CN115951006A - Detection method of kidney-tonifying and turbidity-eliminating traditional Chinese medicine preparation - Google Patents
Detection method of kidney-tonifying and turbidity-eliminating traditional Chinese medicine preparation Download PDFInfo
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Abstract
The invention discloses a detection method of a kidney-tonifying and turbidity-eliminating traditional Chinese medicine preparation, which comprises the following raw materials in parts by weight: 1-75 parts of astragalus root, 1-50 parts of salvia miltiorrhiza, 1-50 parts of schisandra chinensis, 1-30 parts of glabrous greenbrier rhizome and 1-30 parts of sweet wormwood; the detection method comprises the steps of identifying the astragalus, the schisandra and the glabrous greenbrier rhizome in the traditional Chinese medicine preparation by adopting thin-layer chromatography, and determining the content of salvianolic acid B in the salvia miltiorrhiza in the traditional Chinese medicine preparation by adopting high performance liquid chromatography. The invention has the characteristics of simple and convenient detection, shortened detection time and stable and reliable identification.
Description
Technical Field
The invention relates to a traditional Chinese medicine preparation for tonifying kidney and dissolving turbidity, in particular to a detection method of the traditional Chinese medicine preparation for tonifying kidney and dissolving turbidity.
Background
The prescription for tonifying the kidney and dissolving turbidity is a clinical experience prescription of professor Zhanghening of famous Chinese medicine nephropathy experts in China. The prescription is composed of five medicines of astragalus root, salvia root, schisandra fruit, poria cocos and sweet wormwood. The kidney-tonifying and turbid-eliminating granule is prepared based on the theory of traditional Chinese medicine and combines the diagnosis and the pathology of western medicine. The effective prescription for treating chronic nephritis is formed by screening medicine through years of clinical experiments. In the formula, the astragalus, the schisandra and the salvia are used as monarch drugs, the astragalus is used for tonifying spleen and qi and tonifying kidney, especially for tonifying spleen and kidney, the first important drug is used in the weight proportion, and the unprocessed one takes the effect of tonifying qi and strengthening exterior and the one for strengthening exterior and benefits defensive qi to prevent exogenous pathogenic factors. The shizandra berry is sour and warm in nature and can warm and enter kidney, and is a top-quality product for tonifying kidney, namely tonifying kidney and replenishing essence, and also can induce astringency to stop enuresis, thus being the essential herb for treating fine essence loss. The red sage root has the effects of promoting blood circulation and removing blood stasis, is mainly used for promoting blood circulation and has the function of nourishing blood, and is used for treating chronic kidney diseases, blood stasis is caused after long-term diseases and essence and blood are injured, and the red sage root has the effects of invigorating the kidney, tonifying the spleen and promoting blood circulation and is taken as a monarch drug in the prescription by combining the functions of promoting blood circulation and nourishing blood of the red sage root and the astragalus root and the schisandra fruit; in addition, chronic kidney diseases, spleen failure, kidney failure, strangulation, retention of water evil, accumulation of damp turbidity and heat accumulation for a long time are caused, so the formula takes the rhizoma smilacis glabrae as a ministerial drug, is sweet and light, promotes diuresis, clears damp-heat, is a ministerial drug and has the effect of treating symptoms. The sweet wormwood herb is bitter, pungent and cold, is an adjuvant drug, can assist the warm property of the astragalus, the schisandra and the salvia, can clear deficiency heat and promote diuresis and heat, has strict compatibility, is combined with the monarch and minister drugs, assists in treatment, and has the effects of tonifying spleen and kidney, activating blood stasis and promoting diuresis and turbidity. However, at present, the problems of long detection time and low stability exist in the detection of the components of each traditional Chinese medicine in the prescription.
Disclosure of Invention
The invention aims to provide a detection method of a kidney-tonifying and turbidity-eliminating traditional Chinese medicine preparation. The invention has the characteristics of simple and convenient detection, shortened detection time and stable and reliable identification.
The technical scheme of the invention is as follows: the detection method of the kidney-tonifying and turbidity-eliminating traditional Chinese medicine preparation comprises the following raw materials in parts by weight: 1 to 75 portions of astragalus root, 1 to 50 portions of salvia miltiorrhiza, 1 to 50 portions of shizandra berry, 1 to 30 portions of glabrous greenbrier rhizome and 1 to 30 portions of sweet wormwood; the detection method comprises the steps of identifying the astragalus, the schisandra and the glabrous greenbrier rhizome in the traditional Chinese medicine preparation by adopting a thin-layer chromatography, and determining the content of salvianolic acid B in the salvia miltiorrhiza in the traditional Chinese medicine preparation by adopting a high performance liquid chromatography.
In the detection method of the kidney-tonifying and turbid-removing traditional Chinese medicine preparation, the identification method of the astragalus membranaceus comprises the following steps: collecting 2g Chinese medicinal preparation, adding water 20ml, ultrasonic extracting for 10min, filtering, extracting the filtrate with water saturated n-butanol solution for 3 times, each time 20ml, mixing n-butanol solutions, washing with ammonia solution for 2 times, each time 20ml, discarding ammonia solution, concentrating n-butanol solution to dryness, and dissolving the residue with methanol 1ml to obtain sample solution; adding methanol into astragaloside IV reference substance to obtain 1mg solution per 1ml as reference substance solution; and (3) absorbing 5-10 ul of each of the test solution and the reference solution by thin layer chromatography, respectively dropping on the same silica gel G thin layer plate, developing by using chloroform-methanol-water solution as a developing agent in a volume ratio of 13:6:2, taking out, drying in the air, spraying with 10% sulfuric acid ethanol solution, heating at 105 ℃ until spots are clearly developed, and inspecting in the sunlight.
In the detection method of the kidney-tonifying and turbid-removing traditional Chinese medicine preparation, the identification method of the schisandra chinensis comprises the following steps: taking 4g of the traditional Chinese medicine preparation, adding 20ml of trichloromethane, carrying out ultrasonic extraction for 30 minutes, filtering, concentrating the filtrate to be dry, and adding 1ml of trichloromethane into residues to dissolve the residues to be used as a test solution; adding chloroform 20ml into fructus Schisandrae control 1g, reflux extracting for 30 min, filtering, concentrating the filtrate to dryness, and dissolving the residue with chloroform 1ml to obtain control solution; according to thin-layer chromatography, absorbing 5-10 ul of each of the sample solution and the reference solution, respectively dropping on the same silica gel G plate thin-layer plate, taking the petroleum ether-ethyl formate-formic acid upper layer solution as a developing agent according to the volume ratio of 10:6:1, developing, taking out, air drying, and inspecting under an ultraviolet lamp.
In the detection method of the kidney-tonifying and turbid-removing traditional Chinese medicine preparation, the identification method of the glabrous greenbrier rhizome comprises the following steps: taking 2g of the Chinese medicinal preparation, adding 20ml of water, carrying out ultrasonic treatment for 10 minutes, filtering, adding the filtrate into a macroporous adsorption resin column, eluting with 50ml of water, discarding the water solution, eluting with 30% ethanol, discarding the first 75ml, collecting the second 100ml, evaporating to dryness, and dissolving the residue with 5ml of methanol to obtain a sample solution; adding methanol into astilbin reference substance to obtain 0.1mg solution per 1ml as reference substance solution; according to thin layer chromatography, absorbing 5-10 ul of each of the sample solution and the reference solution, respectively dropping on the same silica gel G thin layer plate, developing with toluene-ethyl acetate-formic acid as developing agent at volume ratio of 1.5:3:0.9, taking out, air drying, spraying with aluminum trichloride test solution, standing for 5min, and viewing under ultraviolet lamp.
In the detection method of the kidney-tonifying and turbid-removing traditional Chinese medicine preparation, the content determination method of salvianolic acid B in the salvia miltiorrhiza is as follows: performing high performance liquid chromatography with salvianolic acid B as reference, methanol-acetonitrile-formic acid-water at volume ratio of 28:7:1:63 as mobile phase, detecting wavelength of 284-288 nm, respectively sucking 10 μ l of reference solution and sample solution, injecting into liquid chromatograph, and measuring.
In the detection method of the kidney-tonifying and turbidity-eliminating traditional Chinese medicine preparation, the preparation method of the reference substance solution comprises the following steps: weighing salvianolic acid B reference substance, adding 75% methanol to obtain 65 μ g solution per 1ml, to obtain Saviae Miltiorrhizae radix reference substance solution; the preparation method of the test solution comprises the following steps: taking 0.1g of the traditional Chinese medicine preparation, adding 24ml of 75% methanol, carrying out ultrasonic extraction for 10 minutes, taking out, adding 75% methanol to 25ml of scales, filtering by using a microporous membrane, and taking a subsequent filtrate, namely the test solution.
In the detection method of the kidney-tonifying and turbidity-eliminating traditional Chinese medicine preparation, the traditional Chinese medicine preparation comprises the following raw materials in parts by weight: 75 parts of astragalus root, 50 parts of salvia miltiorrhiza, 50 parts of schisandra fruit, 30 parts of glabrous greenbrier rhizome and 30 parts of sweet wormwood.
In the detection method of the kidney-tonifying and turbidity-eliminating traditional Chinese medicine preparation, the preparation method of the traditional Chinese medicine preparation comprises the following steps: crushing schisandra chinensis, extracting the crushed schisandra chinensis, astragalus mongholicus, salvia miltiorrhiza, rhizoma smilacis glabrae and sweet wormwood with water for three times, wherein each time is 1 hour, filtering an extracting solution, combining filtrates, concentrating the filtrate under reduced pressure to obtain a concentrated solution with the relative density of 1.05-1.10, placing the concentrated solution to room temperature, adding ethanol to ensure that the alcohol content of the concentrated solution is 50%, stirring, standing for 15 hours, and taking a supernatant; centrifuging the remaining precipitate and collecting the supernatant; and combining the two parts of supernatant, recovering ethanol, concentrating under reduced pressure to obtain thick paste with the relative density of 1.35-1.40, and drying under reduced pressure to obtain the traditional Chinese medicine composition.
In the detection method of the kidney-tonifying and turbidity-eliminating traditional Chinese medicine preparation, the dosage form of the traditional Chinese medicine preparation includes any one of granules, tablets, pills, capsules, granules, oral liquid, sprays, injections, suspensions and microsphere preparations.
Compared with the prior art, the invention has the beneficial effects that:
the detection method of the traditional Chinese medicine preparation is to determine the astragalus, the schisandra, the glabrous greenbrier rhizome and the sweet wormwood herb in the preparation by a thin-layer chromatography identification method and determine the content of the salvianolic acid B in the salvia miltiorrhiza by a high performance liquid chromatography. The method for measuring the content of the salvia miltiorrhiza has the advantages of simple operation, low cost, short detection time, good separation effect, sensitivity, accuracy and the like; the method for identifying the astragalus, the schisandra and the rhizoma smilacis glabrae has the advantages of high accuracy, high precision, high recovery rate, high stability, strong specificity, good repeatability, no interference and the like. The invention can effectively detect whether the product is qualified or not and whether the quality is good or bad, ensures the consistency of the drug effect and the stability of the quality, and further ensures the stability of the product quality and the safety and the effectiveness of clinical medication.
Drawings
FIG. 1 is an HPLC chart of a salvianolic acid B control;
FIG. 2 is an HPLC chart of salvianolic acid B;
FIG. 3 is a HPL plot of a Salvia miltiorrhiza negative control sample absent;
FIG. 4 is a HPL profile of a Salvia miltiorrhiza positive control sample;
FIG. 5 is a linear relationship investigation graph of salvianolic acid B;
FIG. 6 is a diagram showing the development effect of thin layers of the radix astragali identification reference substance and the test substance;
FIG. 7 is a diagram of the effect of the specificity investigation of the thin-layer identification of Astragalus membranaceus;
FIG. 8 is a diagram showing the identification effect of the thin layer of Astragalus membranaceus on the thin layer plate of manufacturer 1;
FIG. 9 is a diagram showing the identification effect of the thin layer of Astragalus membranaceus on the thin layer plate of manufacturer 2;
FIG. 10 is a diagram showing the identification effect of the thin layer of Astragalus membranaceus on the thin layer plate of manufacturer 3;
FIG. 11 is a graph showing the identification effect of thin layer of Astragalus membranaceus under the conditions of 20 deg.C and 50% humidity;
FIG. 12 is a diagram of the identification effect of thin layer of Astragalus membranaceus under the conditions of 25 deg.C temperature and 55% humidity;
FIG. 13 is a diagram showing the identification effect of thin layers of Astragalus membranaceus under the conditions of 35 deg.C and 70% humidity;
FIG. 14 is a thin layer development effect diagram of the identification reference substance and test solution of fructus Schisandrae;
FIG. 15 is a diagram of the effect of a specificity investigation test for Chinese magnoliavine fruit thin layer identification;
FIG. 16 is a graph showing the identification effect of fructus Schisandrae chinensis thin layer of the thin layer plate of manufacturer 1;
FIG. 17 is a graph showing the identification effect of fructus Schisandrae chinensis thin layer on the thin layer plate of manufacturer 2;
FIG. 18 is a thin-layer identification effect chart of fructus Schisandrae chinensis of 3 thin-layer plate of manufacturer;
FIG. 19 is a graph showing the identification effect of fructus Schisandrae chinensis thin layer at 20 deg.C and 50% humidity;
FIG. 20 is a diagram showing the identification effect of fructus Schisandrae chinensis thin layer at 25 deg.C and 55% humidity;
FIG. 21 is a diagram of the identification effect of fructus Schisandrae chinensis thin layer at 35 deg.C and 70% humidity;
FIG. 22 is a diagram showing the thin-layer development effect of Smilax glabra identification control and test solutions;
FIG. 23 is a diagram of the effect of a specificity investigation test for rhizoma Smilacis Glabrae thin layer identification;
FIG. 24 is a graph showing the identification effect of rhizoma Smilacis Glabrae thin layer on the thin layer plate of manufacturer 1;
FIG. 25 is a diagram of the identification effect of Smilax glabra Linne thin layer of the thin layer plate of the manufacturer 2;
FIG. 26 is a graph showing the identification effect of rhizoma Smilacis Glabrae thin layer of a thin layer plate of manufacturer 3;
FIG. 27 is a graph showing the identification effect of rhizoma Smilacis Glabrae thin layer at 20 deg.C and 50% humidity;
FIG. 28 is a diagram of identification effect of rhizoma Smilacis Glabrae thin layer at 25 deg.C and 55% humidity;
FIG. 29 is a graph showing the identification effect of rhizoma Smilacis Glabrae thin layer at 35 deg.C and 70% humidity.
Detailed Description
The present invention is further illustrated by the following examples, which are not to be construed as limiting the invention.
Example (b):
the traditional Chinese medicine preparation for tonifying the kidney and dissolving turbidity comprises the following components:
750g of astragalus, 500g of salvia miltiorrhiza, 500g of schisandra chinensis, 300g of rhizoma smilacis glabrae and 300g of sweet wormwood.
The preparation method of the traditional Chinese medicine preparation for tonifying the kidney and eliminating turbid comprises the following steps:
pulverizing the Chinese magnoliavine fruit, adding water to the rest four medicinal materials for extraction for three times, 1 hour each time, filtering an extracting solution, combining filter liquor, concentrating the filter liquor under reduced pressure (T is less than 75 ℃) to obtain a concentrated solution with the relative density of 1.05-1.10 (50 ℃), placing the concentrated solution to room temperature, adding ethanol to ensure that the alcohol content of the concentrated solution is 50%, stirring, standing for 15 hours, and taking supernate; centrifuging the rest precipitate (at centrifugal condition of 10min, 2000r/min), and collecting supernatant; and mixing the two supernatants, recovering ethanol, concentrating under reduced pressure to obtain a thick paste with a relative density of 1.35-1.40 (60 ℃), drying under reduced pressure (T = 70-75 ℃), crushing, sieving with an 80-mesh sieve, adding a proper amount of lactose and aspartame, uniformly mixing, granulating, drying (T = 40-50 ℃), and finishing granules to obtain the traditional Chinese medicine.
The detection method of the kidney-tonifying and turbid-removing traditional Chinese medicine preparation comprises the following steps:
(1) The method for identifying the astragalus comprises the following steps: taking a proper amount of the product particles, grinding, weighing 2g, adding 20ml of water, carrying out ultrasonic extraction for 10 minutes, filtering, extracting the filtrate with water saturated n-butyl alcohol solution for 3 times, 20ml each time, combining the n-butyl alcohol solutions, washing with ammonia test solution for 2 times, 20ml each time, discarding the ammonia test solution, concentrating the n-butyl alcohol solution to dryness, and dissolving the residue with 1ml of methanol to obtain a test solution. And adding methanol into an appropriate amount of astragaloside IV reference substance to obtain a solution containing 1mg per 1ml as reference substance solution. Performing thin-layer chromatography (0502 of the four ministerial rules of the design of the Chinese pharmacopoeia 2020), sucking sample solution and reference solution 5-10 ul respectively, dropping on the same silica gel G thin-layer plate, taking the lower layer solution of chloroform-methanol-water (volume ratio of 13:6: 2) as developing agent, developing, taking out, air drying, spraying 10% sulphuric acid ethanol solution, heating at 105 deg.C until the spots are clearly developed, and inspecting under sunlight. Spots of the same color appear in the chromatogram of the test solution at positions corresponding to those in the chromatogram of the control solution.
(2) The identification method of the schisandra chinensis comprises the following steps: taking a proper amount of the product particles, grinding, weighing 4g, adding 20ml of trichloromethane, carrying out ultrasonic extraction for 30 minutes, filtering, concentrating the filtrate to dryness, and dissolving residues in 1ml of trichloromethane to obtain a test solution. Taking 1g of schisandra chinensis as a reference medicinal material, adding 20ml of chloroform, performing reflux extraction for 30 minutes, filtering filtrate, concentrating to dryness, and adding 1ml of chloroform into residues to dissolve the residues to obtain a reference medicinal material solution. According to a thin-layer chromatography (0502 of the four ministry of the general rules of the design of the Chinese pharmacopoeia 2020), sucking sample solution and reference solution 5-10 ul respectively, respectively dropping on the same silica gel G plate thin layer plate, taking the upper solution of petroleum ether (30-60 ℃) -ethyl formate-formic acid (volume ratio is 10:6: 1) as a developing agent, developing, taking out, airing, and placing under an ultraviolet lamp (365 nm) for inspection. Spots of the same color appear on the chromatogram of the test solution at the positions corresponding to those on the chromatogram of the control solution.
(3) The identification method of the glabrous greenbrier rhizome comprises the following steps: taking a proper amount of the product particles, grinding, weighing 2g, adding 20ml of water, carrying out ultrasonic treatment for 10 minutes, filtering, adding the filtrate into a D-101 macroporous adsorption resin column (the inner diameter is 1.5cm, and the height is 13 cm), eluting with 50ml of water, discarding the water solution, eluting with 30% ethanol, discarding the first 75ml, continuously collecting 100ml, evaporating to dryness, and dissolving the residue with 5ml of methanol to obtain a sample solution. Adding methanol into astilbin control to obtain solution containing 0.1mg per 1ml as control solution. Performing thin-layer chromatography (0502 of the four ministry of the Ministry of the Japan, the & ltChinese pharmacopoeia & gt 2020 edition), sucking the sample solution and the reference solution by 5-10 ul respectively, dropping the sample solution and the reference solution on the same silica gel G thin-layer plate respectively, developing by taking toluene-ethyl acetate-formic acid (the volume ratio is 1.5:3: 9) as a developing agent, taking out, drying, spraying the test solution of aluminum trichloride, placing for 5 minutes, and then placing under an ultraviolet lamp (365 nm) for inspection. In the chromatogram of the test solution, fluorescent spots of the same color appear at the corresponding positions of the chromatogram of the control solution.
(4) The method for measuring the content of the salvianolic acid B in the salvia miltiorrhiza comprises the following steps:
the high performance liquid chromatography is adopted for determination, and the chromatographic conditions of the high performance liquid chromatography are as follows: octadecylsilane chemically bonded silica is used as a filling agent; methanol-acetonitrile-formic acid-water (volume ratio is 28:7:1: 63) is used as a mobile phase, and the flow rate of the mobile phase is 0.9 ml/min-1.1 ml/min; the detection wavelength is 284-288 nm; the temperature of the chromatographic column is 28-35 ℃, the plate number is not less than 3000 according to the salvianolic acid B, and the separation degree is not less than 1.5.
The control solution was prepared as follows: precisely weighing appropriate amount of salvianolic acid B reference substance, and adding 75% methanol to obtain 65 μ g solution per 1 ml;
the preparation of the test solution comprises the following steps: taking 0.1g of the product, precisely weighing, placing in a 25ml measuring flask, adding about 24ml of 75% methanol, ultrasonically extracting for 10 minutes, taking out, placing to room temperature, adding 75% methanol to a scale of 25ml, shaking up, filtering with a microporous membrane (the pore diameter is 0.22 μm), and taking the subsequent filtrate to obtain the sample solution.
The determination method comprises the following steps of; precisely sucking 10 μ l of each of the reference solution and the sample solution, injecting into liquid chromatograph, and measuring.
In order to ensure the scientific, reasonable and feasible detection method, the detection method is researched and investigated.
1. Investigating the content determination methodology of salvianolic acid B in salvia miltiorrhiza:
the salvia miltiorrhiza is a monarch drug in the traditional Chinese medicine preparation, and the salvianolic acid B is a main effective component of the salvianolic acid B, so the salvianolic acid B is selected as an index component of the internal quality of the preparation, 75% methanol is adopted to carry out pretreatment on a sample, and the high performance liquid chromatography is adopted to carry out sample determination.
1. Instruments, reagents and samples
The instrument comprises the following steps: the system comprises an Agilent 1260 high performance liquid chromatograph, an XP205 electronic analytical balance and a KQ-250DB numerical control ultrasonic cleaner;
reagent: methanol (analytical purity of kang scientific and technology Limited, tianjin), acetonitrile (chromatographic purity of kang scientific and technology Limited, tianjin), and formic acid (analytical purity of kang scientific and technology Limited, tianjin);
sample preparation: salvianolic acid B reference (batch No. 111562-201110)
Particles for tonifying kidney and resolving turbidity (20120420, 20120507, 20120502, the preparation of the invention, provided by Guizhou Bailing pharmaceutical products GmbH).
2. Chromatographic condition and system adaptability test
Octadecylsilane chemically bonded silica is used as a filling agent; detection wavelength: λ =286nm; column temperature: 30 ℃; mobile phase: methanol-acetonitrile-formic acid-water (28: 7:1: 63); flow rate: 1ml/min; the theoretical plate number calculated by salvianolic acid B is not less than 3000, and the separation degree is 1.5.
Salvianolic acid B N =5.54 × (t) R /W 1/2h ) 2 ＝5.54×(12.475/0.4611) 2 ＝4056；
R＝2(t R2 －t R1 )/(W 1 +W 2 )＝1.69；
Therefore, it is specified that the number of theoretical plates should not be less than 3000 and the degree of separation should not be less than 1.5.
3. Preparation of control solutions
Weighing appropriate amount of salvianolic acid B reference (lot No. 111562-201110), and adding 75% methanol to obtain solution containing 65 μ g of salvianolic acid B per 1 ml.
4. Preparation of test solution
Preparation of a test solution: taking 0.1g of the product, precisely weighing, placing in a 25ml measuring flask, adding about 24ml of 75% methanol, performing ultrasonic extraction for 10 minutes, taking out, placing to room temperature, adding 75% methanol to 25ml scale, shaking up, and filtering with a microporous membrane (the pore diameter is 0.22 μm) to obtain a test solution.
5. Preparation of Positive solutions
Taking 0.2g of salvia miltiorrhiza powder which is sieved by a 50-mesh sieve, precisely weighing, placing in a triangular flask with a plug, precisely adding 50ml of 75% methanol, weighing, heating and refluxing for 1 hour, cooling, weighing again, supplementing the lost weight with 75% methanol, shaking up, filtering with a microporous membrane, and taking the subsequent filtrate as a positive solution.
6. Preparation of negative control solution
Taking 0.74g of the prescription lacking the salvia miltiorrhiza medicinal material, and preparing a negative solution by the same preparation method of the test solution.
7. Methodology validation
7.1 specificity
Performing specificity test according to the above chromatographic conditions and sample processing method, wherein the liquid chromatogram of salvianolic acid B reference, test sample, positive and negative are shown in figures 1-4 respectively.
7.2 inspection of linearity and Range
Sucking control stock solution (0.3219 mg/ml) 0.5,1,2,4,8, 10ml, placing into 10ml measuring flask, adding 75% methanol to scale, and shaking. Injecting 10 mu l of the solution respectively, drawing a standard curve graph by taking the injection amount as a horizontal coordinate and the peak area as a vertical coordinate, and processing to obtain a regression equation: y =1224.4X-52.715 (Y is peak area, X is concentration), r =0.9995. The results showed that salvianolic acid B showed a good linear relationship with peak area integral in the range of 0.1611-3.222. Mu.g, as shown in FIG. 5.
7.3 precision test
Taking 10 mu l of sample liquid (20120420) under the content measurement item, sampling for 6 times, respectively recording peak area integral values, calculating the content, measuring the peak area integral value, wherein the RSD is 0.41%, and the instrument precision is good.
7.4 repeatability test
And (4) precisely weighing 6 parts of 20120420 samples respectively according to the operation under the content measurement method, and performing content measurement. The average mass fraction of the salvianolic acid B is 13.64mg/g, and the RSD is 1.42 percent, which indicates that the method has better repeatability.
7.5 stability test
And (3) standing the sample solution (20120420) to be injected for 24 hours, injecting the sample solution into a liquid chromatograph for measurement at different times (0, 2,4,8, 12 and 24 hours), recording a peak area integral value, and calculating the content, wherein the RSD value of the salvianolic acid B is 0.87%, which shows that the salvianolic acid B in the test solution has good stability within 24 hours.
7.6 sample recovery test
Taking 9 parts of 20120420 batches of samples, each part of which is about 0.05g, precisely weighing, adding 2ml, 2.5ml and 3ml of stock solutions of reference substances (0.3219 mg/ml) into finished products respectively, treating according to a method for preparing test solution, and determining according to a content determination method. The results are shown in Table 1.
TABLE 1 salvianolic acid B sample recovery test results (n = 9)
Analysis of the results in table 1 shows that the sample recovery mean value is 102.75%, and the RSD is 1.12%, indicating that the method has high accuracy.
7.7 durability test
3 factors of different mobile phase proportions, column temperatures and flow rates in the experiment are selected, and the factors are respectively investigated according to the treatment methods under the items of '4' and '5'.
7.7.1 selection of extraction method, time and particle size
The extraction method comprises an extraction sample granularity extraction test, wherein ultrasonic extraction of samples and samples with different granularities treated by a medicinal material extraction method is considered for 10 minutes, and the content of the salvianolic acid B is determined. The measured contents are shown in Table 2.
TABLE 2 extraction method, sample granulometry results
The test result shows that: compared with the reflux extraction, the ultrasonic extraction is complete, and samples with different particle sizes can be basically completely extracted after the ultrasonic extraction. Therefore, the particles of the test sample are selected for ultrasonic extraction.
And (3) an extraction time investigation test, wherein ultrasonic extraction is carried out on a sample at different times after a solvent is added, and the content is measured, and the result is shown in table 3.
TABLE 3 measurement of extraction time content
The test result shows that: the complete extraction can be realized by ultrasonic treatment for 10 minutes.
7.7.2 examination of the proportions of the mobile phases
2 parts of 20120420 sample is precisely weighed, methanol-acetonitrile-formic acid-water is used as a mobile phase, different proportions of the mobile phase are changed, other conditions are unchanged, the peak area is recorded, the content is calculated, the difference of the measurement result with the original mobile phase proportion is calculated, and the result is shown in table 4.
The test result shows that: the ratio of methanol and water in the mobile phase is slightly changed, and the content measurement RSD =0.883%, so the slight change of the ratio of the mobile phase has no obvious influence on the separation degree of the sample and the measurement result.
TABLE 4 durability examination of different mobile phase ratios
7.7.3 examination of the pH of the mobile phase
Precisely weighing 2 parts of the 20120420 sample, using methanol-acetonitrile-formic acid-water as a mobile phase, changing different concentrations of formic acid, measuring under other conditions, recording peak areas, calculating the content, and calculating the difference between the measurement results and the mobile phase, wherein the results are shown in Table 5.
TABLE 5 results of durability examination of mobile phase pH
The test result shows that: the ratio of formic acid in the mobile phase fluctuates from the set ratio, and the content measurement RSD =2.923%, so that the small change of the concentration of formic acid in the mobile phase has no obvious influence on the measurement result.
7.7.4 inspection of different chromatography columns
2 parts of the 20120420 sample were precisely weighed, and the content was calculated by using methanol-acetonitrile-formic acid-water (28.
TABLE 6 investigation of the durability of different chromatography columns
The test result shows that: the three chromatographic columns have no significant influence on the content determination of the salvianolic acid B.
7.7.5 Effect of column temperature
2 parts of the 20120420 sample is precisely weighed, the column temperature of the chromatographic column is changed, other conditions are unchanged, the peak area is recorded, the content is calculated, the difference of the measurement results under different column temperature conditions is calculated, and the result is shown in table 7.
TABLE 7 durability test results for different column temperatures
The test result shows that: the column temperature is set to fluctuate up and down at 30 ℃, the chromatographic column temperature is in the range of 28-35 ℃, and the content determination RSD =1.621 percent, so the content determination of the salvianolic acid B with slight change of the column temperature has no obvious influence.
7.7.6 influence of flow Rate
2 parts of the 20120420 sample are precisely weighed, the mobile phase is measured at different flow rates under the condition that other conditions are unchanged, the peak area is recorded, the content is calculated, the difference of the measurement results under the conditions of different flow rates is calculated, and the result is shown in Table 8.
The test result shows that: the column temperature is set to fluctuate up and down at 1.0ml/min, the flow rate of the mobile phase is in the range of 0.9ml/min to 1.1ml/min, and the content determination RSD =1.027%, so the content determination of the salvianolic acid B with slight change of the flow rate has no obvious influence.
TABLE 8 durability test results for different flow rates
7.7.7 influence of the detection wavelength
2 parts of the 20120420 sample are precisely weighed, the detection wavelength is subjected to micro-fluctuation, other conditions are unchanged, the peak area is recorded, the content is calculated, the difference of the measurement results under different detection wavelength conditions is calculated, and the result is shown in Table 9.
TABLE 9 durability test results for different detection wavelengths
The test result shows that: the content determination RSD =1.326% in the range of 284-288 nm with the detection wavelength floating up and down at 286nm, so the content determination of the salvianolic acid B with small change of the detection wavelength has no obvious influence.
7.8 and 3 batches of sample content determination results
Injecting 10 μ l of the test solution and the reference solution into a liquid chromatograph under chromatographic conditions defined by content determination, and determining the content of salvianolic acid B. The results of the assay of the three samples are shown in Table 10.
TABLE 10 results of sample measurement
The determination results show that the high performance liquid chromatography for determining the content of the salvianolic acid B in the product has good linear relation and durability, and the sample recovery rate, the precision and the reproducibility meet the requirements, so the high performance liquid chromatography can be used as a method for determining the content of the salvianolic acid B in the kidney-tonifying and turbid-removing particles.
2. Identification of astragalus:
radix astragali is dried root of Astragalus membranaceus (Fisch.) bge. Var. Mongholicus (bge.) Hsiao of Leguminosae. Collected in the 2020 edition of Chinese pharmacopoeia (part I). In the experiment, an astragaloside IV reference substance is selected, water extraction is firstly adopted, then a method of water saturated n-butyl alcohol extraction and ammonia test solution washing is adopted to extract characteristic components of the astragalus in the kidney-tonifying and turbid-removing particles, thin-layer identification is carried out on the astragalus, and the influence of different development media, different sample amounts, thin-layer plates of different manufacturers, different temperatures and different humidities on the thin-layer chromatography of the coptis in the preparation is inspected. The test result shows that: the method takes the astragaloside IV as a reference substance, takes a chloroform-methanol-water (volume ratio is 13.
2.1 instruments, reagents and samples
The invention relates to a KQ-250DB type numerical control ultrasonic cleaner (ultrasonic instruments, inc. of Kunshan), a DGG-9246A electric heating constant temperature blast drying box (Shanghai Qixin scientific instruments, inc.), a one hundred thousand electronic balance (Mettler-Tolyduo instruments, shanghai) and n-butyl alcohol, ammonia test solution, methanol and chloroform are all analytically pure, water is purified water, astragaloside A reference substances (Chinese pharmaceutical biological product institute, no. 110781-200613) are used for tonifying the kidney and eliminating turbidity (20420120, 20120502 and 20120507).
2.2 preparation of test solutions
Taking a proper amount of the product particles, grinding, weighing 2g, adding 20ml of water, carrying out ultrasonic extraction for 10 minutes, filtering, extracting the filtrate with water saturated n-butyl alcohol solution for 20ml each time, combining the n-butyl alcohol solutions, washing with ammonia test solution for 2 times, 20ml each time, discarding the ammonia test solution, concentrating the n-butyl alcohol solution to dryness, and dissolving the residue with 1ml of methanol to obtain a test solution.
2.3 preparation of control solutions
And adding methanol into proper amount of astragaloside IV control to obtain 1mg solution per 1ml as control solution.
2.4 preparation of negative samples
According to the parts of the components in the prescription, taking the other medicines except the astragalus, preparing a negative sample according to the process requirements, weighing 3.2g, and operating according to the method under the item of preparation of the test solution to obtain a negative sample solution.
2.5 deployment System
Chloroform-methanol-water (13.
2.6 color-developing agent
10% sulfuric acid ethanol solution.
2.7 examination of conditions of thin layer chromatography
2.7.1 optimal spot size examination
Respectively sucking 1 μ l, 5 μ l and 10 μ l of control solution, and 1 μ l, 3 μ l, 5 μ l, 8 μ l, 10 μ l and 15 μ l of test solution, respectively dropping on the same thin silica gel G plate, spreading, taking out, air drying, spraying with developer, heating at 105 deg.C until the spots are clearly developed, and inspecting in sunlight. Referring to FIG. 6, in FIG. 6, the sample application amount of the control sample is 1. Mu.l, 5. Mu.l and 10. Mu.l from left to right, and the sample application amount of the test sample is 1. Mu.l, 3. Mu.l, 5. Mu.l, 8. Mu.l, 10. Mu.l and 15. Mu.l from left to right.
As can be seen from FIG. 6, the sample application amount of the sample is 5-10 μ l for the control and the sample, and the sample application amount is 5-10 μ l for the sample because the sample application amount is overloaded when the sample application amount is greater than 10 μ l.
2.8 methodological validation
2.8.1 specialization examination
Three batches of kidney-tonifying and turbidity-eliminating granule samples were subjected to the specificity test as described above, and the results are shown in FIG. 7.
As can be seen from fig. 7, the negative sample was not perturbed; the astragaloside IV has clear spots with the same color at the corresponding positions of the chromatogram of the astragaloside IV reference substance, the spots are round and round, and the separation effect is good. The method is proved to have strong specificity and clear spot color development.
2.8.2 durability test
Three batches of kidney-tonifying and turbidity-eliminating particle samples are respectively compared with 3 thin-layer plates of different manufacturers, and 3 different temperatures and humidities are used as variation factors to carry out experiments according to the identification method.
2.8.2.1 thin layer plate tests from different manufacturers, the results are shown in FIGS. 8-10.
The results of durability experiments of 3 different manufacturers show that the thin-layer identification chart of the astragalus membranaceus in the kidney-tonifying and turbidity-removing particles has good spot separation effect and clear spots, and thin-layer plates of different manufacturers have little influence on main spots of the astragalus membranaceus and have good repeatability.
2.8.2.2 temperature and humidity tests
The various temperature and humidity settings are shown in table 11. The test patterns are shown in FIGS. 11-13.
TABLE 11 examination of different temperatures and relative humidities
The experimental spectrums with different temperatures and relative humidities are examined, so that fluorescent spots with the same color are displayed in the positions, corresponding to the positions of the chromatogram of the reference substance, of the sample chromatogram, the spots are clear, the separation effect is good, the Rf value is moderate, and the negative control is free of interference. It is thus understood that the temperature and relative humidity vary within the normal environmental range, and do not affect the effectiveness of the identification method.
3. Identification of Schisandra chinensis
Fructus Schisandrae is dried mature fruit of Schisandra chinensis (Turcz.) Baill. The producing area is Liaoning. Collected in the 2020 edition of Chinese pharmacopoeia (part I). The schisandra chinensis is one of monarch drugs in a prescription for tonifying the kidney and eliminating turbid pathogen, and the schisandrin is the main active ingredient. Thin-layer identification is carried out by taking a schisandra chinensis control medicinal material as a control, and the influence of different sample amounts, thin-layer plates of different manufacturers, different temperatures and different humidities on the thin-layer chromatography of the schisandra chinensis medicinal material in the kidney tonifying and turbid resolving process is examined. Test results show that by using a schisandra chinensis contrast medicinal material as a contrast and using petroleum ether (30-60 ℃), ethyl formate-formic acid (volume ratio is 10. Therefore, the method can be used as a thin-layer identification method for the schisandra chinensis medicinal material in tonifying the kidney and eliminating turbid pathogen, and is listed as a detection item of the invention.
3.1 instruments, reagents and samples
KQ-250DB type numerical control ultrasonic cleaner (Kunshan ultrasonic instruments, inc.), DGG-9246A electric heating constant temperature blast drying box (Shanghai Qixin scientific instruments, inc.), one hundred thousand electronic balance (Mettler-Tooliduo instruments, shanghai) Inc.), chloroform, petroleum ether (30-60 ℃), ethyl formate and formic acid are all analytically pure. The schisandra chinensis contrast medicinal material (China pharmaceutical biologicals institute, number is 120922-201007), tonifies the kidney and dissolves the turbid (20120420, 20120502, 20120507, the preparation of the invention is provided by pharmaceutical products GmbH of the Guizhou Bailing enterprise group).
3.2 preparation of test solutions
Taking a proper amount of the product particles, grinding, weighing 4g, adding 20ml of trichloromethane, carrying out ultrasonic extraction for 30 minutes, filtering, concentrating the filtrate to dryness, and dissolving residues in 1ml of trichloromethane to obtain a test solution.
3.3 preparation of control solutions
Adding chloroform 20ml into fructus Schisandrae control 1g, reflux extracting for 30 min, filtering, concentrating the filtrate to dryness, and dissolving the residue with chloroform 1ml to obtain control solution.
3.4 preparation of negative samples
According to the components of the prescription, taking the other medicines except the schisandra chinensis, preparing a negative sample according to the process requirements, weighing 3.7g, and operating according to the method under the item of preparation of the test solution to obtain a negative sample solution.
3.5 deployment System
Petroleum ether (30 to 60 ℃) -ethyl formate-formic acid (volume ratio 10.
3.6 optimal spotting inspection
Respectively sucking 5 μ l, 10 μ l and 15 μ l of the test solutions 4 μ l, 5 μ l, 8 μ l, 10 μ l and 15 μ l, respectively dropping on different thin silica gel G plates, inspecting, developing, taking out, air drying, and inspecting under 365nm ultraviolet lamp. See fig. 14. In FIG. 14, the sample application amounts of the control sample are 5. Mu.l, 10. Mu.l and 15. Mu.l from left to right, and the sample application amounts of the test sample are 4. Mu.l, 5. Mu.l, 8. Mu.l, 10. Mu.l and 15. Mu.l from left to right.
As can be seen from FIG. 14, the optimal spot size of the sample is 5-10 μ l, the spots are clear, the separation degree is good, and when the spot size is 15 μ l, the spot size is overloaded and the spots have trailing phenomenon.
3.7 methodological validation
3.7.1 specialization examination
Three batches of kidney-tonifying and turbidity-eliminating granule samples were subjected to the specificity test as described above, and the results are shown in FIG. 15.
As can be seen in fig. 15, the negative sample was not perturbed; the spot with the same color is clear and round at the position corresponding to the chromatogram of the reference substance, and the separation effect is good. The method is proved to have strong specificity and clear spot color.
3.7.2 durability test
Three batches of kidney-tonifying and turbidity-eliminating particle samples are respectively compared with 3 thin-layer plates of different manufacturers, and 3 different temperatures and humidities are used as variation factors to carry out experiments according to the identification method.
3.7.2.1 thin layer plate tests of different manufacturers, the results are shown in FIGS. 16-18.
The durability experiment results of 3 different manufacturers show that the spot separation effect of the thin-layer identification chart of the schisandra chinensis in the kidney-tonifying and turbidity-removing particles is good, spots are clear, the influence of the thin-layer plates of the different manufacturers on the main spots of the schisandra chinensis is small, and the repeatability is good.
3.7.2.2 different temperature and humidity tests
The different temperature and humidity settings are shown in table 12. The test patterns are shown in FIGS. 19 to 21.
TABLE 12 examination of different temperatures and relative humidities
The experimental spectra of different temperatures and relative humidities show that in the chromatogram of the test sample, fluorescent spots with the same color are displayed at the corresponding positions of the chromatogram of the reference sample, the spots are clear, the separation effect is good, the Rf value is moderate, and the negative control is free of interference. It is thus understood that the temperature and relative humidity vary within the normal environmental range, and do not affect the effectiveness of the identification method.
4. Identification of glabrous greenbrier rhizome
Rhizoma Smilacis Glabrae is dried rhizome of Smilax glabra Roxb. Collected in the 2020 edition of Chinese pharmacopoeia (part I). In the experiment, an astilbin reference substance is selected as a reference, water and ultrasonic extraction are firstly adopted to extract characteristic components, and then the characteristic components are washed by water through a D-101 macroporous adsorption resin column to elute impurities; and extracting the characteristic components of the rhizoma smilacis glabrae in the kidney tonifying and turbidity removing process by using a 30% ethanol elution method, and avoiding the influence of impurities on the main components during detection. The thin-layer identification is carried out on the rhizoma smilacis glabrae medicinal material, and the influence of different sample amounts, thin-layer plates of different manufacturers, different temperatures and different humidity on the thin-layer chromatography of the rhizoma smilacis glabrae medicinal material in the preparation is investigated. The test result shows that: the method takes astilbin reference substance as a reference, takes toluene-ethyl acetate-formic acid (volume ratio is 1.5.
4.1 instruments, reagents and samples
The preparation method comprises the following steps of preparing a KQ-250DB type numerical control ultrasonic cleaner (ultrasonic instruments, inc. of Kunshan), a DGG-9246A electric heating constant temperature blast drying box (Shanghai Qixin scientific instruments, inc.), a one hundred thousand electronic balance (Mettler-Tooliduo instruments, shanghai), a D-101 macroporous adsorption resin column, ethanol, toluene, ethyl acetate, formic acid and aluminum trichloride, wherein the D-101 macroporous adsorption resin column, the ethanol, the toluene, the ethyl acetate, the formic acid and the aluminum trichloride are all analytically pure, water is purified water, a astilbin reference substance (a Chinese pharmaceutical and biological product institute, the No. is 111798-200901) is taken, and kidney-tonifying and turbid-removing particles (20120420, 20120502 and 20120507).
4.2 preparation of test solutions
Taking a proper amount of the product particles, grinding, weighing 2g, adding 20ml of water, carrying out ultrasonic treatment for 10 minutes, filtering, adding the filtrate into a D-101 macroporous adsorption resin column (the inner diameter is 1.5cm, and the height is 13 cm), eluting with 50ml of water, discarding the water solution, eluting with 30% ethanol, discarding the first 75ml, continuously collecting 100ml, evaporating to dryness, and adding 5ml of methanol into the residue for dissolving to obtain a sample solution.
4.3 preparation of control solutions
Adding methanol into astilbin reference substance to obtain solution containing 0.1mg per 1ml as reference substance solution.
4.4 preparation of negative samples
According to the components of the prescription, taking the other medicines except the glabrous greenbrier rhizome, preparing a negative sample according to the process requirement, weighing 4.1g, and operating according to the method under the item of preparation of the test solution to obtain a negative sample solution.
4.5 deployment System
Toluene-ethyl acetate-formic acid (volume ratio 1.5.
4.6 color developing agent
And (3) aluminum trichloride test solution.
4.7 examination of conditions of thin layer chromatography
4.7.1 optimal spotting inspection
Respectively sucking control substances 5 μ l, 8 μ l and 10 μ l, respectively dropping test solution 1 μ l, 3 μ l, 5 μ l, 7 μ l, 8 μ l and 10 μ l on silica gel G plate, inspecting, developing, taking out, air drying, spraying with color-developing agent, heating at 105 deg.C for 5min, and inspecting under 365nm ultraviolet lamp. See fig. 22. In FIG. 22, the amounts of the control samples were 5. Mu.l, 8. Mu.l and 10. Mu.l from left to right, and the amounts of the test samples were 1. Mu.l, 3. Mu.l, 5. Mu.l, 7. Mu.l, 8. Mu.l and 10. Mu.l from left to right.
As can be seen from FIG. 22, the sample application amount of 5-10 μ l of the test sample is the most effective, and when the sample application amount is less than 5 μ l, the main spot of the test sample is not clear.
Combining the above experimental results, the optimal spot sample amount of the sample is 5-10 μ l. The thin-layer identification method of the glabrous greenbrier rhizome is determined through preliminary experiment verification, and methodology verification is carried out according to verification guiding principles of a traditional Chinese medicine quality standard analysis method in appendix XVIII A of the edition 2020 of Chinese pharmacopoeia.
4.8 methodological validation
4.8.1 specialization examination
Three batches of kidney-tonifying and turbidity-eliminating granule samples were subjected to the specificity test as described above, and the results are shown in FIG. 23.
As can be seen in fig. 23, the negative sample was not interfered; the spot with the same color is clear and the separation effect is good at the position corresponding to the chromatogram of the reference substance. The method is proved to have strong specificity and clear spot color development.
4.8.2 durability test
Three batches of kidney-tonifying and turbid-removing particle samples are respectively compared with 3 thin-layer plates of different manufacturers, and 3 different temperatures and humidities are used as variation factors to carry out experiments according to the identification method.
4.8.2.1 thin layer plate tests of different manufacturers, the results are shown in FIGS. 24-26.
The durability experiment results of 3 different manufacturers show that the separation effect of the main spots of the thin-layer identification chart of the rhizoma smilacis glabrae in the kidney tonifying and turbidity removing particles is good, the spots are clear, the thin-layer plates of the different manufacturers have little influence on the main spots, and the repeatability is good.
2.8.2.2 temperature and humidity tests
The different temperature and humidity settings are shown in table 13. The test patterns are shown in FIGS. 27 to 29.
TABLE 13 examination of different temperatures and relative humidities
The experimental spectra of different temperatures and relative humidities show that in the chromatogram of the test sample, fluorescent spots with the same color are displayed at the corresponding positions of the chromatogram of the reference sample, the spots are clear, the separation effect is good, the Rf value is moderate, and the negative control is free of interference. It is thus understood that the temperature and relative humidity vary within the normal environmental range, and do not affect the effectiveness of the identification method.
Claims (9)
1. The detection method of the kidney-tonifying and turbidity-eliminating traditional Chinese medicine preparation is characterized by comprising the following steps: the traditional Chinese medicine preparation comprises the following raw materials in parts by weight: 1-75 parts of astragalus root, 1-50 parts of salvia miltiorrhiza, 1-50 parts of schisandra chinensis, 1-30 parts of glabrous greenbrier rhizome and 1-30 parts of sweet wormwood; the detection method comprises the steps of identifying the astragalus, the schisandra and the glabrous greenbrier rhizome in the traditional Chinese medicine preparation by adopting a thin-layer chromatography, and determining the content of salvianolic acid B in the salvia miltiorrhiza in the traditional Chinese medicine preparation by adopting a high performance liquid chromatography.
2. The method for detecting the kidney-tonifying and turbidity-eliminating traditional Chinese medicine preparation as claimed in claim 1, wherein the method comprises the following steps: the identification method of the astragalus comprises the following steps: collecting 2g Chinese medicinal preparation, adding water 20ml, ultrasonic extracting for 10min, filtering, extracting filtrate with water saturated n-butanol solution for 3 times, each time 20ml, mixing n-butanol solutions, washing with ammonia solution for 2 times, each time 20ml, discarding ammonia solution, concentrating n-butanol solution to dryness, dissolving residue with methanol 1ml to obtain sample solution; adding methanol into astragaloside IV reference substance to obtain 1mg solution per 1ml as reference substance solution; absorbing 5-10 ul of each of the test solution and the reference solution by thin layer chromatography, respectively dropping on the same silica gel G thin layer plate, developing with chloroform-methanol-water lower layer solution as developing agent at volume ratio of 13:6:2, taking out, air drying, spraying with 10% sulfuric acid ethanol solution, heating at 105 deg.C until the spots are clearly developed, and inspecting in sunlight.
3. The method for detecting the kidney-tonifying and turbidity-eliminating traditional Chinese medicine preparation as claimed in claim 1, wherein the method comprises the following steps: the identification method of the schisandra chinensis comprises the following steps: taking 4g of the traditional Chinese medicine preparation, adding 20ml of trichloromethane, carrying out ultrasonic extraction for 30 minutes, filtering, concentrating the filtrate to be dry, and adding 1ml of trichloromethane into residues to dissolve the residues to be used as a test solution; adding chloroform 20ml into fructus Schisandrae control 1g, reflux extracting for 30 min, filtering, concentrating the filtrate to dryness, and dissolving the residue with chloroform 1ml to obtain control solution; according to thin-layer chromatography, absorbing 5-10 ul of each of the sample solution and the reference solution, respectively dropping on the same silica gel G plate thin-layer plate, taking the petroleum ether-ethyl formate-formic acid upper layer solution as a developing agent according to the volume ratio of 10:6:1, developing, taking out, air drying, and inspecting under an ultraviolet lamp.
4. The detection method of the kidney-tonifying and turbidity-eliminating traditional Chinese medicine preparation according to claim 1, characterized by comprising the following steps: the identification method of the glabrous greenbrier rhizome comprises the following steps: collecting 2g Chinese medicinal preparation, adding water 20ml, treating with ultrasound for 10min, filtering, introducing the filtrate into macroporous adsorbent resin column, eluting with 50ml water, discarding water solution, eluting with 30% ethanol, discarding the first 75ml, collecting the second 100ml, evaporating to dryness, and dissolving the residue with 5ml methanol to obtain sample solution; adding methanol into astilbin reference substance to obtain 0.1mg solution per 1ml as reference substance solution; according to thin layer chromatography, absorbing 5-10 ul of each of the sample solution and the reference solution, respectively dropping on the same silica gel G thin layer plate, developing with toluene-ethyl acetate-formic acid as developing agent at volume ratio of 1.5:3:0.9, taking out, air drying, spraying with aluminum trichloride test solution, standing for 5min, and viewing under ultraviolet lamp.
5. The method for detecting the kidney-tonifying and turbidity-eliminating traditional Chinese medicine preparation as claimed in claim 1, wherein the method comprises the following steps: the method for measuring the content of the salvianolic acid B in the salvia miltiorrhiza comprises the following steps: adopting high performance liquid chromatography, taking salvianolic acid B as a reference, taking methanol-acetonitrile-formic acid-water with the volume ratio of 28:7:1:63 as a mobile phase, detecting the wavelength of 284-288 nm, respectively sucking 10 mul of reference solution and sample solution, injecting into a liquid chromatograph, and measuring to obtain the final product.
6. The method for detecting the kidney-tonifying and turbidity-eliminating traditional Chinese medicine preparation according to claim 5, wherein the method comprises the following steps: the preparation method of the reference substance solution comprises the following steps: weighing salvianolic acid B reference substance, adding 75% methanol to obtain 65 μ g solution per 1ml, to obtain Saviae Miltiorrhizae radix reference substance solution; the preparation method of the test solution comprises the following steps: taking 0.1g of the traditional Chinese medicine preparation, adding 24ml of 75% methanol, carrying out ultrasonic extraction for 10 minutes, taking out, adding 75% methanol to 25ml, filtering by a microporous membrane, and taking a subsequent filtrate, namely the test solution.
7. The method for detecting the kidney-tonifying and turbidity-eliminating traditional Chinese medicine preparation as claimed in claim 1, wherein the method comprises the following steps: the traditional Chinese medicine preparation comprises the following raw materials in parts by weight: 75 parts of astragalus root, 50 parts of salvia miltiorrhiza, 50 parts of schisandra chinensis, 30 parts of glabrous greenbrier rhizome and 30 parts of sweet wormwood herb.
8. The method for detecting the kidney-tonifying and turbidity-eliminating traditional Chinese medicine preparation as claimed in claim 1, wherein the method comprises the following steps: the preparation method of the traditional Chinese medicine preparation comprises the following steps: crushing schisandra chinensis, extracting the crushed schisandra chinensis, astragalus mongholicus, salvia miltiorrhiza, rhizoma smilacis glabrae and sweet wormwood with water for three times, wherein each time is 1 hour, filtering an extracting solution, combining filtrates, concentrating the filtrate under reduced pressure to obtain a concentrated solution with the relative density of 1.05-1.10, placing the concentrated solution to room temperature, adding ethanol to ensure that the alcohol content of the concentrated solution is 50%, stirring, standing for 15 hours, and taking a supernatant; centrifuging the remaining precipitate and collecting the supernatant; and combining the two parts of supernatant, recovering ethanol, concentrating under reduced pressure to obtain thick paste with the relative density of 1.35-1.40, and drying under reduced pressure to obtain the traditional Chinese medicine composition.
9. The detection method of the kidney-tonifying and turbidity-eliminating traditional Chinese medicine preparation according to claim 1, characterized by comprising the following steps: the dosage form of the traditional Chinese medicine preparation comprises any one of granules, tablets, pills, capsules, medicinal granules, oral liquid, spraying agents, injections, suspensions and microsphere preparations.
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