CN108956835B - Fingerprint detection method of oral medicine for clearing heat from throat - Google Patents
Fingerprint detection method of oral medicine for clearing heat from throat Download PDFInfo
- Publication number
- CN108956835B CN108956835B CN201710358947.3A CN201710358947A CN108956835B CN 108956835 B CN108956835 B CN 108956835B CN 201710358947 A CN201710358947 A CN 201710358947A CN 108956835 B CN108956835 B CN 108956835B
- Authority
- CN
- China
- Prior art keywords
- solution
- fingerprint
- mobile phase
- clearing heat
- throat
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Images
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/88—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/88—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86
- G01N2030/8809—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86 analysis specially adapted for the sample
Landscapes
- Physics & Mathematics (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Other Investigation Or Analysis Of Materials By Electrical Means (AREA)
- Investigating Or Analysing Biological Materials (AREA)
Abstract
The invention discloses a fingerprint detection method of an oral medicine for clearing heat from throat, wherein the detection method comprises the steps of preparation of a reference substance solution, preparation of a test substance solution and detection. Wherein, the high performance liquid detection chromatographic conditions are as follows: the chromatographic column uses octadecylsilane chemically bonded silica as a filler, mobile phase A is acetonitrile, mobile phase B is 0.2% phosphoric acid, gradient elution is carried out, the flow rate of the mobile phase is 1.2ml/min, the column temperature is 35 ℃, and the detection wavelength is as follows: 206 nm.
Description
Technical Field
The invention relates to the field of medicine detection, in particular to a method for detecting a fingerprint spectrum of a Chinese patent medicine by high performance liquid chromatography
Background
The oral liquid for clearing heat from throat and relieving fever for children is a new traditional Chinese medicine which is produced by Sichuan pharmaceutical limited company in the Asia-Bao pharmaceutical industry and is used for treating the lung-stomach excess heat syndrome caused by acute pharyngitis (acute pharyngitis) of children, and comprises 8 traditional Chinese medicinal materials of radix bupleuri, baicalin, Chinese violet, artificial bezoar, endive, houttuynia cordata, rhizoma phragmitis and phaseolus calcaratus, has the effects of clearing heat and removing toxicity, and relieving swelling and sore throat, and is used for treating the lung-stomach excess heat syndrome caused by acute pharyngitis (acute pharyngitis), and the main components and the components of the oral liquid are described in patent CN 1296088C. Clinical practice proves that the oral liquid for clearing heat from throat and clearing heat of children is effective on symptoms and signs of pharyngalgia, dry stool and red and swollen throat of acute pharyngitis (lung-stomach excess heat syndrome) of children, has better safety and is worthy of clinical popularization and application.
No quality detection standard of the variety is recorded in the Chinese pharmacopoeia. The existing standard has the following defects: the method focuses on qualitative identification and quantitative detection of main components of the oral liquid for clearing heat from throat and clearing heat of children, and lacks quality control of the whole medicine. No researchers have conducted overall quality control studies on this variety by review of the literature.
The fingerprint is a chromatogram or spectrum of a certain kind or several kinds of components which are shared by a certain Chinese medicinal material or a Chinese medicinal herb and have characteristics based on the knowledge of the integral action of the Chinese medicinal herb group, and has the characteristics of large information amount, strong characteristics, integrity, ambiguity and the like. At the present stage, under the condition that most of the effective components of the traditional Chinese medicine are not clear, the fingerprint can comprehensively reflect the relative relationship of the chemical components contained in the medicine, embodies the complexity and the relativity of the traditional Chinese medicine components, is adapted to the traditional theory of the traditional Chinese medicine, and can really and effectively characterize, comprehensively evaluate and comprehensively control the internal quality of the traditional Chinese medicine. Has important significance for effectively controlling the quality of the traditional Chinese medicinal materials or the Chinese medicinal materials.
The oral liquid for clearing heat from throat and clearing heat of children belongs to special medicine for children, and the quality control thereof is particularly important. In order to comprehensively control the product quality of the product, it is necessary to establish a new, fast, accurate and suitable quality detection method for the requirement of industrial mass production, so as to meet the overall quality control requirement of the product.
Disclosure of Invention
The invention establishes a fingerprint detection method of the preparation aiming at the problems that the throat-clearing and fever-relieving oral liquid for children has various medicinal flavors and complex components, and the qualitative and quantitative analysis of individual components is difficult to comprehensively reflect the comprehensive information of the medicine, so as to better and more effectively control the overall quality of the throat-clearing and fever-relieving oral liquid for children.
The invention provides a fingerprint detection method of a throat clearing and fever relieving oral preparation for children, which is characterized by comprising the following steps:
(1) preparation of control solutions
Weighing baicalin reference substance, adding methanol to obtain reference substance solution, weighing aesculetin reference substance, and adding 50% methanol to obtain reference substance solution;
(2) preparation of test solution
Weighing the oral liquid for clearing heat from throat and relieving fever of children, adding 70% ethanol for dilution, fixing the volume to scale, shaking up, filtering, and taking the subsequent filtrate, namely the test solution;
(3) the determination method comprises the following steps:
sucking the reference solution and the test solution, injecting into a high performance liquid chromatograph to obtain a chromatogram,
wherein, the chromatographic conditions of the high performance liquid chromatograph are as follows:
the chromatographic column uses octadecylsilane chemically bonded silica as a filler, mobile phase A is acetonitrile, mobile phase B is phosphoric acid solution, and gradient elution is carried out, wherein the gradient procedure is as follows:
the flow rate is 0.8-1.2 ml/min; the detection wavelength is 192-230 nm; the column temperature is 25-40 ℃.
In one embodiment of the assay method of the present invention, the mobile phase B is a 0.2% phosphoric acid solution.
In one embodiment of the assay method of the present invention, the concentration of baicalin in the control solution in step (1) is 0.2mg/ml, and the concentration of aesculetin control solution is 0.08 mg/ml.
In one embodiment of the assay method of the present invention, the pharmaceutical composition in step (2) is added with ethanol at a concentration such that 0.5ml to 2.0ml of the original drug solution is contained per 10ml of the diluted solution.
In one embodiment of the assay method of the present invention, the chromatographic column is a Phenomenex Gemini-C18 column.
In one embodiment of the assay method of the present invention, the column temperature in the chromatographic conditions is preferably 35 ℃.
In one embodiment of the assay method of the present invention, the flow rate of the mobile phase in the chromatographic conditions is preferably 1.2 ml/min.
In one embodiment of the assay method of the present invention, the detection wavelength in the chromatographic conditions is preferably 206 nm.
In a second aspect of the invention, a fingerprint spectrum establishment method of a children oral preparation for clearing heat from throat is provided, which comprises the following steps:
(1) preparation of control solutions
Weighing baicalin reference substance, adding methanol to obtain reference substance solution, weighing aesculetin reference substance, and adding 50% methanol to obtain reference substance solution;
(2) preparation of test solution
Weighing the oral liquid for clearing heat from throat and relieving fever of children, adding 70% ethanol for dilution, fixing the volume to scale, shaking up, filtering, and taking the subsequent filtrate, namely the test solution;
(3) the determination method comprises the following steps:
sucking the reference solution and the test solution, injecting into a high performance liquid chromatograph to obtain a chromatogram,
wherein, the chromatographic conditions of the high performance liquid chromatograph are as follows:
the chromatographic column uses octadecylsilane chemically bonded silica as a filler, mobile phase A is acetonitrile, mobile phase B is phosphoric acid solution, and gradient elution is carried out, wherein the gradient procedure is as follows:
the flow rate is 0.8-1.2 ml/min; the detection wavelength is 192-230 nm; the column temperature is 25-40 ℃;
(4) generating a comparison fingerprint: selecting fingerprint of multiple batches of oral liquid for clearing heat from throat and relieving fever of children, taking baicalin as a reference peak to obtain 10 common peak fingerprints, and generating a comparison fingerprint by using a median calculation method; wherein, the relative retention time of the fingerprint is as follows:
in one embodiment of the fingerprint establishment method of the present invention, the mobile phase B in the step (3) is a 0.2% phosphoric acid solution.
In one embodiment of the fingerprint establishment method of the present invention, the concentration of baicalin in the control solution in step (1) is 0.2mg/ml, and the concentration of the aesculetin control solution is 0.08 mg/ml.
In one embodiment of the fingerprint spectrum establishing method of the present invention, 70% ethanol is added to the pharmaceutical composition in the step (2), so that each 10ml of the diluted solution contains 0.5ml to 2.0ml of the original liquid medicine.
In one embodiment of the fingerprint spectrum establishing method, the chromatographic column is a Phenomenex Gemini-C18 column, the column temperature is preferably 35 ℃, the flow rate of the mobile phase is preferably 1.2ml/min, and the detection wavelength is preferably 206 nm.
The detection conditions selected by the invention are repeatedly compared and verified by experiments, the accuracy is high, and semi-quantitative determination can be realized. The HPLC fingerprint is adopted for the first time to detect the variety, and the main peak of the spectrum is well separated by optimizing methods such as a mobile phase, gradient elution conditions and the like, and the peak shape of a contrast peak is obviously improved, so that the overall quality of the product can be comprehensively controlled, and the requirement of industrial mass production is met.
The invention has the advantages of
(1) The method provided by the invention establishes the children's oral liquid fingerprint for clearing heat from throat, combines the identification of two main components of baicalin and aesculetin, can effectively represent the quality of the product, and is beneficial to comprehensively controlling the quality of the medicine.
(2) The fingerprint emphasizes the front-back sequence and the mutual relation of all the formed fingerprint characteristic peaks and the overall facial features, thereby not only avoiding the one-sidedness of judging the quality of the oral liquid for clearing heat from throat and relieving fever of children due to the determination of individual chemical components, but also reducing the possibility of manual treatment for reaching the quality standard.
(3) The method has the advantages of simplicity, convenience, stability, high precision and good reproducibility.
Drawings
Figure 1 shows the fingerprint of the oral liquid for clearing heat from throat and clearing heat of children, which is established by high performance liquid chromatography analysis with acetonitrile as a mobile phase A and water as a mobile phase B.
Figure 2 shows the fingerprint of the oral liquid for clearing heat from throat and clearing heat for children, which is established by high performance liquid chromatography analysis with acetonitrile as a mobile phase A and 0.2% phosphoric acid solution as a mobile phase B.
Figure 3 shows the fingerprint of the oral liquid for clearing heat from throat and clearing heat of children, which is established by high performance liquid chromatography analysis with acetonitrile as a mobile phase A and 0.05 percent trifluoroacetic acid solution as a mobile phase B.
Figure 4 shows the fingerprint of the oral liquid for clearing heat from throat and clearing heat of children, which is established by high performance liquid chromatography analysis with methanol as a mobile phase A and 0.2% phosphoric acid solution as a mobile phase B.
FIG. 5 shows finger print-contrast print of oral liquid for clearing heat from throat and relieving fever for children, wherein the peak 4 is aesculetin, and the peak 8(S) is baicalin.
Detailed Description
Example 1 fingerprint establishment
1.1 instruments
LC-20AT Shimadzu high performance liquid chromatograph, Agilent 1200 high performance liquid chromatograph, Agilent 1260 high performance liquid chromatograph, chromatographic column (Agilent extended C18, 4.6 × 250mm, 5 μm, S/N: USHR007237), chromatographic column (Phenomenex Gemini-C18, 4.6 × 250mm, 5 μm, S/N:540529-7), chromatographic column (Phenomenex Gemini-C18, 4.6 × 250mm, 5 μm, S/N:540542-28), chromatographic column (Phenomenex Gemini-C18, 4.6 × 250mm, 5 μm, S/N:540608-4), chromatographic column (Kromasil 100-5C18, 4.6 × 250mm, 5 μm, S/N: E59208).
1.2 reagents
Formic acid (AR, national drug group chemical Co., Ltd., lot No. 20130318), phosphoric acid (AR, Beijing chemical Co., Ltd., lot No. 20101107), acetic acid (AR, Fuchen chemical Co., Ltd., Tianjin, lot No. 20130702), ethanol (AR, national drug group chemical Co., Ltd., lot No. 20130801), pure water (Hangzhou Waha group Co., Ltd., lot No. 20130520), acetonitrile (HPLC, SIGMA-ALDRICH, lot No. S92130), methanol (HPLC, Honeywell, lot No. 20130729).
Comparison products: aesculetin reference substance (China food and drug testing institute, batch No. 110741-200506), baicalin reference substance (China food and drug testing institute, batch No. 110715-200815).
1.3 test samples
The oral liquid has the batches of 130201, 130202 and 130203, and the oral liquid has the batches of 18002B, 28008, 28001B, 28019B, 28024B, 38003B, 08004B, 98015B and 08005B, which are produced by the Sichuan pharmaceutical company Limited in the Asia pharmaceutical industry.
1.4 test methods
High performance liquid chromatography (China pharmacopoeia 2015 edition four parts general rules 0512).
Preparation of reference solutions: accurately weighing 10mg of baicalin reference substance, placing in a 50ml measuring flask, adding methanol to dissolve, fixing volume to scale, shaking, filtering, and collecting filtrate to obtain reference substance solution; accurately weighing aesculetin reference substance 10mg, placing in a 50ml measuring flask, adding 50% methanol for dissolving and fixing volume to scale, shaking, accurately weighing 10ml, placing in a 25ml measuring flask, adding 50% methanol for fixing volume to scale, shaking, filtering, and collecting filtrate.
Preparation of a test solution: precisely measuring 1ml of the oral liquid for clearing heat from throat of children, placing the oral liquid in a 10ml measuring flask, adding 70% ethanol for dilution, diluting to a constant volume, shaking up, filtering, and taking a subsequent filtrate, namely the test solution.
Chromatographic conditions are as follows: octadecylsilane chemically bonded silica is used as filler (chromatographic column: Phenomenex Gemini-C18, 250mm × 4.6mm, 5 μm); gradient elution was performed according to the gradient elution procedure of table 2 with acetonitrile as mobile phase a and 0.2% phosphoric acid solution as mobile phase B:
TABLE 2 mobile phase gradient elution conditions
The flow rate is 1.2 ml/min; the detection wavelength is 206 nm; the column temperature was 35 ℃. The theoretical plate number is not less than 10000 calculated according to baicalin peak.
The determination method comprises the following steps: precisely absorbing 10 μ l of reference solution and sample solution respectively, injecting into high performance liquid chromatograph, and recording chromatogram within 80 min.
Example 2 inspection of optimal chromatographic conditions
The apparatus and reagents were the same as in example 1.
2.1 selection of detection wavelength
The control solution and the test solution were prepared as in example 1.
192nm, 206nm, 230nm, 254nm, 278nm, 353nm and 450nm are respectively set for analyzing the finger print of the oral liquid for clearing heat from throat and clearing heat of children, wherein the chromatographic peak separation is better under the wavelength of 192 nm-230 nm. Compared with the spectra under other 6 wavelengths, the 206nm fingerprint spectrum has the largest information quantity, can more fully embody the chemical components of the product, and has small baseline drift, so 206nm is selected as the optimal detection wavelength.
2.2 preparation of test solutions
The reference solution preparation, chromatographic conditions and assay were as in example 1.
Taking the stock solution of the product, 10 times solution diluted by water, 10 times solution diluted by 70% ethanol, and 25 times solution diluted by 70% ethanol, and respectively preparing into test solution.
The main chromatographic peaks in the chromatograms obtained by the different dilution methods are integrated, and the separation degree and the separation effect are compared, and the result shows that the separation degree and the integral separation effect of the sample after being diluted by 10 times by 70% of ethanol are superior to those of other treatment methods, so that the preparation method of the oral liquid for clearing and relieving heat of throat of children is determined as follows: taking 1ml of the product, placing in a 10ml measuring flask, adding 70% ethanol for dilution, fixing the volume to the scale, and shaking up to obtain the product.
2.3 selection of chromatography columns
The packing and packing pattern of the column may have an effect on the separation of the pharmaceutical chemicals. The inventors examined three different brands of C18 columns (5 μm, 250X 4.6mm) from Agilent, Kromasil and Phenomenex. Other chromatographic conditions were the same as in example 1. The separation effect and the peak pattern of the Phenomenex Gemini-C18 column are better than those of the other two columns, so the Phenomenex Gemini-C18 chromatographic column is preferentially selected.
2.4 selection of column temperature
In the chromatographic fingerprint detection, the column temperature usually affects the separation effect, and the inventor analyzes the fingerprint of the children oral liquid for clearing heat from throat and clearing heat to be tested under the four column temperature conditions of 25 ℃, 30 ℃, 35 ℃ and 40 ℃. The result shows that the fingerprint of the sample can be obtained under the condition of four column temperatures, wherein the separation effect of each chromatographic peak is better at 35 ℃, so the 35 ℃ is optimally selected as the detection column temperature.
2.5 investigation of flow Rate
In the chromatographic fingerprint detection, the flow rate can influence the separation effect to a certain extent, and the fingerprint of the oral liquid test solution for clearing heat from throat of children is analyzed under the conditions of three flow rates of 0.8ml/min, 1.0ml/min and 1.2 ml/min. The result shows that the fingerprint of the sample can be obtained under the condition of three flow rates, wherein the separation effect of each chromatographic peak is better when the flow rate is 1.2ml/min, and 1.2ml/min is selected as the most suitable flow rate.
2.6 chromatographic information acquisition time determination
In order to investigate the chromatographic information acquisition time, the product is detected under the following chromatographic conditions: a chromatographic column: phenomenex Gemini-C18 column (250 mm. times.4.6 mm, 5 μm); detection wavelength: 206 nm; column temperature: 35 ℃; flow rate: 1.2 ml/min; sample introduction amount: 10 mu l of the mixture; mobile phase: acetonitrile was used as mobile phase a and 0.2% phosphoric acid solution was used as mobile phase B, and gradient elution was performed as shown in table 3. Collecting chromatographic information within 120 min.
TABLE 3 mobile phase gradient elution conditions
Since there was no absorption peak after 80min, chromatographic information was collected over 80 min.
Example 3 fingerprint method verification
According to the pre-experimental results, different mobile phase systems are investigated, and the chromatographic conditions with the most abundant chromatographic peak information and the best separation degree are screened.
Preparation of a test solution: taking 1ml of the product, placing the product in a 10ml measuring flask, adding 70% ethanol for dilution to a constant volume to scale, shaking up, filtering, and taking a subsequent filtrate as a test solution.
Chromatographic conditions are as follows: 4 systems of acetonitrile-water solution, acetonitrile-0.2% phosphoric acid solution, acetonitrile-0.05% trifluoroacetic acid solution and methanol-0.2% phosphoric acid solution are examined by adopting a Phenomenex Gemini-C18 chromatographic column for testing
(1) The system 1: acetonitrile as mobile phase A and water as mobile phase B, and referring to figure 1
TABLE 4 mobile phase gradient elution conditions
(2) And (3) system 2: acetonitrile as mobile phase A, and 0.2% phosphoric acid solution as mobile phase B, see figure 2
TABLE 5 mobile phase gradient elution conditions
(3) And (3) system: acetonitrile as mobile phase A, and 0.05% trifluoroacetic acid solution as mobile phase B, see figure 3
TABLE 6 mobile phase gradient elution conditions
(4) And (4) system: methanol as mobile phase A and 0.2% phosphoric acid solution as mobile phase B, see figure 4
TABLE 7 mobile phase gradient elution conditions
The atlas of the system can show that the acetonitrile-0.2% phosphoric acid solution system has larger peak information amount and stable baseline, and can better reflect the whole quality information of the oral liquid for clearing heat from throat and clearing heat of children; and the other three systems have serious baseline drift or less peak appearance, and cannot comprehensively express the quality information of the product, so that the acetonitrile-0.2 percent phosphoric acid solution system is determined to be selected as the mobile phase condition of the fingerprint spectrum of the product through screening.
Example 4 fingerprint methodological verification
4.1 blank test
70% ethanol is directly injected according to the chromatographic conditions of the example 1, and a chromatogram map of 80 minutes is recorded, and the result shows that the system has no residue and interference.
4.2 System suitability test
Taking the product, operating according to the method of example 1, preparing a test solution, directly injecting sample for 6 times, taking baicalin with peak 8 as a reference peak, and calculating the relative retention time and the relative peak area of each main chromatographic peak. The result shows that the relative retention time of 10 common peaks is stable, the RSD is less than 2 percent, the relative peak area is relatively constant, and the RSD is less than 5 percent; the system applicability of the method is good. The results are shown in tables 8 and 9.
TABLE 8 relative retention times for precision tests
TABLE 9 relative peak area for precision test
4.3 repeatability test
The experimenter A takes the same batch of the product, prepares 6 parts of test solution according to the operation of the example 1, detects, and calculates the relative retention time and the relative peak area of each main chromatographic peak by taking the baicalin with the No. 8 peak as a reference peak. The result shows that the relative retention time of 10 common peaks is stable, the RSD is less than 2 percent, the relative peak area is relatively constant, and the RSD is less than 5 percent; the method is shown to have good repeatability. The results are shown in tables 10 and 11.
TABLE 10 relative retention times for reproducibility tests
TABLE 11 relative peak area for repeatability tests
4.4 stability test
Taking the product, operating according to the method of example 1, injecting sample for detection at 0, 3, 6, 9, 12, 18, 24, 30 and 36 hours of sample preparation respectively, taking baicalin of No. 8 peak as a reference peak, and calculating the relative retention time and the relative peak area of each main chromatographic peak. The result shows that the relative retention time of 10 common peaks is stable, the RSD is less than 2 percent, the relative peak area is relatively constant, and the RSD is less than 5 percent; the test solution of the product is stable within 36 hours. The results are shown in tables 12 and 13.
TABLE 12 stability test relative Retention time
TABLE 13 relative peak areas for stability testing
4.5 durability test
4.5.1 durability of different instruments
The product is taken and operated according to the method of the embodiment 1, instruments of different models are used for detection, the spectrum of the Agilent 1200 instrument is used as a reference, the similarity of the spectrums between different instruments is calculated by using the reference spectrum, the result shows that the measurement results between different instruments are approximately the same, the similarity of the fingerprint spectrums is greater than 0.99, and the influence of the model of the high performance liquid chromatograph on the fingerprint method is small. The results are shown in Table 14.
TABLE 14 Instrument durability test results
4.5.2 durability of chromatography columns in different batches
The product is taken and operated according to the method of the embodiment 1, different batches of chromatographic columns of the same model are used for detection, the spectrum of one batch of chromatographic column (the serial number 540529-7) is taken as a reference, the similarity is calculated, the result shows that the measurement results of the chromatographic columns of different batches are approximately the same, the similarity of the fingerprint spectrums is more than 0.99, and the batch of the chromatographic columns has little influence on the method. The results are shown in Table 15.
TABLE 15 chromatographic column durability test results
Example 5 establishment of a comparison fingerprint
For the oral liquid for clearing heat from throat and clearing heat of children with the batch numbers of 130201, 130202 and 130203 produced by the pharmaceutical company Limited in Sichuan of Yabao pharmaceutical industry and the oral liquid for clearing heat from throat and clearing heat of children with the batch numbers of 18002B, 28008, 28001B, 28019B, 28024B, 38003B, 08004B, 98015B and 08005B produced by the pharmaceutical company Limited in Beijing of Yabao medicine, 12 batches of samples are counted, respectively detected according to the established fingerprint spectrum measuring method, and a comparison fingerprint spectrum is generated by adopting the matching of the similarity evaluation system of traditional Chinese medicine chromatography fingerprint spectrum (version 2012.0) of the Committee of the national pharmacopoeia, see the attached figure 5.
Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that changes, modifications, substitutions and alterations can be made in these embodiments without departing from the principles and spirit of the invention, the scope of which is defined in the appended claims and their equivalents.
Claims (10)
1. A fingerprint detection method for a throat-clearing and fever-relieving oral preparation for children is characterized by comprising the following steps: (1) preparation of control solutions
Weighing baicalin reference substance, adding methanol to obtain reference substance solution, weighing aesculetin reference substance, and adding 50% methanol to obtain reference substance solution;
(2) preparation of test solution
Weighing the oral liquid for clearing heat from throat and relieving fever of children, adding 70% ethanol for dilution, fixing the volume to scale, shaking up, filtering, and taking the subsequent filtrate, namely the test solution;
(3) the determination method comprises the following steps:
sucking the reference solution and the test solution, injecting into a high performance liquid chromatograph to obtain a chromatogram,
wherein, the chromatographic conditions of the high performance liquid chromatograph are as follows:
the chromatographic column uses octadecylsilane chemically bonded silica as a filler, mobile phase A is acetonitrile, mobile phase B is phosphoric acid solution, and gradient elution is carried out, wherein the gradient procedure is as follows:
the flow rate is 0.8-1.2 ml/min; the detection wavelength is 192-230 nm; the column temperature is 25-40 ℃.
2. The fingerprint detection method according to claim 1, wherein the mobile phase B in step (3) is a 0.2% phosphoric acid solution.
3. The fingerprint detection method according to claim 1, wherein the concentration of baicalin in the control solution in step (1) is 0.2mg/ml, and the concentration of aesculetin control solution is 0.08 mg/ml.
4. The fingerprint detection method according to claim 1, wherein 70% ethanol is added to the oral liquid for clearing heat from the throat and clearing heat of children in the step (2), so that 0.5ml to 2.0ml of original liquid medicine is contained in each 10ml of diluted solution.
5. The fingerprint detection method of claim 1, wherein the chromatographic column is a phenomenex gemini-C18 column, the column temperature is 35 ℃, the flow rate of the mobile phase is 1.2ml/min, and the detection wavelength is 206 nm.
6. A fingerprint spectrum establishment method of a children oral preparation for clearing heat from throat comprises the following steps:
(1) preparation of control solutions
Weighing baicalin reference substance, adding methanol to obtain reference substance solution, weighing aesculetin reference substance, and adding 50% methanol to obtain reference substance solution;
(2) preparation of test solution
Weighing the oral liquid for clearing heat from throat and relieving fever of children, adding 70% ethanol for dilution, fixing the volume to scale, shaking up, filtering, and taking the subsequent filtrate, namely the test solution;
(3) the determination method comprises the following steps:
sucking the reference solution and the test solution, injecting into a high performance liquid chromatograph to obtain a chromatogram,
wherein, the chromatographic conditions of the high performance liquid chromatograph are as follows:
the chromatographic column uses octadecylsilane chemically bonded silica as a filler, mobile phase A is acetonitrile, mobile phase B is phosphoric acid solution, and gradient elution is carried out, wherein the gradient procedure is as follows:
the flow rate is 0.8-1.2 ml/min; the detection wavelength is 192-230 nm; the column temperature is 25-40 ℃;
(4) generating a comparison fingerprint: selecting fingerprint of multiple batches of oral liquid for clearing heat from throat and relieving fever of children, taking baicalin as a reference peak to obtain 10 common peak fingerprints, and generating a comparison fingerprint by using a median calculation method; wherein the relative retention time of the fingerprint is
7. The fingerprint spectrum establishing method according to claim 6, wherein the mobile phase B in the step (3) is 0.2% phosphoric acid solution.
8. The fingerprint spectrum establishing method according to claim 6, wherein the concentration of baicalin in the reference solution in step (1) is 0.2mg/ml, and the concentration of the aesculetin reference solution is 0.08 mg/ml.
9. The fingerprint spectrum establishing method according to claim 6, wherein 70% ethanol is added into the oral liquid for clearing heat from throat and clearing heat of children in the step (2), so that 0.5ml to 2.0ml of original liquid medicine is contained in each 10ml of diluted solution.
10. The fingerprint spectrum establishing method according to claim 6, wherein the chromatographic column is a Phenomenex Gemini-C18 column, the column temperature is 35 ℃, the flow rate of the mobile phase is 1.2ml/min, and the detection wavelength is 206 nm.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710358947.3A CN108956835B (en) | 2017-05-19 | 2017-05-19 | Fingerprint detection method of oral medicine for clearing heat from throat |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710358947.3A CN108956835B (en) | 2017-05-19 | 2017-05-19 | Fingerprint detection method of oral medicine for clearing heat from throat |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN108956835A CN108956835A (en) | 2018-12-07 |
| CN108956835B true CN108956835B (en) | 2022-02-25 |
Family
ID=64462748
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201710358947.3A Active CN108956835B (en) | 2017-05-19 | 2017-05-19 | Fingerprint detection method of oral medicine for clearing heat from throat |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN108956835B (en) |
Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114113437B (en) * | 2021-12-06 | 2024-04-12 | 苏州市药品检验检测研究中心 | HPLC fingerprint spectrum of oral liquid for clearing away heat and toxic materials and application thereof in quality control of oral liquid for clearing away heat and toxic materials |
| CN118275595A (en) * | 2024-05-29 | 2024-07-02 | 广州白云山和记黄埔中药有限公司 | Fingerprint spectrum and multi-component content determination method of nasopharynx toxicity-removing preparation |
Family Cites Families (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1296088C (en) * | 2004-04-30 | 2007-01-24 | 北京中医药大学药厂 | Medicine for treating acute pharyngitis of children and preparation method thereof |
| WO2006024545A1 (en) * | 2004-09-03 | 2006-03-09 | Stichting Voor De Technische Wetenschappen | Fused bicyclic natural compounds and their use as inhibitors of parp and parp-mediated inflammatory processes |
| CN101904948B (en) * | 2010-07-27 | 2011-09-28 | 广东众生药业股份有限公司 | Traditional Chinese medicine preparation of new Zhongsheng pill and preparation method thereof |
| CN105510488B (en) * | 2016-01-25 | 2017-10-03 | 亚宝药业集团股份有限公司 | A kind of finger-print and its quality determining method for relieving pain patch |
| CN105911161B (en) * | 2016-04-12 | 2018-03-27 | 吉林修正药业新药开发有限公司 | A kind of sulfathiazole HPLC fingerprint map construction methods |
-
2017
- 2017-05-19 CN CN201710358947.3A patent/CN108956835B/en active Active
Also Published As
| Publication number | Publication date |
|---|---|
| CN108956835A (en) | 2018-12-07 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN111337589B (en) | Method for establishing orange-shell mixture HPLC fingerprint spectrum | |
| CN111624271B (en) | Liquid chromatography method for detecting corresponding substance of peony and licorice decoction, standard fingerprint spectrum and application | |
| CN109613134B (en) | Construction method and application of UPLC fingerprint spectrum of cortex mori medicinal material | |
| WO2022116972A1 (en) | Method for detecting monosaccharide and oligosaccharide content and fingerprint specturm of compound sophora flavescens injection | |
| CN108956835B (en) | Fingerprint detection method of oral medicine for clearing heat from throat | |
| CN109932441B (en) | Method for establishing HPLC fingerprint of Shuganning injection | |
| CN107576743B (en) | Method for establishing fingerprint of Bining spray and fingerprint thereof | |
| CN110441413B (en) | Construction method and detection method of HPLC fingerprint of Qianbai rhinitis tablets | |
| CN102068553B (en) | Method for constructing high performance liquid chromatographic (HPLC) fingerprint of Mammary lump preparation arising from qi stagnation and blood stasis | |
| CN114216980B (en) | Method for establishing HPLC-ELSD fingerprint of starwort root | |
| CN114965802B (en) | Quality control method of climacteric syndrome relieving tablet | |
| CN113189248B (en) | HPLC fingerprint construction and detection method of Yinhua Miyanling tablets | |
| CN108548885A (en) | The method that two-dimensional liquid chromatography detects compound Nanxing pain paste | |
| CN113917009A (en) | Construction method and application of bupleurum chinense non-saponin component HPLC fingerprint | |
| CN109374771B (en) | Fingerprint spectrum detection method of snow pear syrup | |
| CN114152687A (en) | Fingerprint construction method and application of traditional Chinese medicine compound containing lotus seeds | |
| CN100416271C (en) | Effective liquid phase chromatography for measuring alkali content of dita leaves | |
| CN114577918A (en) | Ultra-high performance liquid chromatography detection method for active ingredient content and fingerprint spectrum of compound sophora flavescens injection | |
| CN111351883A (en) | Method for measuring content of rutin in sophora and scutellaria ointment | |
| CN106404929B (en) | A kind of component analyzing method of Radix Et Caulis Acanthopanacis Senticosi injection | |
| CN110907574B (en) | Quality control method, quality control spectrogram and construction method of traditional Chinese medicine composition | |
| CN111474276B (en) | Quality control method of yang invigorating tablet preparation | |
| CN109541099B (en) | Method for identifying white peony root and rhizome or extract thereof | |
| CN108445104B (en) | Fingerprint spectrum of rabdosia japonica, and establishment method and application thereof | |
| CN113219095B (en) | Construction method of fingerprint of Fukean tablet and fingerprint thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |