CN115047116A - Quality detection method for internally eliminating scrofula tablets - Google Patents
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Abstract
The invention discloses a quality detection method of internally-cured scrofula tablets, which comprises a fingerprint detection method and a content determination method; the fingerprint detection method comprises the following steps: step 1, preparing a test solution for internally eliminating scrofula tablets; step 2, preparing a mixed reference substance solution; step 3, precisely absorbing the solutions of the test sample and the reference substance respectively, injecting the solutions into a liquid chromatogram instrument, and recording the chromatograms; and 4, exporting a fingerprint instrument of the internal scrofula eliminating tablet, importing the fingerprint instrument into a traditional Chinese medicine chromatographic fingerprint similarity evaluation system, carrying out peak point correction on the fingerprint of the sample by a median method, and carrying out data matching analysis to generate a common pattern diagram. The fingerprint spectrum and the content measuring method of the internally-cured scrofula tablet established by the invention can comprehensively and objectively evaluate the quality of the internally-cured scrofula tablet. The method has the advantages of good stability, high accuracy, good repeatability, high precision and the like.
Description
Technical Field
The invention belongs to the technical field of detection of traditional Chinese medicine preparations, and particularly relates to a quality detection method of an internal scrofula eliminating tablet.
Background
The internal eliminating scrofula tablet is a prescription preparation prepared from 15 medicinal materials of figwort root, oyster shell (calcined), selfheal, trichosanthes root, codonopsis pilosula, astragalus root, white mustard seed, common burreed rhizome, zedoary, red paeony root, thunberg fritillary bulb, seaweed, pangolin (processed), goat milk and earthworm, is a hospital preparation (Su medicine Z04000312) of a traditional Chinese and western medicine combined hospital in Nanjing city, and is different from a formula recorded in Chinese pharmacopoeia. Clinically, the internal elimination scrofula tablet and the antituberculous first-line treatment medicament isoniazide are combined to treat pulmonary tuberculosis and multidrug-resistant tuberculosis, and the large side effects of liver function damage and gastrointestinal reaction are reduced; the western medicine is combined with the internal scrofula eliminating pill to treat the tuberculosis of the cervical lymph nodes, so that the clinical curative effect is improved, the sterilization effect is improved, and the prognosis is improved; the internal eliminating scrofula pill is combined with medicines such as levothyroxine sodium and the like to treat benign thyroid nodules, so that adverse reactions of western medicines are reduced, and the treatment compliance of patients is enhanced.
In the prior art, rosmarinic acid is measured by HPLC to control the quality of the internal scrofula tablets, an RP-HPLC method is used to measure the content of naringin in the tablets, and HPLC is used to measure the content of selfheal rosmarinic acid and figwort harpagoside in the prescription. The invention aims to establish an HPLC fingerprint method and multi-component content analysis of the internal scrofula tablet, simultaneously measure the content of 4 main components (paeoniflorin, benzoic acid, calycosin glucoside and rosmarinic acid), form fuzzy and accurate quality combined control of the fingerprint and the multi-component content measurement, further improve the quality control standard of the internal scrofula tablet, and provide a reference basis for evaluating and controlling the quality of the internal scrofula tablet.
Disclosure of Invention
The purpose of the invention is as follows: the invention aims to overcome the defects of the prior art and develop a quality detection method for internally eliminating scrofula tablets.
The technical scheme is as follows: in order to realize the purpose, the invention adopts the technical scheme that:
a quality detection method for internally eliminating scrofula tablets comprises the following steps:
step 1: preparation of internal eliminating scrofula tablet test solution
Taking a proper amount of internal scrofula tablets of different batches, powdering, sieving powder, taking a proper amount of powder, precisely adding methanol, carrying out ultrasonic treatment, filtering, taking subsequent filtrate, evaporating to dryness in a water bath kettle evaporating dish, dissolving the residue in methanol, placing in a volumetric flask, fixing the volume to a scale mark with methanol, shaking up, centrifuging, and filtering through a 0.45 mu m microporous membrane to obtain the medicine;
step 2: preparation of Mixed control solutions
Accurately weighing p-hydroxybenzoic acid, paeoniflorin, benzoic acid, calycosin glucoside, isoquercetin, rosmarinic acid, cinnamic acid, quercetin, and formononetin reference substances respectively to obtain single reference substance solution; precisely sucking the 9 single reference substances, placing in a volumetric flask, adding methanol to constant volume to scale mark, and filtering with 0.45 μm microporous membrane to obtain mixed reference substances;
Respectively sucking the test solution in the step 1 and the mixed reference solution in the step 2, and injecting the solutions into a high performance liquid chromatograph to obtain a chromatogram of the test solution and a chromatogram of the mixed reference solution;
Preferably, in the above-described method for detecting the quality of internal scrofula eliminating tablet, step 1, the method for preparing a test sample solution of an internal scrofula eliminating tablet comprises:
grinding different batches of internal scrofula tablets into powder, sieving the powder with a 60-mesh sieve, taking 1g of the powder, placing the powder in a conical flask with a 50mL mouth grinder, precisely adding 30mL of methanol, sealing, carrying out ultrasonic treatment for 30min, filtering, taking subsequent filtrate, evaporating the filtrate in an evaporating dish at 70 ℃ in a water bath kettle, dissolving the residue in methanol, placing the dissolved residue in a 5mL volumetric flask, adding methanol to a constant volume to reach a scale mark, shaking uniformly, and metering to 12000 r.min -1 Centrifuging for 10min, and filtering with 0.45 μm microporous membrane.
Preferably, in the above method for detecting the quality of internal scrofula tablet, the method for preparing the mixed reference solution in step 2 comprises:
precisely weighing p-hydroxybenzoic acid, paeoniflorin, benzoic acid, calycosin glucoside, isoquercitin, rosmarinic acid, cinnamic acid, quercetin, and formononetin reference substances to obtain reference substances with concentration of 0.882 g.L respectively -1 、0.885g·L -1 、0.880g·L -1 、1.442g·L -1 、0.886g·L -1 、1.122g·L -1 、2.414g·L -1 、0.703g·L -1 、0.126g·L -1 A single control solution of (a); precisely sucking 1mL of each of the 9 single reference substance solutions, placing in a 20mL volumetric flask, adding methanol to a constant volume to scale mark, and filtering with 0.45 μm microporous membrane to obtain a mixed reference substance for use.
Preferably, in the method for detecting the quality of the internally scrofula tablet, the chromatographic condition in the step 3 is YMC-Pack ODS-AC 18 A chromatographic column with the specification of 4.6mm multiplied by 250mm and 5 mu m; mobile phase: the phase A is acetonitrile-phase B is 0.1 percent phosphoric acid water solution, and gradient elution is carried out; flow rate: 1.0 mL/min -1 (ii) a The column temperature is 35 ℃; detection wavelength: 230 nm; sample introduction amount: 10 μ L.
Preferably, in the method for detecting the quality of the internally scrofula tablet, the gradient elution procedure in step 3 is as follows: 0-30 min, 5% A; 30-105 min, 5% -30% A; 105-175 min, 30% -100% A; 175-180 min, 100% -5% A; 180-185 min, 5% A.
The quality detection method of the internally scrofula tablet comprises the step 5 of comparing 14 common peaks of a fingerprint, wherein p-hydroxybenzoic acid is a peak No. 1, the retention time is 24.415min, paeoniflorin is a peak No. 4, the retention time is 61.410min, benzoic acid is a peak No. 5, the retention time is 66.719min, calycosin glucoside is a peak No. 6, the retention time is 71.745min, isoquercitin is a peak No. 7, the retention time is 74.833min, rosmarinic acid is a peak No. 9, the retention time is 85.991min, cinnamic acid is a peak No. 11, the retention time is 95.064min, quercetin is a peak No. 12, the retention time is 98.040min, formononetin is a peak No. 13, and the retention time is 98.040 min.
Optimization of fingerprint detection conditions
1. Selection of detection wavelength
The sample solution is scanned at full wavelength, chromatograms at 210nm, 230nm, 254nm, 280nm, 330nm and 350nm are compared, and the target component is found to have good absorption at about 230nm and good peak shape. 230nm was chosen as the detection wavelength, (e.g., chromatograms at different wavelengths in FIGS. 5 and 6).
2. Selection of mobile phase
The research examines different mobile phases (acetonitrile-0.2% phosphoric acid aqueous solution, methanol-0.1% phosphoric acid aqueous solution and acetonitrile-0.1% formic acid aqueous solution), and as a result, the acetonitrile-0.1% phosphoric acid aqueous system is adopted, the separation effect and the peak shape are good, the base line is relatively stable, and therefore, the acetonitrile-0.1% phosphoric acid aqueous solution is selected as the mobile phase.
3. Screening by gradient elution
The invention screens the following 4 different gradient elution modes, and finally obtains the optimal gradient elution mode of the invention.
Elution gradient one: the chromatogram is shown in FIG. 7.
And (3) elution gradient II: the chromatogram is shown in FIG. 8.
| Time (min) | Volume concentration of acetonitrile | 0.1% volume concentration of phosphoric acid |
| 0 | 5％ | 95％ |
| 30 | 5％ | 95％ |
| 105 | 30％ | 70％ |
| 130 | 100％ | 0％ |
| 135 | 5％ | 95％ |
| 140 | 5％ | 95％ |
Elution gradient three: the chromatogram is shown in FIG. 9.
| Time (min) | Volume concentration of acetonitrile | 0.1% phosphoric acid aqueous |
| 0 | 5％ | 95％ |
| 30 | 5％ | 95％ |
| 105 | 30％ | 70％ |
| 150 | 100％ | 0％ |
| 155 | 5％ | 95％ |
| 160 | 5％ | 95％ |
Elution gradient four: the chromatogram is shown in FIG. 10.
| Time (min) | Volume concentration of acetonitrile | 0.1% phosphoric acid aqueous |
| 0 | 5％ | 95％ |
| 30 | 5％ | 95％ |
| 105 | 30％ | 70％ |
| 175 | 100％ | 0％ |
| 180 | 5％ | 95％ |
| 185 | 5％ | 95％ |
Wherein the elution gradient of four is the best, therefore, the elution gradient of four is the preferred elution condition of the invention.
3. Selection of column temperature and flow rate
This study examined various column temperatures (25, 30, 35 ℃) and flow rates (0.6, 0.8, 1.0 mL. min) -1 The effect on the chromatographic peak shows that the column temperature is 35 ℃ and the flow rate is 1.0 mL/min at the wavelength of 230nm -1 The obtained chromatographic peak has good peak shape, number, separation degree and the like.
The invention has the beneficial effects that:
(1) the invention screens out the best test sample and the best reference sample, screens out the best mobile phase composition and chromatographic conditions such as gradient elution mode through a large number of experiments, establishes the internal scrofula tablet fingerprint and content determination method, not only can effectively characterize the quality of the internal scrofula tablet, but also is beneficial to comprehensively monitoring the quality of the internal scrofula tablet.
(2) The method has the advantages of good stability, high precision, high accuracy, good reproducibility and the like. Can comprehensively, objectively and accurately evaluate and control the quality of the internal scrofula eliminating tablet.
Drawings
FIG. 1 is an HPLC chromatogram of a sample solution.
FIG. 2 is a mixed control HPLC chromatogram.
FIG. 3 shows HPLC finger prints of 10 different batches of internally-eliminated scrofula tablets.
FIG. 4 is an HPLC chromatogram of paeoniflorin, benzoic acid, calycosin glucoside, and rosmarinic acid control.
FIG. 5 shows chromatograms at different wavelengths of 210nm, 230nm and 254 nm.
FIG. 6 shows chromatograms at different wavelengths of 280nm, 330nm and 350 nm.
Figure 7 chromatogram of elution gradient one.
FIG. 8 chromatogram of elution gradient two.
Figure 9 chromatogram of elution gradient three.
Figure 10 chromatogram of elution gradient four.
Detailed Description
Embodiments of the present invention will be described in detail with reference to examples, in which specific conditions are not specified, according to conventional conditions or conditions recommended by manufacturers. The reagents or instruments used are not indicated by the manufacturer, and are all conventional products commercially available.
1 Instrument and reagent
1.1 instrument LC-20AT high performance liquid chromatograph (SPD-M20A diode array detector) (Shimadzu corporation, Japan); model BP211D ten thousandth precision analytical balance (beijing sidoris instruments systems ltd); one hundred thousand precision electronic analytical balance model FA1004N (beijing sidoris instruments systems ltd); HH-6 model digital display constant temperature water bath (Experimental instruments in south of the Yangtze river, Changzhou city); model SHZ-D (III) circulating water type vacuum pump (Ongyu Yunhua instruments, Inc.); KQ5200DE model digital control ultrasonic cleaner (frequency: 40KHz, ultrasonic Instrument Co., Ltd., Kunshan).
1.2 reagent paeoniflorin (batch No. PS000825, content > 98%) was purchased from Doctoria Biotech limited; cinnamic acid (batch No. 180607, > 98%) was purchased from Nanjing Bega Biotech Ltd); p-hydroxybenzoic acid (batch No. 100278-. Acetonitrile is chromatographically pure, water is purified water, and the rest reagents are analytically pure. Internal eliminating scrofula tablet 10 batches (homemade in the hospital combining traditional Chinese medicine and Western medicine in Nanjing, approval number: Su Yao Zhi character Z04000312, batch numbers 09180104, 09180304, 09180601, 09180709, 09180903, 09181104, 09180903, 09190104, 09190203 and 200301).
Example 1
1. A quality detection method for internally eliminating scrofula slices comprises the following steps:
step 1: preparation of internal eliminating scrofula tablet test solution
Taking a proper amount of the internal scrofula tablet samples of the 10 batches, pulverizing, sieving the powder with a 60-mesh sieve, taking 1g, placing the powder in a conical flask with a 50mL grinding mouth stopper, precisely adding 30mL of methanol, sealing, carrying out ultrasonic treatment for 30min, filtering, taking subsequent filtrate, evaporating the filtrate to dryness in an evaporation pan at 70 ℃, dissolving the residue with methanol, placing the solution in a 5mL volumetric flask, adding methanol to a constant volume to a scale line, shaking up, and centrifuging (12000 r.min) -1 And 10min), and filtering through a 0.45 mu m microporous filter membrane to obtain the product.
Step 2: preparation of control solutions
Accurately weighing p-hydroxybenzoic acid, paeoniflorin, benzoic acid, calycosin glucoside, isoquercitin, rosmarinic acid, cinnamic acid, quercetin, and formononetin reference to obtain a solution with a concentration of 0.882 g.L -1 、0.885g·L -1 、0.880g·L -1 、1.442g·L -1 、0.886g·L -1 、1.122g·L -1 、2.414g·L -1 、0.703g·L -1 、0.126g·L -1 And (4) a control solution. Precisely sucking 1mL of each of the 9 reference substances, placing in a 20mL volumetric flask, adding methanol to constant volume to scale lines, and filtering with a 0.45 μm microporous membrane to obtain a mixed reference substance. And (5) refrigerating for later use.
Respectively sucking the test solution in the step 1 and the mixed reference solution in the step 2, and injecting the solutions into a high performance liquid chromatograph to obtain a chromatogram of the test solution and a chromatogram of the mixed reference solution;
the chromatographic conditions are as follows: YMC-Pack ODS-A C18 column (4.6 mm. times.250 mm, 5 μm); mobile phase: acetonitrile (A) -0.1% phosphoric acid water solution (B), gradient elution (0-30 min, 5% A, 30-105 min, 5-30% A, 105-175 min, 30-100% A, 175-180 min, 100-5% A, 180-185 min, 5% A); flow rate: 1.0 mL/min -1 (ii) a The column temperature is 35 ℃; detection wavelength: 230 nm; sample introduction amount: 10 μ L.
TABLE 1 similarity between batches of samples and consensus patterns
The invention utilizes similarity software to analyze 10 batches of internal scrofula tablet fingerprint spectrums, the similarity of each batch of samples is over 0.66, and the similarity of each batch of internal scrofula tablets is good.
And marking chemical components of peaks in the comparison fingerprint spectrum according to the retention time of the mixed comparison product solution chromatogram. The fingerprint contains 14 common peaks, wherein p-hydroxybenzoic acid is peak 1, retention time is 24.415min, paeoniflorin is peak 4, retention time is 61.410min, benzoic acid is peak 5, retention time is 66.719min, calycosin glucoside is peak 6, retention time is 71.745min, isoquercitin is peak 7, retention time is 74.833min, rosmarinic acid is peak 9, retention time is 85.991min, cinnamic acid is peak 11, retention time is 95.064min, quercetin is peak 12, retention time is 98.040min, formononetin is peak 13, and retention time is 98.040 min.
2. HPLC fingerprinting methodology investigation
1. Precision test an internal scrofula tablet sample (lot 09190203) was selected, a test sample solution was prepared according to the method of step 1, samples were continuously injected 6 times according to the chromatographic conditions of step 3, 10 μ L of sample was injected each time, paeoniflorin was used as a reference peak, and retention time and peak area of 14 common peaks were recorded. The retention time RSD of all the common peaks is less than 1.5%, the RSD of the peak area is less than 3%, and the result shows that the precision of the instrument is good.
2. Stability test an internal scrofula sample (batch No. 09190203) is selected, a test sample solution is prepared according to the method in the step 1, sample injection detection is carried out according to the chromatographic condition in the step 3, sample injection is respectively carried out for 0, 4, 8, 16 and 24 hours, the sample injection amount is 10 mu L each time, paeoniflorin is taken as a reference peak, and the retention time and the peak area of 14 common peaks are recorded. The retention time RSD of all the common peaks is less than 0.7%, the RSD of the peak area is less than 3%, and the result shows that the stability of the test sample is good within 24 h.
3. In a repeatability test, 6 internal scrofula sample (batch number 09190203) are selected, a test sample solution is prepared according to the method in the step 1, sample injection detection is carried out according to the chromatographic condition in the step 3, the sample injection amount is 10 mu L each time, paeoniflorin is taken as a reference peak, and the retention time and the peak area of 14 common peaks are recorded. The retention time RSD of each component is less than 0.36%, the RSD of the peak area is less than 3%, and the result shows that the method has good repeatability.
EXAMPLE 2 quantitative analysis of Multi-index Components in internal scrofula eliminating tablet
1. Investigation of linear relationships
Accurately weighing penoniflorin, benzoic acid, calycosin glucoside, and rosmarinic acid as reference substances to obtain concentrations of 0.269 g.L -1 、0.039g·L -1 、0.013g·L -1 、0.034g·L -1 And (4) mixing the reference substance with the solution.
The injection volumes were 2. mu.L, 4. mu.L, 8. mu.L, 12. mu.L, 16. mu.L, 20. mu.L, as determined by the chromatographic conditions of step 3 of example 1. HPLC chromatograms of paeoniflorin, benzoic acid, calycosin glucoside and rosmarinic acid reference are shown in FIG. 4. Linear equations of the 4 components are established by respectively using the peak area (Y) and the sample injection amount (X) of the reference substance as shown in table 2, and the results show that the peak areas and the sample injection amounts of the 4 components have good linear relations.
TABLE 2 Standard Curve
| Name of reference substance | Linear equation of equations | r | Linear Range (μ g) |
| Paeoniflorin | Y＝249635X+21697 | 0.9996 | 0.134～2.69 |
| Benzoic acid | Y＝158962X+24295 | 0.9996 | 0.019～0.39 |
| Calycosin glucoside | Y＝27972X+5035.1 | 0.9996 | 0.006～0.13 |
| Rosmarinic acid | Y＝53944X-63950 | 0.9995 | 0.017～0.34 |
2. The content determination takes the above 10 internal scrofula samples, prepares the test sample solutions of 10 batches of samples according to the same method as the step 1 of the example 1, performs sample injection determination according to the chromatographic conditions of the step 3 of the example 1, records the peak areas of the test sample solutions, substitutes the standard curve equation, and calculates the content of each component, and the specific result is shown in the table 3.
Table 3 internal eliminating scrofula sample 4 kinds of component content determination results (n is 10)
3. Methodology investigation
3.1 precision test
Selecting an internal scrofula sample (batch No. 09190203), preparing a test solution according to the method in the step 1 of the example 1, carrying out sample injection detection according to the chromatographic condition in the step 3 of the example 1, carrying out sample injection for 6 times continuously, carrying out sample injection with the sample injection amount of 10 mu L each time, and recording a chromatogram. The results show that the instrument precision is good, and the RSD of the peak areas of paeoniflorin, benzoic acid, calycosin glucoside and rosmarinic acid are respectively 0.60%, 0.86%, 0.75% and 0.49%.
3.2 stability test
Selecting an internal scrofula sample (batch number 09190203), preparing a test solution according to the method in the step 1 of the example 1, carrying out sample injection detection according to the chromatographic conditions in the step 3 of the example 1, repeatedly carrying out sample injection with 0, 4, 8, 16 and 24 hours respectively, carrying out sample injection with the sample injection amount of 10 mu L each time, and recording a chromatogram. As a result, the peak areas RSD of paeoniflorin, benzoic acid, calycosin glucoside and rosmarinic acid are respectively 0.86%, 0.37%, 1.04% and 0.46%, which indicates that the test sample has good stability within 24 h.
3.3 repeatability test
Selecting 6 internal scrofula sample (batch number 09190203), preparing a test solution according to the method in the step 1 of the example 1, carrying out sample injection detection according to the chromatographic conditions in the step 3 of the example 1, wherein the sample injection amount is 10 mu L each time, and recording a chromatogram, wherein the results show that the peak areas RSD of paeoniflorin, benzoic acid, calycosin glucoside and rosmarinic acid are respectively 2.84%, 2.99%, 2.65% and 1.64%, which shows that the method has good repeatability.
3.4 sample application recovery test
Selecting an internal scrofula sample (batch number 09180104), precisely weighing 1g and 9 parts in total, placing the sample in a 50mL conical flask, preparing a test sample solution according to the method in the step 1 of the example 1, adding a mixed reference substance which is 80%, 100% and 120% of the paeoniflorin content in the sample into the final solid, dissolving the mixed reference substance, placing the mixed reference substance in a 5mL volumetric flask, and fixing the volume to the scale mark by using methanol. The sample was sampled and tested under the chromatographic conditions of step 3 of example 1, and the HPLC chromatograms thereof were measured to record the peak areas of the 4 components, respectively, and the contents of the 4 components and the sample recovery rates were calculated according to the standard curves of Table 2 above, and the average recovery rates of the 4 components were calculated to be 99.76%, 98.87%, 97.37%, 98.91%, and RSD were 2.72%, 1.57%, 2.58%, and 1.75%, respectively. The results of the determination of the recovery of 4 components of paeoniflorin, benzoic acid, calycosin glucoside and rosmarinic acid are shown in Table 4.
Table 4 recovery rate measurement results (n ═ 9)
The experimental results show that the fingerprint spectrum detection method and the content determination method for internally eliminating scrofula tablets established by the invention have good precision, stability and repeatability, can effectively characterize the quality of the internally eliminating scrofula tablets, and are beneficial to comprehensively monitoring the quality of the internally eliminating scrofula tablets.
Claims (8)
1. A quality detection method for internally eliminating scrofula slices is characterized by comprising the following steps:
step 1: preparation of internal eliminating scrofula tablet test solution
Taking a proper amount of internal scrofula tablets of different batches, powdering, sieving powder, taking a proper amount of powder, precisely adding methanol, carrying out ultrasonic treatment, filtering, taking subsequent filtrate, evaporating to dryness in a water bath kettle evaporating dish, dissolving the residue in methanol, placing in a volumetric flask, fixing the volume to a scale mark with methanol, shaking up, centrifuging, and filtering through a 0.45 mu m microporous membrane to obtain the medicine;
step 2: preparation of Mixed control solutions
Accurately weighing p-hydroxybenzoic acid, paeoniflorin, benzoic acid, calycosin glucoside, isoquercetin, rosmarinic acid, cinnamic acid, quercetin, and formononetin reference substances respectively to obtain single reference substance solution; precisely sucking the 9 single reference substances, placing in a volumetric flask, adding methanol to constant volume to scale mark, and filtering with 0.45 μm microporous membrane to obtain mixed reference substances;
step 3, chromatogram determination
Respectively sucking the test solution in the step 1 and the mixed reference solution in the step 2, and injecting the solutions into a high performance liquid chromatograph to obtain a chromatogram of the test solution and a chromatogram of the mixed reference solution;
step 4, exporting the fingerprint of the internal scrofula tablet test solution obtained in the step 3, and importing the fingerprint into a Chinese medicine chromatography fingerprint similarity evaluation system 2004A edition; carrying out peak point correction on the fingerprint of the sample by a median method, carrying out data matching analysis, generating a common mode diagram, establishing a reference fingerprint of a test sample solution of an internal scrofula elimination tablet, and calculating the relative retention time and the relative peak area of each common peak; and marking chemical components of peaks in the comparison fingerprint spectrum according to the retention time of the mixed comparison product solution chromatogram.
2. The method for detecting the quality of internally scrofula tablets according to claim 1, wherein said internally scrofula tablets are prepared by mixing,
step 1, the preparation method of the internal eliminating scrofula tablet test solution comprises the following steps:
grinding different batches of internal scrofula tablets into powder, sieving the powder with a 60-mesh sieve, taking 1g of the powder, placing the powder into a 50mL conical flask with a grinding mouth, precisely adding 30mL of methanol, sealing, carrying out ultrasonic treatment for 30min, filtering, taking subsequent filtrate, evaporating the filtrate to dryness in an evaporating dish at 70 ℃ in a water bath kettle, dissolving the residue in methanol, placing the solution into a 5mL volumetric flask, adding methanol to a constant volume to a scale line, shaking uniformly, and metering the volume to 12000 r.min -1 Centrifuging for 10min, and filtering with 0.45 μm microporous membrane.
3. The method for detecting the quality of internally scrofula tablets according to claim 1, wherein the method for preparing the mixed reference substance solution in the step 2 comprises the following steps:
precisely weighing p-hydroxybenzoic acid, paeoniflorin, benzoic acid, calycosin glucoside, isoquercitin, rosmarinic acid, cinnamic acid, quercetin, and formononetin reference substances to obtain reference substances with concentration of 0.882 g.L respectively -1 、0.885g·L -1 、0.880g·L -1 、1.442g·L -1 、0.886g·L -1 、1.122g·L -1 、2.414g·L -1 、0.703g·L -1 、0.126g·L -1 A single control solution of (a); precisely sucking 1mL of each of the 9 single reference substance solutions, placing in a 20mL volumetric flask, adding methanol to a constant volume to scale mark, and filtering with 0.45 μm microporous membrane to obtain a mixed reference substance for use.
4. The method for detecting the quality of internally scrofula tablets as claimed in claim 1, wherein said method comprisesIn step 3, the chromatographic conditions are YMC-PackODS-AC 18 A chromatographic column with the specification of 4.6mm multiplied by 250mm and 5 mu m; mobile phase: the phase A is acetonitrile-phase B is 0.1 percent phosphoric acid water solution, and gradient elution is carried out; flow rate: 1.0 mL/min -1 (ii) a The column temperature is 35 ℃; detection wavelength: 230 nm; sample introduction amount: 10 μ L.
5. The method for detecting the quality of internally scrofula tablets according to claim 4, wherein the gradient elution procedure of step 3 is as follows: 0-30 min, 5% A; 30-105 min, 5% -30% A; 105-175 min, 30% -100% A; 175-180 min, 100% -5% A; 180-185 min, 5% A.
6. The method of claim 1, wherein the fingerprint spectrum of step 5 contains 14 common peaks, wherein p-hydroxybenzoic acid is peak 1, the retention time is 24.415min, paeoniflorin is peak 4, the retention time is 61.410min, benzoic acid is peak 5, the retention time is 66.719min, calycosin glucoside is peak 6, the retention time is 71.745min, isoquercitin is peak 7, the retention time is 74.833min, rosmarinic acid is peak 9, the retention time is 85.991min, cinnamic acid is peak 11, the retention time is 95.064min, quercetin is peak 12, the retention time is 98.040min, formononetin is peak 13, and the retention time is 98.040 min.
7. The method for detecting the quality of internally scrofula tablets according to claim 1, further comprising the following steps:
establishment of standard curve equation
Accurately weighing penoniflorin, benzoic acid, calycosin glucoside and rosmarinic acid reference substance to obtain concentrations of 0.269 g.L -1 、0.039g·L -1 、0.013g·L -1 、0.034g·L -1 A control mixed solution;
respectively injecting 2 muL, 4 muL, 8 muL, 12 muL, 16 muL and 20 muL of reference substance mixed solution, and establishing a standard curve equation of 4 components according to the peak area Y and the sample injection amount X of the reference substance;
Substituting the peak area of the chromatogram of the test solution into a standard curve equation, and calculating the corresponding content of the effective components in the test.
8. The method for detecting the quality of internally-cured scrofula tablet according to any one of claims 1 to 7, wherein the internally-cured scrofula tablet is prepared from 15 medicinal materials including radix scrophulariae, calcined oyster shell, selfheal, radix trichosanthis, radix codonopsis pilosulae, radix astragali, semen brassicae, rhizoma sparganii, rhizoma zedoariae, radix paeoniae rubra, thunberg fritillary bulb, seaweed, squama manis preparata, goat milk and earthworm.
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| CN118275596A (en) * | 2024-04-23 | 2024-07-02 | 湖南易能生物医药有限公司 | Quality control method of Chinese herbal compound containing rhizoma sparganii and rhizoma curcumae and application thereof |
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| CN115778997A (en) * | 2022-12-28 | 2023-03-14 | 江苏海洋大学 | Preparation method and quality detection method of Qibang granules |
| CN115778997B (en) * | 2022-12-28 | 2023-10-27 | 江苏海洋大学 | Preparation method and quality detection method of astragalus root and burdock granules |
| CN118275596A (en) * | 2024-04-23 | 2024-07-02 | 湖南易能生物医药有限公司 | Quality control method of Chinese herbal compound containing rhizoma sparganii and rhizoma curcumae and application thereof |
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