Quality control method of honeysuckle and radix scutellariae buccal dripping pills based on fingerprint spectrum

Abstract

The invention relates to the technical field of medicine quality control, and particularly discloses a quality control method of honeysuckle and radix scutellariae buccal dripping pills based on a fingerprint spectrum, which comprises three steps of preparation of a test solution, preparation of a reference solution and chromatographic detection. On the basis that only chlorogenic acid and baicalin are detected originally, cryptochlorogenic acid peaks, neochlorogenic acid peaks and baicalein peaks are also included in the quality control standard of the honeysuckle and radix scutellariae buccal dripping pills, so that the problem that in the prior art, the quality control of different batches of honeysuckle and radix scutellariae buccal dripping pills takes the content of chlorogenic acid and baicalin as the current quality standard; and quality control cannot be carried out on products with the same content of chlorogenic acid and baicalin and different curative effects.

Description

Technical Field

[0001] The invention relates to the field of drug quality control, and in particular to a quality control method for Yinhuang Hanhua Dropping Pills based on fingerprints. Background Art

[0002] Yinhuang Hanhua Drops are a modern compound preparation refined from the extracts of honeysuckle and scutellaria. This preparation has the effects of clearing away heat, detoxifying and relieving inflammation. It is commonly used clinically to treat upper respiratory tract infections, acute and chronic tonsillitis, pharyngitis and other exogenous diseases.

[0003] The fingerprint of traditional Chinese medicine is established on the basis of the systematic research on the components of traditional Chinese medicine. It adopts certain analytical methods and analytical instruments to obtain the chromatogram or spectrum that can indicate its chemical characteristics. Through the specificity of the fingerprint, it can effectively identify its quality.

[0004] At present, in the production of Yinhuang Containing Drops, it is necessary to control the fluctuation of the quality of different batches of Yinhuang Containing Drops to maintain the stability of the quality of the products shipped. In the prior art, enterprises use the determination of the content of chlorogenic acid and baicalin as a quality control method, and screen out the same batch of products with chlorogenic acid and baicalin content above the standard as good products, and the same batch of products with chlorogenic acid and baicalin content below the standard as defective products. Although this method can complete the simple judgment of the quality of different batches of Yinhuang Containing Drops, the quality control index is relatively single and only focuses on the main ingredients of the drug. In actual use, it is found that the efficacy of Yinhuang Containing Drops with the same content of chlorogenic acid and baicalin is different, and this difference cannot be controlled by the quality control method of the prior art.

[0005] Regarding the technical problem that chlorogenic acid and baicalin have the same content but different therapeutic effects in the prior art, technicians usually believe that it is caused by the different quality and origin of the raw medicinal materials. Therefore, they control the raw materials and strictly screen and test the raw medicinal materials coming to the factory to ensure the quality and origin of the raw materials, and then ensure the quality of the final product, Yinhuang Containing Drops. However, this method has great limitations. First, this method can only avoid the impact of defective medicinal materials, and cannot control the impact of process fluctuations on Yinhuang Containing Drops; second, the origin and appearance quality of the medicinal materials depend on the experience of the technicians, and the measurability is poor. Therefore, there is a need for a quality control method with high measurability that can directly control the production of Yinhuang Containing Drops. Summary of the invention

[0006] The object of the present invention is to provide a quality control method for Yinhuang oral dropping pills based on fingerprint, so as to solve the technical problem that the quality control of different batches of Yinhuang oral dropping pills in the prior art is to determine the content of chlorogenic acid and baicalin as the current quality standard, and it is impossible to perform quality control on products with the same content of chlorogenic acid and baicalin but different therapeutic effects.

[0007] In order to solve the above problems, the technical solution adopted by the present invention is as follows: a quality control method of Yinhuang Containing Dropping Pills based on fingerprint spectrum, wherein the Yinhuang Containing Dropping Pills are Chinese medicine dropping pills prepared from honeysuckle extract and scutellaria baicalensis extract, comprising the following steps:

[0008] Step 1: Prepare the test solution: grind the extracted Yinhuang Hanhua Dropping Pills sample into powder, and add it into 10% to 90% methanol to prepare the test solution;

[0009] Step 2: Prepare reference solution: weigh appropriate amounts of cryptochlorogenic acid, neochlorogenic acid, chlorogenic acid, baicalin, and baicalein reference substances, add 10% to 90% by volume of methanol to prepare reference solutions with concentrations of cryptochlorogenic acid, neochlorogenic acid, chlorogenic acid, baicalin, and baicalein of 1 to 600 μg / ml, respectively;

[0010] Step 3: Chromatographic detection: The test solution and the reference solution are subjected to content detection and characteristic spectrum construction by HPLC or UPLC, and the determination results are compared with the detection index;

[0011] The chromatographic conditions include: the chromatographic column is filled with octadecylsilane bonded silica gel, the mobile phase A is 0.4% phosphoric acid aqueous solution, the mobile phase B is acetonitrile, and the gradient elution is: 0-25min, 10% B; 25-65min, 10%-24% B; 65-85min 24%-33% B; 50-105min, 33%-40% B; 105-130min, 40%-60% B.

[0012] The beneficial effects of this embodiment are:

[0013] 1. In the prior art, the quality control of Yinhuang Hanhua Dropping Pills is carried out by measuring the content of chlorogenic acid and baicalin, and the products of the same batch with chlorogenic acid and baicalin content above the standard are selected as good products, and the products of the same batch with chlorogenic acid and baicalin content below the standard are selected as defective products. However, in actual use, it is found that the efficacy of Yinhuang Hanhua Dropping Pills with the same chlorogenic acid and baicalin content is different, and the efficacy of the produced Yinhuang Hanhua Dropping Pills cannot be guaranteed to be stable only by the content of chlorogenic acid and baicalin. During the research and development process of this application, the technical personnel believed that the quality control method of the prior art paid too much attention to the main ingredients of the drug, while the theory of traditional Chinese medicine emphasizes the compatibility principle of monarch, minister, adjuvant and envoy. Only focusing on the main ingredients of the drug cannot reflect the influence of the compound compatibility environment on the efficacy during the treatment of traditional Chinese medicine, and is not enough to reflect the "multi-target" and "multi-path" action characteristics of traditional Chinese medicine compound preparations. Therefore, the content of chlorogenic acid and baicalin in Yinhuang Hanhua Drop Pills is the same, but the efficacy is not stable. This application establishes Yinhuang Hanhua Drop Pills H The PLC fingerprint spectrum is combined with chemical pattern recognition to perform multi-dimensional comprehensive statistical analysis, and network pharmacology analysis is used to simulate the compounding environment of Yinhuang Oral Drops. Finally, the cryptochlorogenic acid peak, neochlorogenic acid peak and baicalin peak are also included in the quality control standard of Yinhuang Oral Drops. Therefore, compared with the prior art, the quality control method of the present application controls the production process of Yinhuang Oral Drops more closely to the "multi-target" and "multi-path" action characteristics of Yinhuang Oral Drops in the actual treatment process. Therefore, it is more consistent with the efficacy and the quality control is more precise.

[0014] 2. Since the present application needs to include the cryptochlorogenic acid peak, the neochlorogenic acid peak and the baicalin peak into the quality control standard, the chromatographic conditions of HPLC need to be adjusted to ensure that the cryptochlorogenic acid peak, the neochlorogenic acid peak and the baicalin peak can be well separated. The present application adjusts the test solution and the method for preparing the reference solution, adjusts the mobile phase A to 0.4% phosphoric acid aqueous solution, the mobile phase B to acetonitrile, and adjusts the gradient elution process according to the mobile phase. The results are ( Figure 1 ) proved that the chromatographic conditions of the present application can well separate cryptochlorogenic acid, neochlorogenic acid, chlorogenic acid, baicalin and baicalein during multiple content detection processes, and the separation is stable and the measurability is high.

[0015] Furthermore, in step 1 and step 2, the concentration of methanol is 80%.

[0016] Furthermore, in step 1, ultrasonic treatment is performed after methanol is added.

[0017] Furthermore, the chromatographic conditions also include a column temperature of 35°C.

[0018] Further, the chromatographic conditions include using 235nm ultraviolet wavelength for detection

[0019] Further, when preparing the test solution in step 1, the Yinhuang Hanhua Dropping Pills sample extracted in step 1 is ground and then passed through a No. 4 sieve. After sieving, the sample powder is accurately weighed and placed in a brown volumetric flask, methanol is accurately added, and ultrasonic treatment is performed at 53kHz for 30 minutes. After cooling to room temperature, the lost weight is supplemented with methanol, shaken well, filtered, and the filtrate is passed through a 0.22μm microporous filter membrane to obtain the test solution. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 This is the superposition diagram of the HPLC fingerprint of Yinhuang Hanhua Drop Pills of the present invention.

[0021] Figure 2 for Figure 1 Similarity table of HPLC fingerprints of Zhongyinhuang Hanhua Drop Pills,

[0022] Figure 3 for Figure 1 The HCA dendrogram was made with the 15 common peak areas of the HPLC fingerprint of Zhongyinhuang Hanhua Pills as variables.

[0023] Figure 4 The PCA diagram was made with the 15 common peak areas of the HPLC fingerprint of Yinhuang Hanhua Drop Pills as variables.

[0024] Figure 5 are the three principal component tables extracted from PCA.

[0025] Figure 6 This is the component loading matrix diagram in PCA analysis.

[0026] Figure 7 This is the permutation test plot in PCA analysis.

[0027] Figure 8 This is the OPLS-DA score graph in PCA analysis.

[0028] Fig. 9 This is the VIP value diagram in PCA analysis.

[0029] Fig.10 This is the load scatter plot in PCA analysis.

[0030] Fig.11 To obtain the protein interaction PPI network diagram,

[0031] Fig.12 is the core target table,

[0032] Fig.13 The signal pathway diagram is shown in Figure 2.

[0033] Fig.14 is a component-target-pathway network diagram,

[0034] Fig.15 Heat map of molecular docking scores. DETAILED DESCRIPTION

[0035] The following is further described in detail through specific implementation methods:

[0036] Implementation example Figure 1-15 As shown:

[0037] Example 1

[0038] A quality control method for Yinhuang Containing Pills based on fingerprints, wherein the Yinhuang Containing Pills are Chinese medicine pills prepared from honeysuckle extract and scutellaria baicalensis extract, comprising the following steps:

[0039] Step 1: Prepare the test solution: grind the extracted Yinhuang Hanhua Dropping Pills sample into powder, and add it into 10% to 90% methanol to prepare the test solution;

[0040] When preparing the test solution, the present application screened the selected solvents, wherein the screened solvents included water, 20%, 40%, 60%, and 80% methanol-water solution, and compared the ultrasonic process for different time periods (10, 20, and 30 minutes) to prepare the test solution. After solvent extraction, baicalin was used as a reference for content analysis. The results showed that the 80% methanol extract had the highest baicalin content, and the extraction was basically completed after 30 minutes of ultrasonication. Therefore, the present application ultimately adopted 80% methanol-water solution and ultrasonication for 30 minutes as the preparation method for the test solution.

[0041] During the specific preparation, the extracted Yinhuang Hanhua Drop Pills sample was ground into powder, and then passed through a No. 4 sieve. After sieving, the powder was accurately weighed to about 0.8 g, placed in a 100 ml brown volumetric flask, and 100 ml of 80% methanol was accurately added, weighed, and ultrasonically treated at 53 kHz for 30 minutes. After cooling to room temperature, the lost weight was supplemented with 80% methanol, shaken, filtered, and the filtrate was passed through a 0.22 μm microporous filter membrane to obtain the test solution.

[0042] Step 2: Prepare reference solution: weigh appropriate amounts of cryptochlorogenic acid, neochlorogenic acid, chlorogenic acid, baicalin, and baicalein reference substances, add 10% to 90% by volume of methanol to prepare reference solutions with concentrations of cryptochlorogenic acid, neochlorogenic acid, chlorogenic acid, baicalin, and baicalein of 1 to 600 μg / ml, respectively;

[0043] Accurately weigh appropriate amounts of cryptochlorogenic acid, neochlorogenic acid, chlorogenic acid, baicalin, and baicalein reference substances and add 80% methanol to prepare. The prepared reference substance solution must be stored below 10°C.

[0044] Step 3: Chromatographic detection: The test solution and the reference solution are subjected to content detection and characteristic spectrum construction by HPLC or UPLC, and the determination results are compared with the detection index;

[0045] The chromatographic conditions include: the chromatographic column uses octadecylsilane bonded silica gel as a filler, the selected model is Agilent C18 analytical column (4.6mm×250mm, 5μm), the column temperature is 35°C, and the injection volume is 10μl.

[0046] Mobile phase A is 0.4% phosphoric acid aqueous solution, and mobile phase B is acetonitrile. When selecting the mobile phase, the present application investigated mobile phase elution systems such as methanol-water, acetonitrile-water, methanol-0.4% phosphoric acid aqueous solution, and acetonitrile-0.4% phosphoric acid aqueous solution. Among them, when acetonitrile-0.4% phosphoric acid aqueous solution is used as the elution solvent, the separation effect of each chromatographic peak is the best.

[0047] Gradient elution was: 0-25 min, 10% B; 25-65 min, 10%-24% B; 65-85 min, 24%-33% B; 50-105 min, 33%-40% B; 105-130 min, 40%-60% B.

[0048] The effects of UV wavelengths (235, 280, and 326 nm) were also compared, and it was found that at 235 nm, there were more chromatographic peaks, better separation, and peak shape.

[0049] Related research involved in this application

[0050] 1. Spectrum detection and analysis

[0051] 1.1 Experimental conditions

[0052] In order to find the quality difference between batches of Yinhuang Hanhua Dropping Pills, the present application took 12 batches of Yinhuang Hanhua Dropping Pills samples and prepared test solutions and reference solutions according to the method of the embodiment of the present application and carried out HPLC experimental detection. The spectrum of sample S2 was set as the reference fingerprint spectrum (R), the time window width was 0.1, and the median method was used. In combination with multi-point correction and Marker peak matching, the characteristic spectrum of Yinhuang Hanhua Dropping Pills and the HPLC superimposed fingerprint spectrum of 12 batches of samples were generated. The superimposed fingerprint spectrum is as follows Figure 1 As shown, the results showed that the relative retention time RSD of the 15 common peaks was 0-2.26%, and the relative peak area RSD was 0-41.51%.

[0053] 1.2 Similarity Analysis

[0054] The sample S2 chromatogram was used as the reference spectrum and similarity analysis was performed by computer. Since the peak height and peak area of ​​chromatographic peak No. 11 (baicalin) were significantly different from those of other chromatographic peaks, it played a major role in the fingerprint similarity evaluation, resulting in the similarity calculation results being 1, which affected the accuracy of the similarity calculation results. Therefore, the chromatographic peak No. 11 was removed before calculating the similarity, and the fingerprint similarity table of 12 batches of particles was obtained. The fingerprint similarity table is shown in the figure below: Figure 2 As shown, it shows that there is a high degree of similarity between the batches, and also reflects that using only the chlorogenic acid and baicalin content as the quality control method of Yinhuang Containing Drops is difficult to reflect the batch differences of Yinhuang Containing Drops.

[0055] 2. Chemical recognition pattern analysis

[0056] 2.1 Hierarchical Cluster Analysis (HCA)

[0057] The 15 common peak areas of 12 batches of samples were used as variables, and the HCA dendrogram was drawn using SPSS26.0 software using intergroup linkage and squared Euclidean distance method. The HCA dendrogram is shown in the figure. Figure 3 As shown, the results show that when the square Euclidean distance is between 10, sample S4 can be clustered into Class 1 alone, and the rest of the samples can be clustered into Class 1; when the square Euclidean distance is between 5, sample S4 can be clustered into Class 1 alone, S1, S2 and S11 can be clustered into Class 1, and the rest of the samples can be clustered into Class 1; this indicates that when the production and preparation processes are the same, there is a large difference in the common peak area between S4 and the rest of the samples, while the difference in the common peak area between the Yinhuang Hanhua Dropping Pills samples except S4 is relatively small. Analysis shows that some quality differences caused by different production batches may be related to the production process and the storage of drugs.

[0058] 2.2 Principal Component Analysis (PCA)

[0059] The 15 common peak areas of 12 batches of samples were used as variables, and PCA was performed using SPSS26.0 software, and the PCA graph was drawn using SIMCA 14.1 software. Figure 4 As shown, the PCA graph shows that the remaining sample points except S4 are all within the 95% confidence interval, indicating that there are certain differences in quality between sample S4 and the other 11 batches of Yinhuang Hanhua Dropping Pills, and sample S4 should be grouped separately.

[0060] The main components were extracted with the characteristic value > 1 as the screening condition. The main components can basically reflect the main characteristics of the peak areas of 15 common peaks within a certain range. This application extracted 3 main components, and the results are as follows Figure 5 As shown in Figure 1, the sum of the contribution rates of the principal components is 87.539%. In order to reflect the correlation coefficients between each principal component and the 15 common peaks, i.e., the original variables, the component loading matrix analysis was performed on the data. The results are shown in Figure 1. Figure 6As mentioned above, the greater the absolute value of the load in the component load matrix, the greater the contribution to the principal component. Therefore, it can be seen that principal component 1 mainly reflects the information of peaks 1 (neochlorogenic acid), 2, 3 (chlorogenic acid), 4 (cryptochlorogenic acid), 5 (caffeic acid), 6, and 10; principal component 2 mainly reflects the information of peaks 11 (baicalin), 12, 13, 14 (baicalin), and 15; and principal component 3 mainly reflects the information of peaks 7 and 14 (baicalin).

[0061] 2.3 Orthogonal Partial Least Squares Discriminant Analysis (OPLS-DA)

[0062] On the basis of cluster analysis, the 15 common peak areas of 12 batches of samples were used as variables, and SIMCA14.1 software was used to perform OPLS-DA processing to further analyze the differences in the chemical composition of Yinhuang Hanhua Drop Pills from different production batches. The OPLS-DA score graph was obtained ( Figure 8 )、VIP value chart( Fig. 9 ), 200 permutation test plots ( Figure 7 ) and load scatter plots ( Fig.10 ).

[0063] The parameters of OPLS-DA analysis model stability are R2X=0.827, R2Y=0.914, Q2=0.83, all >0.5, indicating that the established model has high stability and good predictive ability.

[0064] To verify that the model is not overfitting, we randomly permuted 200 times to perform a permutation test, and obtained a permutation test graph. The permutation test graph is as follows: Figure 7 As shown in the figure, the intercept of the Q2 regression line on the Y axis is -0.615<0

[12] , which indicates that the established model does not have overfitting phenomenon and can be used to discriminate and analyze the batch differences of 12 batches of samples.

[0065] From the OPLS-DA score graph, we can see that the OPLS-DA score graph is as follows Figure 8 As shown in the figure, according to the OPLS-DA score, the 12 batches of samples can be clustered into 3 categories, among which sample S4 is clustered into 1 category, S1, S2, and S11 are clustered into 1 category, and the remaining samples are clustered into 1 category. VIP can reflect the contribution of each common peak [8]. Taking VIP>1 as the judgment standard, 5 chromatographic peaks were screened out.

[0066] VIP can reflect the contribution of each common peak, so a VIP value graph is drawn. The VIP value graph is as follows: Fig. 9As shown in the figure, with VIP>1 as the judgment standard, 5 chromatographic peaks were screened out. The identification of chemical reference substances by VIP size was peak 4 (cryptochlorogenic acid), 1 (neochlorogenic acid), 11 (baicalin), peak 2, and peak 3 (chlorogenic acid). The above components may be the main substances causing quality differences between batches of samples, and can be used as the main detection substances for quality control of Yinhuang Hanhua Drop Pills. Cryptochlorogenic acid, neochlorogenic acid, peak 2, and chlorogenic acid are derived from honeysuckle extract, and baicalin is derived from Scutellaria extract.

[0067] Fingerprint spectrum and chemical recognition mode were used to comprehensively predict and analyze Yinhuang oral dropping pills based on the "principle of compound compatibility" and "measurability and specificity". The results showed that the contents of cryptochlorogenic acid, chlorogenic acid, neochlorogenic acid and baicalin in Yinhuang oral dropping pills varied greatly among different production batches. The four compounds may be important markers leading to quality differences between batches. Chlorogenic acid, neochlorogenic acid, cryptochlorogenic acid and baicalin can be used as quality control markers for Yinhuang oral dropping pills.

[0068] In order to explore the effectiveness of cryptochlorogenic acid, chlorogenic acid, neochlorogenic acid, baicalin and baicalin on the treatment process. This application uses network pharmacology alignment to conduct research. The research process is as follows

[0069] 3. Network pharmacology research

[0070] 3.1 Active ingredient target prediction

[0071] The targets of chlorogenic acid, neochlorogenic acid, cryptochlorogenic acid, caffeic acid, baicalin and baicalein were retrieved through the TCMSP database (http: / / old.tcmsp-e.com. / tcmsp.php), and the species was defined as "Homo sapines" in the Uniprot database (https: / / www.uniprot.org), and the target gene names of the components obtained were standardized. Since the target points of the components collected from the TCMSP database were relatively few, the Canonical SMILES numbers of the six components were obtained through the PubChem database (https: / / pubchem.ncbi.nlm.nih.gov / ), and the targets of the six active ingredients were collected using Swiss TargetPrediction (http: / / www.swisstargetprediction.ch / ). The target information found in the TCMSP database was merged, and after removing duplicate targets, a total of 173 drug targets were obtained.

[0072] 3.2 Acquisition of disease targets

[0073] Yinhuang Hanhua Drops are a commonly used drug for the clinical treatment of pharyngitis, acute and chronic tonsillitis, and upper respiratory tract infections. The keywords "upper respiratory tract infection", "pharyngitis", "acute tonsillitis", and "chronic tonsillitis" were entered into the GeneCards and OMIM databases, and all targets collected in the OMIM database were used as disease targets. The disease targets collected in the GeneCards database were taken as the median four times, and after summarizing and removing duplicate targets, a total of 1056 disease targets were obtained.

[0074] The component targets and disease targets were uploaded to the Venny 2.1.0 online platform (https: / / www.liuxiaoyuyuan.com.cn / ) respectively, and a total of 49 targets were obtained after the intersection. The 49 targets were input into the STRING online analysis platform (https: / / cn.string-db.org / ), and the species was set to "Homo sapines", the highest confidence protein interaction parameter score value was >0.9, and the protein interaction data of the free nodes in the network were hidden. The Cytoscape 3.10.2 software was used for visualization to obtain the protein interaction PPI network diagram ( Fig.11 ), perform topological analysis with "Analysis Network", calculate and select the core targets whose betweenness centrality (BC) and closeness centrality (CC) are ≥ median and whose degree value is ≥10, and arrange the PPI network nodes in descending order according to the degree value, and obtain 14 core targets, such as Fig.12 shown.

[0075] The DAVID online analysis platform (https: / / david.ncifcrf.gov / ) was used to perform GO and KEGG enrichment analysis on the intersection targets. A total of 211 entries were obtained from the GO enrichment analysis (P < 0.01), including 172 biological processes (BP), 21 cellular components (CC) and 18 molecular functions (MF). The top 20 signaling pathways were selected based on the P value. The results are as follows: Fig.13 shown.

[0076] 3.3 Construction and analysis of the “drug-ingredient-target-pathway” network

[0077] The 6 identified chemical substances, 49 targets and 20 signal pathways in Yinhuang Hanhua Dropping Pills were mapped using Cytoscape 3.10.2 software to draw a component-target-pathway network. Fig.14 As shown. Fig.14 It can be seen that the six active ingredients of Yinhuang Suction Drops play a role in different signal pathways by acting on multiple targets, and the relationships among ingredients, targets and pathways are complex, which shows the complexity of the mechanism of action of Chinese herbal compound in treating diseases.

[0078] 3.4 Molecular docking analysis

[0079] From the PPI network, we can know that the main targets of Yinhuang Hanhua Dropping Pills are TP53, STAT3, SRC, AKT1 and other targets. The molecular docking results of the six components and the core targets TP53, STAT3, SRC and AKT1 are shown as follows: Fig.15 As shown, the binding energies of the six components with the four targets are all less than 0, indicating that they can spontaneously bind to the components in vivo. Among them, baicalin has a good docking effect with the four targets.

[0080] The above network pharmacology study shows that the six active ingredients in Yinhuang oral drops can exert clinical therapeutic effects through key targets such as TP53, STAT3, SRC, AKT, PI3K / Akt, MAPK and other signaling pathways; baicalin has the highest degree value in the "ingredient-target-pathway" network, suggesting that it may be the key substance for Yinhuang oral drops to treat upper respiratory tract infections, pharyngitis, acute and chronic tonsillitis. Molecular docking binding energy shows that the binding energy of baicalin to the four key targets is <-5kcal, indicating that it has good activity.

[0081] In summary of the above experiments, this application uses baicalein, cryptochlorogenic acid, chlorogenic acid, neochlorogenic acid and baicalin as important markers for the quality control of Yinhuang oral drops, making the quality control between batches more stable and more in line with the "multi-target" and "multi-pathway" action characteristics of traditional Chinese medicine, and its quality control is more precise.

[0082] The above is only an embodiment of the present invention, and the common knowledge such as the known specific structure and characteristics in the scheme is not described in detail here. It should be pointed out that for those skilled in the art, several deformations and improvements can be made without departing from the structure of the present invention, which should also be regarded as the protection scope of the present invention, and these will not affect the effect of the implementation of the present invention and the practicality of the patent. The scope of protection required by this application shall be based on the content of its claims, and the specific implementation methods and other records in the specification can be used to interpret the content of the claims.

Claims

1. A quality control method for Yinhuang Hanhua Dropping Pills based on fingerprint, wherein the Yinhuang Hanhua Dropping Pills are Chinese medicine dropping pills prepared from honeysuckle extract and scutellaria baicalensis extract, characterized in that: The following steps are involved: Step 1: Prepare the test solution: grind the extracted Yinhuang Hanhua Dropping Pills sample into powder, and add it into 10% to 90% methanol to prepare the test solution; Step 2: Prepare reference solution: weigh appropriate amounts of cryptochlorogenic acid, neochlorogenic acid, chlorogenic acid, baicalin, and baicalein reference substances, add 10% to 90% by volume of methanol to prepare reference solutions with concentrations of cryptochlorogenic acid, neochlorogenic acid, chlorogenic acid, baicalin, and baicalein of 1 to 600 μg / ml, respectively; Step 3: Chromatographic detection: The test solution and the reference solution are subjected to content detection and characteristic spectrum construction by HPLC or UPLC, and the determination results are compared with the detection index; The chromatographic conditions include a chromatographic column filled with octadecylsilane bonded silica gel, mobile phase A being 0.4% phosphoric acid aqueous solution, mobile phase B being acetonitrile, and gradient elution being: 0-25 min, 10% B; 25-65 min, 10%-24% B; 65-85 min, 24%-33% B; 50-105 min, 33%-40% B; 105-130 min, 40%-60% B.

2. The quality control method of Yinhuang Hanhua Dropping Pills based on fingerprint spectrum according to claim 1, characterized in that: In step 1 and step 2, the concentration of methanol is 80%.

3. The quality control method of Yinhuang Hanhua Dropping Pills based on fingerprint spectrum according to claim 2 is characterized in that: In the step 1, methanol is added and then ultrasonicated.

4. The quality control method of Yinhuang Hanhua Dropping Pills based on fingerprint spectrum according to claim 1, characterized in that: The chromatographic conditions also include a column temperature of 35°C.

5. The quality control method of Yinhuang Hanhua Dropping Pills based on fingerprint spectrum according to claim 1 is characterized in that: The chromatographic conditions include detection using an ultraviolet wavelength of 235 nm.

6. The quality control method of Yinhuang Hanhua Dropping Pills based on fingerprint spectrum according to claim 3 is characterized in that: When preparing the test solution in step 1, grind the Yinhuang Hanhua Dropping Pill sample extracted in step 1 and pass it through a No. 4 sieve. After sieving, accurately weigh the sample powder, place it in a brown volumetric flask, and accurately add Methanol, 53kHz ultrasonic treatment for 30min, after cooling to room temperature, add methanol to make up the lost weight, shake well, Filter and pass the filtrate through a 0.22 μm microporous filter membrane to obtain the test solution.

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