrometry data are processed using Thermo Xcalibur4.4 and Compound TM Discoverer 3.2, the raw data collected by the mass spectrometry are imported into Compound Discoverer 3.2 software for preliminary calculation, and the main chromatographic peaks are qualitatively analyzed according to the retention time, accurate molecular weight and characteristic fragment ion information of the corresponding secondary mass spectrum of each main chromatographic peak of the test solution.

[0018] Furthermore, 14 compounds of Rhizoma Coptidis were identified, including 9 glycosides, 3 phenanthrenes and 2 organic acid compounds.

[0019] Compared with the prior art, the present invention also has the following technical effects:

[0020] (1) Based on the UHPLC-Q-Exactive Plus Orbitrap-HRMS technology, 14 compounds of Bletilla ochracea Schltr. were identified in this invention. Combining with network pharmacology analysis, a visualized graph of "active ingredient-target-pathway-disease" was constructed for the 14 compounds of Bletilla ochracea Schltr., revealing how the active ingredients of Bletilla ochracea Schltr. exert their pharmacodynamic effects through the combined regulation of "multiple targets, multiple pathways, and multiple ways". Results: The protein targets were ranked from large to small according to the degree value. The top 5 core targets with the highest degree values were TNF (degree value 37), IL6 (degree value 35), AKT1 (degree value 35), CASP3 (degree value 34), and MMP9 (degree value 32). The higher the degree value of these targets, the more critical the target is in the treatment of gastric ulcer. Therefore, the method of this invention provides a new idea for elucidating the mechanism of action of Bletilla ochracea Schltr. in treating gastric ulcer, and these systems biology information will be verified in future research, further providing a preliminary research basis for the prevention and treatment of gastric ulcer with Bletilla ochracea Schltr.

[0021] (2) Based on the UHPLC-Q-Exactive Plus Orbitrap-HRMS technology combined with network pharmacology method, the research method for the diseases affected by the active pharmacodynamic ingredients of Bletilla ochracea Schltr. is networked and systematic. Compared with the traditional research method, the R & D scope is narrowed and the R & D purpose is made more precise, thus saving the drug R & D cost. At the same time, the efficiency of drug screening and prediction is greatly improved, which provides a new reference for the R & D of active ingredients of other Chinese medicinal materials and has a breakthrough significance for the development and industrialization of China's rich traditional Chinese medicine resources. Description of the Drawings

[0022] Figure 1 The total ion current chromatograms of the test solution of Bletilla ochracea Schltr. under the positive ion mode (A) and negative ion mode (B) of this invention (A: the total ion current chromatogram of Bletilla ochracea Schltr. under the positive ion mode; B: the total ion current chromatogram of Bletilla ochracea Schltr. under the negative ion mode);

[0023] Figure 2 The pie chart of the proportion of the identification of the pharmacodynamic ingredients of Bletilla ochracea Schltr. based on the UHPLC-Q-Exactive Plus Orbitrap-HRMS technology of this invention;

[0024] Figure 3 The Venn diagram of Bletilla ochracea Schltr. and GU targets of this invention;

[0025] Figure 4 The PPI network diagram of the core targets of Bletilla ochracea Schltr. of this invention;

[0026] Figure 5 The bubble chart of the KEGG enrichment analysis of Bletilla ochracea Schltr. of this invention. Detailed implementation mode

[0027] The following is a further detailed description through specific implementation modes:

[0028] This embodiment: A method for analyzing the association between Bletilla ochracea and diseases based on UHPLC-Q-Exactive Plus Orbitrap-HRMS combined with network pharmacology, includes the following steps:

[0029] 1.1 Instruments and reagents:

[0030] 1.1.1 Instruments:

[0031] Ultra-high performance liquid chromatograph (Thermo Scientific Vanquish UHPLC; Thermo Fisher Scientific, USA); High-resolution mass spectrometer (Q-Exactive-Plus; Thermo Fisher Scientific, USA)

[0032] 1.1.2 Reagents:

[0033] Acetonitrile was purchased from Merck KGaA, batch number 1.06035.2508; Formic acid was purchased from Thermo Fisher Scientific (China) Co., Ltd., batch number A117-50

[0034] 2.2 UHPLC-Q-Exactive Plus Orbitrap-HRMS test

[0035] 2.2.1 Preparation of test solution:

[0036] (1) Take 10 g of Bletilla ochracea, add 100 ml of distilled water, soak for 30 min, to obtain the soaked Bletilla ochracea;

[0037] (2) Boil the soaked Bletilla ochracea over high heat and then simmer for 30 min, take the decoction of Bletilla ochracea and place it separately;

[0038] (3) Take the residue of Bletilla ochracea, boil it over high heat and then simmer for 15 min, combine the decoctions, take 20 mL, evaporate to dryness in an evaporating dish, add 20 mL of methanol, stir to dissolve, shake well, filter, accurately absorb 1 mL of the filtrate and place it in a 10 mL volumetric flask, add methanol to volume to the scale, filter with a 0.22 μm microporous filter membrane, and take the subsequent filtrate.

[0039] 2.2.2 Chromatographic analysis:

[0040] (1) Chromatographic conditions: Chromatographic column: Hypersil GOLD C 18(100 mm × 2.1 mm, 1.9 um); Mobile phase A is 0.1% formic acid acetonitrile, B is 0.1% formic acid aqueous solution, and the elution gradient is 0 - 1 min, 3% A; 1 - 3 min, 3% → 8% A; 3 - 4 min, 8% → 20% A; 4 - 7 min, 20% → 35% A; 7 - 10 min, 35% → 45% A; 10 - 12 min, 45% → 75% A; 12 - 13 min, 75% → 97% A; 13 - 15 min, 97% → 97% A; 15 - 16 min, 97% → 3% A; 16 - 18 min, 3% A; Column temperature is 45°C; Flow rate is 0.30 mL / min; Injection volume is 1 μL.

[0041] (2) Mass spectrometry conditions: HESI ion source collects data in positive and negative ion modes, with positive and negative ion voltages of 2.5 kV and 3.5 kV; Collision energies are 20, 40, 60 eV; Sheath gas flow rate is 30 Arb; Auxiliary gas flow rate is 10 Arb; Capillary temperature is 320°C; Scanning range is m / z 100 - 1500; Scanning mode is Full MS / ddMS 2 ; Full MS first - level mass spectrometry resolution is 70000; Second - level mass spectrometry resolution is 17500; Probe heater temperature is 350°C.

[0042] 2.2.3 Data processing:

[0043] Mass spectrometry data is processed using Thermo Xcalibur 4.4 and Compound TM TMDiscoverer 3.2. The original data collected by mass spectrometry is imported into the Compoud Discoverer 3.2 software for preliminary calculation. According to the retention time, exact molecular weight of each main chromatographic peak in the test solution, and the characteristic fragment ion information of the corresponding secondary mass spectrometry, qualitative analysis is performed on each main chromatographic peak.

[0044] 3.3 Network pharmacology analysis:

[0045] 3.3.1 Collection of active components and component targets of Bletilla ochracea Schltr.:

[0046] The pharmacodynamic components of Bletilla ochracea Schltr. are respectively imported into TCMIP (http: / / www.tcmip.cn / TCMIP / index.php / Home / Login / login.html), TCMSP (https: / / tcmsp - e.com / tcmsp.php) and PharmMapper. At the same time, combined with relevant documents, the active components of Bletilla ochracea Schltr. and the corresponding targets of the active components are collected. After deleting the common targets of the active components of Bletilla ochracea Schltr., an active component dataset is constructed with the targets of the active components of Bletilla ochracea Schltr.

[0047] 3.3.2 Collection of GU-related targets and common targets:

[0048] In the Genecard database (https: / / genecards.org / ), the keyword "Gastric ulcer" was used to retrieve the target information related to GU. Then, the intersection mapping was performed between the targets of the active components of Bletilla ochracea and the targets of GU, and the intersection of the two was taken to find the common potential targets between the active components of Bletilla ochracea and the disease.

[0049] 3.3.3 PPI network and core targets:

[0050] The common potential targets between the active components of Bletilla ochracea and the disease were imported into the String database to construct a PPI network. Cytoscape 3.6.1 was used to perform network topology analysis and visualization on the "tsv" file obtained from the PPI network, and the software was used to analyze the topological structure characteristic parameters of the network nodes to determine the core target screening conditions and conduct subsequent screening and analysis.

[0051] 3.3.4 GO analysis and KEGG analysis

[0052] The core targets obtained from the PPI network analysis were imported into the David database, and the species was selected as "Homo sapiens". GO analysis and KEGG enrichment analysis were performed on the core targets, and the corresponding charts were output to construct a visualization graph of "active ingredient - target - pathway - disease".

[0053] 3.4 Results and discussion

[0054] See the attached Figure 1 The total ion current chromatograms of the test solution of Bletilla ochracea under the positive ion mode (A) and negative ion mode (B) shown: According to the obtained accurate molecular weight and corresponding secondary ion fragments, the cleavage rules were analyzed, and a total of 14 compounds in Bletilla ochracea were identified.

[0055] Combined with Table 1 and Table 2 shown: Among the 14 compounds in Bletilla ochracea, there are 9 glycoside compounds, 3 phenanthrene compounds and 2 organic acid compounds. The specific proportions are shown in Figure 2 shown, indicating that the above compounds jointly exerted the anti-gastric ulcer pharmacological effect of Bletilla striata.

[0056]

[0057] Table 1 Table 2 Chemical characterization of B.ochracea Schltr by UHPLC-Q-exactive-MS / MS in positive ion

[0058]

[0059] Table 2

[0060] Based on UHPLC-Q-Exactive Plus Orbitrap-HRMS technology, 14 compounds were identified from Bletilla ochracea, and 8 active components were screened out, including Dactylorhin A, Gymnoside I, Gastrodin, 4,7-Dihydroxy-2-methoxy-9,10-dihydrophenanthrene, Gymnoside IX, α-Isobutylmalic acid, 4,7-Dihydroxy-1-p-hydoxybenzyl-2-methoxy-9,10-dihydophenanthrene and Militarine. These compounds all have good biological activities. By querying TCMSP, TCMIP, PharmMapper databases and related literature, 290 potential action targets of Bletilla ochracea were obtained after deleting duplicate targets, as shown in Table 3 for details.

[0061]

[0062] Table 3

[0063] See Figure 3 As shown, “Gastric ulcer” was input into the GeneCard database to search for GU-related targets, and targets with “Relevance score” greater than 10 were screened out, obtaining a total of 586 GU targets. The GU targets and the targets corresponding to the components of Bletilla ochracea were imported into Venny 2.1.0, and 44 targets for Bletilla ochracea to treat GU were obtained.

[0064] The 44 common targets were imported into the String database to construct a PPI network to explore the interaction relationships between the targets. Then, the “tsv” file obtained from String was imported into Cytoscape 3.6.1 for network visualization, and the “csv” file was exported. Targets with a value greater than the median of Degree were selected, and finally 21 targets were obtained for visualization, as shown in Figure 4The protein targets were arranged from largest to smallest according to the degree value. The top 5 core targets with the highest degree values were TNF (degree value 37), IL6 (degree value 35), AKT1 (degree value 35), CASP3 (degree value 34), and MMP9 (degree value 32). The higher the degree value of these targets, the more crucial the target is in the treatment of gastric ulcer. The results are shown in Table 4.

[0065] Name Degree Betweenness Centrality Closeness Centrality TNF 37 0.0735 0.8776 IL6 35 0.0446 0.8431 AKT1 35 0.0460 0.8431 CASP3 34 0.0453 0.8269 MMP9 32 0.0194 0.7963 IL1B 32 0.0318 0.7963 PTGS2 31 0.0205 0.7818 ESR1 31 0.0406 0.7818 EGF 30 0.0218 0.7544 SRC 28 0.0161 0.7288 PPARG 28 0.0106 0.7414 IL2 27 0.0124 0.7288 IGF1 27 0.0206 0.7288 RHOA 26 0.0206 0.7167 HRAS 25 0.0187 0.7049 MAPK14 24 0.0105 0.6935 PLAU 23 0.0140 0.6719 IGF1R 22 0.0070 0.6615 HGF 22 0.0084 0.6615 JAK2 22 0.0063 0.6615 STAT1 21 0.0022 0.6515

[0066] Table 4

[0067] The results of GO analysis showed that there were 259 biological processes, 21 cellular components, and 32 molecular functions involved in the anti-GU effect of Bletilla ochracea. Taking PValue < 0.01, 119 biological processes, 12 cellular components, and 12 molecular functions were obtained. According to the enrichment results, the biological processes were related to the positive regulation of cell migration, the positive regulation of smooth muscle cell proliferation, the positive regulation of phosphatidylinositol 3-kinase signaling, and the positive regulation of the MAPK cascade. The cellular components were related to glutamatergic synapses, foreign components on the cytoplasmic side of the plasma membrane, caveolae, extracellular regions, extracellular spaces, etc. The molecular functions were related to enzyme binding, growth factor activity, identical protein binding, and insulin receptor binding. The results of KEGG enrichment analysis showed that there were 130 pathways involved in the treatment of GU by Bletilla ochracea. These pathways included the IL-17 signaling pathway, TNF signaling pathway, MAPK signaling pathway, and Rap1 signaling pathway, etc. See specifically Figure 5 , and most of these pathways were related to targets such as IL-6, TNF, CASP3, and IL-1β.

[0068] The above are only examples of the present invention. Specific technical solutions and / or common knowledge such as characteristics well known in the art are not described in detail herein. It should be noted that for those skilled in the art, without departing from the technical solution of the present invention, several modifications and improvements can be made, which should also be regarded as the protection scope of the present invention, and these will not affect the implementation effect of the present invention and the practicality of the patent. The protection scope required by this application should be based on the content of its claims, and the specific implementation manners described in the specification can be used to interpret the content of the claims.

Claims

1. A method for analyzing the association between Rhizoma Coptidis and diseases based on UHPLC-Q-Exactive Plus Orbitrap-HRMS combined with network pharmacology, characterized in that: The following steps are involved: Step S1: Analyze and identify the active ingredients in Rhizoma Bletillae Sinensis using UHPLC-Q-Exactive Plus Orbitrap-HRMS, and identify 14 active ingredients in Rhizoma Bletillae Sinensis Sinensis. Step S2: Import the 14 active ingredients of Huanghuabai in step S1 into TCMIP, TCMSP and PharmMapper respectively. At the same time, combine the relevant files to collect the targets corresponding to Huanghuabai and the active ingredients, delete the duplicate targets of Huanghuabai and the active ingredients, and construct the active ingredient dataset using the targets of Huanghuabai and the active ingredients. Step S3: In the Genecard database, "Gastric ulcer" was used as a keyword to retrieve target information related to GU. This information was then intersected with the targets of R. chinensis and its active ingredients obtained in step S2 and the GU targets. The intersection of the two was used to identify potential common targets between R. chinensis and its active ingredients and diseases. Step S4: Import the common potential targets between Huanghuabai and its active ingredients and diseases in step S3 into the String database to construct a PPT network. Use Cytoscape software to analyze the topological structural characteristic parameters of the network nodes, determine the core target screening conditions and conduct subsequent screening analysis. Step S5: Import the core targets obtained from the PPI network analysis in step S4 into the David database, select "Homosapiens" as the species, perform GO analysis and KEGG enrichment analysis on the core targets, output the corresponding charts, and construct a "active ingredient-target-pathway-disease" visualization graph.

2. The method for analyzing the association between Rhizoma Coptidis and diseases based on UHPLC-Q-Exactive Plus Orbitrap-HRMS combined with network pharmacology according to claim 1, characterized in that: In step S1, the analysis and identification of the medicinal ingredients in Rhizoma Coptidis includes preparing a test solution, mass spectrometry analysis, and data processing, wherein the preparation of the test solution includes the following steps: (1) Take 10 g of yellow flower and white flower, add 100 mL of distilled water, and soak for 30 min to obtain the soaked yellow flower and white flower; (2) Boil the soaked chrysanthemum root with high heat and then simmer for 30 minutes over low heat. Set aside the chrysanthemum root decoction; (3) Take the white of the yellow flower and the filter residue, boil them over high heat, then simmer for 15 minutes over low heat. Combine the decoction and take 20 mL. Evaporate to dryness in an evaporating dish, add 20 mL of methanol, stir to dissolve, shake well, filter, accurately pipette 1 mL of the filtrate into a 10 mL volumetric flask, add methanol to make up to the mark, filter with a 0.22 μm microporous filter membrane, and take the filtrate.

3. The method for analyzing the association between Rhizoma Coptidis and diseases based on UHPLC-Q-Exactive Plus Orbitrap-HRMS combined with network pharmacology according to claim 2, characterized in that: The chromatographic analysis conditions are as follows: Chromatographic column: HypersilGOLD C 18 (100mm×2.1mm, 1.9um); mobile phase A was 0.1% formic acid in acetonitrile, and mobile phase B was 0.1% formic acid in water. The elution gradient was: 0-1min, 3% A; 1-3min, 3%→8% A; 3-4min, 8%→20% A; 4-7min, 20%→35% A; 7-10min, 35%→45% A; 10-12min, 45%→75% A; 12-13min, 75%→97% A; 13-15min, 97%→97% A; 15-16min, 97%→3% A; 16-18min, 3% A; column temperature 45°C; flow rate 0.30mL / min; injection volume 1μL.

4. The method for analyzing the association between Rhizoma Coptidis and diseases based on UHPLC-Q-Exactive Plus Orbitrap-HRMS combined with network pharmacology according to claim 2, characterized in that: The mass spectrometry conditions were as follows: HESI ion source was used to collect data in positive and negative ion modes, with positive and negative ion voltages of 2.5 kV and 3.5 kV, respectively; collision energies of 20, 40, and 60 eV; sheath gas flow rate of 30 Arb; auxiliary gas flow rate of 10 Arb; capillary temperature of 320°C; scan range of m / z 100-1500; and scan mode of Full MS / ddMS. 2 ; FullMS primary mass spectrometer resolution 70000; secondary mass spectrometer resolution 17500; probe heater temperature 350℃.

5. The method for analyzing the association between Rhizoma Coptidis and diseases based on UHPLC-Q-Exactive Plus Orbitrap-HRMS combined with network pharmacology according to claim 2, characterized in that: The data processing is as follows: mass spectrometry data are processed using ThermoXcalibur 4.4 and Compound TM Discoverer 3.2, the raw data collected by the mass spectrometry are imported into Compound Discoverer 3.2 software for preliminary calculation, and the main chromatographic peaks are qualitatively analyzed based on the retention time, accurate molecular weight and corresponding characteristic fragment ion information of the secondary mass spectrum of each main chromatographic peak of the test solution.

6. The method for analyzing the association of Rhizoma Coptidis and its diseases based on UHPLC-Q-Exactive Plus Orbitrap-HRMS combined with network pharmacology according to claim 1, characterized in that: Fourteen compounds were identified in Rhizoma Coptidis, including 9 glycosides, 3 phenanthrenes and 2 organic acids.

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