CN1679870A - Quality Control Method of Compound Danshen Dripping Pills - Google Patents

Abstract

The invention relate to quality control method of compound Danshen root drops. The method comprises: measuring the fingerprint pattern of the compound Danshen root drops control sample by high efficiency liquid chromatography; measuring the fingerprint pattern of product to be measured of the compound Danshen root drops by the same method and in the same condition; comparing two results. When the absorption peak numbers of two results achieve the value described in the invention, the product to be measured is qualified.

Description

Quality Control Method of Compound Danshen Dripping Pills

technical field

The invention relates to a quality control method for compound danshen dripping pills, in particular to controlling the quality of compound danshen dropping pills by using a fingerprint detection method.

Background technique

Cardiovascular and cerebrovascular diseases are common diseases that seriously endanger human beings. In recent years, due to social development, changes in work, life, diet and environment, cardiovascular and cerebrovascular diseases are on the rise. For the treatment of cardiovascular diseases, although the single-target effect of traditional Chinese medicine is lower than that of western medicine, its multi-channel, multi-target, dynamic overall treatment, and less toxic and side effects are far beyond the reach of western medicine. Chinese patent medicine with definite curative effect The comprehensive treatment effect of traditional Chinese medicine is more than that of western medicine. Compound Danshen preparations are mostly used to treat cardiovascular and cerebrovascular diseases, such as Compound Danshen Tablets, Compound Danshen Dropping Pills, Guanxin Danshen Dropping Pills, etc. These compound salvia miltiorrhiza preparations (both containing salvia miltiorrhiza and Panax notoginseng) have different therapeutic effects because of their different formulas, or different formula ratios, or different extraction and refining methods, or different dosage forms. In addition, because the quality control methods of these compound Danshen preparations are not comprehensive enough, it is difficult to fully characterize their physicochemical characteristics. Therefore, improving the extraction and refining methods and quality control methods of the compound Danshen preparation has become a subject of active research.

Salvia miltiorrhiza is the dry root and rhizome of Salvia miltiorrhiza Bge. Produced in most parts of the country. Since the quality of Salvia miltiorrhiza is uneven due to different varieties, origins, and harvesting periods, it is difficult to guarantee the stability of the quality of its finished products. At present, for the identification of Danshen and its compound preparations, one or two active components or index components of Danshen are usually selected for content determination, and the quality is judged by the content. For example, the quality of Danshen medicinal materials can be identified by the content of tanshinone IIA (page 58 of the Chinese Pharmacopoeia 2000 edition) or the content of danshensu (Zhang Youqin et al., Chinese Medical Journal, 2000, 28 (3): 68); Variety and place of origin (Qiu Feijun, Chinese Modern Applied Pharmacy, 1998, 15(5): 16; Hu Shilin et al., Chinese Journal of Traditional Chinese Medicine, 1999, 24(12): 721); content of Danshensu (Yan Changkai et al., Chinese Hospital Pharmaceutical Journal, 2000, 20 (10): 600), protocatechualdehyde content (Zheng Mojing et al., China Pharmaceutical Affairs, 2000, 14 (4): 254) or tanshinone IIA content (Lin Weizhong, etc., Chinese patent medicine, 2000, 22 (11): 766) etc. to judge the quality of compound Danshen preparations, and use the content of tanshinone IIA to distinguish the quality of compound Danshen tablets produced by different manufacturers (Shao Shuijuan, China Pharmaceutical Industry, 2000, 9 (7): 27) etc. There are dozens of known chemical components of Salvia miltiorrhiza, and most of its fat-soluble components are quinone-type red-yellow substances, such as tanshinone I, IIA, IIB, tanshinone A, B, C, potassium tanshinate, isotanshinone I, II , Cryptotanshinone, etc. Most of its water-soluble components are phenolic aldehydes, phenolic acids, and diterpene acids, such as succinic acid, salvianolic acid A, B, C, 3,4-dihydroxybenzoic acid, etc. In addition, β-sitosterol, vitamin E, etc. were also isolated (Edited by Wang Boxiang, Hepatobiliary Diseases of Traditional Chinese Medicine, first edition, China Medical Science and Technology Press, 1993, page 96). Panax notoginseng is the dried root of Panax notoginseng (Burk.) FH Chen, a plant of the Araliaceae family. It is mainly produced in Yunnan, Guangxi, Sichuan and other places. It is a precious specialty medicinal material in my country and is commonly used in traditional Chinese medicine. It is sweet in taste, slightly bitter, warm in nature, and belongs to the liver and kidney channels. It has the effects of dispelling blood stasis, stopping bleeding, reducing swelling and relieving pain. It is traditionally used to treat traumatic injuries and various bleeding diseases. Studies have shown that Panax notoginseng mainly contains chemical components such as saponins, polysaccharides, amino acids, etc., among which saponins are the material basis for the application of the effect of promoting blood circulation and removing blood stasis of Panax notoginseng, and are the main active ingredients of Panax notoginseng. Panax notoginseng saponins contain ginsenosides R _{b1 ,} R _b2 , R _c , R _d , Re , R _f , R _g1 , R _g2 , B _h1 , notoginseng saponins R ₁ , R ₂ , R ₃ , R ₄ , R ₆ and more than 20 kinds of saponins. These components all belong to Dammarsarane type [Dammarsarane type] tetracyclic triterpene saponins, among which ginsenosides R _b1 , R _g1 , and notoginseng saponin R ₁ are the three components with the highest content, and notoginseng saponin R ₁ is the most abundant Representatively characterized compounds.

The fingerprint of traditional Chinese medicine refers to the chromatogram or spectrum of a certain type or several types of components that are common in certain Chinese medicinal materials or Chinese patent medicines. In the case that most of the active ingredients of traditional Chinese medicine are not clear at this stage, the fingerprint of traditional Chinese medicine is of great significance for effectively controlling the quality of Chinese medicinal materials or Chinese patent medicines. In the 1980s, the major manufacturers of Japanese Kampo medicines had already adopted HPLC fingerprinting to control the quality within the enterprise. During the joint development of Ginkgo biloba extract in Germany and France, it was found that the medical effect of Ginkgo biloba extract is the result of the overall action of the substance group obtained from the extract, and the quality control of such a whole also uses HPLC fingerprinting method. In the plant and herbal medicine guidelines formulated by the US FDA in recent years, fingerprinting has been clearly used as a quality control method for mixed substance groups (FDA. Guidance of Industry: Botanical Drug (Draft). 2000 August). With the deepening of research, people found that as a product of the practice of TCM theory, TCM, especially compound TCM, none of the components contained in it can represent its overall curative effect. It is gradually realized that the current quality standards referring to the quality control mode of western medicine (synthetic medicine) cannot properly reflect the inherent quality of traditional Chinese medicine. From the perspective of development trend, it is a development trend from the current quality control mode to a combination of comprehensive, macroscopic and quantifiable identification and content determination of main active ingredients.

The current standard for quality evaluation of traditional Chinese medicine is to identify and measure one or several active ingredients, active ingredients or index ingredients by means of spectroscopy or chromatography, as well as the routine inspection items stipulated in the Pharmacopoeia. For example, the 2000 edition of the Chinese Pharmacopoeia [Part 1] contains a total of 602 kinds of medicinal materials and patent medicines. Among them, there are 992 TLC identifications, 308 varieties have content determination [volumetric method, spectroscopic method, liquid chromatography, gas chromatography and TLC scanning method], and most varieties have general inspection items. Obviously, these quality standards are set to imitate the model of chemicals. Other countries such as the United Kingdom, India, the American Herbal Pharmacopoeia, the Chinese medicine in the Japanese Pharmacopoeia, and the German Herbal Medicine Monograph edited by the German Commission E also adopted basically the same content. For chemical drugs, their active ingredients are single compounds with clear structure and clear structure-activity relationship, and their content and purity directly express their efficacy and safety. However, the characteristic of traditional Chinese medicine is compound compatibility, and the content of any single effective or active ingredient cannot express its overall curative effect. For example, astragaloside IV contained in Astragalus membranaceus is currently the most common target selected as the identification and content determination of quality standards, but there is no evidence to prove the clear connection between astragaloside IV and the functions and indications of Astragalus membranaceus. Similarly, Coptidis Rhizoma, Phellodendron Phellodendron, and Sankezhen all contain trabeculine, which is generally used as the target of detection, but the functions and indications of the three are completely different. The situation with compound preparations is more complicated. The non-one-to-one non-linear theory and practice of traditional Chinese medicine indicate that the quality of traditional Chinese medicine should adopt some kind of macroscopic comprehensive quality evaluation method.

Compound Danshen Dropping Pills is a drop pill preparation made of Danshen and Panax notoginseng as the main raw materials. It is clinically used to treat various diseases caused by cardiovascular and cerebrovascular diseases, coronary heart disease, angina pectoris, myocardial ischemia, and microcirculation disorders. Its therapeutic effect has been clinically verified, and whether the quality of the drug and the content of active ingredients in Compound Danshen Dripping Pills can be guaranteed are the basis for determining the curative effect of Compound Danshen Dripping Pills. If one or two active components of Danshen are used to describe the internal quality of Compound Danshen Dripping Pills, it is somewhat one-sided, not to mention the non-effective index components. To control the efficacy of Compound Danshen Dripping Pills, it is not enough to characterize and control only one or two of its chemical components, and it is necessary to control its substance group as a whole. Therefore, in addition to "micro analysis", some kind of "macro analysis" method should be used to effectively characterize the quality of traditional Chinese medicine as a whole. As a quality control method for Chinese herbal medicines and their extracts, fingerprinting has become an international consensus. Now, there are many determination methods for the active components in Danshen such as tanshinone and danshensu, and for the determination of active components in Panax notoginseng such as notoginseng saponin R _g1 and ginsenoside R _g1 , etc. The macroscopic quality control method for the quality control of Compound Danshen Dripping Pill preparations has not been reported yet.

Contents of the invention

The purpose of the present invention is to provide a kind of method of compound danshen dripping pill quality control, by this kind of method, can control the quality of compound danshen dripping pill preparation.

The purpose of the present invention is to establish a method for quality control of Chinese medicine compound preparations. The present invention uses a large number of experiments under certain conditions to compare the fingerprints of the compound Danshen dripping pills with the fingerprints of the Sanqi medicinal material fingerprints and the compound Danshen dripping pills intermediate. Compare the fingerprints, observe the reproducibility of its absorption peaks under the same chromatographic test conditions, and determine whether most of the effective chemical components in the medicinal material of Panax notoginseng enter the intermediate of compound danshen dripping pills and compound danshen drops through fingerprints pill control samples. Prove by experiment, contain most of chemical components in the Radix Notoginseng medical material in the compound danshen dripping pill control sample in the present invention, therefore measure the fingerprint of the salvia miltiorrhiza chemical component in the compound danshen dropping pill, and compare it with the corresponding control fingerprint spectrum By comparison, the quality of Compound Danshen Dripping Pills can be controlled.

The present invention can be implemented through the following steps:

(a). Establishment of the control fingerprint of Compound Danshen Dripping Pills

Preparation of Compound Danshen Dripping Pills Control Sample Solution:

It is called Compound Danshen Dripping Pills, dissolved in 1-20mL of 1-8% ammonia water, ultrasonically dissolved, passed through a 0.30-0.60μm filter membrane, and 2-10mL of the filtrate was passed through a packed C-18 column, 5-40mL 8-35 Rinse with 1% methanol and 1-8% ammonia solution, discard the rinse solution, then wash with 5-35ml water, discard the water wash solution, and then elute with methanol, collect the methanol precipitated solution in a 1-15mL volumetric flask, dilute to volume, and centrifuge for later use;

Determination of Fingerprint of Compound Danshen Dripping Pills:

Draw the above-mentioned control sample solution and inject it into a liquid chromatograph, and use high-performance liquid chromatography to measure it to obtain the fingerprint of the compound Danshen dripping pill. , mobile phase A is glacial acetic acid aqueous solution, mobile phase B is glacial acetic acid acetonitrile solution; detection wavelength is 200-210nm;

(b). Determination of the fingerprint of the product to be tested in Compound Danshen Dripping Pills:

Get the compound danshen dripping pill product to be tested, and measure the fingerprint of the product to be tested according to the steps and conditions described in the above (a);

(c). Comparing the fingerprint of the compound danshen dripping pills to be tested with the control fingerprint of the compound danshen dripping pills to identify the number of absorption peaks to determine whether the product quality is qualified.

The preferred invention can be carried out through the following steps:

The preparation method of Compound Danshen Dropping Pills reference sample solution in the (a) step is to claim Compound Danshen Dropping Pills, be dissolved in 5-15mL 2-6% ammonia water, ultrasonically dissolve, pass through a 0.35-0.55 μm filter membrane, take 2- 10mL of the filtrate was passed through the packed C-18 small column, washed with 10-30mL of 10-30% methanol and 2-6% ammonia solution, discarded the rinse solution, then washed with 10-30ml water, discarded the water wash solution, and eluted with methanol, collected Put the methanol eluent in a 1-10mL volumetric flask, dilute to volume, and centrifuge for later use;

Chromatographic conditions: the chromatographic column is octadecylsilane bonded silica gel as filler; gradient precipitation is adopted, mobile phase A is 0.01% glacial acetic acid aqueous solution, mobile phase B is acetonitrile solution containing 0.01% glacial acetic acid; gradient elution procedure as follows:

At 0 minutes, mobile phase A was 80% 0.01% glacial acetic acid aqueous solution, and mobile phase B was 20% 0.01% glacial acetic acid acetonitrile solution;

At 15 minutes, mobile phase A was 65% 0.01% glacial acetic acid aqueous solution, and mobile phase B was 35% 0.01% glacial acetic acid acetonitrile solution;

At 25 minutes, mobile phase A was 65% 0.01% glacial acetic acid aqueous solution, and mobile phase B was 35% 0.01% glacial acetic acid acetonitrile solution;

At 40 minutes, mobile phase A was 57% 0.01% glacial acetic acid aqueous solution, and mobile phase B was 43% 0.01% glacial acetic acid acetonitrile solution;

At 50 minutes, mobile phase A was 57% 0.01% glacial acetic acid aqueous solution, and mobile phase B was 43% 0.01% glacial acetic acid acetonitrile solution;

At 65 minutes, mobile phase A was 42% 0.01% glacial acetic acid aqueous solution, and mobile phase B was 58% 0.01% glacial acetic acid acetonitrile solution;

At 75 minutes, mobile phase A was 25% 0.01% glacial acetic acid aqueous solution, and mobile phase B was 75% 0.01% glacial acetic acid acetonitrile solution;

The injection volume is 5-45 μL; the flow rate is 0.8 mL·min ^-1 ; the detection wavelength is 201-205 nm; the column temperature is 25-35°C.

The establishment of the best standard fingerprint of Compound Danshen Dripping Pills adopts the following method:

The preparation method of Compound Danshen Dropping Pills contrast sample solution in the described (a) step is, claims Compound Danshen Dropping Pills 0.4～1.5g, is dissolved in 10mL 4% ammoniacal liquor, ultrasonic dissolves, crosses 0.45 μm filter membrane, gets 5mL filtrate and crosses 500mg Filled C-18 small column, wash with 20mL 20% methanol 4% ammonia solution, discard the washing solution, then wash with 20ml water, discard the water washing solution, then elute with methanol, collect the methanol eluate in a 5mL volumetric flask, and constant volume , centrifuge for standby, the injection volume is 20 μL;

Chromatographic conditions: injection volume 20 μL; flow rate 0.8 mL·min ^-1 ; detection wavelength 203 nm; column temperature 30°C.

In the establishment of the fingerprint of the control compound Danshen dripping pills, the preparation ratio of the mobile phase A solution was prepared according to the volume ratio, and the preparation ratio of the mobile phase B solution was prepared according to the volume ratio.

In the establishment of the comparison fingerprint of Compound Danshen Dripping Pills, there are 14 absorption peaks in the comparison fingerprint of Compound Danshen Dripping Pills obtained by measuring according to high performance liquid chromatography, and the absorption peaks whose single peak area exceeds 10% of the total peak area have 14 absorption peaks. 3, namely:

For peak No. 2, the average retention time RT is 12.12min, the RSD is 0.34%, the peak area is 2723.73, and the RSD is 12.11%;

Peak No. 3, average retention time RT is 20.34min, RSD is 0.23%, peak area is 1684.49, RSD is 18.59%;

For peak No. 12, the average retention time RT is 47.88min, the RSD is 0.25%, the peak area is 1547.80, and the RSD is 13.25%.

In the establishment of the reference fingerprint of Compound Danshen Dropping Pills, there are 14 absorption peaks in the obtained Compound Danshen Dropping Pills reference fingerprint according to high performance liquid chromatography, and the absorption peaks whose single peak area exceeds 5% of the total peak area have 14 absorption peaks. 7, namely:

For peak No. 2, the average retention time RT is 12.12min, the RSD is 0.34%, the peak area is 2723.73, and the RSD is 12.11%;

Peak No. 3, average retention time RT is 20.34min, RSD is 0.23%, peak area is 1684.49, RSD is 18.59%;

For peak No.9, the average retention time RT is 42.94min, the RSD is 0.21%, the peak area is 665.38, and the RSD is 11.82%;

For peak No. 11, the average retention time RT is 45.68min, the RSD is 0.23%, the peak area is 947.39, and the RSD is 16.06%;

For peak No. 12, the average retention time RT is 47.88min, the RSD is 0.25%, the peak area is 1547.80, and the RSD is 13.25%;

For peak No. 13, the average retention time RT is 68.21min, the RSD is 0.13%, the peak area is 622.91, and the RSD is 10.39%;

For peak No. 14, the average retention time RT is 69.43min, the RSD is 0.12%, the peak area is 820.24, and the RSD is 12.94%.

In the establishment of the reference fingerprint of Compound Danshen Dropping Pills, there are 14 absorption peaks in the obtained Compound Danshen Dropping Pills reference fingerprint according to high performance liquid chromatography, of which the single peak area exceeds 2% of the total peak area. 13, respectively;

For peak No. 1, the average retention time RT is 10.87min, the RSD is 0.31%, the peak area is 506.92, and the RSD is 13.03%;

For peak No. 2, the average retention time RT is 12.12min, the RSD is 0.34%, the peak area is 2723.73, and the RSD is 12.11%;

Peak No. 3, average retention time RT is 20.34min, RSD is 0.23%, peak area is 1684.49, RSD is 18.59%;

Peak No. 5, the average retention time RT is 23.29min, RSD is 0.25%, peak area is 565.78, RSD is 13.80%;

For peak No. 6, the average retention time RT is 24.48min, the RSD is 0.28%, the peak area is 309.39, and the RSD is 18.98%;

For peak No. 7, the average retention time RT is 29.00min, the RSD is 0.62%, the peak area is 345.32, and the RSD is 14.28%;

Peak No. 8, the average retention time RT is 41.21min, RSD is 0.25%, peak area is 436.06, RSD is 10.88%;

For peak No.9, the average retention time RT is 42.94min, the RSD is 0.21%, the peak area is 665.38, and the RSD is 11.82%;

For peak No. 10, the average retention time RT is 44.16min, the RSD is 0.21%, the peak area is 472.84, and the RSD is 14.59%;

For peak No. 11, the average retention time RT is 45.68min, the RSD is 0.23%, the peak area is 947.39, and the RSD is 16.06%;

For peak No. 12, the average retention time RT is 47.88min, the RSD is 0.25%, the peak area is 1547.80, and the RSD is 13.25%;

For peak No. 13, the average retention time RT is 68.21min, the RSD is 0.13%, the peak area is 622.91, and the RSD is 10.39%;

For peak No. 14, the average retention time RT is 69.43min, the RSD is 0.12%, the peak area is 820.24, and the RSD is 12.94%.

The present invention adopts following method to control the quality of compound danshen dripping pills, takes the product of compound danshen dropping pills to be tested, adopts the same method as the establishment method, chromatographic conditions, and measuring methods of the comparison fingerprint of compound danshen dropping pills to measure, and obtains the fingerprints , the compound danshen dropping pills product fingerprint to be tested is compared with the compound danshen dropping pill control fingerprint, and when the two fingerprints have 3 or more identical absorption peaks, the quality of the compound danshen dropping pill to be tested is considered qualified.

The preferred compound danshen dripping pills product fingerprint to be tested is compared with the compound danshen dropping pill contrast fingerprint, and when the two fingerprints have 5 or more identical absorption peaks, it is considered that the quality of the compound danshen dripping product to be tested is qualified.

Further preferred compound danshen dripping pills product fingerprints to be tested are compared with compound danshen dripping pills contrast fingerprints, when the two fingerprints have 7 or more identical absorption peaks, it is considered that the compound danshen dripping products to be tested Quality standards.

It is further preferred to compare the fingerprint of the product to be tested with the Compound Danshen Dripping Pills and the comparison of the fingerprints of the Compound Danshen Dripping Pills. When the absorption peaks of the fingerprints of the two fingerprints exceed 13 or more than 7 identical absorption peaks, it is considered that the Compound Danshen Dripping Pills The quality of the product to be tested is qualified.

The best compound danshen dripping pills product fingerprint is compared with the compound danshen dropping pill control fingerprint, and when the two fingerprints have 14 identical absorption peaks, the quality of the compound danshen dropping pill product to be tested is considered qualified.

The present invention measures the notoginseng chemical components in the notoginseng medicinal material and the compound danshen dripping pill intermediate, the method is as follows:

The method for obtaining the HPLC fingerprint of the chemical components of Radix Notoginseng in the Radix Notoginseng medicinal material is as follows:

The preparation of the notoginseng medicinal material test sample: take the medicinal material in the reflux bottle, add distilled water to reflux to extract, filter, add distilled water to the filter residue, reflux, combine the filtrate, dissolve in it, centrifugal filter, and set aside;

Precisely draw the solution of the test product and inject it into the liquid chromatograph, measure according to the high-performance liquid chromatography, and obtain the fingerprint of the chemical composition of the notoginseng components;

In the fingerprint spectrum, there are 5 absorption peaks of the component chemical components of the Panax notoginseng medical material test product, and there are 3 absorption peaks with a single peak area exceeding 10% of the total peak area, which are respectively:

For peak No. 1, the average retention time RT is 10.81min, the RSD is 0.06%, the peak area is 539.57, and the RSD is 17.73%;

For peak No. 2, the average retention time RT is 12.03min, the RSD is 0.02%, the peak area is 2482.50, and the RSD is 8.74%;

For peak No. 3, the average retention time RT is 20.16min, the RSD is 0.11%, the peak area is 1514.5, and the RSD is 12.02%.

A preferred method for obtaining the HPLC fingerprint of the chemical components of Radix Notoginseng in Radix Notoginseng, the method is as follows:

Preparation of Notoginseng medical material test sample: Weigh 2.5g of medicinal material in a reflux bottle, add 50mL of distilled water, reflux for 1.5 hours, filter, add 50mL of distilled water to the filter residue, reflux for 1 hour, combine the filtrate, and dissolve it in a 100mL measuring bottle. Centrifugal filtration, spare; 2 parallel, injection volume 10μL;

Precisely draw the solution of the test product and inject it into the liquid chromatograph, measure according to the high-performance liquid chromatography, and obtain the fingerprint of the chemical composition of the notoginseng components;

In the fingerprint spectrum, there are 5 absorption peaks of the component chemical components of the Panax notoginseng medical material test product, and there are 4 absorption peaks with a single peak area exceeding 5% of the total peak area, which are respectively:

For peak No. 1, the average retention time RT is 10.81min, the RSD is 0.06%, the peak area is 539.57, and the RSD is 17.73%;

For peak No. 2, the average retention time RT is 12.03min, the RSD is 0.02%, the peak area is 2482.50, and the RSD is 8.74%;

For peak No. 3, the average retention time RT is 20.16min, the RSD is 0.11%, the peak area is 1514.5, and the RSD is 12.02%;

For peak No. 5, the average retention time RT is 28.42min, the RSD is 0.27%, the peak area is 356.20, and the RSD is 15.30%.

The best method for obtaining high performance liquid chromatography fingerprints of the chemical components of Panax notoginseng in medicinal materials, the method is as follows:

Preparation of Notoginseng medical material test sample: Weigh 2.5g of medicinal material in a reflux bottle, add 50mL of distilled water, reflux for 1.5 hours, filter, add 50mL of distilled water to the filter residue, reflux for 1 hour, combine the filtrate, and dissolve it in a 100mL measuring bottle. Centrifugal filtration, spare; 2 parallel, injection volume 10μL;

Precisely draw the solution of the test product and inject it into the liquid chromatograph, measure according to the high-performance liquid chromatography, and obtain the fingerprint of the chemical composition of the notoginseng components;

In the fingerprint spectrum, there are 5 absorption peaks of the component chemical components of the Panax notoginseng medical material test product, and wherein there are 5 absorption peaks with a single peak area exceeding 2% of the total peak area, which are respectively:

For peak No. 1, the average retention time RT is 10.81min, the RSD is 0.06%, the peak area is 539.57, and the RSD is 17.73%;

For peak No. 2, the average retention time RT is 12.03min, the RSD is 0.02%, the peak area is 2482.50, and the RSD is 8.74%;

For peak No. 3, the average retention time RT is 20.16min, the RSD is 0.11%, the peak area is 1514.5, and the RSD is 12.02%;

Peak No. 4, the average retention time RT is 23.04min, RSD is 0.07%, peak area is 250.63, RSD is 18.25%;

For peak No. 5, the average retention time RT is 28.42min, the RSD is 0.27%, the peak area is 356.20, and the RSD is 15.30%.

The method for obtaining the HPLC fingerprint of the chemical components of Sanqi in the compound Danshen dripping pill intermediate is as follows:

Compound Danshen Dropping Pill Intermediate Preparation of the test sample: Weigh the Compound Danshen Dropping Pill Intermediate, dissolve in 1-20mL of 1-8% ammonia water, ultrasonically dissolve, pass through a 0.30-0.60μm filter membrane, and take 2-10mL Pass the filtrate through the packed C-18 small column, wash with 5-40mL 8-35% methanol and 1-8% ammonia solution, discard the washing solution, then wash with 5-35ml water, discard the washing solution, elute with methanol, and collect methanol Put the eluate in a 1-15mL volumetric flask, constant volume, centrifuge for later use, and make 2 copies in parallel, the injection volume is 20μL;

The method for obtaining high-performance liquid phase fingerprints of the chemical components of notoginseng in the preferred compound danshen dripping pill intermediate is as follows:

Compound Danshen dripping pill intermediate preparation: Weigh 0.2g of compound Danshen dripping pill intermediate, dissolve in 5-15mL 2-6% ammonia water, dissolve by ultrasonic, pass through 0.35-0.55μm filter membrane, take 2-10mL filtrate Packed C-18 small column, wash with 10-30mL 10-30% methanol 2-6% ammonia solution, discard the washing solution, then wash with 10-30ml water, discard the water washing solution, then elute with methanol, collect the methanol for washing Deliquify in a 1-10mL volumetric flask, constant volume, centrifuge for later use, make 2 parallel portions, and the injection volume is 20μL;

The best method for obtaining HPLC fingerprints of the chemical components of Sanqi in the intermediate of compound Danshen dripping pills is as follows:

The preparation of Compound Danshen Dropping Pill intermediate for testing: Weigh 0.1～0.5g of Compound Danshen Dropping Pill intermediate, dissolve in 10mL 4% ammonia water, dissolve by ultrasonic, pass through 0.45μm filter membrane, take 5mL filtrate and pass through the C- For 18 small columns, wash with 20mL of 20% methanol and 4% ammonia solution, discard the washing solution, wash with 20ml of water, discard the water sink solution, and then elute with methanol, collect the methanol eluate in a 5mL volumetric flask, dilute to volume, and centrifuge for later use , in parallel, with an injection volume of 20 μL;

Precisely draw the solution of the test product and inject it into the liquid chromatograph, measure according to the high-performance liquid chromatography, and obtain the fingerprint of the chemical composition of the notoginseng components;

There are 12 absorption peaks of the chemical components of Sanqi component in the intermediate of Compound Danshen Dripping Pills, among which there are 11 absorption peaks whose single peak area exceeds 2% of the total peak area, which are:

For peak No. 1, the average retention time RT is 10.85min, the RSD is 0.08%, the peak area is 557.29, and the RSD is 15.67%;

For peak No. 2, the average retention time RT is 12.10min, the RSD is 0.06%, the peak area is 2728.83, and the RSD is 16.66%;

For peak No. 3, the average retention time RT is 20.33min, the RSD is 0.08%, the peak area is 3704.31, and the RSD is 14.15%;

Peak No. 4, the average retention time RT is 23.86min, RSD is 0.07%, peak area is 272.69, RSD is 12.54%;

Peak No. 5, the average retention time RT is 23.28min, RSD is 0.08%, peak area is 369.12, RSD is 21.52%;

For peak No. 7, the average retention time RT is 28.82min, the RSD is 1.36%, the peak area is 289.16, and the RSD is 17.71%;

Peak No. 8, average retention time RT is 42.93min, RSD is 0.07%, peak area is 239.14, RSD is 18.84%;

Peak No. 9, the average retention time RT is 45.66min, RSD is 0.06%, peak area is 445.17, RSD is 19.56%;

For peak No. 10, the average retention time RT is 47.85min, the RSD is 0.06%, the peak area is 723.59, and the RSD is 14.24%;

For peak No. 11, the average retention time RT is 68.17min, the RSD is 0.03%, the peak area is 407.66, and the RSD is 10.86%;

For peak No. 12, the average retention time RT is 69.4min, the RSD is 0.03%, the peak area is 476.31, and the RSD is 15.46%.

There are 12 absorption peaks of the chemical components of Sanqi component in the intermediate of Compound Danshen Dripping Pills, among which there are 4 absorption peaks whose single peak area exceeds 5% of the total peak area, which are:

For peak No. 1, the average retention time RT is 10.85min, the RSD is 0.08%, the peak area is 557.29, and the RSD is 15.67%;

For peak No. 2, the average retention time RT is 12.10min, the RSD is 0.06%, the peak area is 2728.83, and the RSD is 16.66%;

For peak No. 3, the average retention time RT is 20.33min, the RSD is 0.08%, the peak area is 3704.31, and the RSD is 14.15%;

For peak No. 10, the average retention time RT is 47.85min, the RSD is 0.06%, the peak area is 723.59, and the RSD is 14.24%.

There are 12 absorption peaks of the chemical components of Sanqi component in the intermediate of Compound Danshen Dripping Pills, among which there are 2 absorption peaks whose single peak area exceeds 10% of the total peak area, which are:

For peak No. 2, the average retention time RT is 12.10min, the RSD is 0.06%, the peak area is 2728.83, and the RSD is 16.66%;

For peak No. 3, the average retention time RT is 20.33min, the RSD is 0.08%, the peak area is 3704.31, and the RSD is 14.15%;

For peak No. 10, the average retention time RT is 47.85min, the RSD is 0.06%, the peak area is 723.59, and the RSD is 14.24%.

The present invention finds by measuring the fingerprints of Compound Danshen Dripping Pills, Compound Danshen Dripping Pills intermediates, and Panax notoginseng medicinal materials that the number of absorption peaks among the three is not exactly the same, indicating that the fingerprints of Compound Danshen Dripping Pills are used as Compound Danshen Dripping Pills The quality control standard can more objectively reflect the presence and content of the internal components of the product than the fingerprint of the three seven ingredients alone as the quality standard, so as to fully control the quality of the product.

The advantages of the present invention are as follows:

(1). The HPLC standard fingerprint chromatogram established with the main active ingredient Sanqi chemical components in Compound Danshen Dripping Pills as an index represents most of the pharmacological activities of Compound Danshen Dripping Pills, and can effectively characterize the quality of Compound Danshen Dripping Pills. In this way, the detection of the most possible chemical components of the compound Danshen dripping pills is realized, which is beneficial to the overall monitoring of the quality of traditional Chinese medicines.

(2). Treat the fingerprints of the active ingredients of Sanqi in Compound Danshen Dripping Pills as a whole, pay attention to the sequence and interrelationship of each characteristic peak of the fingerprint, and pay attention to the overall appearance characteristics, which avoids the need to measure only one or two The one-sidedness of judging the overall quality of Compound Danshen Dripping Pills based on the chemical composition also reduces the possibility of artificial processing to meet the quality standards. The invention provides a new control standard for complete and accurate evaluation of the quality of the compound danshen dripping pills, and contributes to improving the quality and curative effect of the compound danshen dropping pills and other preparations with salvia miltiorrhiza and notoginseng as main components.

(3). The present invention has the characteristics of simple, stable, high precision, good reproducibility and easy mastery.

The present invention provides a method for identification of Compound Danshen Dripping Pills. The present invention is a practical method obtained through a large number of experiments. In the present invention, the three compounds in Compound Danshen Dripping Pills obtained by high-efficiency liquid phase and under the same test conditions The high-efficiency liquid phase standard fingerprint of the seven chemical components has the characteristics of good repeatability. Therefore, the fingerprint of the compound Danshen dripping pill established by the above-mentioned determination method can be used as a standard fingerprint. Standard fingerprints for ingredient determination to identify its active ingredients.

For those skilled in the art, based on the technical contents disclosed in the present invention, other implementations of the present invention will be apparent to those skilled in the art, and the embodiments of the present invention are only examples. Various changes and improvements can be made to the present invention without departing from the spirit and scope of the present invention. For example, the measurement results obtained by using different detection instruments may be different, but as long as the quality control method described in the present invention is used, it is within the protection scope of the present invention.

Description of drawings

The fingerprints of Panax notoginseng medicinal materials, the fingerprints of the active ingredients of Panax notoginseng in the intermediate of compound Danshen dripping pills, the fingerprints of the active ingredients of Panax notoginseng in compound Danshen dripping pills, and the comparison of the HPLC fingerprints of the chemical components of Panax notoginseng from different sources are given below , is intended to further illustrate the present invention, but does not constitute a limitation to the present invention.

Figure 1 Fingerprints of the active ingredients of Panax notoginseng medicinal material

Fig. 2 Fingerprint of the active ingredient of notoginseng in compound danshen dripping pill intermediate

Fig. 3 Fingerprints of active ingredients of Sanqi in Compound Danshen Dripping Pills

Figure 4 Comparison of the HPLC fingerprints of the chemical components of Panax notoginseng from different sources

Among them: 1 is compound Danshen dripping pills

2 is the intermediate of Compound Danshen Dripping Pills

  3 is notoginseng medicinal material

Detailed ways

The following examples of preparation are listed to further describe the present invention in detail, and the examples are only used to illustrate the present invention and do not limit the present invention.

Embodiment one (preparation example)

Weigh 41.06g of Salvia miltiorrhiza and 8.03g of Panax notoginseng, add water to decoct twice, measure water 4 times for 2 hours for the first time, measure water 3 times for 1 hour for the second time, filter, combine the filtrates, concentrate to a specific gravity of 1.19-1.20( 75±1°C), add ethanol with a concentration of about 90% until the ethanol content is 65% (20°C), let it stand for 12 hours, separate the supernatant, recover the ethanol, concentrate until the relative density of the liquid is 1.37 (55-60 ℃), to obtain the Radix Notoginseng Extract. Take the above-mentioned Danshen Sanqi extract and 0.46g of borneol, mix it evenly with 18g of polyethylene glycol-6000, heat it to a temperature of 85°C, and after 80 minutes, move it to a dripping tank of a dripping machine whose tank temperature is kept at 86°C . Drop the drug liquid into liquid paraffin at 8°C, take out the dropping pills, remove the oil, and select the pills through a sieve to obtain the product.

Embodiment two (preparation example)

Weigh 59.36g of Salvia miltiorrhiza and 6.38g of Panax notoginseng, add potassium carbonate of 1.0% of the total amount of medicinal materials, decoct twice, the first time 4 times the amount of water for 2.5 hours, the second time 3 times the amount of water for 1.5 hours, filter, and combine the filtrates , concentrated to a specific gravity of 1.19-1.20 (75±1°C), add ethanol with a concentration of about 85% until the ethanol content is 70% (20°C), let it stand for 10 hours, separate the supernatant, recover the ethanol, and concentrate to the pharmaceutical The relative density of the liquid is 1.35 (55-60°C), and the extract of Danshen Sanqi is obtained. Take the above-mentioned Danshen Sanqi extract and 0.34g of borneol, mix it with 23g of polyethylene glycol-6000 evenly, heat it to a temperature of 89°C, and after 100 minutes, transfer it to a dripping tank of a dripping machine whose tank temperature is kept at 85°C . Drop the liquid medicine into methyl silicone oil at 8°C, take out the dropping pills, remove the oil, and select the pills through a sieve to obtain the product.

Embodiment three (preparation example)

Weigh 31.12g of salvia miltiorrhiza and 9.21g of notoginseng, add 0.5% sodium hydroxide of the total amount of medicinal materials, decoct twice, the first time 4 times the amount of water for 1.5 hours, the second time 3 times the amount of water for 1.5 hours, filter, and the filtrate Combine and concentrate until the specific gravity is 1.19-1.20 (75±1°C), add ethanol with a concentration of about 88% until the ethanol content is 66% (20°C), let stand for 10 hours, separate the supernatant, reclaim the ethanol, and concentrate to The relative density of the liquid medicine is 1.40 (55-60° C.), and the extract of Danshen Sanqi is obtained. Take the above-mentioned Danshen Sanqi extract, 0.50 g of borneol, 90 g of mannitol, 15 g of edetate calcium sodium and 15 ml of distilled water, mix the above components, freeze-dry, and make a powder injection.

Embodiment four (preparation example)

Weigh 116.35g of Salvia miltiorrhiza and 58.21g of Panax notoginseng, add water to decoct twice, measure 4 times of water for 2 hours for the first time, and 1.5 hours for 3 times of water for the second time, filter, combine the filtrates, and concentrate until the specific gravity is 1.19-1.20( 75±1°C), add ethanol with a concentration of about 88% until the ethanol content is 66% (20°C), let it stand for 10 hours, separate the supernatant, recover the ethanol, concentrate until the relative density of the liquid is 1.40 (55-60 ℃), to obtain the Radix Notoginseng Extract. Take 1.8g of the above-mentioned Danshen Sanqi extract and balm oil, mix them with 40g of microcrystalline cellulose, add 3% povidone ethanol solution to make a soft material, pass through a 18-mesh sieve to make granules, dry at 60°C for 35 minutes, and granulate. Add 4g of talcum powder, mix well, fill in capsules, and get ready.

Embodiment five (preparation example)

Weigh 116.35g of Salvia miltiorrhiza and 58.21g of Panax notoginseng, add 2.0% sodium bicarbonate of the total amount of medicinal materials, decoct twice, the first time 4 times the amount of water for 2 hours, the second time 3 times the amount of water for 1.5 hours, filter, and the filtrate Combine and concentrate until the specific gravity is 1.19-1.20 (75±1°C), add ethanol with a concentration of about 88% until the ethanol content is 66% (20°C), let stand for 10 hours, separate the supernatant, reclaim the ethanol, and concentrate to The relative density of the liquid medicine is 1.40 (55-60° C.), and the extract of Danshen Sanqi is obtained. Take above-mentioned Danshen Sanqi extract and 0.9 g of borneol, mix with 120 g of microcrystalline cellulose, 40 g of hydroxypropyl methylcellulose, 5 g of xylitol, and 2 g of magnesium stearate, and press into tablets to obtain the product.

Embodiment six (preparation example)

Weigh 140.35g of Salvia miltiorrhiza and 36.42g of Panax notoginseng, decoct twice with water, measure 4 times of water for 2 hours for the first time, and 1.5 hours for 3 times of water for the second time, filter, combine the filtrates, and concentrate until the specific gravity is 1.19-1.20( 75±1°C), add ethanol with a concentration of about 90% until the ethanol content is 65% (20°C), let it stand for 8 hours, separate the supernatant, recover the ethanol, concentrate until the relative density of the liquid is 1.35 (55-60 ℃), to obtain the Radix Notoginseng Extract. Take 1.0 g of the above-mentioned Danshen Sanqi extract and borneol, mix it with 46 g of microcrystalline cellulose, add 3% povidone ethanol solution to make a soft material, pass through a 18-mesh sieve to make granules, dry at 60°C for 30 minutes, granulate, add 4g of talcum powder, mixed evenly, and pressed into tablets.

Embodiment seven (compound Danshen dripping pill notoginseng component fingerprint detection example)

1. Instruments and reagents

Instrument: Agilent 1100 liquid chromatography, including quaternary pump, online degassing device, automatic sampler, DAD detector, column thermostat, Chemstation workstation; BS210S electronic balance (1/10 ^-4 g) (Beijing Sedoli Sri Lanka), METTLERAE240 electronic balance (1/10 ^-4 g or 1/10 ^-5 g) (Mettler-Toledo (Shanghai) Co., Ltd.), LD4-2 centrifuge (4000r/min) (Beijing Medical Centrifuge machine factory), digital constant temperature water bath (Tianjin Changfeng Co., Ltd.), RE-52AA rotary evaporator (Shanghai Yarong Biochemical Instrument Factory), SHE-(III) circulating water vacuum pump (Gongyi Yingyu Yuhua Instrument Factory) , KQ-250B ultrasonic cleaner (Kunshan Ultrasonic Instrument Co., Ltd.), HENGAO T&D filter (HENGGAO T&D), synthetic fiber filter membrane (pore size 0.45μm) (Shanghai Xingya Purification Material Factory); C ₁₈ small column (filler is C ₁₈ H ₁₇ liquid chromatography stationary phase, 50-100 mesh, produced by Tianjin Chemical Reagent No. 2 Factory. Dry packing 600mg, column inner diameter 1cm).

Reagents: acetonitrile (chromatographically pure, Merck, USA), acetic acid (premium grade), Wahaha purified water.

2. Preparation of Compound Danshen Dripping Pills Control Sample

The preparation of Compound Danshen Dropping Pills test sample: take by weighing 1.0g of each batch of Compound Danshen Dropping Pills in Example 1, dissolve in 10ml 4% ammonia water, ultrasonically dissolve, cross 0.45 μm filter membrane, get 5ml filtrate and cross C-18 small Column (500mg filler), 20ml 20% methanol and ammonia water for elution, collect methanol eluate in 5ml volumetric flask, constant volume, centrifuge for later use; parallel 2, injection volume 20μl.

The preparation of compound Danshen dripping pill intermediate test sample: take by weighing 0.2g of each batch of Danshen Sanqi extract in embodiment one, be dissolved in 10ml 4% ammonia water, ultrasonically dissolve, cross 0.45 μm filter membrane, get 5ml filtrate and cross C -18 small column, wash with 15ml of 20% methanol and 4% ammonia solution first, then wash with 20ml of water, collect the methanol eluate in a 5ml volumetric flask, dilute to volume, and centrifuge for later use. Two parallel copies were made, and the injection volume was 20 μl.

The preparation of the notoginseng medical material test sample: take by weighing 2.5g of the medicinal material in a reflux bottle, add 50ml of distilled water, reflux for 1.5 hours, filter, add 50ml of distilled water in the filter residue, reflux for 1 hour, combine the filtrate, and dissolve it in a 100ml measuring bottle. Centrifugal filtration, spare. Two parallel copies were made, and the injection volume was 10 μl.

3. HPLC analysis conditions

Agilent SB-C ₁₈ analytical column (4.6mm×250mm, Zorbax SB USCL010304); mobile phase: Phase A is 0.01% (V/V) glacial acetic acid aqueous solution, and phase B is acetonitrile solution containing 0.01% glacial acetic acid. The flow rate is 0.8ml·min ^-1 ; the detection wavelength is 203nm; the column temperature is 30°C.

The gradient elution program is as follows: Time A: 0.01% HAC-H2O (%) (v/v) B: 0.01% HAC-CH3CN (%) (v/v) 0152540506575 80656557574225 20353543435875

Compound Danshen Dripping Pill Notoginseng Fingerprint Component Name: Peak 1- Peak 2- Peak 3- Peak 4- Peak 5- Peak 6- Peak 7- Notoginsenoside R ₁ Ginsenoside R _e +R _g1 Ginsenoside R _b1 Notoginsenoside R ₂ Ginsenoside R _h1 Ginsenoside Rhliso.(F1) Ginsenoside R _d Peak 8- Peak 9- Peak 10- Peak 11- Peak 12- Peak 13- Peak 14- Notoginsenoside R ₂ -H2O Notoginsenoside R ₂ -H2O Ginsenoside R _g6 /F ₄ Ginsenoside R _k3 /R _h4 (R _k3 ) Ginsenoside R _k3 /R _h4 (R _h4 ) Ginsenoside R _k1 /R _g5 (R _k1 ) Ginsenoside R _k1 /R _g5 (R _g5 )

A total of more than 200 batches of compound Danshen extract were analyzed for the fingerprints of Danshen and Panax notoginseng, and the similarity was above 90%. Now the retention time of the chromatogram, the average value of the peak area and the RSD value are summarized as follows:

Panax notoginseng fingerprint part peak number average retention time RSD% of retention time average peak area RSD% of peak area Single peak as a percentage of total peak area 1 10.85 0.08 557.29 15.67 5.36% 2 12.1 0.06 2728.83 16.66 26.23% 3 20.33 0.08 3704.31 14.15 35.61% 4 20.86 0.07 272.69 12.54 2.62% 5 23.28 0.08 369.12 21.52 3.55% 6 24.48 0.08 190.23 17.45 1.83% 7 28.82 1.36 289.16 17.71 2.78% 8 42.93 0.07 239.14 18.84 2.30% 9 45.66 0.06 445.17 19.56 4.28% 10 47.85 0.06 723.59 14.24 6.96% 11 68.17 0.03 407.66 10.86 3.92% 12 69.4 0.03 476.31 15.46 4.58%

A total of more than 200 batches of compound Danshen dripping pills were analyzed for the fingerprints of Danshen and Panax notoginseng, and the similarity was above 90%. Now the retention time of the chromatogram, the average value of the peak area and the RSD value are summarized as follows:

Panax notoginseng fingerprint part peak number average retention time Retention time RSD% average peak area Peak area RSD% Single peak as a percentage of total peak area 1 10.87 0.45 506.92 13.03 4.27% 2 12.12 0.34 2723.73 12.11 22.93% 3 20.34 0.23 1684.49 18.95 14.18% 4 20.86 0.16 231.84 18.99 1.95% 5 23.29 0.25 565.78 13.8 4.76% 6 24.48 0.28 309.39 18.98 2.60% 7 29 0.62 345.32 14.28 2.91% 8 41.21 0.25 436.06 10.88 3.67% 9 42.94 0.21 665.38 11.82 5.60% 10 44.16 0.21 472.84 14.59 3.98% 11 45.68 0.23 947.39 16.06 7.97% 12 47.88 0.25 1547.8 13.25 13.03% 13 68.21 0.13 622.91 10.39 5.24% 14 69.43 0.12 820.24 12.93 6.90%

Embodiment eight (compound Danshen dripping pills control fingerprint establishment)

1. Instruments and reagents

Instrument: Agilent 1100 liquid chromatography, including quaternary pump, online degassing device, automatic sampler, DAD detector, column thermostat, Chemstation workstation; BS210S electronic balance (1/10 ^-4 g) (Beijing Sedoli Sri Lanka), METTLERAE240 electronic balance (1/10 ^-4 g or 1/10 ^-5 g) (Mettler-Toledo (Shanghai) Co., Ltd.), LD4-2 centrifuge (4000r/min) (Beijing Medical Centrifuge machine factory), digital constant temperature water bath (Tianjin Changfeng Co., Ltd.), RE-52AA rotary evaporator (Shanghai Yarong Biochemical Instrument Factory), SHE-(III) circulating water vacuum pump (Gongyi Yingyu Yuhua Instrument Factory) , KQ-250B ultrasonic cleaner (Kunshan Ultrasonic Instrument Co., Ltd.), HENGAO T&D filter (HENGGAO T&D), synthetic fiber filter membrane (pore size 0.45μm) (Shanghai Xingya Purification Material Factory); C ₁₈ small column (filler is C ₁₈ H ₁₇ liquid chromatography stationary phase, 50-100 mesh, produced by Tianjin Chemical Reagent No. 2 Factory. Dry packing 600mg, column inner diameter 1cm).

Reagents: acetonitrile (chromatographically pure, Merck, USA), acetic acid (premium grade), Wahaha purified water.

2. Preparation of the test article

Weigh 1.0g of Compound Danshen Dripping Pills, dissolve in 10mL of 4% ammonia water, ultrasonically dissolve, pass through a 0.45μm filter membrane, take 5mL of filtrate and pass through a C-18 small column with 500mg filler, rinse with 20mL of 20% methanol and 4% ammonia solution, discard and rinse Then wash with 20ml of water, discard the washing solution, and then elute with methanol, collect the methanol eluate in a 5mL volumetric flask, constant volume, centrifuge for later use, parallel 2 parts, injection volume 20μL;

The preparation of compound Danshen dripping pill intermediate test sample: take by weighing 0.2g of each batch of Danshen Sanqi extract in embodiment two, be dissolved in 10mL 4% ammonia water, ultrasonically dissolve, cross 0.45 μm filter membrane, get 5mL filtrate and cross 500mg Packed C-18 small column, washed with 20mL of 20% methanol and 4% ammonia solution, discarded the washing solution, then washed with 20ml of water, discarded the water washing solution, and then precipitated with methanol, collected the methanol eluate in a 5mL volumetric flask, constant volume, Centrifuge for standby, 2 parallel copies, injection volume 20μL;

The preparation of the notoginseng medical material test sample: take by weighing 2.5g of the medicinal material in a reflux bottle, add 50ml of distilled water, reflux for 1.5 hours, filter, add 50ml of distilled water in the filter residue, reflux for 1 hour, combine the filtrate, and dissolve it in a 100ml measuring bottle. Centrifugal filtration, spare. Two parallel copies were made, and the injection volume was 10 μl.

3. HPLC analysis conditions

Agilent SB-C ₁₈ analytical column (4.6mm×250mm, Zorbax SB USCL010304); mobile phase: Phase A is 0.01% (V/V) glacial acetic acid aqueous solution, and phase B is acetonitrile solution containing 0.01% glacial acetic acid. The flow rate is 0.8ml·min ^-1 ; the detection wavelength is 203nm; the column temperature is 30°C.

The gradient elution program is as follows: Time A: 0.01% HAC-H2O (%) B: 0.01% HAC-CH3CN (%) 0152540506575 80656557574225 20353543435875

Determination of 40 batches of Compound Danshen Dripping Pills, Panax notoginseng medicinal materials, and Compound Danshen Dripping Pills intermediates according to the above method to obtain fingerprints:

In the fingerprint spectrum, there are 14 absorption peaks of the chemical components of the notoginseng component in the compound Danshen dripping pills, among which there are 13 absorption peaks whose single peak area exceeds 2% of the total peak area, which are respectively:

For peak No. 1, the average retention time RT is 10.87min, the RSD is 0.31%, the peak area is 506.92, and the RSD is 13.03%;

For peak No. 2, the average retention time RT is 12.12min, the RSD is 0.34%, the peak area is 2723.73, and the RSD is 12.11%;

Peak No. 3, average retention time RT is 20.34min, RSD is 0.23%, peak area is 1684.49, RSD is 18.59%;

Peak No. 5, the average retention time RT is 23.29min, RSD is 0.25%, peak area is 565.78, RSD is 13.80%;

For peak No. 6, the average retention time RT is 24.48min, the RSD is 0.28%, the peak area is 309.39, and the RSD is 18.98%;

For peak No. 7, the average retention time RT is 29.00min, the RSD is 0.62%, the peak area is 345.32, and the RSD is 14.28%;

Peak No. 8, the average retention time RT is 41.21min, RSD is 0.25%, peak area is 436.06, RSD is 10.88%;

For peak No.9, the average retention time RT is 42.94min, the RSD is 0.21%, the peak area is 665.38, and the RSD is 11.82%;

For peak No. 10, the average retention time RT is 44.16min, the RSD is 0.21%, the peak area is 472.84, and the RSD is 14.59%;

For peak No. 11, the average retention time RT is 45.68min, the RSD is 0.23%, the peak area is 947.39, and the RSD is 16.06%;

For peak No. 12, the average retention time RT is 47.88min, the RSD is 0.25%, the peak area is 1547.80, and the RSD is 13.25%;

For peak No. 13, the average retention time RT is 68.21min, the RSD is 0.13%, the peak area is 622.91, and the RSD is 10.39%;

For peak No. 14, the average retention time RT is 69.43min, the RSD is 0.12%, the peak area is 820.24, and the RSD is 12.94%;

In the fingerprint spectrum, there are 5 absorption peaks of the component chemical components of the Panax notoginseng medical material test product, and wherein there are 5 absorption peaks with a single peak area exceeding 2% of the total peak area, which are respectively:

For peak No. 1, the average retention time RT is 10.81min, the RSD is 0.06%, the peak area is 539.57, and the RSD is 17.73%;

For peak No. 2, the average retention time RT is 12.03min, the RSD is 0.02%, the peak area is 2482.50, and the RSD is 8.74%;

For peak No. 3, the average retention time RT is 20.16min, the RSD is 0.11%, the peak area is 1514.5, and the RSD is 12.02%;

The average retention time RT of No. 4 peak is 23.04min, RSD is 0.07%, peak area is 250.63, RSD is 18.25%;

Peak No. 5, the average retention time RT is 28.42min, RSD is 0.27%, peak area is 356.20, RSD is 15.30%;

There are 12 absorption peaks of the chemical components of Sanqi component in the intermediate of Compound Danshen Dripping Pills, among which there are 11 absorption peaks whose single peak area exceeds 2% of the total peak area, which are:

For peak No. 1, the average retention time RT is 10.85min, the RSD is 0.08%, the peak area is 557.29, and the RSD is 15.67%;

For peak No. 2, the average retention time RT is 12.10min, the RSD is 0.06%, the peak area is 2728.83, and the RSD is 16.66%;

For peak No. 3, the average retention time RT is 20.33min, the RSD is 0.08%, the peak area is 3704.31, and the RSD is 14.15%;

Peak No. 4, the average retention time RT is 23.86min, RSD is 0.07%, peak area is 272.69, RSD is 12.54%;

Peak No. 5, the average retention time RT is 23.28min, RSD is 0.08%, peak area is 369.12, RSD is 21.52%;

For peak No. 7, the average retention time RT is 28.82min, the RSD is 1.36%, the peak area is 289.16, and the RSD is 17.71%;

Peak No. 8, average retention time RT is 42.93min, RSD is 0.07%, peak area is 239.14, RSD is 18.84%;

Peak No. 9, the average retention time RT is 45.66min, RSD is 0.06%, peak area is 445.17, RSD is 19.56%;

For peak No. 10, the average retention time RT is 47.85min, the RSD is 0.06%, the peak area is 723.59, and the RSD is 14.24%;

For peak No. 11, the average retention time RT is 68.17min, the RSD is 0.03%, the peak area is 407.66, and the RSD is 10.86%;

For peak No. 12, the average retention time RT is 69.4min, the RSD is 0.03%, the peak area is 476.31, and the RSD is 15.46%.

Claims (11)

1. a quality control method of compound danshen dripping pill, is characterized in that comprising the steps: (a). Establishment of the control fingerprint of Compound Danshen Dripping Pills Preparation of Compound Danshen Dripping Pills Control Sample Solution: It is called Compound Danshen Dripping Pills, dissolved in 1-20mL of 1-8% ammonia water, ultrasonically dissolved, passed through a 0.30-0.60μm filter membrane, and 2-10mL of the filtrate was passed through a packed C-18 column, 5-40mL of 8-35% ammonia Rinse with methanol 1-8% ammonia solution, discard the washing solution, then wash with 5-35ml water, discard the water washing solution, then elute with methanol, collect the methanol eluate in a 1-15mL volumetric flask, dilute to volume, and centrifuge for later use; Determination of Fingerprint of Compound Danshen Dripping Pills: Draw the above-mentioned control sample solution and inject it into a liquid chromatograph, and use high-performance liquid chromatography to measure it to obtain the fingerprint of the compound Danshen dripping pill. , mobile phase A is glacial acetic acid aqueous solution, mobile phase B is glacial acetic acid acetonitrile solution; detection wavelength is 200-210nm; (b). Determination of the fingerprint of the product to be tested in Compound Danshen Dripping Pills: Get the compound danshen dripping pill product to be tested, and measure the fingerprint of the product to be tested according to the steps and conditions described in the above (a); (c). Comparing the fingerprint of the compound danshen dripping pills to be tested with the control fingerprint of the compound danshen dripping pills to identify the number of absorption peaks to determine whether the product quality is qualified. 2. the quality control method of compound danshen dripping pills as claimed in claim 1, is characterized in that: The preparation method of Compound Danshen Dripping Pills reference sample solution in the (a) step is to claim Compound Danshen Dripping Pills, dissolve in 5-15mL of 2-6% ammonia water, ultrasonically dissolve, pass through a 0.35-0.55 μm filter membrane, and take 2-10mL Pass the filtrate through a packed C-18 small column, wash with 10-30mL of 10-30% methanol and 2-6% ammonia solution, discard the washing solution, then wash with 10-30ml of water, discard the water washing solution, then elute with methanol, collect the methanol for washing Remove the liquid in a 1-10mL volumetric flask, constant volume, and centrifuge for later use; Chromatographic conditions: the chromatographic column is octadecylsilane bonded silica gel as filler; gradient elution is adopted, mobile phase A is 0.01% glacial acetic acid aqueous solution, mobile phase B is acetonitrile solution containing 0.01% glacial acetic acid; gradient elution The procedure is as follows: At 0 minutes, mobile phase A was 80% 0.01% glacial acetic acid aqueous solution, and mobile phase B was 20% 0.01% glacial acetic acid acetonitrile solution; At 15 minutes, mobile phase A was 65% 0.01% glacial acetic acid aqueous solution, and mobile phase B was 35% 0.01% glacial acetic acid acetonitrile solution; At 25 minutes, mobile phase A was 65% 0.01% glacial acetic acid aqueous solution, and mobile phase B was 35% 0.01% glacial acetic acid acetonitrile solution; At 40 minutes, mobile phase A was 57% 0.01% glacial acetic acid aqueous solution, and mobile phase B was 43% 0.01% glacial acetic acid acetonitrile solution; At 50 minutes, mobile phase A was 57% 0.01% glacial acetic acid aqueous solution, and mobile phase B was 43% 0.01% glacial acetic acid acetonitrile solution; At 65 minutes, mobile phase A was 42% 0.01% glacial acetic acid aqueous solution, and mobile phase B was 58% 0.01% glacial acetic acid acetonitrile solution; At 75 minutes, mobile phase A was 25% 0.01% glacial acetic acid aqueous solution, and mobile phase B was 75% 0.01% glacial acetic acid acetonitrile solution; The injection volume is 5-45μL; the flow rate is 0.8mL·min-1; the detection wavelength is 201-205nm; the column temperature is 25-35°C. 3. the quality control method of compound danshen dripping pills as claimed in claim 2, is characterized in that: The preparation method of Compound Danshen Dropping Pills reference sample solution in the (a) step is as follows: claim 0.4～1.5g of Compound Danshen Dropping Pills, dissolve in 10mL4% ammonia water, ultrasonically dissolve, cross 0.45 μm filter membrane, get 5mL filtrate and cross 500mg filler For the C-18 small column, wash with 20mL of 20% methanol and 4% ammonia solution, discard the washing solution, then wash with 20ml of water, discard the water washing solution, and then elute with methanol, collect the methanol eluate in a 5mL volumetric flask, dilute to volume, and centrifuge For standby, the injection volume is 20 μL; Chromatographic conditions: injection volume 20 μL; flow rate 0.8 mL·min ^-1 ; detection wavelength 203 nm; column temperature 30°C. 4. as claimed in claim 1 or 2 or 3 described compound danshen dropping pills quality control method, it is characterized in that in contrast to the establishment of compound danshen dropping pills fingerprint, the preparation ratio of its mobile phase A solution is to prepare by volume ratio, flow The preparation ratio of phase B solution is prepared by volume ratio. 5. the product quality control method of compound danshen dripping pill as claimed in claim 1, 2 or 3 is characterized in that, there are 14 absorption peaks in the contrast fingerprint of described compound danshen dripping pill, and wherein single peak area exceeds total There are 3 absorption peaks with a peak area of 10%, which are: For peak No. 2, the average retention time RT is 12.12min, the RSD is 0.34%, the peak area is 2723.73, and the RSD is 12.11%; Peak No. 3, average retention time RT is 20.34min, RSD is 0.23%, peak area is 1684.49, RSD is 18.59%; For peak No. 12, the average retention time RT is 47.88min, the RSD is 0.25%, the peak area is 1547.80, and the RSD is 13.25%. 6. the compound danshen dropping pill product quality control method as claimed in claim 1, 2 or 3, it is characterized in that there are 14 absorption peaks in the contrast fingerprint collection of described compound danshen dropping pill, wherein single peak area exceeds total peak area There are 7 absorption peaks for 5%, which are: For peak No. 2, the average retention time RT is 12.12min, the RSD is 0.34%, the peak area is 2723.73, and the RSD is 12.11%; Peak No. 3, average retention time RT is 20.34min, RSD is 0.23%, peak area is 1684.49, RSD is 18.59%; For peak No.9, the average retention time RT is 42.94min, the RSD is 0.21%, the peak area is 665.38, and the RSD is 11.82%; For peak No. 11, the average retention time RT is 45.68min, the RSD is 0.23%, the peak area is 947.39, and the RSD is 16.06%; For peak No. 12, the average retention time RT is 47.88min, the RSD is 0.25%, the peak area is 1547.80, and the RSD is 13.25%; For peak No. 13, the average retention time RT is 68.21min, the RSD is 0.13%, the peak area is 622.91, and the RSD is 10.39%; For peak No. 14, the average retention time RT is 69.43min, the RSD is 0.12%, the peak area is 820.24, and the RSD is 12.94%. 7. the product quality control method of compound danshen dripping pill as claimed in claim 1,2 or 3, it is characterized in that, there are 8 absorption peaks in the contrast fingerprint of described compound danshen dripping pill, and its single peak area all exceeds There are 13 absorption peaks at 2% of the total peak area, which are: For peak No. 1, the average retention time RT is 10.87min, the RSD is 0.31%, the peak area is 506.92, and the RSD is 13.03%; For peak No. 2, the average retention time RT is 12.12min, the RSD is 0.34%, the peak area is 2723.73, and the RSD is 12.11%; Peak No. 3, average retention time RT is 20.34min, RSD is 0.23%, peak area is 1684.49, RSD is 18.59%; Peak No. 5, the average retention time RT is 23.29min, RSD is 0.25%, peak area is 565.78, RSD is 13.80%; For peak No. 6, the average retention time RT is 24.48min, the RSD is 0.28%, the peak area is 309.39, and the RSD is 18.98%; For peak No. 7, the average retention time RT is 29.00min, the RSD is 0.62%, the peak area is 345.32, and the RSD is 14.28%; Peak No. 8, the average retention time RT is 41.21min, RSD is 0.25%, peak area is 436.06, RSD is 10.88%; For peak No.9, the average retention time RT is 42.94min, the RSD is 0.21%, the peak area is 665.38, and the RSD is 11.82%; For peak No. 10, the average retention time RT is 44.16min, the RSD is 0.21%, the peak area is 472.84, and the RSD is 14.59%; For peak No. 11, the average retention time RT is 45.68min, the RSD is 0.23%, the peak area is 947.39, and the RSD is 16.06%; For peak No. 12, the average retention time RT is 47.88min, the RSD is 0.25%, the peak area is 1547.80, and the RSD is 13.25%; For peak No. 13, the average retention time RT is 68.21min, the RSD is 0.13%, the peak area is 622.91, and the RSD is 10.39%; For peak No. 14, the average retention time RT is 69.43min, the RSD is 0.12%, the peak area is 820.24, and the RSD is 12.94%. 8. the compound danshen dripping pill quality control method as claimed in claim 1, 2 or 3, is characterized in that, described compound danshen dripping pill product fingerprint to be tested compares with described compound danshen dripping pill contrast fingerprint, both When there are 3 or more identical absorption peaks in the fingerprint spectrum, it is considered that the quality of the compound Danshen dripping pill to be tested is qualified. 9. the quality control method of compound danshen dripping pills as claimed in claim 8, is characterized in that, described compound danshen dropping pills product fingerprint to be tested compares with described compound danshen dropping pill contrast fingerprint, both fingerprints have When there are 5 or more identical absorption peaks, it is considered that the quality of the compound Danshen dripping pills to be tested is qualified. 10. the quality control method of compound danshen dripping pills as claimed in claim 8, is characterized in that, described compound danshen dropping pills product fingerprint spectrum to be tested compares with described compound danshen dripping pill contrast fingerprint, both fingerprints have When there are 7 or more identical absorption peaks, it is considered that the quality of the compound Danshen dripping pills to be tested is qualified. 11. the method for quality control of compound danshen dripping pills as claimed in claim 8, is characterized in that tested compound danshen dropping pills product fingerprints compare with contrast compound danshen dropping pills fingerprints, both fingerprints have 13 or 13 When more than same absorption peak, think that the quality of described Compound Danshen Dripping Pills product to be tested is qualified. 12. The quality control method of Compound Danshen Dripping Pills as claimed in claim 8, characterized in that the fingerprints of the tested Compound Danshen Dripping Pills are compared with the fingerprints of the reference Compound Danshen Dripping Pills, and the two fingerprints have a total of 14 peaks. Just think that the Compound Danshen Dripping Pill product to be tested is a qualified product.

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