CN113759043A - Characteristic spectrum of motherwort fruit and construction method thereof, method for measuring content of stachydrine hydrochloride, motherwort fruit formula granules and preparation method thereof - Google Patents
Characteristic spectrum of motherwort fruit and construction method thereof, method for measuring content of stachydrine hydrochloride, motherwort fruit formula granules and preparation method thereof Download PDFInfo
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Abstract
The invention belongs to the technical field of traditional Chinese medicine detection, and particularly relates to a characteristic spectrum of motherwort fruit, a construction method of the characteristic spectrum, a stachydrine hydrochloride content measuring method, motherwort fruit formula particles and a preparation method of the motherwort fruit formula particles. By controlling the gradient program, the characteristic spectrum of the motherwort fruit can be constructed by a specific method, and the characteristic spectrum is reliable, stable and good in repeatability, so that a more scientific, reasonable and detailed basis is provided for quality control of the motherwort fruit. The construction method changes the peak time by adjusting the flow rate, shortens the operation time, is easy to operate and realize, can fully reflect the information of characteristic peaks of the motherwort fruits, displays the chemical component characteristics of the motherwort fruits, keeps good separation degree and peak shape between main components, can comprehensively evaluate the motherwort fruits of different production places and the quality thereof, and is quick and effective.
Description
Technical Field
The invention belongs to the technical field of traditional Chinese medicine detection, and particularly relates to a characteristic spectrum of motherwort fruit and a construction method thereof, a stachydrine hydrochloride content measuring method, motherwort fruit formula particles and a preparation method thereof; more particularly, the invention relates to a characteristic spectrum of motherwort fruit formula particles, decoction pieces, freeze-dried powder or medicinal materials and a construction method thereof, a method for measuring the content of threonine hydrochloride in motherwort fruit, the motherwort fruit formula particles and a preparation method thereof.
Background
The motherwort fruit is dried mature fruit of Leonurus japonicus Houtt. of Labiatae, has the efficacies of mainly activating blood and regulating menstruation, and clearing liver and improving vision, is a commonly used gynecological traditional Chinese medicine, and is distributed in most regions of the country. In autumn, the overground part of the fruit is harvested when the fruit is ripe, the fruit is dried in the sun, and impurities are removed. According to the reports of the existing documents, the analysis research on the composition of the motherwort fruit by some students at present shows that the motherwort fruit mainly contains chemical components such as alkaloid, fatty oil, flavone, volatile oil and the like, and is rich in various amino acids, mineral elements and the like. Wherein, the stachydrine hydrochloride is monomer alkaloid with high content and strong activity, and has the effects of promoting blood circulation, regulating menstruation, inducing diuresis, relieving swelling, contracting uterus and the like. The content of stachydrine hydrochloride in motherwort fruit medicinal materials is specified in 'Chinese pharmacopoeia' of 2020 edition, but the sample preparation method is complex and has large energy loss; in addition, the method for evaluating the quality of motherwort fruit in the prior art needs to be further improved, and the motherwort fruit with different production areas and the quality thereof cannot be comprehensively evaluated.
Disclosure of Invention
Therefore, the invention provides a characteristic spectrum of motherwort fruit formula particles, decoction pieces, freeze-dried powder or medicinal materials, a construction method thereof and a method for measuring the content of threonine hydrochloride in the motherwort fruit, and aims to overcome the defects that the prior art cannot comprehensively evaluate the quality of the motherwort fruit in different production places, and the method for measuring the content of threonine hydrochloride in the motherwort fruit is complex and has large energy consumption.
Furthermore, the invention also provides a basis for preparing the motherwort fruit formula particles meeting the quality standard in the field, and further provides the motherwort fruit formula particles and the preparation method thereof.
Therefore, the invention provides the following technical scheme.
The invention provides a method for constructing a characteristic spectrum of motherwort fruit formula particles, decoction pieces, freeze-dried powder or medicinal materials, which comprises the following steps,
(1) preparing a motherwort fruit test solution;
(2) detecting the motherwort fruit sample solution by adopting a high performance liquid chromatography, taking a potassium dihydrogen phosphate solution as a mobile phase, and performing gradient elution, wherein the gradient elution procedure comprises the following steps: 0-16min, and the flow rate of the mobile phase is 0.6-0.8 ml/min; 16-17min, the flow rate is 0.6-0.8 → 0.9-1.1 ml/min; 17-41min, and the flow rate of the mobile phase is 0.9-1.0 ml/min; 41-41.1min, the flow rate of the mobile phase is 0.9-1.0 → 0.8 ml/min; 41.1min, and the flow rate of the mobile phase is 0.8 ml/min.
Preferably, an ion exchange method among high performance liquid chromatography is used.
The chromatographic conditions of the high performance liquid chromatography further comprise: the detection wavelength is 190-194nm, and the column temperature is 28-32 ℃;
preferably, the potassium dihydrogen phosphate solution also comprises triethylamine and phosphoric acid;
preferably, the volume fraction of triethylamine in the potassium dihydrogen phosphate solution is 0.04-0.07%, and the volume fraction of phosphoric acid is 0.12-0.1%;
the concentration of the potassium dihydrogen phosphate solution is 13-16 mmol/L;
0-16min, and the flow rate of the mobile phase is 0.8 ml/min; 16-17min, the flow rate of the mobile phase is 0.8-1.0 ml/min; 17-41min, and the flow rate of the mobile phase is 1.0 ml/min; 41-41.1min, and the flow rate of the mobile phase is 1.0-0.8 ml/min; 41.1min, and the flow rate of the mobile phase is 0.8 ml/min.
The construction method also comprises a step of preparing a reference substance by adopting stachydrine hydrochloride, and a step of detecting a reference substance solution according to the construction method of claim 1 or 2 to obtain a reference substance characteristic map;
preferably, the preparation method of the control solution comprises the following steps: taking stachydrine hydrochloride reference substance, and adding solvent to make into stachydrine hydrochloride 0.1-0.3mg per 1 ml.
The preparation method of the motherwort fruit test sample solution comprises the steps of taking a test sample, adding a solvent, and extracting to obtain a test sample solution;
preferably, the solvent for extraction is at least one of aqueous ethanol, methanol, ethyl acetate and water;
the extraction mode is one of ultrasonic, shaking and refluxing;
the extraction time is not less than 20min, preferably 20-40 min.
When the sample to be detected is freeze-dried powder, decoction pieces or medicinal materials, the preparation method of the test solution specifically comprises the following steps: taking the test sample powder, precisely weighing, adding an ethanol-containing aqueous solution, extracting, and filtering to obtain a subsequent filtrate, namely the test sample solution. Wherein the extraction method is preferably ultrasonic extraction.
When the sample to be tested is a formula particle, the preparation method of the test solution specifically comprises the following steps: taking a test sample, precisely weighing, adding water, extracting, and filtering to obtain a subsequent filtrate, namely a test sample solution. Wherein the extraction method is preferably ultrasonic extraction.
The invention provides a motherwort fruit characteristic spectrum obtained by the construction method.
The invention also provides a motherwort fruit characteristic spectrum, wherein the characteristic spectrum at least has 5 characteristic peaks;
taking the No. 3 peak as a reference peak, the relative retention time of each characteristic peak is within +/-10% of a specified value, and the specified value is as follows: 0.585 (peak 1), 0.613 (peak 2), 1.077 (peak 4), 1.288 (peak 5).
The invention provides a method for measuring the content of threonine hydrochloride in motherwort fruit, which comprises the following steps,
(1) preparing a stachydrine hydrochloride reference product solution and a motherwort fruit test product solution;
(2) determining the characteristic spectrums of the leonurus fruit reference product and the leonurus fruit test product according to the construction method of the leonurus fruit characteristic spectrum, and obtaining the content of the stachydrine hydrochloride in the leonurus fruit according to the China pharmacopoeia 2020 edition rule 0512.
In addition, the invention provides a preparation method of motherwort fruit formula granules, which comprises the following steps,
(1) extracting motherwort fruit medicinal materials or decoction pieces to obtain a motherwort fruit extracting solution;
(2) concentrating the fructus Leonuri extractive solution to obtain fructus Leonuri concentrated solution with relative density of 1.05-1.1;
(3) preheating the motherwort fruit concentrated solution, performing spray drying to obtain spray dried powder, and granulating to obtain the motherwort fruit formula granules.
The extraction rate of the motherwort fruit extracting solution is 4-8%;
preferably, the concentration temperature is 50-80 ℃, and the concentration time is not higher than 24 h;
more preferably, the inlet air temperature for spray drying is 170-185 ℃.
The extraction comprises the steps of extracting a motherwort fruit medicinal material or decoction pieces by a water decoction method to obtain a motherwort fruit extracting solution;
preferably, the motherwort fruit medicinal material or decoction pieces are decocted and extracted by at least two times to obtain a motherwort fruit extracting solution;
more preferably, 9-13 times of water is added into the motherwort fruit medicinal material or decoction pieces, boiling extraction is carried out for 60-90min, and the extraction is repeated at least twice to obtain the motherwort fruit extracting solution.
In the spray drying process, adding a first auxiliary material into the concentrated solution, and performing spray drying, wherein the addition amount of the first auxiliary material is 0-3% of the weight of the decoction pieces.
And adding a second auxiliary material into the spray-dried powder, and granulating to obtain a formula granule, wherein the adding amount of the second auxiliary material is 0-3% of the amount of the decoction pieces.
Further, the invention provides the motherwort fruit formula particle prepared by the method.
The technical scheme of the invention has the following advantages:
1. the method for constructing the characteristic spectrum of the motherwort fruit comprises the following steps of (1) preparing a motherwort fruit test product solution; (2) detecting the motherwort fruit sample solution by adopting a high performance liquid chromatography, taking a potassium dihydrogen phosphate solution as a mobile phase, and performing gradient elution, wherein the gradient elution procedure comprises the following steps: 0-16min, and the flow rate of the mobile phase is 0.6-0.8 ml/min; 16-17min, the flow rate is 0.6-0.8 → 0.9-1.1 ml/min; 17-41min, and the flow rate of the mobile phase is 0.9-1.0 ml/min; 41-41.1min, the flow rate of the mobile phase is 0.9-1.0 → 0.8 ml/min; 41.1min, and the flow rate of the mobile phase is 0.8 ml/min. The construction method provided by the invention can be used for constructing the motherwort fruit characteristic spectrum which is reliable, stable and good in repeatability, and provides a more scientific, reasonable and detailed basis for the quality control of the motherwort fruit. The construction method changes the peak time by adjusting the flow rate, shortens the operation time, is easy to operate and realize, can fully reflect the information of characteristic peaks of the motherwort fruits, displays the chemical component characteristics of the motherwort fruits, keeps good separation degree and peak shape between main components, can comprehensively evaluate the motherwort fruits of different production places and the quality thereof, and is quick and effective.
2. The method for measuring the content of the threonine hydrochloride in the motherwort fruit provided by the invention is wider in application range, higher in accuracy and stronger in applicability.
3. According to the preparation method of the motherwort fruit formula particles, provided by the invention, the effective ingredients in the motherwort fruit decoction pieces can be converted into the formula particles, so that the loss is reduced, and the powder yield of the prepared formula particles is high by optimizing the extraction process.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, and it is obvious that the drawings in the following description are some embodiments of the present invention, and other drawings can be obtained by those skilled in the art without creative efforts.
FIG. 1 is a feature spectrum of 17 batches of motherwort fruit freeze-dried powder in example 1 of the present invention;
FIG. 2 is a control profile in example 1 of the present invention;
FIG. 3 is a characteristic map obtained by the gradient elution program A of section 1.1 in Experimental example 1 of the present invention;
FIG. 4 is a characteristic map obtained by the gradient elution program B of section 1.1 in Experimental example 1 of the present invention;
FIG. 5 is a characteristic map obtained by the gradient elution program C of section 1.1 in Experimental example 1 of the present invention;
FIG. 6 is a feature map of section 2.1 of the investigation of specificity in Experimental example 2 of the present invention;
FIG. 7 is a feature map of the invention in Experimental example 2, section 2.1 for specificity study;
FIG. 8 is a feature map of 2.2 sections of delay test in Experimental example 2 of the present invention;
FIG. 9 is a linear graph of section 2.7 in Experimental example 2 of the present invention.
Detailed Description
The following examples are provided to further understand the present invention, not to limit the scope of the present invention, but to provide the best mode, not to limit the content and the protection scope of the present invention, and any product similar or similar to the present invention, which is obtained by combining the present invention with other prior art features, falls within the protection scope of the present invention.
The examples do not show the specific experimental steps or conditions, and can be performed according to the conventional experimental steps described in the literature in the field. The reagents or instruments used are not indicated by manufacturers, and are all conventional reagent products which can be obtained commercially.
Reagent and apparatus
Reagent: potassium dihydrogen phosphate: the top grade is pure; phosphoric acid: carrying out chromatographic purification; triethylamine: carrying out chromatographic purification; acetonitrile: carrying out chromatographic purification; methanol: carrying out chromatographic purification; ethanol: analyzing and purifying; the water is distilled water of Drech.
The instrument comprises the following steps: ML204T electronic balance (Mettler Torlo), XY2000-2C electronic balance (Changzhou lucky electronics Co., Ltd.), MSA125P-1CE-DI electronic balance (Sadous scientific instruments (Beijing) Co., Ltd.), Waters Acquity Arc high performance liquid chromatograph.
A chromatographic column:
SCX (4.6X 250mm) chromatography column
Reagent testing: stachydrine hydrochloride reference (batch No. 110712) -201614, China institute for testing food and drug).
The batch numbers of the motherwort fruit freeze-dried powder are respectively as follows: YP 190801-015000-01; YP190801-473500-02, YP 190907-024300-03; YP 190915-236000-04; YP 190915-473000-05; YP 190915-236000-06; YP 190915-237200-07; YP 190915-237200-08; YP 190915-237200-09; YP 190915-237200-10; YP 190915-237200-11; YP 190915-237200-12; YP 190927-014100-14; YP 190927-014100-15; YP 190927-014100-16; YP 190927-014100-17; YP 190927-014100-18; corresponding in turn to S1-S17.
The motherwort fruit decoction piece batch number and the production place are 190801-; 190801 473500-02, new county of south Yang city, Henan province; 190907 024300-03, and Aohan flag in the city of Rifeng in autonomous region of inner Mongolia; 190915-; 190915 + 473000-05, south-Yang City of Henan province; 190915-; 190915 237200-07, Hooshan City, Anhui province; 190915-; 190915 + 130000-09, Liaoning province; 190915, 230000-10, Anhui; 190915, 230000-11, Anhui; 190915 + 130000-12, Jilin province; 190927-014100-14, the right flag of Germet of city in autonomous region of inner Mongolia; 190927-4750-15 of Asclepias edulis, Kalopanax pictus county of south-south China; 190927-471600-16, Luoyang city, Henan province; 190927-152300-17, Hailon city, Suizhihua city, Heilongjiang province; 190927-629000-18, Tunning City and mountain areas of Sichuan province.
Example 1
The embodiment provides a method for constructing a characteristic spectrum of motherwort fruit freeze-dried powder, which comprises the following steps,
(1) preparation of control solutions: taking stachydrine hydrochloride, adding 70% ethanol to prepare a reference substance solution containing 0.2mg of stachydrine hydrochloride per 1 ml;
preparation of a test solution: collecting 0.1g of fructus Leonuri lyophilized powder, grinding, precisely weighing, placing in a conical flask, precisely adding 10ml of 70% ethanol water solution, sealing, weighing, performing ultrasonic treatment at power of 500W and frequency of 40kHz for 30min, cooling, weighing again, supplementing loss by 70% ethanol water solution, shaking, filtering with 0.22 μm microporous membrane, and collecting filtrate.
(2) Respectively and precisely sucking 10 mu L of test solution and reference solution, injecting into a high performance liquid chromatograph, and measuring under the chromatographic conditions: to be provided with
SCX (4.6X 250mm, 5 μm) column, 15mmol/L potassium dihydrogen phosphate solution (containing 0.06% triethylamine and 0.14% phosphoric acid) as mobile phase, gradient elution: 0-16min, and the flow rate of the mobile phase is 0.8 ml/min; 16-17min, the flow rate of the mobile phase is 0.8-1.0 ml/min; 17-41min, and the flow rate of the mobile phase is 1.0 ml/min; 41-41.1min, and the flow rate of the mobile phase is 1.0-0.8 ml/min; 41.1min, and the flow rate of the mobile phase is 0.8 ml/min; the column temperature is 30 ℃; the detection wavelength was 192 nm.
The embodiment also provides a method for measuring the content of stachydrine hydrochloride in the motherwort fruit freeze-dried powder, which comprises the following steps,
the characteristic spectrums of the stachydrine hydrochloride reference substance solution and the test substance solution are obtained according to the method, the content of the stachydrine hydrochloride in the motherwort fruit is obtained according to the China pharmacopoeia 2020 edition convention 0512, and the calculation formula is as follows:
wherein, WTest articleRepresenting the peak area of stachydrine hydrochloride in the characteristic spectrum of the sample, CReference substanceRepresents the concentration of the stachydrine hydrochloride reference solution, VReference substanceRepresents hydrochloric acidVolume of stachydrine control solution, WReference substanceRepresents the peak area of the stachydrine hydrochloride control, CTest articleRepresents the concentration of the test solution, VTest articleRepresents the volume of the stachydrine hydrochloride control solution.
Example 2
The embodiment provides a method for constructing a characteristic spectrum of motherwort fruit freeze-dried powder, which comprises the following steps,
(1) preparation of control solutions: taking stachydrine hydrochloride, adding 70% ethanol to prepare a reference substance solution containing 0.23mg of stachydrine hydrochloride per 1 ml;
preparation of a test solution: collecting 0.1g of fructus Leonuri lyophilized powder, grinding, precisely weighing, placing in a conical flask, precisely adding 10ml of 70% ethanol water solution, sealing, weighing, performing ultrasonic treatment at power of 500W and frequency of 40kHz for 30min, cooling, weighing again, supplementing loss by 70% ethanol water solution, shaking, filtering with 0.22 μm microporous membrane, and collecting filtrate.
(2) Respectively and precisely sucking 10 mu L of test solution and reference solution, injecting into a high performance liquid chromatograph, and measuring under the chromatographic conditions: to be provided with
SCX (4.6X 250mm) column, 15mmol/L potassium dihydrogen phosphate solution (containing 0.06% triethylamine and 0.14% phosphoric acid) as mobile phase, gradient elution: 0-16min, and the flow rate of the mobile phase is 0.8 ml/min; 16-17min, the flow rate of the mobile phase is 0.8-1.0 ml/min; 17-41min, and the flow rate of the mobile phase is 1.0 ml/min; 41-41.1min, and the flow rate of the mobile phase is 1.0-0.8 ml/min; 41.1min, and the flow rate of the mobile phase is 0.8 ml/min; the column temperature was 28 ℃; the detection wavelength was 190 nm.
Example 3
The embodiment provides a method for constructing a characteristic spectrum of motherwort fruit decoction pieces, which comprises the following steps,
(1) preparation of control solutions: taking stachydrine hydrochloride, adding 70% ethanol to prepare a reference substance solution containing 0.2mg of stachydrine hydrochloride per 1 ml;
preparation of a test solution: weighing 1g of motherwort fruit decoction pieces powder, precisely weighing, placing in a conical flask, precisely adding 10ml of 70% ethanol aqueous solution, sealing, weighing, performing ultrasonic treatment at power of 500W and frequency of 40kHz for 30min, cooling, weighing again, supplementing lost weight with 70% ethanol aqueous solution, shaking, filtering with 0.22 μm microporous membrane, and collecting filtrate.
(2) Respectively and precisely sucking 10 mu L of test solution and reference solution, injecting into a high performance liquid chromatograph, and measuring under the chromatographic conditions: to be provided with
SCX (4.6X 250mm, 5 μm) column, 15mmol/L potassium dihydrogen phosphate solution (containing 0.06% triethylamine and 0.14% phosphoric acid) as mobile phase, gradient elution: 0-16min, and the flow rate of the mobile phase is 0.8 ml/min; 16-17min, the flow rate of the mobile phase is 0.8-1.0 ml/min; 17-41min, and the flow rate of the mobile phase is 1.0 ml/min; 41-41.1min, and the flow rate of the mobile phase is 1.0-0.8 ml/min; 41.1min, and the flow rate of the mobile phase is 0.8 ml/min; the column temperature is 30 ℃; the detection wavelength was 192 nm.
The embodiment also provides a method for measuring the content of stachydrine hydrochloride in the motherwort fruit decoction pieces, which comprises the following steps,
the characteristic spectrums of the stachydrine hydrochloride reference substance solution and the test substance solution are obtained according to the method, the content of the stachydrine hydrochloride in the motherwort fruit is obtained according to the China pharmacopoeia 2020 edition convention 0512, and the calculation formula is as follows:
wherein, WTest articleRepresenting the peak area of stachydrine hydrochloride in the characteristic spectrum of the sample, CReference substanceRepresents the concentration of the stachydrine hydrochloride reference solution, VReference substanceRepresents the volume of the stachydrine hydrochloride control solution, WReference substanceRepresents the peak area of the stachydrine hydrochloride control, CTest articleRepresents the concentration of the test solution, VTest articleRepresents the volume of the stachydrine hydrochloride control solution.
Example 4
The embodiment provides a method for constructing a characteristic spectrum of motherwort fruit formula particles, which comprises the following steps,
(1) preparation of control solutions: taking stachydrine hydrochloride, adding 70% ethanol to prepare a reference substance solution containing 0.2mg of stachydrine hydrochloride per 1 ml;
preparation of a test solution: collecting 0.1g fructus Leonuri granule, grinding, precisely weighing, placing in a conical flask, precisely adding 10ml water, sealing, weighing, performing ultrasonic treatment at power of 500W and frequency of 40kHz for 30min, cooling, weighing, supplementing with 70% ethanol water solution, shaking, filtering with 0.22 μm microporous membrane, and collecting filtrate.
(2) Respectively and precisely sucking 10 mu L of test solution and reference solution, injecting into a high performance liquid chromatograph, and measuring under the chromatographic conditions: to be provided with
SCX (4.6X 250mm, 5 μm) column, 15mmol/L potassium dihydrogen phosphate solution (containing 0.06% triethylamine and 0.14% phosphoric acid) as mobile phase, gradient elution: 0-16min, and the flow rate of the mobile phase is 0.8 ml/min; 16-17min, the flow rate of the mobile phase is 0.8-1.0 ml/min; 17-41min, and the flow rate of the mobile phase is 1.0 ml/min; 41-41.1min, and the flow rate of the mobile phase is 1.0-0.8 ml/min; 41.1min, and the flow rate of the mobile phase is 0.8 ml/min; the column temperature is 30 ℃; the detection wavelength was 192 nm.
The embodiment also provides a method for measuring the content of stachydrine hydrochloride in the motherwort fruit formula particles, which comprises the following steps,
the characteristic spectrums of the stachydrine hydrochloride reference substance solution and the test substance solution are obtained according to the method, the content of the stachydrine hydrochloride in the motherwort fruit is obtained according to the China pharmacopoeia 2020 edition convention 0512, and the calculation formula is as follows:
wherein, WTest articleRepresenting the peak area of stachydrine hydrochloride in the characteristic spectrum of the sample, CReference substanceRepresenting stachydrine hydrochloride reference solutionsConcentration, VReference substanceRepresents the volume of the stachydrine hydrochloride control solution, WReference substanceRepresents the peak area of the stachydrine hydrochloride control, CTest articleRepresents the concentration of the test solution, VTest articleRepresents the volume of the stachydrine hydrochloride control solution.
Experimental example 1 creation of characteristic map
Preparation of control solutions: taking stachydrine hydrochloride, adding 70% ethanol to prepare a reference substance solution containing 0.2mg of stachydrine hydrochloride per 1 ml;
preparation of a test solution: collecting 0.1g of fructus Leonuri lyophilized powder, grinding, precisely weighing, placing in a conical flask, precisely adding 10ml of 70% ethanol water solution, sealing, weighing, performing ultrasonic treatment at power of 500W and frequency of 40kHz for 30min, cooling, weighing again, supplementing loss by 70% ethanol water solution, shaking, filtering with 0.22 μm microporous membrane, and collecting filtrate.
Precisely sucking 10 μ L of each of the test solution and the reference solution, injecting into a high performance liquid chromatograph, and measuring under the chromatographic conditions: to be provided with
SCX (4.6X 250mm) column, 15mmol/L potassium dihydrogen phosphate solution (containing 0.06% triethylamine and 0.14% phosphoric acid) as mobile phase, gradient elution: 0-16min, and the flow rate of the mobile phase is 0.8 ml/min; 16-17min, the flow rate of the mobile phase is 0.8-1.0 ml/min; 17-41min, and the flow rate of the mobile phase is 1.0 ml/min; 41-41.1min, and the flow rate of the mobile phase is 1.0-0.8 ml/min; 41.1min, and the flow rate of the mobile phase is 0.8 ml/min; the column temperature is 30 ℃; the detection wavelength was 192 nm.
Obtaining the characteristic spectrum of the motherwort fruit freeze-dried powder and the reference product solution of the stachydrine hydrochloride, and selecting 5 characteristic peaks according to the principles of good stability, separation degree and peak shape of relative retention time.
The characteristic map has 5 characteristic peaks, the No. 3 peak is taken as a reference peak, the relative retention time of each characteristic peak is within +/-10% of a specified value, and the specified value is as follows: 0.585 (peak 1), 0.613 (peak 2), 1.077 (peak 4), 1.288 (peak 5); wherein, the peak No. 3 is stachydrine hydrochloride.
17 batches of motherwort fruit freeze-dried powder (S1-S17) are subjected to characteristic spectrum obtaining according to the method and content of stachydrine hydrochloride according to the method of the embodiment 1, analysis is carried out by software of a traditional Chinese medicine chromatography fingerprint spectrum similarity evaluation system (2012 edition), a No. 3 peak is taken as a reference substance peak, after multi-point correction, automatic matching is carried out to obtain 17 batches of sample superposition spectrums and comparison characteristic spectrums thereof, 5 common peaks are determined, as shown in figures 1 and 2, S1-S17 are sequentially corresponded to in figure 1 from bottom to top, the similarity evaluation result is shown in Table 1, and the similarity value of 17 batches of motherwort fruit samples is between 0.981 and 1.000. The content of stachydrine hydrochloride in 17 batches of motherwort fruit freeze-dried powder is shown in table 2, unit mg/g represents the mass of the stachydrine hydrochloride contained in each 1g of motherwort fruit freeze-dried powder.
Table 117 motherwort fruit Lonicera evaluation results
TABLE 217 fructus Leonuri lyophilized powder with stachydrine hydrochloride content
| Batch number | Stachydrine hydrochloride content mg/g |
| 190801-015000-01 | 33.4 |
| 190801-473500-02 | 20.3 |
| 190907-024300-03 | 31.2 |
| 190915-236000-04 | 27.7 |
| 190915-473000-05 | 29.6 |
| 190915-236000-06 | 20.1 |
| 190915-237200-07 | 29.9 |
| 190915-237200-08 | 32.7 |
| 190915-237200-09 | 21.3 |
| 190915-237200-10 | 42.8 |
| 190915-237200-11 | 27.2 |
| 190915-237200-12 | 15.1 |
| 190927-014100-14 | 20.4 |
| 190927-014100-15 | 20.1 |
| 190927-014100-16 | 28.2 |
| 190927-014100-17 | 34.3 |
| 190927-014100-18 | 24.7 |
1.1 examination of gradient elution procedure
4 parts of the same sample solution is prepared, and the other parts are the same as the experimental example 1 except for different gradient elution procedures, and the characteristic spectrum of the sample is obtained through detection.
A. The gradient elution procedure was: the detection result is shown in figure 3 by isocratic elution with 15mmol/L potassium dihydrogen phosphate solution (containing triethylamine with volume fraction of 0.06% and phosphoric acid with volume fraction of 0.14%) as mobile phase.
B. The gradient elution procedure was: gradient elution is carried out by taking 15mmol/L potassium dihydrogen phosphate solution (containing 0.06% triethylamine and 0.14% phosphoric acid) as mobile phase A and acetonitrile as mobile phase B; the flow rate is 0.8mL/min, and the column temperature is 30 ℃; wherein, the gradient elution procedure is as follows: 0-22min, the mobile phase A is 100%, and the mobile phase B is 0%; 22-25min, the mobile phase A is 100-90%, and the mobile phase B is 0-10%; 25-40min, 90% for mobile phase A and 10% for mobile phase B, the results are shown in FIG. 4.
C. The gradient elution procedure was: taking 15mmol/L potassium dihydrogen phosphate solution (containing 0.06% triethylamine and 0.14% phosphoric acid) as mobile phase A, and gradient eluting; the flow rate is 0.8 mL/min; wherein, the gradient elution procedure is as follows: 0-16min, the flow rate is 0.8 ml/min; 16-17min, and the flow rate is 0.8-1.0 ml/min; 17-42min, with flow rate of 1 ml/min; 42-43min, with flow rate of 1.0-0.8 ml/min; 43-45min, flow rate of 0.8ml/min, and results are shown in FIG. 5.
D. The gradient elution procedure was as in example 1.
The characteristic spectrum obtained by the gradient elution program A has a larger solvent peak in 2-6min and a large peak in 48-50 min; the gradient elution procedure B found that the addition of mobile phase B (acetonitrile) caused a large shift in chromatographic peak retention time, which was not feasible; the chromatographic peak of the characteristic spectrum obtained under the chromatographic condition C has a good separation effect, no chromatographic peak appears after 41min, and the chromatographic peak obtained through the gradient elution procedure D has a good separation degree effect, rich information and high reproducibility.
Experimental example 2 methodological examination
2.1 specificity
Taking 70% ethanol water solution as a blank sample, sampling stachydrine hydrochloride reference substance solution, and sampling by adopting the method of the embodiment 1 to obtain the characteristic maps of the blank sample and the reference substance solution, and referring to the figures 6 and 7, and discharging the interference of the ethanol water solution.
2.2 retardation
The test solution was injected as in example 1 and subjected to the delayed test, as shown in FIG. 8, and no other peak appeared after 40 min.
2.3 repeatability
Motherwort fruit lyophilized powder (batch No. 190907-. The results in tables 3-4 show that the relative retention time RSD of 5 characteristic peaks is 0.0% -0.1%, and the RSD of the relative peak area is 0.0% -4.7%, which indicates that the repeatability of the characteristic spectrum is good; the results in table 5 show that the RSD value of the stachydrine hydrochloride measurement result is 2.2%, which meets the requirement of the fourth part 9101 (drug quality standard analysis method verification guideline) of 2020 edition in the "chinese pharmacopoeia" on the repeatability of the method.
TABLE 3 Retention time and relative Retention time of the individual characteristic peaks
TABLE 4 Peak area and relative Peak area of each characteristic Peak
TABLE 5 stachydrine hydrochloride repeatability test results
2.4 intermediate precision
Different analysts perform intermediate precision tests at different times by using another Waters ultra high performance liquid chromatograph (2998 detector), prepare 6 parts of test sample solution in parallel according to the method of example 1, measure according to the chromatographic conditions, calculate the relative retention time and the relative peak area of each common peak in the characteristic map by using the peak 3 (stachydrine hydrochloride) as the reference peak (S), see tables 6 and 7, and the results show that the relative retention time RSD of 5 identification peaks is 0.0-0.1%, and the relative peak area RSD is 0.0-6.4%, which indicates that the precision of the characteristic map is better. Comparing the samples under the intermediate precision term with the samples under the repeatability term, see tables 8 and 9, the relative retention time RSD is in the range of 0.0-2.2%, and the relative peak area RSD is in the range of 0.0-29.2%, which shows that the intermediate precision of the characteristic spectrum is better.
TABLE 6 retention time and relative retention time of characteristic peak of intermediate precision of fructus Leonuri
TABLE 7 peak area and relative peak area of characteristic peak of intermediate precision of fructus Leonuri
TABLE 8 repeatability and intermediate precision relative retention time
TABLE 9 repeatability and intermediate precision relative peak areas
Intermediate precision tests were performed by different analysts at different times using another waters high performance liquid chromatograph (PDA detector). 6 portions of motherwort fruit freeze-dried powder (batch number: 190907-.
TABLE 10 results of intermediate precision experiments
2.5 stability
Taking a test sample solution (the preparation method is the same as that of example 1), and determining a characteristic spectrum and the peak area of stachydrine hydrochloride according to the method of example 1 at 0 hour, 2 hour, 4 hour, 6 hour, 8 hour, 10 hour, 12 hour, 24 hour, 36 hour and 48 hour respectively, wherein the relative retention time and the relative peak area of each characteristic peak in the characteristic spectrum are shown in tables 11 and 12; meanwhile, the reference solution of stachydrine hydrochloride is taken to measure the peak areas of the reference solution of stachydrine hydrochloride in the steps of 0 hour, 2 hours, 4 hours, 6 hours, 8 hours, 10 hours, 12 hours, 24 hours, 36 hours and 48 hours according to the method in the example 1, and the peak areas of the stachydrine hydrochloride in the test solution and the reference solution are shown in the table 13.
TABLE 11 retention time and relative retention time of the individual characteristic peaks
TABLE 12 Peak area and relative Peak area of each characteristic Peak
TABLE 13 Peak areas of stachydrine hydrochloride in test solution and control at different times
| Time | Peak area of stachydrine hydrochloride in test sample | Peak area of control |
| 0h | 931110 | 598977 |
| 2h | 932025 | 595367 |
| 4h | 939630 | 599855 |
| 6h | 939247 | 600298 |
| 8h | 920596 | 600171 |
| 12h | 940278 | 601613 |
| 24h | 939054 | 584945 |
| 36h | 932449 | 584666 |
| 48h | 942427 | 590494 |
| RSD | 0.7％ | 1.3％ |
In tables 11 and 12, by examining the solution stability for 48 hours, the relative retention time RSD of the characteristic peak of the test solution is in the range of 0.1-0.6%, and the relative peak area RSD% is in the range of 0.0-2.9%. In Table 13, the RSD value of the peak area of the stachydrine hydrochloride reference substance within 48h is 1.3%, and the RSD value of the peak area of the stachydrine hydrochloride peak in the test substance is 0.7%, which indicates that the test substance solution is stable within 48 h.
2.6 accuracy
Preparation of control solutions: precisely weighing appropriate amount of stachydrine hydrochloride reference substance, precisely weighing, and respectively adding 70% ethanol to obtain reference substance solution A containing 0.201mg of stachydrine hydrochloride per 1ml, reference substance solution B containing 0.1419mg of stachydrine hydrochloride per 1ml, and reference substance solution C containing 0.060mg of stachydrine hydrochloride per 1 ml.
Preparation of sample solution: precisely weighing 9 parts of motherwort fruit freeze-dried powder, wherein each part is about 0.05g, precisely weighing the motherwort fruit freeze-dried powder to serve as a test sample, precisely adding 10ml of each of reference substance solutions A-C and 3 parts of each of reference substance solutions A-C into the test sample to obtain 9 parts of sample solutions, detecting according to the method of example 1, and calculating the recovery rate according to the following formula, wherein the results are shown in Table 10;
wherein the measured value refers to the content of stachydrine hydrochloride in the sample solution; the added amount of the control is the amount of the control in the control solution added when the sample solution is prepared.
The recovery rate range of the stachydrine hydrochloride measured by the recovery rate test in the table 14 is 92.03-100.88%, the average recovery rate is 97.69%, and the recovery rate conforms to the regulation of the four parts 9101 (drug quality standard analysis method verification guide principle) in the 2020 edition of Chinese pharmacopoeia, which indicates that the established content measurement method has good accuracy.
TABLE 14 recovery test results
2.7 Linear survey
Reference for precisely weighing stachydrine hydrochlorideAdding 6.993mg of product into a 10ml volumetric flask, adding 70% ethanol water solution to dissolve the product, fixing the volume to the scale, shaking up to obtain stachydrine hydrochloride reference substance solution A, respectively taking 0.5ml, 1ml, 2ml and 4ml of the stachydrine hydrochloride reference substance solution A, placing the solution into the 10ml volumetric flask, fixing the volume to the scale line by using 70% ethanol water solution, and shaking up to obtain four stachydrine hydrochloride reference substance solutions B-E with different concentrations; respectively filtering the stachydrine hydrochloride reference substance solutions A-E with 0.22 mu m microfiltration membranes, taking subsequent filtrates, injecting samples according to the method, wherein the injection amount is 10 mu L, obtaining the characteristic map of the stachydrine hydrochloride reference substance solution, injecting samples twice for each reference substance solution, taking the average value of the peak areas of the two stachydrine hydrochloride characteristic peaks as an ordinate, taking the concentration of the stachydrine hydrochloride reference substance solution as an abscissa, and performing linear regression by a least square method to obtain a regression equation of Y-3479617.3X-11034.0, r-1.0, and the linear range of the regression equation is 0.034965-0.6993 mg/ml-1。
TABLE 15 concentration of stachydrine hydrochloride control solution and characteristic peak area of stachydrine hydrochloride
| Standard curve | A | B | C | D | E |
| Peak area 1 | 2422006 | 103036 | 237082 | 476499 | 961743 |
| |
2421204 | 105136 | 238480 | 477258 | 963753 |
| Mean value of | 2421605 | 104086 | 237781 | 476879 | 962748 |
| Concentration mg.ml-1 | 0.6993 | 0.034965 | 0.06993 | 0.13986 | 0.27972 |
Example 5
The embodiment provides a preparation method of motherwort fruit formula granules, which comprises the following steps,
(1) collecting fructus Leonuri decoction pieces, adding 12 times of water into the first decoction, boiling and extracting for 60min, adding 10 times of water into the second decoction, boiling and extracting for 60min, filtering the medicinal liquid, and mixing to obtain fructus Leonuri extractive solution.
(2) Concentrating the obtained fructus Leonuri extractive solution at 65 deg.C to obtain concentrated solution with relative density of 1.10 (measured at 60 deg.C), and refrigerating for use, wherein the concentration extraction time is no more than 24 hr.
(3) Preheating the obtained concentrated solution to 60 deg.C, adding dextrin as first adjuvant in an amount of 2% of the decoction pieces, spray drying at 175 deg.C, and collecting spray dried powder; adding a second auxiliary material into the spray-dried powder, wherein the adding amount is 2% of the amount of the decoction pieces, uniformly mixing, performing dry granulation, and controlling the compression roller gap of a granulator to be 0.1-1.3 mm; the rotating speed of the press roll is 20 rpm; and (3) granulating under the pressure of 60-90 bar by using a 16-mesh sieve and a 60-mesh sieve, and packaging the granules with a medicinal composite membrane to obtain a finished product.
Experimental example 3
3.1 determination of the extraction Rate of the motherwort fruit extract
Taking 17 batches of motherwort fruit decoction pieces, each 200g, adding 8 times of water into each decoction piece, soaking for 30min, heating to boil, decocting for 30min, filtering while hot by using 150-mesh filter cloth, decocting for 20min while boiling with 6 times of water, filtering while hot by using 150-mesh filter cloth, combining the two filtrates, cooling the liquid medicine in water bath to room temperature to obtain a motherwort fruit extracting solution, and weighing the total weight of the extracting solution liquid medicine. Weighing 1/10 weight parts, placing in evaporating dish (parallel sample), evaporating in water bath, drying at 105 deg.C for 5 hr, cooling in desiccator for 30min, and precisely weighing (M)1) Drying at 105 deg.C for 1 hr, cooling in a desiccator for 30min, weighing to constant weight, and precisely weighing (M)2And the unit is g) the extraction rate of the motherwort fruit extract is calculated according to the formula (1).
In the middle, W is the mass of the decoction pieces, and the unit is g; n is the total mass of the motherwort fruit extracting solution, and the unit is g; m is 1/10 of the total mass of the motherwort fruit extract, and the unit is g; m0The mass of the evaporating dish is given in g.
Concentrating the rest fructus Leonuri extractive solution at 65 deg.C under reduced pressure until the ratio of fluid extract to decoction pieces is about 1:1, transferring to a lyophilizing tray, vacuum lyophilizing (at-35 deg.C, specifically drying at-35 deg.C for 4h, drying at-30 deg.C for 4h, drying at-20 deg.C for 4h, drying at-10 deg.C for 4h, drying at 0 deg.C for 3h, drying at 10 deg.C for 3h, drying at 20 deg.C for 3h, drying at 30 deg.C for 10h, drying at 35 deg.C for 10h, and vacuum degree of 10Pa), collecting lyophilized powder, preparing fructus Leonuri sample solution according to the method of example 1, measuring the content of stachydrine hydrochloride in the sample solution, calculating stachydrine hydrochloride transfer rate according to formula (2), and calculating the stachydrine hydrochloride transfer rate of each batch of fructus Leonuri decoction pieces according to Table 16.
TABLE 16 transfer rate of stachydrine hydrochloride
According to the above experiment results, the average motherwort fruit yield of 17 batches is + -10% as the range of the motherwort fruit standard decoction yield, namely 4.2% -7.8%.
3.2 examination of extraction Process parameters
The influence of the extraction water addition amount, the extraction times and the extraction time on the cream yield and the stachydrine hydrochloride content is examined by adopting an orthogonal test. Performing L by orthogonal experiment with water addition amount, extraction frequency and extraction time as indexes9(34) And performing orthogonal experiment to determine the optimal extraction process condition. The specific experimental steps comprise that 9 parts of motherwort fruit decoction pieces (190907-. The factor levels are shown in Table 17, the experimental protocol is shown in Table 18, and the experimental results are shown in Table 19.
TABLE 17 orthogonal experiment factor horizon
Note: the water addition amount for the second and third extractions is A-2, and A represents the multiple of the water addition amount for the first extraction.
Table 18 orthogonal experimental protocol table
TABLE 19 orthogonal experimental results table
| Test No | Percentage of cream discharged (%) | Stachydrine hydrochloride content (mg/g) | Transfer Rate (%) |
| 1 | 4.4 | 37.5 | 56.9 |
| 2 | 7.1 | 33.1 | 81 |
| 3 | 8.7 | 29.9 | 89.7 |
| 4 | 5.6 | 35.4 | 68.4 |
| 5 | 6.4 | 32.1 | 70.8 |
| 6 | 7.3 | 34.0 | 85.6 |
| 7 | 5.9 | 33.7 | 68.6 |
| 8 | 6.7 | 33.9 | 78.30 |
| 9 | 8.5 | 32.7 | 95.8 |
Note: the content of stachydrine hydrochloride in the decoction pieces is 2.9 mg/g.
The results of the extraction rate and the stachydrine hydrochloride transfer rate in the table 14 are input into Design-expert10.0.4.0 for processing, the results of the motherwort fruit extract are referred to, various factors such as the loss in the comprehensive production process are taken as target values for comprehensive evaluation, and the extraction process combination closest to the standard decoction is preferably as follows: extracting for 2 times, decocting with 12 times of water, boiling and extracting for 1 hr; decocting twice with 10 times of water, boiling and extracting for 1 hour.
3.3 investigation of concentration temperature
Collecting 200g of motherwort fruit decoction pieces (190907-, the content of stachydrine hydrochloride was obtained as shown in Table 20.
TABLE 20 stachydrine hydrochloride content (mg/g) in the different samples
| Temperature of |
50 |
60℃ | 70℃ | 80℃ |
| 0h | 37.48 | 37.48 | 37.48 | 37.48 |
| 3h | 36.76 | 37.03 | 36.94 | 37.32 |
| 6h | 36.98 | 37.29 | 37.27 | 37.84 |
| 9h | 37.19 | 37.41 | 37.66 | 38.39 |
| 12h | 38.19 | 38.40 | 38.25 | 38.56 |
| 24h | 37.34 | 37.57 | 38.57 | 39.32 |
The experimental result shows that the content of the stachydrine hydrochloride has small change amplitude within 24 hours of heating under different conditions of 50-80 ℃, and can be considered to be basically kept unchanged; the content of stachydrine hydrochloride tends to increase or decrease at 24h, so the concentration time is recommended to be controlled within 24 h.
3.4 investigation of spray drying parameters
Collecting fructus Leonuri decoction pieces 3000g, adding water 8 times the decoction pieces into the first decoction, soaking for 30min, boiling and decocting for 30min, adding water 6 times the decoction pieces into the second decoction, boiling and decocting for 20min, filtering the extractive solution with 150 mesh filter cloth, mixing filtrates, concentrating the filtrate at 50 deg.C under reduced pressure to certain density, weighing the concentrated solution, and measuring the solid content. In the production process of the motherwort fruit formula particles, a spray drying mode is adopted, and the optimal spray drying parameters are determined by investigating parameters such as air inlet temperature, the adding amount of the first auxiliary material and the like.
Experimental group 1: concentrating the concentrated solution until the relative density is 1.1, adding no first adjuvant, spray drying at 170 deg.C, setting fan at 35, and peristaltic pump at 20rpm to obtain spray dried powder.
Experimental group 2: concentrating the concentrated solution to relative density of 1.1, adding a first adjuvant (dextrin) into the concentrated solution, wherein the dextrin is 2% of the decoction pieces, the air inlet temperature of spray drying is 170 deg.C, the fan setting is 35, and the peristaltic pump speed is 20rpm, to obtain spray dried powder.
Experimental group 3: concentrating the concentrated solution to relative density of 1.1, adding 2% dextrin into the concentrated solution, spray drying at air inlet temperature of 180 deg.C, setting fan at 35, and peristaltic pump at 20rpm to obtain spray dried powder.
Experimental group 4: concentrating the concentrated solution to relative density of 1.05, adding 2% dextrin into the concentrated solution, spray drying at air inlet temperature of 180 deg.C, setting fan at 35, and peristaltic pump at 20rpm to obtain spray dried powder.
Experimental group 5: concentrating the concentrated solution to relative density of 1.05, adding 4% dextrin into the concentrated solution, spray drying at air inlet temperature of 180 deg.C, setting fan at 35, and peristaltic pump at 20rpm to obtain spray dried powder.
The powder collection rate of the spray-dried powder obtained in experimental groups 1 to 5 was calculated, and the moisture and the dissolution rate in the dried powder were measured. Meanwhile, a test solution was prepared according to the method of example 1, and the content of stachydrine hydrochloride was measured, and the results are shown in Table 21.
Table 21 motherwort fruit spray drying process investigation result
| Experimental group | Percentage of collected pollen (%) | Stachydrine hydrochloride content (mg/g) | Phenomenon of spray drying |
| 1 | 79.62 | 18.6 | The tower sticking phenomenon occurs at the bottom of the |
| 2 | 82.17 | 18.3 | No wall sticking, good powder fluidity |
| 3 | 84.47 | 24.4 | Has slight tower adhesion phenomenon, and the powder is easy to collect |
| 4 | 87.19 | 18.1 | Slight tower adhesion and |
| 5 | 90.10 | 11.9 | The phenomena of tower adhesion and powder leakage are reduced, and the collection is easy |
The experimental groups 2 and 3 show that the powder yield is not obviously different when the air inlet temperature is 170 ℃ and 180 ℃; the experimental group can see that the powder yield is not obviously different when the density is 1.10 and 1.05; in experimental groups 4 and 5, the powder yield is only improved by 2.91% when 4% of auxiliary materials are added, the principle of low auxiliary materials is followed, and the historical process is to add 2% of auxiliary materials, so that the proportion of 2% of auxiliary materials is still maintained.
In summary, when preparing the motherwort fruit formula particles, the air inlet temperature of spray drying is 170-.
It should be understood that the above examples are only for clarity of illustration and are not intended to limit the embodiments. Other variations and modifications will be apparent to persons skilled in the art in light of the above description. And are neither required nor exhaustive of all embodiments. And obvious variations or modifications of the invention may be made without departing from the spirit or scope of the invention.
Claims (11)
1. A method for constructing a feature spectrum of motherwort fruit formula granules, decoction pieces, freeze-dried powder or medicinal materials is characterized by comprising the following steps of,
(1) preparing a motherwort fruit test solution;
(2) detecting the motherwort fruit sample solution by adopting a high performance liquid chromatography, taking a potassium dihydrogen phosphate solution as a mobile phase, and performing gradient elution, wherein the gradient elution procedure comprises the following steps: 0-16min, and the flow rate of the mobile phase is 0.6-0.8 ml/min; 16-17min, the flow rate is 0.6-0.8 → 0.9-1.1 ml/min; 17-41min, and the flow rate of the mobile phase is 0.9-1.0 ml/min; 41-41.1min, the flow rate of the mobile phase is 0.9-1.0 → 0.8 ml/min; 41.1min, and the flow rate of the mobile phase is 0.8 ml/min.
2. The method for constructing according to claim 1, wherein the chromatographic conditions of the high performance liquid chromatography further comprise: the detection wavelength is 190-194nm, the column temperature is 28-32 ℃, and the sample injection amount is 8-12 mu l;
preferably, the potassium dihydrogen phosphate solution also comprises triethylamine and phosphoric acid;
preferably, the volume fraction of triethylamine in the potassium dihydrogen phosphate solution is 0.04-0.07%, and the volume fraction of phosphoric acid is 0.12-0.1%;
the concentration of the potassium dihydrogen phosphate solution is 13-16 mmol/L;
preferably, the gradient elution procedure comprises: 0-16min, and the flow rate of the mobile phase is 0.8 ml/min; 16-17min, the flow rate of the mobile phase is 0.8-1.0 ml/min; 17-41min, and the flow rate of the mobile phase is 1.0 ml/min; 41-41.1min, and the flow rate of the mobile phase is 1.0-0.8 ml/min; 41.1min, and the flow rate of the mobile phase is 0.8 ml/min.
3. The construction method according to claim 1 or 2, further comprising a step of preparing a reference substance from stachydrine hydrochloride, and a step of detecting a reference substance solution according to the construction method according to claim 1 or 2 to obtain a reference substance characteristic map;
preferably, the preparation method of the control solution comprises the following steps: taking stachydrine hydrochloride reference substance, and adding solvent to make into stachydrine hydrochloride 0.1-0.3mg per 1 ml.
4. The method according to any one of claims 1-3, wherein the fructus Leonuri sample solution is prepared by extracting a sample with a solvent to obtain a sample solution;
preferably, the solvent for extraction is at least one of aqueous ethanol, methanol, ethyl acetate and water;
the extraction mode is one of ultrasonic, shaking and refluxing;
the extraction time is not less than 20min, preferably 20-40 min.
5. A characteristic spectrum of motherwort fruit obtained by the construction method according to any one of claims 1 to 4.
6. A characteristic spectrum of motherwort fruit is characterized in that the characteristic spectrum at least has 5 characteristic peaks;
taking the No. 3 peak as a reference peak, the relative retention time of each characteristic peak is within +/-10% of a specified value, and the specified value is as follows: 0.585 (peak 1), 0.613 (peak 2), 1.077 (peak 4), 1.288 (peak 5).
7. A method for measuring the content of hydrochloric acid threonine alkali in motherwort fruit is characterized by comprising the following steps,
(1) preparing a stachydrine hydrochloride reference product solution and a motherwort fruit test product solution;
(2) the characteristic spectrums of the leonurus fruit characteristic spectrum reference product and the leonurus fruit test product are determined according to the construction method of the leonurus fruit characteristic spectrum of any one of claims 1 to 4, and the content of the stachydrine hydrochloride in the leonurus fruit is obtained according to the standard 0512 of the year-old edition of Chinese pharmacopoeia 2020.
8. A preparation method of motherwort fruit formula particles is characterized by comprising the following steps,
(1) extracting motherwort fruit medicinal materials or decoction pieces to obtain a motherwort fruit extracting solution;
(2) concentrating the fructus Leonuri extractive solution to obtain fructus Leonuri concentrated solution with relative density of 1.05-1.1;
(3) preheating the motherwort fruit concentrated solution, performing spray drying to obtain spray dried powder, and granulating to obtain the motherwort fruit formula granules.
9. The method according to claim 8, wherein the motherwort fruit extract has a ratio of 4-8%;
preferably, the concentration temperature is 50-80 ℃, and the concentration time is not higher than 24 h;
more preferably, the inlet air temperature for spray drying is 170-185 ℃.
10. The method according to claim 8 or 9, wherein the extracting comprises extracting fructus Leonuri or decoction pieces with water to obtain fructus Leonuri extractive solution;
preferably, the motherwort fruit medicinal material or decoction pieces are decocted and extracted by at least two times to obtain a motherwort fruit extracting solution;
more preferably, 9-13 times of water is added into the motherwort fruit medicinal material or decoction pieces, boiling extraction is carried out for 60-90min, and the extraction is repeated at least twice to obtain the motherwort fruit extracting solution.
11. Motherwort fruit formula prepared according to the process of any one of claims 8 to 10.
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| CN1973855A (en) * | 2006-10-16 | 2007-06-06 | 成都市时代第一药物研究所有限公司 | Motherwort injection |
| CN111272905A (en) * | 2020-03-26 | 2020-06-12 | 广东一方制药有限公司 | Characteristic spectrum construction method and detection method of motherwort fruit medicinal material |
| CN112946134A (en) * | 2021-02-04 | 2021-06-11 | 成都第一制药有限公司 | HPLC fingerprint spectrum of motherwort injection and establishing method thereof |
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| CN1973855A (en) * | 2006-10-16 | 2007-06-06 | 成都市时代第一药物研究所有限公司 | Motherwort injection |
| CN111272905A (en) * | 2020-03-26 | 2020-06-12 | 广东一方制药有限公司 | Characteristic spectrum construction method and detection method of motherwort fruit medicinal material |
| CN112946134A (en) * | 2021-02-04 | 2021-06-11 | 成都第一制药有限公司 | HPLC fingerprint spectrum of motherwort injection and establishing method thereof |
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| XIN-DAN LIU 等: "Quality assessment of crude and processed Leonuri Fructus by chemical and color analysis combined with chemometric method" * |
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