Pharmaceutical composition as well as construction method and detection method of characteristic chromatogram of pharmaceutical composition

Abstract

The invention provides a pharmaceutical composition and a construction method of a characteristic spectrum of the pharmaceutical composition. The pharmaceutical composition has better flowability and stability, meanwhile, the characteristic peak in the characteristic chromatogram constructed by the method is adopted as a measurement index, the pharmaceutical composition can be accurately identified, the component content of the pharmaceutical composition can be determined, the specificity is high, and quality control of the pharmaceutical composition is facilitated. Moreover, the construction method is simple and convenient to operate, has the advantages of good reproducibility, high precision, strong stability and the like, and is suitable for popularization and application.

Description

Technical Field

[0001] The present invention relates to the field of medicine, and in particular to a pharmaceutical composition in the field of traditional Chinese medicine and a construction method and a detection method of its characteristic spectrum. Background Art

[0002] For some products in the field of traditional Chinese medicine, the improvement and improvement of their preparation process may have an impact on product quality, including properties, physical and chemical identification, thin layer identification, routine inspection, content determination, etc. The quality standards of different batches of products need to be unified and need to be further improved. Therefore, establishing a comprehensive and quantifiable comprehensive ingredient quality monitoring method is important for improving product quality stability and consistency. Summary of the invention

[0003] The present invention aims to solve at least one of the technical problems existing in the prior art to a certain extent. To this end, the present invention proposes a method for constructing a characteristic spectrum of a pharmaceutical composition, a method for identifying the pharmaceutical composition, a method for detecting the pharmaceutical composition, and a method for quality control of the pharmaceutical composition. The characteristic peaks in the characteristic spectrum of the pharmaceutical composition constructed by the method of the present application are used as a measurement index, which can accurately identify the pharmaceutical composition and determine the component content of the pharmaceutical composition, has strong specificity, and is helpful for the quality control of the pharmaceutical composition. In addition, the construction method is simple to operate, has the advantages of good reproducibility, high precision and strong stability, and is suitable for popularization and application.

[0004] In one aspect of the present invention, the present invention proposes a method for constructing a characteristic spectrum of a pharmaceutical composition, characterized in that it includes: providing a plurality of different batches of pharmaceutical composition samples; pre-treating each sample under the same conditions to obtain a plurality of test solutions; performing high performance liquid chromatography on the plurality of test solutions under the same conditions to obtain a plurality of chromatograms; comparing and analyzing the plurality of chromatograms to determine common characteristic peaks, so as to obtain the characteristic spectrum of the pharmaceutical composition; wherein the mobile phase used in the high performance liquid chromatography detection includes: mobile phase A, the mobile phase A is a 0.05-0.15 mass % phosphoric acid solution; mobile phase B, the mobile phase B is acetonitrile; The elution mode of the mobile phase is gradient elution; wherein the pharmaceutical composition contains active ingredients, and the active ingredients are made from the following Chinese medicinal raw materials: puffball, zedoaria, honeysuckle, peach kernel, Scrophularia, Trigonella, safflower, salvia miltiorrhiza, isatis root, ophiopogon, thunbergii, oriental water plantain, chicken's gizzard lining, cicada shell, oroxylum butterfly, and dandelion; preferably, the weight ratio of the puffball to any one of the raw materials selected from zedoaria, peach kernel, Trigonella, safflower, and chicken's gizzard lining is 1:2, the weight ratio of the puffball to any one of the raw materials selected from salvia miltiorrhiza, fritillaria thunbergii, oriental water plantain, cicada shell, and oroxylum butterfly is 1:3, the weight ratio of the puffball to any one of the raw materials selected from honeysuckle, Scrophularia, isatis root, and dandelion is 1:5, and the weight ratio of the puffball to the weight of ophiopogon is 1:4. Specifically, in the pharmaceutical composition, the active ingredient is made of the following Chinese medicinal raw materials in the following weight percentages: based on the total weight of the Chinese medicinal raw materials, 2% puffball, 4% zedoaria, 10% honeysuckle, 4% peach kernel, 10% figwort, 4% trillium, 4% safflower, 6% salvia miltiorrhiza, 10% isatis root, 8% ophiopogon, 6% thunbergii, 6% oriental water plantain, 4% chicken gizzard lining, 6% cicada shell, 6% oroxylum, and 10% dandelion.

[0005] The inventors of the present invention used high performance liquid chromatography to detect the pharmaceutical composition and found that the composition and elution conditions of the mobile phase would significantly affect the separation effect. When gradient elution was performed using 0.05-0.15 mass % phosphoric acid solution as mobile phase A and acetonitrile as mobile phase B, more chromatographic peaks with high separation and good peak shape could be obtained.

[0006] Furthermore, the inventors performed the same pretreatment and high-performance liquid chromatography detection on multiple different batches of samples, and determined the common 7 common characteristic peaks by comparing and analyzing the multiple chromatograms obtained. These 7 common characteristic peaks constitute the characteristic spectrum of the pharmaceutical composition, and the characteristic spectrum can be used to identify the pharmaceutical composition, determine the component content of the pharmaceutical composition and whether the quality meets the standard, and achieve quality control. In addition, the method is simple to operate, has the advantages of good reproducibility, strong specificity, high precision and strong stability, and is suitable for promotion and application.

[0007] According to a specific embodiment of the present invention, the elution mode of the mobile phase is as follows:

[0008] Time (min) Mobile phase A (%) Mobile phase B (%) 0～20 90～85 10～15 20～25 85～80 15～20 25～55 80～65 20～35 55～65 65～45 35～55

[0009] The inventors obtained the above-mentioned preferred elution mode and flow rate through a large number of experiments, thereby all common characteristic peaks can be detected simultaneously, and a characteristic spectrum with high separation and good peak shape can be obtained.

[0010] According to a specific embodiment of the present invention, the flow rate of the mobile phase used in the high performance liquid chromatography detection is 0.8-1.2 mL / min, the column temperature is 25-35°C, and the wavelength is 320-330 nm. The inventors obtained the above-mentioned preferred chromatographic conditions through a large number of experiments, thereby all common characteristic peaks can be detected at the same time, and a characteristic spectrum with high separation and good peak shape can be obtained.

[0011] According to a specific embodiment of the present invention, the chromatographic column used in the high performance liquid chromatography detection is selected from a C18 chromatographic column. @ C 18 (4.6mm×250mm, 5μm), PFchrom C 18 (4.6mm×250mm, 5μm), SinoChrom ODS-BP C 18 (4.6mm×250mm, 5μm), Welch Blossmate @ C 18 (4.6mm×250mm 5μm), Agilent 5TC C 18 (4.6mm×250mm 5μm) found that Welch Xtimate @ C 18 The separation effect of the characteristic spectrum of the pharmaceutical composition of the present invention is better using a column or an Agilent 5TC-C18 column.

[0012] According to a specific embodiment of the present invention, the pretreatment comprises: mixing the pharmaceutical composition with methanol and then performing ultrasonic treatment, supplementing the weight of the resulting mixed solution with methanol, shaking, filtering, collecting the filtrate, and obtaining a test solution. The inventors obtained the above-mentioned preferred pretreatment conditions through a large number of experiments, thereby effectively extracting the characteristic components corresponding to the common characteristic peaks with few impurities, thereby helping to obtain a characteristic spectrum with high separation and good peak shape.

[0013] According to a specific embodiment of the present invention, the ultrasonic treatment is carried out at a power of 220 to 280 W and a frequency of 35 to 45 kHz for 10 to 30 minutes, so as to fully extract the components of the pharmaceutical composition of the present invention.

[0014] According to a specific embodiment of the present invention, the concentration of methanol is 90-100% by volume, so as to fully extract the components of the pharmaceutical composition of the present invention.

[0015] According to a specific embodiment of the present invention, the method further comprises: under the same conditions, respectively subjecting the reference solution and the plurality of test solution to high performance liquid chromatography detection, preferably, the reference solution is selected from Solution. As a reference, the resolution is good.

[0016] According to a specific embodiment of the present invention, the method for preparing the reference solution includes: taking an appropriate amount of a reference substance (also called a "reference substance"), accurately weighing it, and dissolving it in methanol to prepare a solution containing approximately 0.1 mg per 1 mL, as the reference solution.

[0017] According to a specific embodiment of the present invention, the number of the multiple different batches is at least 5 (eg, 5, 6, 7, 8, 9 or 10), so as to ensure the accuracy of the constructed characteristic spectrum.

[0018] According to a specific embodiment of the present invention, the characteristic spectrum includes 7 common characteristic peaks, with the second characteristic peak as the reference peak, and the ratios of the relative retention times of the 7 common characteristic peaks to the retention time of the reference peak are 0.63±10.0%, 1, 2.57±10.0%, 2.98±10.0%, 3.05±10.0%, 3.12±10.0%, and 3.48±10.0%, respectively.

[0019] refer to Figure 1 The characteristic spectrum of the pharmaceutical composition of the present invention includes 7 common characteristic peaks, wherein the common characteristic peaks are: Peak 1 is a common characteristic peak, and the substance corresponding to Peak 2 has the following structure:

[0020]

[0021] Peak 3 is a common characteristic peak, peak 4 is a common characteristic peak, peak 5 is a common characteristic peak, and the substance corresponding to peak 6 has the following structure:

[0022]

[0023] Peak 7 is a common characteristic peak. Peak 2 is the reference peak S; the relative retention time of each characteristic peak and the S peak is within the range of ±10.0% of the specified value; the specified values ​​are: 0.63 (peak 1), 1.00 (peak 2, S), 2.57 (peak 3), 2.98 (peak 4), 3.05 (peak 5), 3.12 (peak 6), 3.48 (peak 7).

[0024] In another aspect of the present invention, the present invention proposes a method for identifying the pharmaceutical composition of the present invention, characterized in that it comprises: pre-treating the sample to be tested to obtain a sample solution; performing high performance liquid chromatography on the sample solution to obtain a chromatogram; analyzing the chromatogram with the characteristic spectrum obtained by the above method to determine whether the sample to be tested is the pharmaceutical composition of the present invention; the detection conditions of the high performance liquid chromatography are as defined in the method for constructing the characteristic spectrum. Thus, the pharmaceutical composition of the present invention can be accurately identified by using the identification method according to the present invention.

[0025] According to a specific embodiment of the present invention, the preprocessing is as defined in the method for constructing the characteristic map described above.

[0026] According to a specific embodiment of the present invention, if the similarity between the chromatogram and the characteristic spectrum is not less than 0.9, the sample to be tested is identified as the pharmaceutical composition of the present invention. Specifically, the similarity is obtained by using the fingerprint similarity evaluation software of the "Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System" V2.0 developed by the Chinese Pharmacopoeia Committee.

[0027] It should be noted that the features and advantages described above for the method for constructing the characteristic spectrum are also applicable to the method for identifying the pharmaceutical composition of the present invention and will not be described in detail here.

[0028] In another aspect of the present invention, the present invention proposes a method for detecting the pharmaceutical composition of the present invention, characterized in that it includes: configuring a known concentration solution of a characteristic component corresponding to at least one characteristic peak in the characteristic spectrum obtained by the method for constructing the characteristic spectrum as a reference solution; pretreating the sample to obtain a test solution; performing high performance liquid chromatography on the reference solution and the test solution to obtain a chromatogram; analyzing the chromatogram of the reference solution and the chromatogram of the test solution to determine the content of at least one characteristic component in the sample; wherein the high performance liquid chromatography detection conditions are as defined in the method for constructing the characteristic spectrum as described above. Thus, the method of the embodiment of the present invention can accurately determine the content of characteristic components in the pharmaceutical composition of the present invention, and has the advantages of good reproducibility, strong specificity, high precision and strong stability, and is suitable for popularization and application.

[0029] According to a specific embodiment of the present invention, the preprocessing is as defined in the method for constructing the characteristic map described above.

[0030] It should be noted that the features and advantages described above for the method for constructing the characteristic spectrum are also applicable to the method for detecting the pharmaceutical composition of the present invention and will not be described in detail here.

[0031] In another aspect of the present invention, the present invention proposes a method for quality control of the pharmaceutical composition of the present invention, characterized in that it includes: pre-treating a sample of the pharmaceutical composition to obtain a test solution; subjecting the test solution to high performance liquid chromatography to obtain a chromatogram; analyzing the chromatogram with the characteristic spectrum obtained by the method for constructing the characteristic spectrum described above to determine whether the quality of the sample to be tested meets the standard; wherein the high performance liquid chromatography detection conditions are as defined in the method for constructing the characteristic spectrum described above. Thus, the method according to an embodiment of the present invention can accurately determine whether the sample of the pharmaceutical composition of the present invention meets the requirements, thereby effectively controlling the quality.

[0032] According to a specific embodiment of the present invention, the preprocessing is as defined in the method for constructing the characteristic map described above.

[0033] According to a specific embodiment of the present invention, if the similarity between the chromatogram and the characteristic spectrum is not less than 0.9, the quality of the pharmaceutical composition is deemed to be up to standard.

[0034] It should be noted that the features and advantages described above for the method for constructing the characteristic spectrum are also applicable to the method for quality control of the pharmaceutical composition and will not be repeated here.

[0035] In another aspect of the present invention, the present invention provides a pharmaceutical composition, characterized in that the pharmaceutical composition contains active ingredients, and the active ingredients are made from the following Chinese medicinal raw materials: puffball, zedoaria, honeysuckle, peach kernel, Scrophularia, Trillium, safflower, salvia miltiorrhiza, isatis root, ophiopogon, thunbergii, oriental water plantain, chicken's gizzard lining, cicada shell, oroxylum butterfly, and dandelion; preferably, the weight ratio of the puffball to any one of the raw materials selected from zedoaria, peach kernel, Trillium, safflower, and chicken's gizzard lining is 1:2, the weight ratio of the puffball to any one of the raw materials selected from salvia miltiorrhiza, fritillaria thunbergii, oriental water plantain, cicada shell, and oroxylum butterfly is 1:3, the weight ratio of the puffball to any one of the raw materials selected from honeysuckle, Scrophularia, isatis root, and dandelion is 1:5, and the weight ratio of the puffball to the weight of ophiopogon is 1:4. Specifically, in the pharmaceutical composition, the active ingredient is made of the following Chinese medicinal raw materials in the following weight percentages: based on the total weight of the Chinese medicinal raw materials, 2% puffball, 4% zedoaria, 10% honeysuckle, 4% peach kernel, 10% figwort, 4% trifoliate lanceolate, 4% safflower, 6% salvia miltiorrhiza, 10% isatis root, 8% ophiopogon, 6% thunbergii, 6% oriental water plantain, 4% chicken gizzard lining, 6% cicada shell, 6% oroxylum, and 10% dandelion;

[0036] The pharmaceutical composition further comprises the auxiliary material silicon dioxide. Specifically, the pharmaceutical composition comprises 0.2%-2% (preferably 0.3%-1.5%) silicon dioxide, based on the total weight of the pharmaceutical composition. Preferably, the pharmaceutical composition is a granule or a capsule.

[0037] According to a specific embodiment of the present invention, the similarity between the HPLC chromatogram of the pharmaceutical composition and the characteristic spectrum obtained by the method of the first aspect of the present invention is not less than 0.9. Preferably, the HPLC chromatogram of the pharmaceutical composition is obtained using the HPLC detection conditions and pretreatment operations defined in the first aspect of the present invention.

[0038] Specifically, the pharmaceutical composition has a characteristic spectrum constructed by the method of the first aspect of the present invention, which includes 7 characteristic peaks, and the substance corresponding to the second characteristic peak has the following structure:

[0039]

[0040] The substance corresponding to the sixth characteristic peak has the following structure:

[0041]

[0042] Among them, taking the second characteristic peak as the reference peak, the ratios of the relative retention times of the seven characteristic peaks to the retention time of the reference peak were 0.63±10.0%, 1.00, 2.57±10.0%, 2.98±10.0%, 3.05±10.0%, 3.12±10.0%, and 3.48±10.0%, respectively.

[0043] Or, specifically, in the characteristic spectrum of the pharmaceutical composition of the present invention, the relative peak areas of the seven characteristic peaks are: 0.127, 1.000, 0.528, 0.259, 0.189, 0.080, 0.100, respectively.

[0044] In another aspect of the present invention, the present invention also provides a method for preparing the pharmaceutical composition of the present invention, which comprises (or consists of the following):

[0045] (1) Take some amount of honeysuckle, fritillaria thunbergii, safflower and puffball, crush, sieve and mix;

[0046] (2) taking all the remaining 12 herbs (Zhuzhu, peach kernel, Scrophularia ningpoensis, Tripterygium wilfordii, Salvia miltiorrhiza, Radix Isatidis, Radix Ophiopogonis, Rhizoma Alismatis, Endothelium chicken gizzardii, Periostracum cicadae, Oroxylum indica, Taraxacum officinale) and the remaining 4 herbs (Flos lonicerae, Fritillaria thunbergii, Carthamus tinctorius, Puffball) and boiling them in water, combining the decoctions, filtering, and concentrating the filtrate;

[0047] (3) The powder obtained in step (1) and the thick paste obtained in step (2) are mixed and granulated.

[0048] According to a specific embodiment of the present invention, in the preparation method of the present invention, the granulation in step (3) is to directly perform boiling granulation on the mixed paste powder (i.e., the powder obtained in step (1) and the thick paste obtained in step (2)) and water.

[0049] According to a specific embodiment of the present invention, in the preparation method of the present invention, the partial amount of honeysuckle, Fritillaria thunbergii, safflower and puffball in step (1) is 15% to 75% of the total amount of each medicinal material, preferably 20% to 70% of the total amount of each medicinal material, and more preferably 25% to 65% of the total amount of each medicinal material.

[0050] According to a specific embodiment of the present invention, in the preparation method of the present invention, the pulverizing in step (1) is pulverizing the medicinal material into fine powder; and / or, according to a specific embodiment of the present invention, in the preparation method of the present invention, the sieving in step (1) is sieving through a 100-120 mesh sieve.

[0051] According to a specific embodiment of the present invention, in the preparation method of the present invention, the number of decoctions in step (2) is 1-5 times, preferably 2-4 times; and / or the decoction time in step (2) is 1-5 hours / time, preferably 2-4 hours / time.

[0052] According to a specific embodiment of the present invention, in the preparation method of the present invention, the filtrate in step (2) is concentrated to a thick paste with a relative density of 1.28-1.32 at a temperature of 70°C-90°C, preferably concentrated to a thick paste with a relative density of 1.28-1.32 at a temperature of 75°C-85°C.

[0053] In another aspect of the present invention, the present invention provides use of the pharmaceutical composition of the present invention in the preparation of a medicament for treating or preventing pharyngitis such as chronic laryngitis, vocal cord leukoplakia, reflux pharyngitis and / or chronic pharyngitis; preferably, the chronic laryngitis is selected from vocal cord nodules, vocal cord polyps, and / or, thickening of the vocal cord mucosa, and the hoarseness symptoms caused thereby.

[0054] Beneficial effects:

[0055] 1. The method for constructing the characteristic spectrum of the present invention overcomes the defect that the quality of the relevant dosage form cannot be fully detected from multiple angles due to the interference caused by the complex chemical composition of the pharmaceutical composition of the present invention; the present invention obtains 7 common characteristic peaks and realizes the effective separation of the 7 common characteristic peaks, with good peak shape and no interference, thereby realizing the quality detection of the effectiveness of the pharmaceutical composition of the present invention from multiple angles and in an all-round manner, and further realizing the quality control of the pharmaceutical composition of the present invention, and the quality detection method is simple to operate, has good reproducibility and stability, reliable recovery rate, low detection cost, and high detection efficiency after methodological verification, and the application of this method plays a vital role in the product quality stability of the pharmaceutical composition of the present invention.

[0056] 2. The present invention has established a set of rapid, comprehensive and highly specific quality detection methods for the pharmaceutical composition of the present invention.

[0057] 3. The characteristic spectrum constructed by the present invention can perform quality control on the pharmaceutical composition of the present invention from multiple angles and in all directions, providing more powerful guarantee for the authenticity identification and large-scale production of the pharmaceutical composition of the present invention.

[0058] 4. The present invention detects corresponding characteristic peaks in the characteristic spectra of different batches, and the similarity of the fingerprint spectra is greater than 0.9, indicating that the product quality is uniform and stable.

[0059] Additional aspects and advantages of the present invention will be given in part in the following description and in part will be obvious from the following description, or will be learned through practice of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0060] The above and / or additional aspects and advantages of the present invention will become apparent and easily understood from the description of the embodiments in conjunction with the following drawings, in which:

[0061] Figure 1 The characteristic spectrum of the pharmaceutical composition according to the embodiment of the present invention is shown;

[0062] Figure 2 The calibration result of the common characteristic fingerprint peaks in the characteristic spectrum according to the embodiment of the present invention is shown;

[0063] Figure 3 The chromatogram of the test sample for investigating the attribution of characteristic peaks in the investigation experiment of the HPLC characteristic spectrum according to the embodiment of the present invention is shown. The chromatogram in the figure is, from top to bottom, the test sample solution, the structural formula of solution, Chromatogram of a solution of

[0064] Figure 4 The stability test results in the methodological investigation experiment according to the embodiment of the present invention are shown;

[0065] Figure 5 The precision test results in the methodological investigation experiment according to the embodiment of the present invention are shown;

[0066] Figure 6 The results of the repeatability test in the methodological investigation experiment according to the embodiment of the present invention are shown;

[0067] Figure 7 The intermediate precision Shimadzu-10AT VP test results of the methodological investigation experiment according to the embodiment of the present invention are shown;

[0068] Figure 8 The results of the precision ThermoU3000-1 test in the methodological investigation experiment according to the embodiment of the present invention are shown;

[0069] Fig. 9 The results of the precision ThermoU3000-2 test in the methodological investigation experiment according to the embodiment of the present invention are shown;

[0070] Fig.10 The results of the reproducibility test in the methodological investigation experiment according to the embodiment of the present invention are shown in the laboratory 1Agilent5TC C 18 Chromatographic columns;

[0071] Fig.11 The results of the reproducibility test in the methodological investigation experiment according to the embodiment of the present invention are shown in the laboratory 2Agilent588925-902C 18 Chromatographic columns;

[0072] Fig.12 The chromatogram of the test sample using gradient elution condition 1 in the investigation experiment of the HPLC characteristic spectrum according to the embodiment of the present invention is shown;

[0073] Fig.13 The chromatogram of the test sample using gradient elution condition 2 in the investigation experiment of the HPLC characteristic spectrum according to the embodiment of the present invention is shown;

[0074] Fig.14 The chromatogram of the test sample using gradient elution condition 3 in the investigation experiment of the HPLC characteristic spectrum according to the embodiment of the present invention is shown;

[0075] Fig.15 The chromatogram of the test sample using gradient elution condition 4 in the investigation experiment of the HPLC characteristic spectrum according to the embodiment of the present invention is shown;

[0076] Fig.16 The chromatogram of the test sample using gradient elution condition 5 in the investigation experiment of the HPLC characteristic spectrum according to the embodiment of the present invention is shown;

[0077] Fig.17 The chromatogram of the test sample using gradient elution condition 6 in the investigation experiment of the HPLC characteristic spectrum according to the embodiment of the present invention is shown;

[0078] Fig.18 The chromatogram of the test sample using gradient elution condition 7 in the investigation experiment of the HPLC characteristic spectrum according to the embodiment of the present invention is shown;

[0079] Fig.19 Shows the chromatogram of the test sample extracted with 0.1% methanol in the test sample solution preparation extraction solvent investigation in the investigation experiment of the HPLC characteristic spectrum according to the embodiment of the present invention;

[0080] Fig. 20 The chromatogram of the test sample extracted by methanol in the test sample solution preparation extraction solvent investigation in the HPLC characteristic spectrum investigation experiment according to the embodiment of the present invention is shown. DETAILED DESCRIPTION

[0081] The scheme of the present invention will be explained below in conjunction with the embodiments. It will be appreciated by those skilled in the art that the following embodiments are only used to illustrate the present invention and should not be considered as limiting the scope of the present invention. Where specific techniques or conditions are not indicated in the embodiments, the techniques or conditions described in the literature in this area or the product specifications are used. The reagents or instruments used are not indicated by the manufacturer and are all conventional products that can be obtained commercially.

[0082] 1. Preparation Example

[0083] Preparation Example 1

[0084] Formula:

[0085] 25 parts of puffballs, 50 parts of zedoaria, 125 parts of honeysuckle, 50 parts of peach kernels, 125 parts of figworts, 50 parts of safflower, 75 parts of salvia miltiorrhiza, 125 parts of isatis root, 100 parts of ophiopogon, 75 parts of thunbergii bulbs, 75 parts of orientalis, 50 parts of chicken's gizzard lining, 75 parts of cicada shells, 75 parts of oroxylum butterfly, 125 parts of dandelions;

[0086] The preparation method of the pharmaceutical composition is as follows:

[0087] (1) Take 1 / 2 of honeysuckle, 1 / 2 of Fritillaria thunbergii, 1 / 2 of safflower, and 1 / 5 of puffball, grind into fine powder, sieve, and mix well;

[0088] (2) taking all the remaining 12 herbs (Zhuzhu, peach kernel, Scrophularia, Tripterygium wilfordii, Danshen, Isatis indigotica, Ophiopogon japonicus, Alisma orientalis, Gallus gallus domestica, Cicadae exuviae, Oroxylum indica, Taraxacum officinale) and the remaining 4 herbs (Lonicera japonica, Fritillaria thunbergii, Carthamus tinctorius, Puffball) and boiling them twice, each time for 2 hours, combining the decoctions, filtering, and concentrating the filtrate to a thick paste having a relative density of 1.28-1.32 at 80° C.;

[0089] (3) The fine powder obtained in step (1) and the thick paste obtained in step (2) are mixed and directly granulated with water by boiling to obtain granules, or the granules are loaded into capsules to obtain capsules.

[0090] Preparation Example 2

[0091] Formula:

[0092] 25 parts of puffballs, 50 parts of zedoaria, 125 parts of honeysuckle, 50 parts of peach kernels, 125 parts of figworts, 50 parts of safflower, 75 parts of salvia miltiorrhiza, 125 parts of isatis root, 100 parts of ophiopogon, 75 parts of thunbergii bulbs, 75 parts of orientalis, 50 parts of chicken's gizzard lining, 75 parts of cicada shells, 75 parts of oroxylum butterfly, 125 parts of dandelions;

[0093] The preparation method of the pharmaceutical composition is as follows:

[0094] (1) Take 1 / 2 of honeysuckle, 1 / 2 of Fritillaria thunbergii, 1 / 2 of safflower, and 1 / 5 of puffball, grind into fine powder, sieve, and mix well;

[0095] (2) taking all the remaining 12 herbs (Zhuzhu, peach kernel, Scrophularia, Tripterygium wilfordii, Danshen, Isatis indigotica, Ophiopogon japonicus, Alisma orientalis, Gallus gallus domestica, Cicadae exuviae, Oroxylum indica, Taraxacum officinale) and the remaining 4 herbs (Lonicera japonica, Fritillaria thunbergii, Carthamus tinctorius, Puffball) and boiling them twice, each time for 2 hours, combining the decoctions, filtering, and concentrating the filtrate to a thick paste having a relative density of 1.28-1.32 at 80° C.;

[0096] (3) The fine powder obtained in step (1) and the thick paste obtained in step (2) are mixed, and the mixture is directly granulated by boiling with water. 0.2% (based on the total weight of the granules) of silicon dioxide is added and mixed to obtain granules, or the granules can be loaded into capsules to obtain capsules.

[0097] Preparation Example 3

[0098] Formula:

[0099] 25 parts of puffballs, 50 parts of zedoaria, 125 parts of honeysuckle, 50 parts of peach kernels, 125 parts of figworts, 50 parts of safflower, 75 parts of salvia miltiorrhiza, 125 parts of isatis root, 100 parts of ophiopogon, 75 parts of thunbergii bulbs, 75 parts of orientalis, 50 parts of chicken's gizzard lining, 75 parts of cicada shells, 75 parts of oroxylum butterfly, 125 parts of dandelions;

[0100] The preparation method of the pharmaceutical composition is as follows:

[0101] (1) Take 1 / 2 of honeysuckle, 1 / 2 of Fritillaria thunbergii, 1 / 2 of safflower, and 1 / 5 of puffball, grind into fine powder, sieve, and mix well;

[0102] (2) taking all the remaining 12 herbs (Zhuzhu, peach kernel, Scrophularia, Tripterygium wilfordii, Danshen, Isatis indigotica, Ophiopogon japonicus, Alisma orientalis, Gallus gallus domestica, Cicadae exuviae, Oroxylum indica, Taraxacum officinale) and the remaining 4 herbs (Lonicera japonica, Fritillaria thunbergii, Carthamus tinctorius, Puffball) and boiling them twice, each time for 2 hours, combining the decoctions, filtering, and concentrating the filtrate to a thick paste having a relative density of 1.28-1.32 at 80° C.;

[0103] (3) The fine powder obtained in step (1) and the thick paste obtained in step (2) are mixed, and the mixture is directly granulated by boiling with water. 0.4% (based on the total weight of the granules) of silicon dioxide is added and mixed to obtain granules, or the granules can be loaded into capsules to obtain capsules.

[0104] Preparation Example 4

[0105] Formula:

[0106] 25 parts of puffballs, 50 parts of zedoaria, 125 parts of honeysuckle, 50 parts of peach kernels, 125 parts of figworts, 50 parts of safflower, 75 parts of salvia miltiorrhiza, 125 parts of isatis root, 100 parts of ophiopogon, 75 parts of thunbergii bulbs, 75 parts of orientalis, 50 parts of chicken's gizzard lining, 75 parts of cicada shells, 75 parts of oroxylum butterfly, 125 parts of dandelions;

[0107] The preparation method of the pharmaceutical composition is as follows:

[0108] (1) Take 1 / 2 of honeysuckle, 1 / 2 of Fritillaria thunbergii, 1 / 2 of safflower, and 1 / 5 of puffball, grind into fine powder, sieve, and mix well;

[0109] (2) taking all the remaining 12 herbs (Zhuzhu, peach kernel, Scrophularia, Tripterygium wilfordii, Danshen, Isatis indigotica, Ophiopogon japonicus, Alisma orientalis, Gallus gallus domestica, Cicadae exuviae, Oroxylum indica, Taraxacum officinale) and the remaining 4 herbs (Lonicera japonica, Fritillaria thunbergii, Carthamus tinctorius, Puffball) and boiling them twice, each time for 2 hours, combining the decoctions, filtering, and concentrating the filtrate to a thick paste having a relative density of 1.28-1.32 at 80° C.;

[0110] (3) The fine powder obtained in step (1) and the thick paste obtained in step (2) are mixed, and the mixture is directly granulated by boiling with water. 1.5% (based on the total weight of the granules) of silicon dioxide is added and mixed to obtain granules, or the granules can be loaded into capsules to obtain capsules.

[0111] Preparation Example 5

[0112] Formula:

[0113] 25 parts of puffballs, 50 parts of zedoaria, 125 parts of honeysuckle, 50 parts of peach kernels, 125 parts of figworts, 50 parts of safflower, 75 parts of salvia miltiorrhiza, 125 parts of isatis root, 100 parts of ophiopogon, 75 parts of thunbergii bulbs, 75 parts of orientalis, 50 parts of chicken's gizzard lining, 75 parts of cicada shells, 75 parts of oroxylum butterfly, 125 parts of dandelions;

[0114] The preparation method of the pharmaceutical composition is as follows:

[0115] (1) Take 1 / 2 of honeysuckle, 1 / 2 of Fritillaria thunbergii, 1 / 2 of safflower, and 1 / 5 of puffball, grind into fine powder, sieve, and mix well;

[0116] (2) taking all the remaining 12 herbs (Zhuzhu, peach kernel, Scrophularia, Tripterygium wilfordii, Danshen, Isatis indigotica, Ophiopogon japonicus, Alisma orientalis, Gallus gallus domestica, Cicadae exuviae, Oroxylum indica, Taraxacum officinale) and the remaining 4 herbs (Lonicera japonica, Fritillaria thunbergii, Carthamus tinctorius, Puffball) and boiling them twice, each time for 2 hours, combining the decoctions, filtering, and concentrating the filtrate to a thick paste having a relative density of 1.28-1.32 at 80° C.;

[0117] (3) The fine powder obtained in step (1) and the thick paste obtained in step (2) are mixed, and the mixture is directly granulated by boiling with water. 2.0% (based on the total weight of the granules) of silicon dioxide is added and mixed to obtain granules, or the granules can be loaded into capsules to obtain capsules.

[0118] Preparation Example 6

[0119] Formula:

[0120] 25 parts of puffballs, 50 parts of zedoaria, 125 parts of honeysuckle, 50 parts of peach kernels, 125 parts of figworts, 50 parts of safflower, 75 parts of salvia miltiorrhiza, 125 parts of isatis root, 100 parts of ophiopogon, 75 parts of thunbergii bulbs, 75 parts of orientalis, 50 parts of chicken's gizzard lining, 75 parts of cicada shells, 75 parts of oroxylum butterfly, 125 parts of dandelions;

[0121] The preparation method of the pharmaceutical composition is as follows:

[0122] (1) Take 1 / 2 of honeysuckle, 1 / 2 of Fritillaria thunbergii, 1 / 2 of safflower, and 1 / 5 of puffball, grind into fine powder, sieve, and mix well;

[0123] (2) taking all the remaining 12 herbs (Zhuzhu, peach kernel, Scrophularia, Tripterygium wilfordii, Danshen, Isatis indigotica, Ophiopogon japonicus, Alisma orientalis, Gallus gallus domestica, Cicadae exuviae, Oroxylum indica, Taraxacum officinale) and the remaining 4 herbs (Lonicera japonica, Fritillaria thunbergii, Carthamus tinctorius, Puffball) and boiling them twice, each time for 2 hours, combining the decoctions, filtering, and concentrating the filtrate to a thick paste having a relative density of 1.28-1.32 at 80° C.;

[0124] (3) The fine powder obtained in step (1) and the thick paste obtained in step (2) are mixed, and the mixture is directly granulated by boiling with water. 2.3% (based on the total weight of the granules) of silicon dioxide is added and mixed to obtain granules, or the granules can be loaded into capsules to obtain capsules.

[0125] 2. Drug Composition Prescription and Parameter Screening

[0126] The inventors tested the appearance, properties, particle size, angle of repose, moisture and other parameters of the pharmaceutical compositions prepared in Preparation Examples 1-6. The specific results are as follows:

[0127]

[0128] Particle size*: The sum of particles that cannot pass through the No. 1 sieve and particles that can pass through the No. 5 sieve shall not exceed 15%;

[0129] Angle of repose**: For the pharmaceutical composition of the present invention, an angle of repose within the range of 24°-30° is considered to have very good fluidity, an angle of repose within the range of 31°-35° is considered to have good fluidity, and an angle of repose ≥36° is considered to have poor fluidity;

[0130] Moisture content: not more than 8%.

[0131] Summary: From the above data results, it can be seen that the appearance, properties, particle size, angle of repose, and moisture content of the pharmaceutical compositions obtained in Preparation Examples 2-5 all meet the quality control requirements, while the pharmaceutical composition obtained in Preparation Example 1 (without silicon dioxide) is not suitable for use as granules or capsules due to poor fluidity. It can be seen that for the present invention, a certain amount of silicon dioxide needs to be added to obtain a pharmaceutical composition with good fluidity.

[0132] The inventors further confirmed the influence of the addition of silicon dioxide on the quality of the pharmaceutical composition through the results of stability study.

[0133]

[0134]

[0135] Particle size*: The sum of particles that cannot pass through the No. 1 sieve and particles that can pass through the No. 5 sieve shall not exceed 15%;

[0136] Moisture content**: not more than 8%.

[0137] Summary: Through the analysis of the results of the 3-month accelerated stability study on the pharmaceutical compositions obtained in Preparation Examples 1-6, the pharmaceutical composition obtained in Preparation Example 1 had slightly agglomerated particles under the accelerated test for 3 months, and the sum of the particle sizes that could not pass through the No. 1 sieve and the particle sizes that could pass through the No. 5 sieve was close to the upper limit. The filling amount difference of the pharmaceutical composition obtained in Preparation Example 6 did not meet the regulations (the difference exceeded ±10%), while the results of the pharmaceutical compositions obtained in Preparation Examples 2-5 all met the regulations.

[0138] 3. Experimental Examples

[0139] The test instruments, reagents, samples and reference substances used in the embodiments of the present invention are shown in Tables 1-4.

[0140] Table 1 Test instruments

[0141]

[0142]

[0143] Table 2 Test reagents

[0144] name batch number level Manufacturer Methanol 20065185 HPLC TEDIA Acetonitrile 211003317G104 HPLC OCEANPAK Phosphoric acid 20210902 HPLC Tianjin Tianli Chemical Reagent Ultrapure water / First-class water self made

[0145] Table 3 Test samples

[0146]

[0147] Table 4 Test reference substances

[0148]

[0149]

[0150] Example 1

[0151] Take 10 pellets of the sample contents from 6 batches (JA121060771, JA121070783, JA121070793, JA121070803, JA121070813, JA121070823) for testing and perform the following operations in Laboratory 1:

[0152] 1) Chromatographic conditions: Octadecyl bonded silica gel was used as the filler, and the chromatographic column was selected from Agilent 5TC-C 18 , column length 25cm, inner diameter 4.6mm, particle size 5μm; 0.1 mass% phosphoric acid as mobile phase A, acetonitrile as mobile phase B, gradient elution as specified in the table below, equilibrate for 10 minutes before injection; flow rate 1.0mL per minute; column temperature 30℃; detection wavelength 327nm; theoretical plate number calculated based on reference peak (peak of reference substance 1) should be no less than 10000;

[0153] Table 5 Elution conditions

[0154] Time (minutes) Mobile phase A (%) Mobile phase B (%) 0～20 90～85 10～15 20～25 85～80 15～20 25～55 80～65 20～35 55～65 65～45 35～55

[0155] 2) Preparation of test solution: Take the sample content, grind it into fine powder, take 0.2g of fine powder, weigh it accurately, put it into a stoppered conical flask, add 25mL of 100% methanol accurately, weigh it, treat it with ultrasound (250W, 40KHz) for 20 minutes, let it cool to room temperature, weigh it again, make up the loss with 100% methanol, shake it well, filter it, and take the filtrate to obtain;

[0156] 3) Preparation of reference solution: Take an appropriate amount of reference substance 1, weigh accurately, and dissolve in methanol to make a solution containing about 0.1 mg per 1 mL, which is used as the reference solution;

[0157] 4) Determination: Accurately pipette 10 μL of reference solution and test solution respectively, inject into liquid chromatograph, measure, and record the chromatogram.

[0158] Example 2 Determining the characteristic spectrum

[0159] The chromatograms of the test solutions of the six batches of samples used in the experiment were analyzed and compared, and the "Chinese Medicine Fingerprint Similarity Software" 2.0 was used for calibration to determine their common peaks.

[0160] Results: More than 20 chromatographic peaks were separated in the chromatograms of the 6 batches of test solution, of which 19 were common peaks, and the common peak area accounted for more than 80% of the total peak area. The results are shown in Table 6-8. Figure 2 .

[0161] Taking the peak of reference substance 1 as the reference peak, calculate the relative retention time and peak area ratio. The calculation formula is as follows:

[0162]

[0163] Table 6 Common peak matching results

[0164]

[0165]

[0166] Table 7 Relative retention time of common peaks

[0167]

[0168] Table 8 Common peak relative peak area

[0169]

[0170]

[0171] 1. Attribution of characteristic peaks

[0172] Positioning reference solution: Take (also referred to herein as "reference 1"), (also referred to herein as "reference 2"), An appropriate amount of reference substance (also referred to herein as "reference substance 3") is accurately weighed and dissolved in methanol to prepare a solution containing approximately 0.1 mg of each substance per 1 mL as a positioning reference substance solution.

[0173] The positioning reference solution was measured using the chromatographic conditions of Example 1, and the peaks in the chromatogram of the test solution (batch number: JA121060771) were compared to attribute the characteristic peaks. The results are shown in Table 9.

[0174] Results: Chromatogram of the test solution (see Figure 3 ) in which three known components are identified by characteristic peaks, among which the peaks of reference 1 and reference 2 are more obvious and can be used as characteristic peaks. The absorption of reference 3 is weak at 327nm, and this characteristic spectrum fails to reflect the chromatographic peak of reference 3.

[0175] Table 9 Attribution of characteristic peaks in the chromatogram of the test sample

[0176]

[0177]

[0178] 2. Investigation of reference solution:

[0179] The peaks of known components detected in the test solution were more obvious than those of reference 1 and reference 2, so they were used as references for investigation.

[0180] Take an appropriate amount of reference substance (also called "reference material"), weigh it accurately, and dissolve it in chromatographic methanol to make a 1 mL solution containing about 0.1 mg of each substance, which serves as the reference material solution.

[0181] The reference solution was chromatographically detected according to the chromatographic conditions in Example 1. The results showed that the peak retention time of reference 1 was 11.825 minutes, and the separation effect was good, so it was used as a reference solution. The peak retention time of reference 2 was 42.540 minutes, and its proportion in the chromatogram was relatively low, and its peak was relatively late, so it was not suitable as a reference, and was only used as a positioning solution.

[0182] 3. Methodological investigation

[0183] With reference to the 2020 edition of the Chinese Pharmacopoeia, Part IV, General Rules 9101, "Guidelines for Validation of Analytical Methods" and "Technical Requirements for Fingerprint Research of Traditional Chinese Medicine Injections", the method for constructing the characteristic spectrum of Example 1 was investigated for characteristic spectrum / fingerprint methodology, mainly including solution stability, precision, repeatability, intermediate precision, durability and reproducibility tests. The acceptable standards for validation are mainly based on the acceptable range of the content and precision of the components to be tested in the sample in the "Technical Requirements for Fingerprint Research of Traditional Chinese Medicine Injections" and the 2020 edition of the Chinese Pharmacopoeia, General Rules 9101, "Guidelines for Validation of Analytical Methods".

[0184] (1) Stability test

[0185] The test solution (batch number: JA121060771) was prepared by referring to the test solution preparation method in Example 1, 10 μL of the test solution was accurately pipetted, and the samples were placed at room temperature and protected from light for 0, 8, 20, 24, and 34 h for measurement (the measurement conditions were the same as in Example 1), and the relative retention time and peak area ratio of the common peaks were calculated. At the same time, the similarity evaluation was performed using the "Similarity Evaluation System for Chromatographic Fingerprints of Traditional Chinese Medicines" V2.0 version to investigate the stability of the test solution.

[0186] Verification requirements: The relative retention time RSD of the characteristic peak in the chromatogram of the test solution is ≤1.0%, the relative peak area RSD is ≤5.0%, and the similarity is >0.9. The test solution can be considered to be stable.

[0187] Verification results: In the test solution, the RSD of the relative retention time of each characteristic peak is ≤0.35%, the RSD of the relative peak area is ≤3.54%, and the similarity is greater than 0.9, which indicates that the test solution is stable within 34 hours. The results are shown in Table 10-11. Figure 4 .

[0188] Table 10 Solution stability similarity evaluation

[0189]

[0190] Table 11 Stability test results

[0191]

[0192]

[0193] (2) Precision test

[0194] The test solution (batch number: JA121060771) was prepared by referring to the test solution preparation method in Example 1, 10 μL of the test solution was aspirated for determination (the determination conditions were the same as in Example 1), and the sample was injected 6 times continuously. The relative retention time and relative peak area of ​​the common peaks in the characteristic spectrum were calculated. At the same time, the similarity evaluation was performed using the "Similarity Evaluation System for Chromatographic Fingerprints of Traditional Chinese Medicine" V2.0 version to examine the precision of the instrument.

[0195] Verification requirements: The relative retention time RSD of the characteristic peaks in the chromatogram of the test solution is ≤1.0%, the relative peak area RSD is ≤3.0%, and the similarity is >0.9.

[0196] Verification results: 6 test results, relative retention time RSD ≤ 0.26%, relative peak area RSD ≤ 0.639% (except for peak 10, peak 10 RSD = 3.84%, mainly due to its small proportion), similarity is greater than 0.9, indicating that the instrument precision is good. The results are shown in Table 12-13. Figure 5 .

[0197] Table 12 Precision similarity evaluation

[0198]

[0199] Table 13 Precision test results

[0200]

[0201]

[0202] (3) Repeatability test

[0203] 6 portions of the test solution (batch number: JA121060771) were prepared by referring to the test solution preparation method in Example 1, and 10 μL of each solution was injected for measurement (the measurement conditions were the same as in Example 1). The relative retention time and relative peak area of ​​the common peaks in the characteristic spectrum were calculated. At the same time, the similarity evaluation was performed using the "Similarity Evaluation System for Chromatographic Fingerprints of Traditional Chinese Medicine" V2.0 version to examine the reproducibility of the experimental method.

[0204] Verification requirements: The relative retention time RSD of the common characteristic peaks in the chromatogram of the test solution is ≤2.0%, the relative peak area RSD is ≤15.0%, and the similarity is >0.9.

[0205] Verification results: 6 test results, relative retention time RSD ≤ 0.18%, relative peak area RSD ≤ 10.38%, similarity greater than 0.9, indicating that the repeatability of the determination method is good. The results are shown in Table 14-15. Figure 6 .

[0206] Table 14 Repeatability similarity evaluation

[0207]

[0208] Table 15 Repeatability test results

[0209]

[0210]

[0211] (4) Intermediate precision test

[0212] Take the contents of this product (batch number: JA121060771), and different personnel prepare the test solution according to the preparation method of the test solution in Example 1, and take 10 μL of it in Shimadzu-10AT VP , the sample was injected and analyzed on a Thermo U3000 high performance liquid chromatograph (the measurement conditions were the same as in Example 1), the relative retention time of the characteristic peaks in the fingerprint spectrum was calculated, and at the same time, the similarity evaluation was carried out against the spectrum using the "Similarity Evaluation System for Chromatographic Fingerprints of Traditional Chinese Medicine" V2.0 version with a repeatability test (batch number: JA121060771) to examine the intermediate precision of the experimental method.

[0213] Verification requirements: In the characteristic spectrum, the relative deviation of the characteristic peak relative retention time is specified to be within 5% and the similarity is greater than 0.9.

[0214] Verification results: The common characteristic peaks were detected by different personnel and instruments, the RSD of relative retention time was ≤1.91%, and the similarity was >0.9, indicating that the intermediate precision of the determination method was good. The results are shown in Table 16-17. Figure 7-9 .

[0215] Table 16 Intermediate precision density similarity evaluation (Mark peak)

[0216]

[0217] Table 17 Intermediate precision test results

[0218]

[0219] (5) Reproducibility test

[0220] Take the test sample of this product (batch number: JA121060771) and measure it in laboratory 2 (the measurement conditions are the same as those in Example 1), draw a characteristic spectrum, and examine the reproducibility of the method.

[0221] Verification requirements: The relative retention time of the characteristic peaks in the characteristic spectrum is within the specified range and the separation meets the requirements.

[0222] Verification results: Laboratory 2 used (Agilent 588925-902, 4.6mm×250mm, 5μm) for measurement, and the sample characteristic spectrum was consistent with that of Laboratory 1 Agilent 5TC-C 18 The standard characteristic spectrum is consistent, but the separation of peaks 3 and 5 does not completely reach the baseline separation. The analytical effects of different chromatographic columns and instruments are quite different. It is recommended that peaks 3 and 5 are not listed as characteristic peaks. Other peaks fully meet the requirements, indicating that the method is reproducible and feasible, and the durability meets the requirements. The results are shown in Table 18. Figure 10-11 .

[0223] Table 18 Separation characteristic spectrum results of laboratory 2

[0224]

[0225] The common peaks of the characteristic spectra of 6 batches of samples for the test were analyzed (the measurement conditions were the same as in Example 1). In the corresponding Table 5, the separation effect of peaks 1, 6, 8, and 9 was poor, peaks 2, 4, and 12 were unstable, and the proportion of peaks 7 and 19 in the total peaks was small, so they were not used as common characteristic peaks. The analysis effects of peaks 10 and 13 were quite different due to different chromatographic columns and instruments, and the reproducibility was poor, so they were not finally used as common characteristic peaks; peaks 3, 5, 11, 14, 15, 16, and 18 accounted for 2.27% to 27.4% of the total peaks, and the chromatographic separation was relatively good, so they were intended to be used as common characteristic peaks.

[0226] 4. Proposed feature map

[0227] The characteristic spectrum of the test product should show 9 characteristic peaks. The second characteristic peak is used as the reference peak (S peak). The relative retention time of each characteristic peak and S peak is calculated. Figure 6 .

[0228] The relative retention times of the 9 characteristic peaks of this product are: 0.59 (peak 1), 1.00 (peak S), 2.89 (peak 3), 2.97 (peak 4), 3.20 (peak 5), 3.50 (peak 6), 3.57 (peak 7), 3.61 (peak 8), and 4.06 (peak 9).

[0229] 5. Revision scope of characteristic spectrum limit

[0230] Based on the above results, the revised feature map is:

[0231] The characteristic spectrum of the test product should show 7 common characteristic peaks. The second characteristic peak is used as the reference peak (S peak). The relative retention time of each characteristic peak and the S peak is calculated. It should be within ±10.0% of the specified value. The specified value is:

[0232] 0.63 (peak 1), 1.00 (peak S), 2.57 (peak 3), 2.98 (peak 4), 3.05 (peak 5), 3.12 (peak 6), 3.48 (peak 7), Peak (S): Peak (6):

[0233] 6. Similarity evaluation

[0234] The characteristic spectrum test data of 6 batches (JA121060771, JA121070783, JA121070813, JA121070793, JA121070803, and JA121070823) were imported into the traditional Chinese medicine fingerprint spectrum similarity software. After calculation by the traditional Chinese medicine fingerprint spectrum similarity software, the control characteristic spectrum of the characteristic spectrum was obtained. The characteristic spectrum test data of each batch was calculated with the MARK peak (characteristic peak) similarity, and the similarity of each batch of samples was compared.

[0235] Results Characteristic peaks were detected in the fingerprints of samples from different batches, and the similarities of MARK peaks (characteristic peaks) were all ≥0.9, with no significant differences. The results are shown in Table 19.

[0236] Table 19 Fingerprint similarity evaluation results of different batches of samples

[0237]

[0238]

[0239] 7. System suitability investigation

[0240] The method of Example 1 was adopted to examine the suitability of the system with the separation effect of the reference peak and the adjacent peaks in the chromatogram of the test solution as an indicator. The results are shown in Table 20 below.

[0241] Table 20 System suitability

[0242]

[0243] Example 3

[0244] In this embodiment, the characteristic map is constructed according to the following method:

[0245] 1. Chromatographic conditions: Octadecyl bonded silica gel was used as filler, and the chromatographic column was selected from Agilent 5TC-C 18Or an equivalent chromatographic column, with a column length of 25 cm, an inner diameter of 4.6 mm, and a particle size of 5 μm; 0.1 mass% phosphoric acid as mobile phase A, acetonitrile as mobile phase B, gradient elution as specified in Tables 21-27, and equilibration for 10 minutes before injection; flow rate of 1.0 mL per minute; column temperature of 30°C; detection wavelength of 327 nm; theoretical plate number calculated based on the reference peak should be no less than 10,000;

[0246] 2. Preparation and determination of test solution and reference solution: Same as in Example 1, the characteristic spectra obtained under different gradient elution conditions are shown in Figure 12-18 .

[0247] Table 21 Gradient elution conditions 1

[0248]

[0249] Table 22 Gradient elution conditions 2

[0250]

[0251] Table 23 Gradient elution conditions 3

[0252]

[0253] Table 24 Gradient elution conditions 4

[0254]

[0255] Table 25 Gradient elution conditions 5

[0256]

[0257]

[0258] Table 26 Gradient elution conditions 6

[0259]

[0260] Table 27 Gradient elution conditions 7

[0261]

[0262] Conclusion: In the system of acetonitrile and 0.1 mass% phosphoric acid, the mobile phase gradient elution with different proportions was investigated. When the proportion of acetonitrile reached 55%, the main components in the prescription could be almost completely eluted out of the column. Finally, the gradient elution condition 7 in Table 27 was determined as the optimal chromatographic condition. The separation effect of the chromatographic peaks in the test solution was good, and the baseline separation was basically achieved, which could meet the detection requirements.

[0263] Example 4

[0264] In this embodiment, the characteristic map is constructed according to the following method:

[0265] 1. Chromatographic conditions: Octadecyl bonded silica gel is used as filler, and the chromatographic column is selected from Welch Xtimate @ C 18 Or an equivalent chromatographic column, with a column length of 25 cm, an inner diameter of 4.6 mm, and a particle size of 5 μm; 0.05 mass % phosphoric acid as mobile phase A, acetonitrile as mobile phase B, gradient elution (same as in Example 1), equilibration for 10 minutes before injection; flow rate of 0.8 mL per minute; column temperature of 25° C.; detection wavelength of 320 nm; theoretical plate number calculated based on the reference peak should be not less than 10,000.

[0266] 2. Preparation of test solution: Take the sample content, grind it into fine powder, take 0.2g of fine powder, weigh it accurately, put it in a stoppered conical flask, add 25mL of 90% methanol accurately, weigh it, treat it with ultrasound (220W, 35KHz) for 10 minutes, let it cool to room temperature, weigh it again, make up the loss with 90% methanol, shake it well, filter it, and take the filtrate to obtain;

[0267] 3. Preparation and determination of reference solution: Same as Example 1, record the chromatogram.

[0268] The chromatogram of the test sample showed 7 common characteristic peaks. Taking the second characteristic peak as the reference peak, the ratios of the relative retention time of the 7 common characteristic peaks to the retention time of the reference peak were 0.63±10.0%, 1, 2.57±10.0%, 2.98±10.0%, 3.05±10.0%, 3.12±10.0%, and 3.48±10.0%, respectively. The separation degree of adjacent chromatographic peaks was greater than 1.5, and the baseline separation was achieved between the characteristic peaks, which met the requirements.

[0269] Example 5

[0270] In this embodiment, the characteristic map is constructed according to the following method:

[0271] 1. Chromatographic conditions: Octadecyl bonded silica gel is used as filler, and the chromatographic column is selected from Agilent 5TC-C18 or equivalent chromatographic column, with a column length of 25 cm, an inner diameter of 4.6 mm, and a particle size of 5 μm; 0.15 mass% phosphoric acid is used as mobile phase A, acetonitrile is used as mobile phase B, and gradient elution is performed (same as in Example 1), and equilibrium is performed for 10 minutes before injection; the flow rate is 1.2 mL per minute; the column temperature is 35°C; the detection wavelength is 330 nm; the theoretical plate number calculated based on the reference peak should be not less than 10,000.

[0272] 2. Preparation of test solution: Take the sample content, grind it into fine powder, take 0.2g of fine powder, weigh it accurately, put it in a stoppered conical flask, add 25mL of 95% methanol accurately, weigh it, treat it with ultrasound (280W, 45KHz) for 30 minutes, let it cool to room temperature, weigh it again, make up the loss with 95% methanol, shake it well, filter it, and take the filtrate to obtain;

[0273] 3. Preparation and determination of reference solution: Same as Example 1, record the chromatogram.

[0274] The chromatogram of the test sample showed 7 common characteristic peaks. Taking the second characteristic peak as the reference peak, the ratios of the relative retention time of the 7 common characteristic peaks to the retention time of the reference peak were 0.63±10.0%, 1, 2.57±10.0%, 2.98±10.0%, 3.05±10.0%, 3.12±10.0%, and 3.48±10.0%, respectively. The separation degree of adjacent chromatographic peaks was greater than 1.5, and the baseline separation was achieved between the characteristic peaks, which met the requirements.

[0275] Example 6

[0276] In this embodiment, the characteristic map is constructed according to the following method:

[0277] 1. Chromatographic conditions: same as in Example 1.

[0278] 2. Preparation of test solution: Take the sample content, grind it into fine powder, take 0.2g of fine powder, weigh it accurately, put it in a stoppered conical flask, add 25mL of 50% methanol accurately, weigh it, treat it with ultrasound (250W, 40KHz) for 20 minutes, let it cool to room temperature, weigh it again, make up the loss with methanol, shake it well, filter it, and take the filtrate to obtain;

[0279] 3. Preparation and determination of reference solution: same as in Example 1.

[0280] The characteristic spectra obtained by using 100% methanol and 50% methanol as extraction solvents (see Fig.19 and Fig. 20 ) have the same number of peaks and similar characteristics, with no significant difference, but there is no water in 100% methanol, while the water content in 50% methanol is high. Therefore, compared with 50% methanol, the test solution prepared with 100% methanol is easier to filter, so the preferred extraction solvent is 100% methanol.

[0281] summary

[0282] The characteristic spectrum / fingerprint spectrum developed by this method has a certain specificity. The characteristic peaks basically reflect the main ingredients with ultraviolet absorption and known effective ingredients in the prescription. The method has good durability and reproducibility, and can be used as a basis for quality control of the pharmaceutical composition, to evaluate the uniformity, excellence and stability of the preparation quality, and to compare the consistency of product quality before and after the change.

[0283] The characteristic spectrum / fingerprint detection results of each batch of samples show that the corresponding characteristic peaks are detected in samples from different batches, and the fingerprint spectrum similarity is greater than 0.9, indicating that the product quality is uniform and stable.

[0284] Although the embodiments of the present invention have been shown and described above, it is to be understood that the above embodiments are exemplary and are not to be construed as limitations of the present invention. A person skilled in the art may change, modify, replace and vary the above embodiments within the scope of the present invention.

Claims

1. A method for constructing a characteristic spectrum of a pharmaceutical composition, characterized in that: include: Providing a plurality of different batches of pharmaceutical composition samples; Under the same conditions, each sample was pretreated separately to obtain multiple test sample solutions; Under the same conditions, a plurality of the test sample solutions are subjected to high performance liquid chromatography to obtain a plurality of chromatograms; By comparing and analyzing a plurality of the chromatograms, common characteristic peaks are determined to obtain a characteristic spectrum of the pharmaceutical composition; Wherein, the mobile phase used in the high performance liquid chromatography detection includes: Mobile phase A, wherein the mobile phase A is a 0.05-0.15 mass % phosphoric acid solution; Mobile phase B, wherein the mobile phase B is acetonitrile; The elution mode of the mobile phase is gradient elution; Wherein, the pharmaceutical composition contains active ingredients, and the active ingredients are made from the following Chinese medicinal raw materials: puffball, zedoaria, honeysuckle, peach kernel, Scrophularia, Trigonella, safflower, salvia miltiorrhiza, isatis root, ophiopogon, thunbergii, oriental water plantain, chicken's gizzard lining, cicada shell, oroxylum indicum, and dandelion; preferably, the weight ratio of the puffball to any one of the raw materials selected from zedoaria, peach kernel, Trigonella, safflower, and chicken's gizzard lining is 1:2, the weight ratio of the puffball to any one of the raw materials selected from salvia miltiorrhiza, fritillaria thunbergii, oriental water plantain, cicada shell, and oroxylum indicum is 1:3, the weight ratio of the puffball to any one of the raw materials selected from honeysuckle, Scrophularia, isatis root, and dandelion is 1:5, and the weight ratio of the puffball to the weight of ophiopogon is 1:

4.

2. The method according to claim 1, characterized in that The elution mode of the mobile phase is as follows: 。 3. The method according to claim 1, characterized in that The mobile phase used in the HPLC detection has a flow rate of 0.8 to 1.2 mL / min, a column temperature of 25 to 35° C., and a wavelength of 320 to 330 nm; and / or, the chromatographic column is selected from a C18 chromatographic column, preferably a Welch Xtimate @ C 18 column or Agilent 5TC-C18 column.

4. The method according to claim 1, characterized in that: The pre-processing comprises: The pharmaceutical composition and methanol are mixed and then ultrasonically treated, the resulting mixture is supplemented with methanol to make up the weight, shaken well, filtered, and the filtrate is collected to obtain a test solution; Optionally, the ultrasonic treatment is carried out at a power of 220 to 280 W and a frequency of 35 to 45 kHz for 10 to 30 minutes; Optionally, the concentration of the methanol is 90 to 100 volume %.

5. The method according to claim 1, characterized in that Further including: Under the same conditions, the reference solution and the plurality of test solution are subjected to high performance liquid chromatography detection, preferably, the reference solution is selected from Solution; Optionally, the number of batches of the plurality of different batches is at least 5; Optionally, the characteristic spectrum includes 7 common characteristic peaks, among which the second characteristic peak is used as the reference peak, and the ratios of the relative retention times of the 7 common characteristic peaks to the retention time of the reference peak are 0.63±10.0%, 1.00, 2.57±10.0%, 2.98±10.0%, 3.05±10.0%, 3.12±10.0%, and 3.48±10.0%, respectively.

6. A method for identifying the pharmaceutical composition defined in claim 1, characterized in that: include: Pre-treating the sample to be tested to obtain a sample solution; Performing high performance liquid chromatography on the sample solution to obtain a chromatogram; Analyzing the chromatogram with the characteristic spectrum obtained by the method according to any one of claims 1 to 5 to determine whether the sample to be tested is the pharmaceutical composition defined in claim 1; The detection conditions of the high performance liquid chromatography are as defined in the method according to any one of claims 1 to 3 and 5.

7. The method according to claim 6, characterized in that The pretreatment is as defined in the method of claim 4; If the similarity between the chromatogram and the characteristic spectrum is not less than 0.9, the sample to be tested is determined to be the pharmaceutical composition defined in claim 1.

8. A method for detecting the pharmaceutical composition defined in claim 1, characterized in that: include: A solution of a characteristic component of known concentration corresponding to at least one characteristic peak in the characteristic spectrum obtained by the method according to any one of claims 1 to 5 is prepared as a reference solution; Pre-treating the sample to obtain a test solution; The reference solution and the test solution are respectively subjected to high performance liquid chromatography to obtain a chromatogram; Analyzing the chromatogram of the reference solution and the chromatogram of the test solution to determine the content of at least one characteristic component in the sample; Wherein, the HPLC detection conditions are as defined in the method according to any one of claims 1-3 and 5.

9. The method according to claim 8, characterized in that The pre-treatment is as defined in the method of claim 4.

10. A method for quality control of the pharmaceutical composition defined in claim 1, characterized in that: include: Pretreating the pharmaceutical composition sample to obtain a test solution; The test solution is subjected to high performance liquid chromatography to obtain a chromatogram; Analyze the chromatogram with the characteristic spectrum obtained by the method according to any one of claims 1 to 5 to determine whether the quality of the sample to be tested meets the standard; Wherein, the HPLC detection conditions are as defined in the method according to any one of claims 1 to 3 and 5; Optionally, the pretreatment is as defined in the method of claim 4; Optionally, if the similarity between the chromatogram and the characteristic spectrum is not less than 0.9, the quality of the pharmaceutical composition is deemed to be up to standard.

11. The pharmaceutical composition according to claim 1, characterized in that The pharmaceutical composition comprises the auxiliary material silicon dioxide. Specifically, the pharmaceutical composition comprises 0.2%-2% (preferably 0.4%-1.5%) of silicon dioxide, based on the total weight of the pharmaceutical composition. Preferably, the pharmaceutical composition is a granule or a capsule.

12. The pharmaceutical composition according to claim 11, characterized in that The similarity between the high performance liquid chromatogram of the pharmaceutical composition and the characteristic spectrum constructed by the method of any one of claims 1 to 5 is not less than 0.

9.

13. Use of the pharmaceutical composition according to any one of claims 11-12 in the preparation of a medicament for treating or preventing pharyngitis such as chronic laryngitis, vocal cord leukoplakia, reflux pharyngitis and / or chronic pharyngitis; preferably, the chronic laryngitis is selected from vocal cord nodules, vocal cord polyps, and / or, thickening of the vocal cord mucosa, and the hoarseness symptoms caused thereby.

Images