Traditional Chinese medicine composition, pharmaceutical preparation and application

Abstract

The invention relates to the technical field of traditional Chinese medicine, in particular to a traditional Chinese medicine composition, a pharmaceutical preparation and application. According to the invention, the crebanine and the rhizoma acori graminei volatile oil with the effect of returning to the heart channel are combined for use for the first time, so that the crebanine keeps the cumulative permeation amount and can maintain the effective blood concentration for a long time, and the transdermal absorption amount is increased to meet the treatment requirement; compared with single use of crebanine and combination of the crebanine and the rhizoma acori graminei volatile oil, the percutaneous penetration amount is increased, the peak reaching time is shortened, the percutaneous blood entering content of the crebanine is increased, it is verified that the rhizoma acori graminei volatile oil can play a role in promoting medicine to enter a body, and therefore the traditional Chinese medicine composition is safer, more effective, milder and lasting, the action mechanism is multi-target and multi-effect, and the effect is remarkable. According to the invention, the treatment effect of the crebanine is improved, a more effective and safer treatment strategy is provided for patients with arrhythmia and myocardial ischemia, and a thought and a basis are also provided for developing new drugs for treating cardiovascular diseases.

Description

Technical Field

[0001] The present invention relates to the technical field of traditional Chinese medicine, and in particular to a traditional Chinese medicine composition, a pharmaceutical preparation and uses. Background Art

[0002] Cardiovascular disease (CVD) is one of the major issues in human health, with high morbidity and mortality, affecting most adults over 60 years old worldwide. The main manifestations are malignant arrhythmias, Adams-Stokes syndrome, ventricular tachycardia, ventricular fibrillation and death from ischemic heart disease. Myocardial ischemia-reperfusion (MI / R) is an inevitable part of the treatment of myocardial ischemia, but reperfusion can cause more severe myocardial damage, even exceeding its beneficial effects. Currently, cardiovascular therapeutic drugs will produce a certain degree of liver and kidney damage and drug resistance, and single-channel blockers are more likely to cause side effects. Most of the current clinical use is single-channel blockers, and multi-ion channel blockers are considered to have higher effectiveness and safety.

[0003] Crebanine (Cre) is an aporphyrin-type alkaloid compound that can be isolated from Stephanina Yunnanensis HSLo of the Menispermaceae family and several other plants of the same genus (such as long-petaled crebanine, small-leaved crebanine, Hainan crebanine, medicinal crebanine, white-line potato and tooth-leaved crebanine, etc.). Cre has strong cardiovascular activity and can counteract a variety of experimental arrhythmias in animals caused by drugs (aconitine, barium chloride and chloroform, etc.) and surgery (myocardial ischemia-reperfusion), and its effect is better than that of the positive drug verapamil hydrochloride. However, in clinical practice, it is mainly oral and injection. Crebanine injection has a small antiarrhythmic therapeutic index and a relatively narrow safety range. Improper dosage may lead to poisoning or death. Although crebanine administered by intravenous or oral administration has a rapid onset of arrhythmia, it lasts for a short time and is prone to recurrence. Kebanin can exert anti-arrhythmic and anti-myocardial ischemia-reperfusion effects through multiple targets and multiple pathways, and is suitable for the preparation of transdermal drug delivery systems, but its transdermal permeation is limited, and in-depth research is needed on the synergistic enhancement of drug-adjuvant dual-use transdermal permeation and anti-myocardial ischemia activity mechanisms based on traditional Chinese medicine theory. Therefore, the development of transdermal drug delivery preparations that are effective in treating cardiovascular diseases and have multi-target synergistic effects has important theoretical and practical significance and can fill the clinical demand. Summary of the invention

[0004] In order to overcome the above technical defects, the present invention selects a specific proportion of crebanine and Acorus calamus volatile oil to exert the synergistic effect of percutaneous penetration and heart meridian opening, and reduces heart damage and apoptosis of cardiomyocytes by inhibiting endoplasmic reticulum stress, oxidative stress, and inflammatory response. It is used for the treatment of arrhythmia and myocardial ischemia, playing the role of "drug-adjuvant combination".

[0005] One of the purposes of the present invention is to provide a traditional Chinese medicine composition, comprising crebanine and Acorus calamus volatile oil.

[0006] Acorus tatarinowii Schott is derived from the dried rhizome of Acorus tatarinowii Schott, a plant of the Araceae family. It has the effects of improving the vitality of cardiomyocytes, protecting vascular endothelial cells, reducing ischemia / reperfusion injury of primary cardiomyocytes, inhibiting adriamycin-induced cardiomyocyte injury and apoptosis, preventing cardiomyocyte calcium overload, and reducing myocardial tissue damage in rats with acute myocardial infarction. Acorus tatarinowii Schott volatile oil can inhibit rapid arrhythmias caused by drugs such as aconitine, epinephrine, and barium chloride, and reduce the beating frequency of cardiomyocytes and enhance the vitality of cardiomyocytes.

[0007] In traditional Chinese medicine theory, Acorus calamus belongs to the heart meridian, and has the effects of removing dampness and stimulating appetite, opening the mind and removing phlegm, and refreshing the mind and improving intelligence. However, there are few reports on the application of Acorus calamus volatile oil combined with clebanine in the treatment of arrhythmia and myocardial ischemia. Therefore, the present invention uses Acorus calamus volatile oil and clebanine as research objects, and mixes them in a specific ratio after screening, so as to play a synergistic effect of percutaneous penetration and opening the heart meridian, thereby achieving the effect of "drug-adjuvant combination".

[0008] Further, the composition comprises: 1-30 parts by weight of crebanine; such as 5-30 parts by weight, or 10-30 parts by weight, or 15-30 parts by weight, or 20-30 parts by weight; and

[0009] 1-100 parts by weight of Acorus calamus volatile oil; such as 10-100 parts by weight, or 30-100 parts by weight, or 50-100 parts by weight, or 80-100 parts by weight.

[0010] The second object of the present invention is to provide a pharmaceutical preparation, which is composed of the above composition and pharmaceutical excipients acceptable in the art.

[0011] The pharmaceutical preparation is any one of tablets, capsules, pills, powders, granules, syrups, solutions, emulsions, injections, sprays, aerosols, and patches; the pharmaceutical preparation is administered through gastrointestinal and parenteral routes.

[0012] Particularly, the non-gastrointestinal administration route is selected from injection, respiratory tract administration, skin administration, mucosal administration or cavity administration.

[0013] Among them, the parenteral preparation is selected from injections, sprays, aerosols, patches and the like.

[0014] Particularly, the gastrointestinal administration preparation is selected from tablets, capsules, powders, granules, pills, solutions, emulsions or syrups.

[0015] Furthermore, the pharmaceutical excipient is one or more of a gel matrix, an oil phase matrix, a moisturizer, and an emulsifier;

[0016] The gel matrix is ​​selected from one or more of methyl cellulose, ethyl cellulose, hydroxyethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, sodium carboxymethyl cellulose, carbomer 940, gelatin, and sodium alginate, preferably, selected from carbomer 940;

[0017] The oil phase matrix is ​​selected from one or more of stearic acid, paraffin, beeswax, higher alcohol, liquid paraffin, vaseline or vegetable oil; preferably, selected from one or more of soybean lecithin, cholesterol, lanolin and stearic acid;

[0018] The moisturizing agent is selected from one or more of glycerol, 1,3-butylene glycol, propylene glycol, polyethylene glycol 300, polyethylene glycol 400, and sorbitol; preferably, selected from one or more of propylene glycol, glycerol, and polyethylene glycol 400;

[0019] The emulsifier is preferably one or more of soaps, sodium lauryl sulfate, fatty acid esters of polyols (such as glyceryl monostearate), polysorbates, and polyethylene glycol ethers, preferably one or more of sodium deoxycholate, Tween 80, and Span 80;

[0020] Furthermore, the pharmaceutical preparation is any one of an ointment, a cream, a gel, a patch, a gel patch, and a gel cream.

[0021] Furthermore, the pharmaceutical preparation is a gel, comprising the above composition, a gel matrix, and a moisturizer;

[0022] The invention prepares a gel with the best performance by optimizing the ratio of crebanine and calamus calamus volatile oil and on this basis.

[0023] The gel matrix is ​​selected from one or more of methyl cellulose, ethyl cellulose, hydroxyethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, sodium carboxymethyl cellulose, carbomer 940, gelatin, and sodium alginate, preferably, selected from carbomer 940;

[0024] The humectant is selected from one or more of glycerol, 1,3-butylene glycol, propylene glycol, polyethylene glycol 300, polyethylene glycol 400, and sorbitol; preferably, selected from one or more of propylene glycol, glycerol, and polyethylene glycol 400.

[0025] The above preparations can be prepared according to methods known in the art into suitable administration forms or dosage forms for human use.

[0026] In addition, if necessary, colorants, preservatives, perfumes, flavoring agents, pH adjusters, sweeteners or other materials may be added to the pharmaceutical preparations.

[0027] Furthermore, the pharmaceutical preparation is a liposome gel, comprising the above composition, a gel matrix, a moisturizer, an oil phase matrix, water, and an emulsifier; the amount of the composition added is 1-15%wt of the liposome gel; preferably, the amount added is 2-10%wt;

[0028] The invention prepares a liposome gel with optimal performance by adjusting the proportion of crebanine and calamus calamus volatile oil and on this basis.

[0029] The oil phase matrix is ​​selected from one or more of stearic acid, paraffin, beeswax, higher alcohol, liquid paraffin, vaseline or vegetable oil; preferably, selected from one or more of soybean lecithin, cholesterol, lanolin and stearic acid;

[0030] The emulsifier is preferably one or more of soaps, sodium lauryl sulfate, fatty acid esters of polyols (such as glyceryl monostearate), polysorbates, and polyethylene glycol ethers, preferably one or more of sodium deoxycholate, Tween 80, and Span 80.

[0031] Furthermore, the liposome gel is prepared by the following method: adding crebanine, soybean lecithin, cholesterol, vitamin E, and an emulsifier in a mass ratio of 1: (15-30): (1-2): (0.1-1.5): (5-15) to an organic solvent, dissolving them to obtain a clear and transparent solution, removing the organic solvent to obtain a uniform lipid film; dissolving the lipid film in a hydrating liquid to obtain crebanine liposomes; mixing carbomer 940 and glycerol to obtain a blank gel, and then adding crebanine liposomes and Acorus calamus volatile oil to obtain a finished product.

[0032] The prepared crebanine liposomes are spherical, evenly distributed, and have a complete structure without obvious aggregation. The encapsulation rate is greater than 90%. The average particle size of the liposomes is 73.01±1.66nm. The average potential of the liposomes is -34.75±2.71mV. The liposomes are mixed with calamus volatile oil to prepare a liposome gel with optimal performance.

[0033] The dosage of the pharmaceutical composition of the present invention depends on many factors, such as the nature and severity of the allergic asthma to be prevented or treated, the sex, age, weight and individual response of the patient or animal, the route of administration and the number of administrations, etc. The above dosage can be administered in a single dosage form or divided into several, such as two, three or four dosage forms. The dosage level must be selected based on the specific route of administration, the severity of the condition to be treated, and the condition and previous medical history of the patient to be treated. However, the practice in the art is to start the dosage from a level lower than that required to obtain the desired therapeutic effect and gradually increase the dosage until the desired effect is obtained.

[0034] The third object of the present invention is to provide a pharmaceutical composition and a pharmaceutical preparation for use in the prevention and / or treatment of cardiovascular and cerebrovascular diseases.

[0035] Furthermore, the mechanism of action is manifested in up-regulating the expression levels of Bax and Cx43, down-regulating the expression levels of ATF-6, GRP78, and CHOP proteins, reducing the release of MDA and LDH, and increasing the level of SOD.

[0036] The present invention combines crebanine with Acorus tatarinowii volatile oil having a "returning to the heart meridian" effect for the first time, so that crebanine maintains a cumulative penetration amount and can maintain an effective blood drug concentration for a long time, thereby increasing the transdermal absorption amount to meet treatment needs; compared with the use of crebanine alone, the transdermal penetration amount is increased and the peak time is shortened after the combination with Acorus tatarinowii volatile oil, and the transdermal blood content of crebanine is increased, which verifies that Acorus tatarinowii volatile oil can play a "drug entry-promoting" role, the expression levels of related factors Bax and Cx43 are increased, the expression levels of ATF-6, GRP78 and CHOP proteins are decreased, the release of MDA and LDH is reduced, and the level of SOD is increased. It can be seen that the Chinese medicine composition of the present invention is safer, more effective, milder and more long-lasting, and has a multi-target and multi-effect mechanism of action, which improves the therapeutic effect of crebanine. At the same time, the TCM meridian theory is supported by pharmacokinetics, tissue distribution and pharmacodynamic studies, providing theoretical support for the research of TCM meridian theory, providing a more effective and safer treatment strategy for patients with arrhythmia and myocardial ischemia, and also providing ideas and basis for the development of new drugs for the treatment of cardiovascular diseases. BRIEF DESCRIPTION OF THE DRAWINGS

[0037] Figure 1 The present invention is to TC-induced H 9 C 2 Cellular effects;

[0038] Where 1a is H 9 C 2 Cell survival rate diagram; 1b is H 9 C 2Cell apoptosis rate diagram; compared with the Model group, *P<0.05, **P<0.01, ***P<0.001; compared with the Cre group, ns means no significant difference; compared with the Control group, #P<0.05, ##P<0.01, ###P<0.001, compared with the Cre-H+VOA-H group, &P<0.05, &&P<0.01, &&&P<0.001; Figure 2 Each TC induced H 9 C 2 Effect diagram of antioxidant enzyme activity of cells; compared with Model group, *P<0.05, **P<0.01, ***P<0.001; compared with Cre group, ns means no significant difference; compared with Control group, #P<0.05, ##P<0.01, ###P<0.001, compared with Cre-H+VOA-H group, &P<0.05, &&P<0.01, &&&P<0.001;

[0039] Figure 3 Each TC induced H 9 C 2 Expression of apoptosis-related proteins in cells; compared with the Model group, *P<0.05, **P<0.01, ***P<0.001; compared with the Cre group, ns means no significant difference; compared with the Control group, #P<0.05, ##P<0.01, ###P<0.001, compared with the Cre-H+VOA-H group, &P<0.05, &&P<0.01, &&&P<0.001;

[0040] Figure 4 Figure 2 is a graph of serum factors of rats with arrhythmia in the examples; compared with the Model group, *P<0.05, **P<0.01, ***P<0.001; compared with the Cre group, ns indicates no significant difference, #P<0.05, ##P<0.01, ###P<0.001; Figure 5 The expression diagram of related enzymes in myocardial tissue of rats with arrhythmia in the examples; compared with the Model group,

[0041] *P<0.05, **P<0.01, ***P<0.001; compared with the Cre group, ns means no significant difference, #P<0.05, P<0.01, ###P<0.001;

[0042] Figure 6 The TTC staining images of myocardial tissues of rats with myocardial ischemia-reperfusion in each embodiment;

[0043] Among them, 6A is the TTC slice of myocardial tissue; 6B is the statistical diagram of the infarct area of ​​each drug-treated group; compared with the Model group, *P<0.05, **P<0.01, ***P<0.001; compared with the Cre group, ns means no significant difference, #P<0.05, ##P<0.01, ###P<0.001;

[0044] Figure 7 HE slices of myocardial tissues of rats with myocardial ischemia-reperfusion in each embodiment;

[0045] Figure 8 The expression diagram of serum myocardial injury markers in rats with myocardial ischemia-reperfusion in the examples; compared with the Model group, *P<0.05, **P<0.01, ***P<0.001; compared with the Cre group, ns means no significant difference, #P<0.05, ##P<0.01, ###P<0.001;

[0046] Fig. 9 The expression graphs of apoptosis-related proteins in rats with myocardial ischemia-reperfusion in the examples are shown; compared with the Model group, *P<0.05, **P<0.01, ***P<0.001; compared with the Cre group, ns indicates no significant difference, #P<0.05, ##P<0.01, ###P<0.001;

[0047] Fig.10 The expression graphs of endoplasmic reticulum stress-related proteins in rats with myocardial ischemia-reperfusion in the examples are shown; compared with the Model group, *P<0.05, **P<0.01, ***P<0.001; compared with the Cre group, ns indicates no significant difference, #P<0.05, ##P<0.01, ###P<0.001;

[0048] Fig.11 The expression diagram of serum oxidative stress-related factors in rats with myocardial ischemia-reperfusion in the examples; compared with the Model group, *P<0.05, **P<0.01, ***P<0.001; compared with the Cre group, ns means no significant difference, #P<0.05, ##P<0.01, ###P<0.001;

[0049] Fig.12 The expression diagram of inflammation-related proteins in rats with myocardial ischemia-reperfusion in the examples; compared with the Model group, *P<0.05, **P<0.01, ***P<0.001; compared with the Cre group, ns means no significant difference, #P<0.05, ##P<0.01, ###P<0.001;

[0050] Fig.13 The pharmacokinetic parameters of each rat after administration in the examples are measured;

[0051] Fig.14 This is a graph of the crebanine content in each tissue of each rat in the example over time. DETAILED DESCRIPTION

[0052] In order to make those skilled in the art better understand the technical solution of the present invention, the present invention is described in detail below in conjunction with specific embodiments. The experimental methods in the following examples that do not specify specific conditions are usually based on conventional conditions or conditions recommended by the manufacturer. Unless otherwise specified, percentages and parts are calculated by weight. Unless otherwise defined, all professional and scientific terms used in the text have the same meanings as those familiar to those skilled in the art. In addition, any method and material similar or equivalent to the recorded content can be applied to the present invention. The preferred implementation methods and materials described in the text are for demonstration purposes only.

[0053] The endpoints and any values ​​of the ranges disclosed in this article are not limited to the precise ranges or values, and these ranges or values ​​should be understood to include values ​​close to these ranges or values. For numerical ranges, the endpoint values ​​of each range, the endpoint values ​​of each range and the individual point values, and the individual point values ​​can be combined with each other to obtain one or more new numerical ranges, which should be considered as specifically disclosed in this article.

[0054] The present invention is based on the medicinal value of crebanine in cardiovascular and cerebrovascular diseases, selects and uses the volatile oil of Acorus calamus with osmotic activity in combination, exerts the synergistic effect of percutaneous penetration and heart meridian opening, promotes the drug to enter the target organ or tissue through various physiological barriers such as the blood-brain barrier, mucous membrane, skin, etc., achieves the purpose of alleviating heart damage, reducing the apoptosis of myocardial cells, and plays the role of "drug-adjuvant combination".

[0055] According to the present invention, on one hand, a traditional Chinese medicine composition is provided, comprising crebanine and Acorus calamus volatile oil.

[0056] In traditional Chinese medicine theory, Acorus tatarinowii belongs to the heart meridian and has the effects of removing dampness and stimulating appetite, opening the mind and removing phlegm, refreshing the mind and improving intelligence. Acorus tatarinowii has the effects of improving the vitality of cardiomyocytes, protecting vascular endothelial cells, reducing ischemia / reperfusion injury of primary cardiomyocytes, inhibiting adriamycin-induced cardiomyocyte injury and apoptosis, preventing cardiomyocyte calcium overload, and reducing myocardial tissue damage in rats with acute myocardial infarction. Acorus tatarinowii volatile oil can inhibit rapid arrhythmias caused by drugs such as aconitine, epinephrine, and barium chloride, and reduce the beating frequency of cardiomyocytes and enhance the vitality of cardiomyocytes.

[0057] However, there are few reports on the application of Acorus calamus volatile oil and clebanine in the treatment of arrhythmia and myocardial ischemia. The present invention uses Acorus calamus volatile oil and clebanine as research objects, and mixes them in a specific ratio after screening to play a synergistic role of percutaneous penetration and heart meridian, thereby achieving the effect of "drug-adjuvant combination".

[0058] In some specific embodiments, the composition comprises: 1-50 parts by weight of crebanine; such as 5-50 parts by weight, or 10-40 parts by weight, or 15-30 parts by weight, or 20-30 parts by weight; and

[0059] 1-100 parts by weight of Acorus calamus volatile oil; such as 10-100 parts by weight, or 30-100 parts by weight, or 50-100 parts by weight, or 80-100 parts by weight.

[0060] It is understood by those skilled in the art that the "parts by weight" of each Chinese herbal medicine refers to the dosage ratio of each Chinese herbal medicine, rather than the actual unit of mass. According to the actual situation, the unit part by weight can be any mass, such as 1 part by weight can be 1g, 500g or 1kg, or even 15g, 30g, etc.

[0061] In a second aspect, the present invention provides a pharmaceutical preparation, which is composed of the above-mentioned composition and pharmaceutical excipients acceptable in the art.

[0062] The pharmaceutical excipients of the present invention can be selected from one or more of the following: solvents, propellants, solubilizers, cosolvents, emulsifiers, colorants, adhesives, disintegrants, fillers, lubricants, wetting agents, osmotic pressure regulators, stabilizers, glidants, flavoring agents, preservatives, suspending agents, coating materials, fragrances, anti-adhesives, integrities, penetration enhancers, pH regulators, buffers, plasticizers, surfactants, foaming agents, defoamers, thickeners, inclusion agents, humectants, absorbents, diluents, flocculants and deflocculating agents, filter aids, and release retardants.

[0063] In some specific embodiments, the pharmaceutical excipient is one or more of a gel matrix, an oil phase matrix, a moisturizer, and an emulsifier;

[0064] Based on the present invention, the pharmaceutical preparation is any one of an ointment, a cream, a gel, a patch, a gel patch, and a gel cream.

[0065] In some specific embodiments, the gel matrix is ​​selected from one or more of methyl cellulose, ethyl cellulose, hydroxyethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, sodium carboxymethyl cellulose, carbomer 940, gelatin, and sodium alginate, preferably, selected from carbomer 940;

[0066] In some specific embodiments, the humectant is selected from one or more of glycerol, 1,3-butylene glycol, propylene glycol, polyethylene glycol 300, polyethylene glycol 400, and sorbitol; preferably, selected from one or more of propylene glycol, glycerol, and polyethylene glycol 400.

[0067] Based on the present invention, the pharmaceutical preparation is a liposome gel, comprising the above composition, a gel matrix, a moisturizer, an oil phase matrix, water, and an emulsifier;

[0068] In some specific embodiments, the oil phase matrix is ​​selected from one or more of stearic acid, paraffin, beeswax, higher alcohols, liquid paraffin, vaseline or vegetable oil; preferably, selected from one or more of soybean lecithin, cholesterol, lanolin and stearic acid;

[0069] In some specific embodiments, the emulsifier is preferably one or more of soaps, sodium lauryl sulfate, fatty acid esters of polyols (such as glyceryl monostearate), polysorbates, and polyethylene glycol ethers, preferably, one or more selected from sodium deoxycholate, Tween 80, and Span 80.

[0070] Those skilled in the art will appreciate that the above preparations can be prepared according to methods known in the art into suitable administration forms or dosage forms for human use.

[0071] In addition, if necessary, colorants, preservatives, perfumes, flavoring agents, sweeteners or other materials may be added to the pharmaceutical preparations.

[0072] Based on the present invention, the liposome gel is prepared by the following method: adding crebanine, soybean lecithin, cholesterol, vitamin E and emulsifier in a mass ratio of 1: (15-30): (1-2): (0.1-1.5): (5-15) into an organic solvent, dissolving to obtain a clear and transparent solution, removing the organic solvent to obtain a uniform lipid film; dissolving the lipid film in a hydrating liquid to obtain crebanine liposomes; mixing carbomer 940 and glycerol to obtain a blank gel, and then adding crebanine liposomes and calamus volatile oil to obtain a finished product.

[0073] In a third aspect, the present invention provides use of pharmaceutical compositions and pharmaceutical preparations in the prevention and / or treatment of cardiovascular and cerebrovascular diseases.

[0074] Based on the present invention, the mechanism of action is manifested in up-regulating the expression levels of Bax and Cx43, down-regulating the expression levels of ATF-6, GRP78 and CHOP proteins, reducing the release of MDA and LDH, and increasing the level of SOD.

[0075] The embodiments of the present invention will be described in detail below with reference to the examples, but those skilled in the art will appreciate that the following examples are only used to illustrate the present invention and should not be considered to limit the scope of the present invention. If no specific conditions are specified in the examples, the conditions are carried out according to conventional conditions or conditions recommended by the manufacturer.

[0076] Reagents or instruments without indicating the manufacturer are all conventional products that can be obtained through commercial purchase.

[0077] Example 1 Chinese medicine composition

[0078] 20 parts by weight of crebanine and 10 parts by weight of Acorus calamus volatile oil.

[0079] Example 2 Chinese medicine composition

[0080] 30 parts by weight of gramineous rice and 30 parts by weight of volatile oil of Acorus calamus.

[0081] Example 3 Chinese medicine composition

[0082] 40 parts by weight of gramineous rice and 80 parts by weight of volatile oil of Acorus calamus.

[0083] Example 4 Chinese medicine composition

[0084] 15 parts by weight of gramineous rice and 50 parts by weight of volatile oil of Acorus calamus.

[0085] Example 5 Preparation of Banning Liposome Suspension

[0086] Weigh 6 mg of crebanine, 120 mg of soy lecithin, 10 mg of cholesterol, and 6 mg of vitamin E, add appropriate amount of chloroform to dissolve, and stir with a magnetic stirrer to dissolve as the organic phase. Separately, take 30 mg of sodium deoxycholate and 50 mg of propylene glycol and add them to 5 mL of PBS with pH = 7.4 to dissolve as the aqueous phase; then inject the aqueous phase into the organic phase at a uniform speed, stir to remove the chloroform, and obtain.

[0087] Example 6 Preparation of Banning Liposome Suspension

[0088] Weigh 6 mg of crebanine, 180 mg of soy lecithin, 6 mg of cholesterol, and 9 mg of vitamin E, add appropriate amount of chloroform to dissolve, and stir with a magnetic stirrer to dissolve as the organic phase. Separately, take 60 mg of sodium deoxycholate and 50 mg of propylene glycol, add them into 5 mL of PBS with pH = 7.4 and dissolve as the aqueous phase; then, inject the aqueous phase into the organic phase at a uniform speed, stir to remove the chloroform, and obtain.

[0089] Example 7 Preparation of Banning Liposome Suspension

[0090] Weigh 6 mg of crebanine, 90 mg of soy lecithin, 12 mg of cholesterol, and 0.5 mg of vitamin E, add appropriate amount of chloroform to dissolve, and stir with a magnetic stirrer to dissolve as the organic phase. Separately, take 90 mg of sodium deoxycholate and 50 mg of propylene glycol, add them into 5 mL of PBS with pH = 7.4 to dissolve as the aqueous phase; then, inject the aqueous phase into the organic phase at a uniform speed, stir to remove the chloroform, and obtain.

[0091] Example 8 Liposome Preparation

[0092] Weigh 0.1 g of Carbomer 940 and 2 g of glycerol in an appropriate amount of deionized water, swell overnight at room temperature to obtain a blank gel, add 5 ml of the crebanine liposome suspension prepared in Example 1 and 0.1 g of Acorus calamus volatile oil, adjust the pH to 7, and obtain.

[0093] Example 9 Liposome Preparation

[0094] Weigh 0.1 g of Carbomer 940 and 2 g of glycerol in an appropriate amount of deionized water, swell overnight at room temperature to obtain a blank gel, add 5 ml of the crebanine liposome suspension prepared in Example 2 and 0.1 g of Acorus calamus volatile oil, adjust the pH to 7, and obtain.

[0095] Example 9 Liposome Preparation

[0096] Weigh 0.1 g of Carbomer 940 and 2 g of glycerol in an appropriate amount of deionized water, swell overnight at room temperature to obtain a blank gel, add the crebanine liposome suspension prepared in Example 3 and 0.1 g of Acorus calamus volatile oil, adjust the pH to 7, and obtain.

[0097] Example 10 Preparation of gel formulation

[0098] Weigh 0.1 g of Carbomer 940 and 1.5 mL of moisturizing agent glycerol in an appropriate amount of deionized water, swell overnight (12 h) at room temperature, add triethanolamine to adjust the pH to 6.8-7.0 to obtain a blank gel, and weigh 6 mg of crebanine and 0.1 g of Acorus calamus volatile oil. Dissolve the crebanine in an appropriate amount of 1% citric acid solution, and add the blank gel and Acorus calamus volatile oil to obtain a gel.

[0099] Example 11: Effect of Chinese herbal medicine composition on TG-induced H 9 C 2 Synergistic protective effect of endoplasmic reticulum stress injury in cardiomyocytes

[0100] Cultivate H 9 C 2 Cardiomyocytes were randomly divided into five groups (n=3 in each group), namely, a normal group (Control), a model group (TG), Crebanine (Cre), a volatile oil of Acorus tatarinowii (VOA), and the Chinese medicine composition of Example 2 (Cre+VOA). After the cells were grouped, the corresponding drugs were given 24 h in advance. After the drug pretreatment, thapsigargin (TG) was used to induce endoplasmic reticulum stress apoptosis for modeling. After the modeling, cells or serum were collected for cell viability detection, cell apoptosis detection, SOD, MDA and LDH level detection, GRP78, CHOP, Bax, Bcl-2, Cx43, ATF6, Caspase3 protein expression detection.

[0101] Experimental results: Flow cytometry results showed (see Figure 1), compared with the blank group, the apoptosis rate of cells in the TG group was significantly increased (P<0.01); compared with the TG group, the apoptosis rate of cells in each group was significantly decreased (P<0.01); compared with the Cre group and the VOA group, the apoptosis rate of cells in the Cre+VOA group was significantly decreased, and the difference was statistically significant (P<0.05);

[0102] from Figure 2-3 It can be seen that each drug-treated group can also significantly down-regulate the expression of endoplasmic reticulum stress and apoptosis-related proteins ATF-6, GRP78, CHOP, Caspase-3, Bcl-2, and up-regulate the expression of Bax and Cx43, reduce the release of MDA and LDH, and increase the level of SOD (P>0.05); compared with the Cre group, the Cre+VOA group can significantly enhance the regulatory effect of the above indicators (P>0.05). This shows that the combination of Acorus tatarinowii volatile oil and crebanine can significantly reduce TG-induced H by regulating the expression of ATF-6 / GRP78 / CHOP pathway and its downstream apoptosis proteins. 9 C 2 The combined use of the two is significantly better than either of them alone in terms of endoplasmic reticulum stress injury and cell apoptosis in cardiomyocytes; it can be seen that Acorus tatarinowii volatile oil can significantly enhance the protective effect of crebanine on cardiomyocytes.

[0103] Example 12 Investigating the synergistic therapeutic effect of the Chinese medicine composition on arrhythmia rat model

[0104] Experimental animals: SPF grade SD rats, male, weighing 160-200g, were purchased from Guangdong Medical Experimental Animal Center, with animal quality certificate number: No.44007200077920, license number: SCXK (Guangdong) 2018-0002. The animals were raised in the barrier environment of the experimental animal room of the School of Traditional Chinese Medicine of Guangzhou University of Chinese Medicine, and the experimental unit use license number is: SYXK (Guangdong) 2019-0202. Rats were raised in special IVC cages, with an environmental temperature of 24℃-26℃, a humidity of 40%-60%, and a lighting duration of 12h (8:00-20:00). Rats were given normal SPF feed and sterile water, and the bedding and cages were regularly replaced and disinfected. The experiment was conducted after 7 days of adaptive feeding.

[0105] Statistical analysis: SPSS 25.0 software was used for data processing. The experimental data were expressed as mean ± standard deviation. P < 0.05 was considered statistically different, and P < 0.01 was considered statistically significant.

[0106] Experimental animals and groups: Healthy SD male rats were randomly divided into 7 groups (n=6), namely, Crebanine group (Cre), the composition of Example 1 (Cre-VOA-L), the composition of Example 2 (Cre-VOA-M), the composition of Example 3 (Cre-VOA-H), model control group (Model), positive drug group (VP), and normal control group (Con).

[0107] Kebanning group: 50 mg kg per day -1 Administration by intragastric gavage;

[0108] Cre-VOA-L group: 75 mg kg / day -1 Administration by intragastric gavage;

[0109] Cre-VOA-M group: 75 mg kg / day -1 Administration by intragastric gavage;

[0110] Cre-VOA-H group: 150 mg kg / day -1 Administration by intragastric gavage;

[0111] Positive drug group: 40 mg kg / day -1 Administration by intragastric gavage;

[0112] Blank group: given an equal amount of normal saline every day;

[0113] Model group: given an equal amount of normal saline every day;

[0114] The experiment was conducted after 7 days of adaptive feeding of rats in each group. Rats in each group were fasted and deprived of water for 24 hours before administration. Each experimental group was gavaged 30 minutes in advance, and the blank group and the model group were given an equal amount of normal saline. After anesthesia, the rats were fixed in a supine position and the electrocardiogram was observed. Rats with normal electrocardiogram were selected to establish an arrhythmia model (modeling was performed by injection of aconitine or barium chloride into the tail vein). Blood was collected from the abdominal aorta, and serum was separated to detect changes in the levels of BNP, CK, and LDH in the serum of rats and the levels of CaMKⅡ, CaM, and Na in the heart tissue. + -K + -Changes in ATPase concentration.

[0115] Experimental results: From Figure 4-5 It can be seen that compared with the Model group, each drug-treated group effectively prevented the expression of BNP, LDH, CK in the serum of the Model group rats and CaMKⅡ, CaM, Na in the myocardial tissue + -K + -ATPase increased; compared with the Cre group, each Cre+VOA group could downregulate BNP, LDH, CK in serum of model rats and CaMKⅡ, CaM, Na in myocardial tissue + -K +-ATPase (P>0.05), indicating that Acorus tatarinowii volatile oil can cooperate with crebanine to counteract arrhythmia induced by aconitine and barium chloride in rats and the myocardial damage caused by it through sodium, potassium and calcium multiple ion channels, and Acorus tatarinowii volatile oil can significantly enhance the protective effect of crebanine on arrhythmia in rats.

[0116] Example 13 Investigating the synergistic therapeutic effect of the Chinese medicine composition on myocardial ischemia-reperfusion rat model

[0117] Experimental animals: SPF grade SD rats, male, weighing 160-200g, were purchased from Guangdong Medical Experimental Animal Center, with animal quality certificate number: No.44007200077920, license number: SCXK (Guangdong) 2018-0002. The animals were raised in the barrier environment of the experimental animal room of the School of Traditional Chinese Medicine of Guangzhou University of Chinese Medicine, and the experimental unit use license number is: SYXK (Guangdong) 2019-0202. Rats were raised in special IVC cages, with an environmental temperature of 24℃-26℃, a humidity of 40%-60%, and a lighting duration of 12h (8:00-20:00). Rats were given normal SPF feed and sterile water, and the bedding and cages were regularly replaced and disinfected. The experiment was conducted after 7 days of adaptive feeding.

[0118] Statistical analysis: SPSS 25.0 software was used for data processing. The experimental data were expressed as mean ± standard deviation. P < 0.05 was considered statistically different, and P < 0.01 was considered statistically significant.

[0119] Experimental animals and groups: Healthy SD male rats were randomly divided into 7 groups (n=14), namely, sham operation group (Sham), model control group (Model), Crebanine group (Cre), the composition of Example 1 (Cre-VOA-L), the composition of Example 2 (Cre-VOA-M), the composition of Example 3 (Cre-VOA-H) and positive drug group (VP).

[0120] Sham group: given an equal amount of normal saline every day;

[0121] Model control group (Model): given an equal amount of normal saline every day;

[0122] Crebanine group (Cre): 50 mg kg / day -1 Administration by intragastric gavage;

[0123] Cre-VOA-L group: 75 mg kg / day -1 Administration by intragastric gavage;

[0124] Cre-VOA-M group: 75 mg kg / day -1 Administration by intragastric gavage;

[0125] Cre-VOA-H group: 150 mg kg / day -1 Administration by intragastric gavage;

[0126] Positive drug (VP): 40 mg kg per day -1 Administration by intragastric gavage;

[0127] All groups were given drugs by gavage 5 days in advance. After 5 days, the left anterior descending artery was ligated to establish the myocardial ischemia-reperfusion model in rats. After 30 minutes of ischemia, the suture was loosened and blood perfusion was restored for 120 minutes. After the perfusion, blood was collected from the abdominal aorta to detect the levels of CK, CK-MB, cTnl, LDH, MDA, SOD, TNF-α, IL-6 and IL-1β (n=6 in each group), part of the heart was used for TTC (n=5 in each group), HE (n=6 in each group), Tunel (n=6 in each group) detection, and the other part was used for Western blot detection (n=3 in each group).

[0128] Test results: Figure 6 The TTC results showed that the myocardial infarction area in the Model group reached 42.05%, and the infarction areas in each drug-treated group were: Cre group: 28.54%, Cre-VOA-L group: 26.61%, Cre-VOA-M group: 15.45%, Cre-VOA-H group: 8.50%, VP group: 7.91%. After treatment in each drug-treated group, the myocardial infarction area caused by myocardial ischemia-reperfusion was significantly reduced, and there was a significant difference compared with the model group (P>0.001).

[0129] Figure 7 It can be seen that the HE pathological sections of the Model group showed that the myocardial fibers were partially loose, broken and disorganized, and the cells were divided and dissolved. After drug treatment, although the myocardial tissue still showed loose and broken fibers, the degree of damage was greatly reduced. Figure 8-12It can be seen that compared with the Model group, each drug-treated group could significantly inhibit the expression of GRP-78, ATF-6, PERK, CHOP, Caspase-3, Caspase-9, Bcl-2, p-38 and p-p65 proteins in the myocardial tissue of the model rats and up-regulate the expression of Bax protein, down-regulate the expression levels of TNF-α, IL-6, IL-1β, MDA, LDH, CK, CK-MB, cTnl in the serum of the model rats and increase the activity of SOD (P>0.05). Compared with the Cre group, the Cre+VOA group could significantly enhance the regulatory effect of crebanine on the above indicators (P>0.05), improve the protective effect of crebanine on model rats, and the volatile oil of Acorus tatarinowii could inhibit myocardial ischemia-reperfusion-induced inflammation and oxidative stress injury by down-regulating the MAPK / NF-κB / TNF-α pathway, and inhibit the endoplasmic reticulum stress and cardiomyocyte apoptosis induced by myocardial ischemia-reperfusion through the GRP-78 / PERK / CHOP and GRP-78 / ATF-6 / CHOP pathways. It shows that the volatile oil of Acorus tatarinowii could reduce the damage of the heart and the apoptosis of myocardial cells by inhibiting the endoplasmic reticulum stress, oxidative stress and inflammatory response, and the volatile oil of Acorus tatarinowii could significantly enhance the therapeutic effect of crebanine on myocardial ischemia-reperfusion injury in rats.

[0130] Example 14 Determination of pharmacokinetic parameters of crebanine preparation after administration

[0131] Experimental animals and grouping: The abdomen of healthy male SD rats was depilated and cleaned and dried 24 hours before administration. The depilated rats were randomly divided into three groups (n=6): Cre-Banning gel group (Cre gel group), Cre-Banning liposome gel group without Acorus calamus volatile oil (Cre liposome gel group), and Example 8 preparation (Cre-VOA liposome gel group). The transdermal dose of the gel was 160 mg / kg, and the animals were fasted but not watered 12 hours before the experiment. At 1, 3, 5, 7, 9, 11, 13, 24, 28, 32, 36, and 48 hours after administration, 0.5 mL of blood was collected from the rats' orbits and placed in an anticoagulant blood collection tube. The tube was centrifuged at 6000 r / min for 10 minutes, and the upper plasma in the blood collection tube was aspirated for determination. The determination results are shown as follows: Fig.13 shown.

[0132] Fig.13It can be seen that the pharmacokinetic parameters of the Cre liposome gel group were improved compared with the Cre gel group, but there was no statistical significance. Compared with the Cre gel group alone, the pharmacokinetic parameters of the Cre-VOA liposome gel group increased by 1.49, 1.37, and 1.24 times, respectively, except for the shortening of Tmax, and Cmax, AUC0-t, and AUC0-∞, respectively, and Cmax and AUC0-t had significant differences (P < 0.01). Among them, the Cmax and AUC0-t of the Cre-VOA liposome gel group were 1.30 ± 0.17 mg / L and 20.21 ± 4.15 mg / L*h, respectively, which were significantly higher than those of the Cre liposome gel group (P < 0.01). The experimental results showed that compared with the Cre gel group, the transdermal permeability and transdermal absorption of crebanine increased to varying degrees after being prepared as liposome gel; at the same time, compared with the Cre liposome gel group, the transdermal permeation amount of the Cre-VOA liposome gel group was increased, the peak time was shortened, and the transdermal blood content of crebanine increased, verifying that Acorus calamus volatile oil can play a role in "promoting drug entry into the body".

[0133] Example 15 Determination of the content distribution of crebanine preparation in various tissues after administration

[0134] Experimental animals and grouping: The abdomen of healthy SD rats was depilated 24 hours before administration, and the rats were cleaned and dried. The depilated rats were divided into two groups (n=6): a group without Crebanine liposome gel containing Acorus calamus volatile oil (Cre liposome gel group) and a group with Crebanine liposome gel containing Acorus calamus volatile oil (Cre-VOA liposome gel group). The transdermal dose was 160 mg / kg. The animals were fasted but not allowed to drink water 12 hours before the experiment, and fasted but not allowed to drink water throughout the experiment. The rats were killed 3 hours, 7 hours, and 24 hours after administration, and the heart, liver, spleen, lung, kidney, and brain tissues were quickly taken to determine the Crebanine content. The results are shown in the table below. Fig.13 shown.

[0135] Experimental results: Fig.14 As shown in a, 24 h after administration of Cre liposome gel group, the order of distribution of Crebanine in each tissue was lung>liver>kidney>spleen>brain>heart. Fig.14 As shown in b, 24 hours after administration of Cre-VOA liposome gel group, the order of crebanine distribution in various tissues was lung>liver>spleen>kidney>heart>brain. The results showed that the crebanine content in various tissues of rats in Cre-VOA liposome gel group was higher than that in Cre liposome gel group, and the differences in crebanine content in heart, spleen and lung were significant (P<0.05), indicating that Acorus calamus volatile oil can increase the distribution of crebanine liposomes in myocardial tissue, indicating that Acorus calamus volatile oil has the effect of "leading medicine into the heart".

[0136] Finally, it should be noted that the above description is only a preferred embodiment of the present invention. Under the guidance of the present invention, those skilled in the art can make a variety of similar expressions without violating the purpose and claims of the present invention, and such changes all fall within the protection scope of the present invention.

Claims

1. A Chinese medicine composition, characterized in that: Including crebanine and Acorus calamus volatile oil.

2. The Chinese medicine composition according to claim 1, characterized in that: The composition comprises: 1-50 parts by weight of crebanine; such as 5-50 parts by weight, or 10-40 parts by weight, or 15-30 parts by weight, or 20-30 parts by weight; and 1-100 parts by weight of Acorus calamus volatile oil; such as 10-100 parts by weight, or 30-100 parts by weight, or 50-100 parts by weight, or 80-100 parts by weight.

3. A pharmaceutical preparation, characterized in that The composition is composed of the composition according to claim 1 or 2 and pharmaceutical excipients acceptable in the art.

4. The pharmaceutical preparation according to claim 3, characterized in that The pharmaceutical excipient is one or more of a gel matrix, an oil phase matrix, a moisturizer, and an emulsifier; The gel matrix is ​​selected from one or more of methyl cellulose, ethyl cellulose, hydroxyethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, sodium carboxymethyl cellulose, carbomer 940, gelatin, and sodium alginate, preferably, selected from carbomer 940; The oil phase matrix is ​​selected from one or more of stearic acid, paraffin, beeswax, higher alcohol, liquid paraffin, vaseline or vegetable oil; preferably, selected from one or more of soybean lecithin, cholesterol, lanolin and stearic acid; The moisturizing agent is selected from one or more of glycerol, 1,3-butylene glycol, propylene glycol, polyethylene glycol 300, polyethylene glycol 400, and sorbitol; preferably, selected from one or more of propylene glycol, glycerol, and polyethylene glycol 400; The emulsifier is preferably one or more of soaps, sodium lauryl sulfate, fatty acid esters of polyols (such as glyceryl monostearate), polysorbates, and polyethylene glycol ethers, preferably one or more of sodium deoxycholate, Tween 80, and Span 80.

5. The pharmaceutical preparation according to claim 3, characterized in that The pharmaceutical preparation is any one of tablets, capsules, pills, powders, granules, syrups, solutions, emulsions, injections, sprays, aerosols, and patches; the pharmaceutical preparation is administered through gastrointestinal and parenteral routes.

6. The pharmaceutical preparation according to claim 5, characterized in that The pharmaceutical preparation is a gel, comprising the composition according to claim 1 or 2, a gel matrix, and a moisturizer.

7. The pharmaceutical preparation according to claim 5, characterized in that The pharmaceutical preparation is a liposome gel, comprising the composition according to claim 1 or 2, a gel matrix, a moisturizer, an oil phase matrix, water, and an emulsifier.

8. The pharmaceutical preparation according to claim 7, characterized in that: The pharmaceutical preparation is prepared by the following method: adding crebanine, soybean lecithin, cholesterol, vitamin E and an emulsifier in a mass ratio of 1: (15-30): (1-2): (0.1-1.5): (5-15) into an organic solvent to dissolve to obtain an organic phase, dissolving the organic phase into a hydrating liquid to obtain a crebanine liposome suspension; mixing carbomer 940 and glycerol to obtain a blank gel, and then adding crebanine liposomes and calamus volatile oil to obtain a finished product.

9. Use of the composition according to claims 1-2 and the pharmaceutical preparation according to any one of claims 3-8 in the prevention and / or treatment of cardiovascular and cerebrovascular diseases.

10. The use according to claim 9, characterized in that: The mechanism of action is manifested in up-regulating the expression levels of Bax and Cx43, down-regulating the expression levels of ATF-6, GRP78, and CHOP proteins, reducing the release of MDA and LDH, and increasing the level of SOD.

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