CN110898093B - Application of fructus xanthil and fructus cannabis composition in preparation of lipid-lowering drugs - Google Patents
Application of fructus xanthil and fructus cannabis composition in preparation of lipid-lowering drugs Download PDFInfo
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- A61K2236/30—Extraction of the material
- A61K2236/33—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones
- A61K2236/331—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones using water, e.g. cold water, infusion, tea, steam distillation, decoction
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Abstract
The invention provides an application of a xanthium and ephedra composition in preparing a lipid-lowering drug. The xanthium and ephedra herb composition comprises 5-20 parts of rhizoma atractylodis and 5-20 parts of ephedra herb in parts by weight. When the composition of the rhizoma atractylodis and the ephedra is applied to preparing the lipid-lowering medicine, tests prove that the prepared lipid-lowering medicine has the outstanding effects of lowering triglyceride and cholesterol and promoting fat metabolism, and the range of indications of the rhizoma atractylodis and the ephedra is expanded.
Description
Technical Field
The invention relates to the technical field of lipid-lowering medicines, and particularly relates to an application of a xanthium and ephedra composition in preparation of a lipid-lowering medicine.
Background
Hyperlipidemia (Hyperlipidemia) is a disorder of lipid metabolism, is mainly characterized by elevated blood Triglycerides (TG), cholesterol (TC), low-density lipoproteins (LDL-C) and reduced high-density lipoproteins (HDL-C), and can cause a series of diseases such as arteriosclerosis, stroke, coronary heart disease, myocardial infarction, diabetes, renal failure, and the like. Recent studies have shown that hyperlipidemia is closely associated with the occurrence of neurodegenerative diseases such as Alzheimer's Disease (AD), Parkinson's Disease (PD). With the improvement of living standard and the change of life style of people, the incidence rate of hyperlipidemia is increasing year by year. Hyperlipemia has become an important public health problem at present, and has a remarkable rising trend in all age groups, thus arousing wide attention of scholars.
Lipid-lowering drugs clinically used at present mainly comprise statins, fibrates and the like, and although the curative effect is obvious, in recent years, researches show that the statins can possibly cause adverse reactions such as liver injury, rhabdomyolysis, new tumors, diabetes and the like. Therefore, the development of new lipid-lowering drugs is urgently required.
Disclosure of Invention
The invention mainly aims to provide application of a xanthium and ephedra composition in preparation of a lipid-lowering drug, so as to solve the problem of great side effect of a clinical lipid-lowering drug in the prior art.
In order to achieve the purpose, according to one aspect of the invention, the invention provides a use of a xanthium and ephedra composition in preparation of a lipid-lowering drug, wherein the xanthium and ephedra composition comprises 5-20 parts of rhizoma atractylodis and 5-20 parts of ephedra according to parts by weight.
Further, the xanthium and ephedra composition comprises 10-15 parts of rhizoma atractylodis and 8-12 parts of ephedra according to parts by weight.
Furthermore, the lipid-lowering medicine is an oral preparation.
Further, the oral preparation is decoction, oral liquid, powder, capsule or tablet.
Furthermore, each 100mL of decoction or oral liquid contains 25-100 μ L of fructus xanthil water extract, the fructus xanthil water extract is 200mL of water extract obtained by water extraction of fructus xanthil composition, and the fructus xanthil composition comprises 5-20 g of rhizoma atractylodis and 5-20 g of herba ephedrae.
Furthermore, each 100mL of decoction or oral liquid contains 100 μ L of fructus Xanthii water extract.
Furthermore, the lipid-lowering medicine also comprises a pharmaceutically acceptable carrier.
Further, the lipid-lowering medicine is a xanthium and ephedra pill.
Further, the application comprises a preparation method of the lipid-lowering medicine, and the preparation method comprises the following steps: carrying out water extraction or alcohol extraction on the xanthium and hemp composition to obtain an extracting solution; the extract is used as a raw material to prepare the lipid-lowering medicine.
Further, the step of preparing the lipid-lowering medicine by using the extract as a raw material comprises the following steps: concentrating the extracting solution to obtain a concentrate; mixing the concentrate with pharmaceutically acceptable carrier to obtain the lipid-lowering medicine.
When the technical scheme of the invention is applied to the preparation of the lipid-lowering medicine, the experiment proves that the prepared lipid-lowering medicine has the outstanding effects of lowering triglyceride, lowering cholesterol and promoting fat metabolism, and the scope of indications of the rhizoma atractylodis and the ephedra herb is expanded.
Drawings
The accompanying drawings, which are incorporated in and constitute a part of this application, illustrate embodiments of the invention and, together with the description, serve to explain the invention and not to limit the invention. In the drawings:
FIG. 1 shows a photomicrograph of the blood fat analysis area (dotted box area) after staining zebra fish oil red O according to example 2 of the present invention;
FIG. 2 shows a typical pattern of vascular staining of the tail of zebra fish after treatment in each experimental group of example 2;
figure 3 shows the blood fat reduction results of the xanthane square treated zebrafish of example 2 at different concentrations, wherein p <0.01 and p <0.001, compared to the model control group;
FIG. 4 shows a typical pattern of fluorescent staining of zebra fish fat after treatment in each experimental group of example 4;
fig. 5 shows the blood fat reduction curves of the xanthane squares treated zebrafish of example 4 at different concentrations, wherein p is <0.01 compared to the model control group.
Detailed Description
It should be noted that the embodiments and features of the embodiments in the present application may be combined with each other without conflict. The present invention will be described in detail below with reference to the embodiments with reference to the attached drawings.
As analyzed in the background art of the present application, statins, fibrates and the like in the prior art have significant efficacy, but in recent years, studies find that statins may cause adverse reactions such as liver injury, rhabdomyolysis, new tumors, diabetes and the like, and in order to solve the problem, the present application provides an application of a traditional Chinese medicine composition in preparation of lipid-lowering drugs.
The traditional Chinese medicine research shows that the rhizoma atractylodis and the ephedra are combined with lung and spleen to be concocted, and the platycodon grandiflorum and the radish seed are used for diffusing qi and transforming cold-dampness, so that the clinical indication is that phlegm-dampness is in the interior and heat-condition is not obvious, namely, cold-dampness is in the interior. From the six meridians, the indication of the cang ma wan is phlegm-damp accumulation in the category of taiyin disease. From the zang-fu organs, it pertains to the lung and spleen due to phlegm-dampness and cold-dampness. Clinically, cold phlegm-dampness and internal pathogens are manifested by excessive and sticky phlegm, fullness and stuffiness in the chest and epigastrium, poor appetite, weakness of limbs, abdominal distension, sticky stool, thirst-free and thick and greasy tongue coating, which are typical indications of the cang-ma pills. At present, the rhizoma atractylodis and the ephedra are clinically applied to eliminating phlegm, relieving cough and removing dampness.
The applicant discovers that the rhizoma atractylodis and the ephedra herb have the effect of reducing blood fat when researching the pharmacological effect of the rhizoma atractylodis and the ephedra herb, and therefore, in a typical embodiment of the application, the application provides the application of the fructus xanthil and ephedra herb composition in preparing the lipid-reducing medicine, wherein the fructus xanthil and ephedra herb composition comprises 5-20 parts of the rhizoma atractylodis and 5-20 parts of the ephedra herb in parts by weight.
When the composition of the rhizoma atractylodis and the ephedra is applied to preparing the lipid-lowering medicine, tests prove that the prepared lipid-lowering medicine has the outstanding effects of lowering triglyceride and cholesterol and promoting fat metabolism, and the scope of indications of the rhizoma atractylodis and the ephedra is expanded.
In order to further exert the synergistic effect of the rhizoma atractylodis and the ephedra herb, the rhizoma atractylodis and ephedra herb composition comprises 10-15 parts of the rhizoma atractylodis and 8-12 parts of the ephedra herb by weight.
The lipid-lowering medicine can be prepared into the dosage form of the conventional lipid-lowering medicine at present, and preferably, the lipid-lowering medicine is an oral preparation. So as to promote the absorption of the effective components and exert the drug effect.
The oral preparation can be the conventional dosage form of the traditional Chinese medicine at present, and preferably, the oral preparation is decoction, oral liquid, powder, capsule or tablet.
In order to further improve the blood fat reducing effect of the rhizoma atractylodis composition and not generate any obvious toxic or side effect, preferably, each 100mL of the decoction or the oral liquid contains 25-100 μ L of the fructus xanthil water extract, the fructus xanthil water extract is 200mL of water extract obtained by water extraction of the fructus xanthil composition, and the fructus xanthil composition comprises 5-20 g of rhizoma atractylodis and 5-20 g of ephedra. Preferably, each 100mL of decoction or oral liquid contains 100 μ L of fructus Xanthii water extract.
For convenience of taking, the lipid-lowering medicine can be prepared into a Chinese patent medicine, and preferably, the lipid-lowering medicine also comprises a pharmaceutically acceptable carrier. Such as sweeteners, dispersants, and the like.
When the xanthium and ephedra composition is used for preparing the lipid-lowering medicine, the xanthium and ephedra pills clinically used at present can be directly applied to the lipid-lowering medicine.
Of course, in addition to the fructus xanthil pills, the fructus xanthil composition can be prepared into other forms for preparing lipid-lowering medicines, and preferably, the use comprises a preparation method of the lipid-lowering medicines, wherein the preparation method comprises the following steps: carrying out water extraction or alcohol extraction on the xanthium and hemp composition to obtain an extracting solution; the extract is used as a raw material to prepare the lipid-lowering medicine. The effective components in the fructus xanthil composition are extracted by water extraction or alcohol extraction to obtain an extracting solution, and the obtained extracting solution can be directly taken as a lipid-lowering medicament or can be processed according to the conventional operation of a traditional Chinese medicine preparation to form the lipid-lowering medicament.
In order to facilitate storage and circulation of the lipid-lowering drug, preferably, the step of preparing the lipid-lowering drug by using the extract as a raw material comprises: concentrating the extracting solution to obtain a concentrate; mixing the concentrate with pharmaceutically acceptable carrier to obtain the lipid-lowering medicine.
The advantageous effects of the present application will be further described below with reference to examples and comparative examples.
Laboratory animal
1. Melanin allele mutant translucent Albino strain zebrafish, in a natural pairwise mating breeding mode. The age was 7 days after fertilization, total 330, and 30 in each experimental group (for Maximum Tolerated Concentration (MTC) determination of herbal decoction).
2. Melanin allele mutant translucent Albino strain zebrafish, in a natural pairwise mating breeding mode. The age is 5 days after fertilization, 360 tails in total, and 30 tails in each experimental group (used for evaluating the effect of the traditional Chinese medicine decoction on reducing blood fat).
3. Melanin allele mutant translucent Albino strain zebrafish, in a natural pairwise mating breeding mode. The age is 7 days after fertilization, 360 tails in total, and 30 tails in each experimental group (used for evaluating the blood fat reducing effect (biochemical index) of the traditional Chinese medicine decoction).
4. Melanin allele mutant translucent Albino strain zebrafish, in a natural pairwise mating breeding mode. The age is 5 days after fertilization, 360 tails in total, and 30 tails in each experimental group (used for the influence of traditional Chinese medicine decoction on fat catabolism).
The zebra fish are all raised in water for fish culture at 28 ℃ (water quality: 200mg of instant sea salt is added into per 1L of reverse osmosis water, the conductivity is 480-510 muS/cm, the pH is 6.9-7.2, and the hardness is 53.7-71.6 mg/L CaCO3) The license number for experimental animals is as follows: SYXK (Zhe) 2012-0171. The feeding management meets the requirements of international AAALAC certification.
Experimental sample
Raw formula of cang ma: wherein the preparation and packaging are provided by Beijing Chinese medical hospital affiliated to the university of capital medical science, and the extracting solution is used for experiments.
Adding fructus Xanthii and fructus Cannabis, wherein rhizoma Atractylodis 12g, herba Ephedrae 10g, Raphani semen 30g, radix Platycodi 12g, decocting with water to obtain brown extractive solution, 200 mL/bag × 14, collecting and registering in 2019 in 03 month and 19 days, storing at 4 deg.C, and preparing and packaging the extractive solution provided by Beijing Chinese medicine hospital affiliated to capital medical university.
Simvastatin, a white powder with lot number UZDBF-2K, purchased from echiei (shanghai) chemical industry development limited. The stock solution is prepared into 4mg/mL mother solution by 100% dimethyl sulfoxide (DMSO) for standby at the time of clinical application, and is diluted by 100% DMSO as required, and the DMSO concentration in the final working solution is 0.1%.
Resveratrol, white granules, lot number 36794, was purchased from Shanghai Allantin Biotechnology Ltd. The stock solution was prepared at the time of use with 100% DMSO at 14.4mg/mL, final DMSO concentration was 0.1%.
Instruments and reagents
Fluorescence stereomicroscope (Nikon AZ 100, Japan); dissecting microscopes (SZX7, OLYMPUS, Japan); a camera (VertA1) connected to the microscope; precision electronic balances (CP214, OHAUS, America); multifunctional microplate readers (Mithras LB940, Berthold Technologies); 6-well plates (Nest Biotech, China); 96-well microplate (costar, USA); methylcellulose (Sigma, USA); egg yolk powder (Beijing Kaiyuan, lot number 20170626); oil Red O (Oil Red O, lot SLBP5248V, Sigma); dimethyl sulfoxide (DMSO, Sigma, France); nile red (CAS number 7385-67-3, Sigma); triglyceride (TG) test box (lot No. 20190320, buika beijing); total cholesterol (T-CHO) test kit (batch No. 20190321, Nanjing Keoho).
Example 1: maximum Tolerated Concentration (MTC) assay
1. Concentration group
The dosage of the original formula of fructus Xanthii and fructus Xanthii flavoring refers to the concentration of diluted stock solution with distilled water, for example, 25 μ L/mL of the original formula of fructus Xanthii refers to 22g/200mL of extract containing 25 μ L per 100mL of liquid.
2. Experimental methods
And randomly selecting 330 black pigment allele mutant type semitransparent Albino strain zebra fishes 7 days (7dpf) after fertilization in a six-well plate, and treating 30 zebra fishes in each well (experimental group). The fructus Xanthii original prescription is respectively dissolved in water and is given with the concentrations of 25 muL/mL, 50 muL/mL, 100 muL/mL, 200 muL/mL and 400 muL/mL, and fructus Xanthii is flavored with the concentrations of 6.25 muL/mL, 12.5 muL/mL, 25 muL/mL, 50 muL/mL and 100 muL/mL, and a normal control group is set, and the volume of each hole (experimental group) is 3 mL. After each experimental group is treated for 48 hours, the phenotype and death condition of the zebra fish are observed and recorded, and the MTC of the two traditional Chinese medicine decoctions for the zebra fish is respectively determined.
3. Results of the experiment
The Cannabis sativa L original prescription does not produce any obvious toxic or side effect on the zebra fish and does not cause the death of the zebra fish when the concentration is 25-100 mu L/mL, induces 6.7% (2/30 tails) of the zebra fish to turn on one side when the concentration is 200 mu L/mL, and induces 40% (12/30 tails) of the zebra fish to die when the concentration is 400 mu L/mL. Therefore, the ratio of the cang-ma original prescription to the zebra fish MTC is determined to be 100 mu L/mL.
When the fructus xanthil and fructus cannabis are flavored at the concentration of 6.25-25 mu L/mL, no obvious toxic or side effect is generated on the zebra fish, the zebra fish is not killed, and 100% (30/30 tails) of the zebra fish are killed when the concentration of 50 mu L/mL and 100 mu L/mL are both generated. Therefore, the content of the xanthium sibiricum added with the flavor to the zebra fish MTC is determined to be 25 muL/mL. See table 1 for details.
TABLE 1 "concentration-lethality" results after treatment of Chinese medicinal decoction (n ═ 30)
Example 2: evaluation of blood lipid-lowering Effect
1. Concentration group
2. Basis of concentration determination
According to concentration groping experiments, the MTC of the xanthium sibiricum prescription to the zebra fish is 100 mu L/mL. The experimental concentration for evaluating the blood fat reducing effect is set as follows: 6.25 μ L/mL (1/16MTC), 12.5 μ L/mL (1/8MTC), 25 μ L/mL (1/4MTC), 50 μ L/mL (1/2MTC), and 100 μ L/ML (MTC).
According to concentration groping experiments, the MTC of the xanthium sibiricum flavored to the zebra fish is 25 muL/mL. The experimental concentration for evaluating the blood fat reducing effect is set as follows: 1.5625. mu.L/mL (1/16MTC), 3.125. mu.L/mL (1/8MTC), 6.25. mu.L/mL (1/4MTC), 12.5. mu.L/mL (1/2MTC) and 50. mu.L/ML (MTC).
3. Making of models
Feeding 5dpf melanin allele mutant type translucent Albino zebra fish by using egg yolk powder to establish a zebra fish hyperlipidemia model.
4. Experimental methods
Randomly selecting 5dpf melanin allele mutant type translucent Albino strain zebra fish in a beaker, feeding the zebra fish with yolk powder to establish a hyperlipidemia model, randomly selecting 360 model zebra fish in a six-hole plate after the model is established, and treating 30 zebra fish in each hole (experimental group). The xanthium and ephedra original prescription is respectively dissolved in water and is given with the concentration of 6.25 mu L/mL, 12.5 mu L/mL, 25 mu L/mL, 50 mu L/mL and 100 mu L/mL, the xanthium and ephedra flavor is given with the concentration of 1.5625 mu L/mL, 3.125 mu L/mL, 6.25 mu L/mL, 12.5 mu L/mL and 50 mu L/mL, the positive control drug simvastatin is given with the concentration of 40ng/mL, and meanwhile, a model control group is set, and the volume of each hole (experimental group) is 3 mL. After each experimental group is treated for 48 hours, dyeing is carried out by using oil red O, 10 zebra fish in each group are randomly selected after dyeing and photographed under an anatomical microscope, Image-Pro Plus 6.0 advanced Image processing software is used for carrying out Image analysis and collecting data, specifically shown in fig. 1 and fig. 2, the sum (S) of the blood lipid optical density of the blood vessel at the tail part of the zebra fish is analyzed and counted, and the blood lipid reducing effect of the two traditional Chinese medicine decoctions is respectively evaluated according to the statistical result of the sum of the optical density. The statistical treatment result is expressed by mean + -SE, and the calculation formula of the hypolipidemic effect is as follows:
statistical analysis using analysis of variance and Dunnett's T-test indicated significant differences with p < 0.05.
5. Results of the experiment
The positive control medicine simvastatin 40ng/mL concentration group zebra fish tail blood vessel blood lipid optical density sum (6962 pixels) is compared with the model control group (9966 pixels) to p <0.01, the effect of reducing the blood lipid of the zebra fish is 30%, and the result shows that 40ng/mL simvastatin has the effect of reducing the blood lipid.
The sum of the blood lipid optical density of the tail blood vessels of the zebra fish in the concentration groups of 6.25 muL/mL, 12.5 muL/mL, 25 muL/mL, 50 muL/mL and 100 muL/mL of the xanthina original prescription is 10442 pixels, 8104 pixels, 6649 pixels, 5951 pixels and 6604 pixels respectively, the blood lipid reduction effect is-5%, 19%, 33%, 40% and 34% respectively, compared with a model control group, the concentration groups of 6.25 muL/mL and 12.5 muL/mL p are both >0.05, the concentration groups of 25 muL/mL, 50 muL/mL and 100 muL/mL p are <0.01& p <0.001& p <0.001, and the result shows that the xanthina original prescription has the blood lipid reduction effect on the zebra fish hyperlipidemia induced by the yolk powder under the concentration conditions of 25 muL/mL, 50 muL/mL and 100 muL/mL.
The sum of the optical densities of blood lipids of tail blood vessels of the zebra fish in concentration groups of 1.5625 muL/mL, 3.125 muL/mL, 6.25 muL/mL, 12.5 muL/mL and 25 muL/mL of the siberian fish is 10191 pixels, 10433 pixels, 10645 pixels, 10512 pixels and 10534 pixels respectively, and the blood lipid reduction effects are-2%, -5%, -7%, -5% and-6% respectively, compared with a model control group, the p of each concentration group is greater than 0.05, which indicates that the blood lipid reduction effect of the siberian fish induced by the yolk powder under the experimental concentration condition is not obvious. See table 2 for details.
TABLE 2 blood lipid lowering analysis of treated zebrafish (n 10)
Note: p <0.01, p <0.001, compared to model control group
Example 3 evaluation of hypolipidemic Effect (Biochemical index)
1. Concentration group
2. Basis of concentration determination
According to concentration grope experiments, the MTC of the xanthium and ephedra original prescription to the zebra fish is 100 and 25 mu L/mL. The experimental concentration of the evaluation (biochemical index) of the blood fat reducing effect is set as follows: 6.25 μ L/mL (1/16MTC), 12.5 μ L/mL (1/8MTC), 25 μ L/mL (1/4MTC), 50 μ L/mL (1/2MTC), and 100 μ L/ML (MTC).
According to concentration groping experiments, the MTC of the xanthium sibiricum flavored to the zebra fish is 25 muL/mL. The experimental concentration of the evaluation (biochemical index) of the blood fat reducing effect is set as follows: 1.5625. mu.L/mL (1/16MTC), 3.125. mu.L/mL (1/8MTC), 6.25. mu.L/mL (1/4MTC), 12.5. mu.L/mL (1/2MTC) and 50. mu.L/ML (MTC).
3. Making of models
Feeding 7dpf melanin allele mutant type translucent Albino zebra fish by using egg yolk powder to establish a zebra fish hyperlipidemia model.
4. Experimental methods
Randomly selecting 7dpf melanin allele mutant type semitransparent Albino strain zebra fish in a beaker, feeding the zebra fish with yolk powder to establish a hyperlipidemia model, randomly selecting 360 model zebra fish in a six-hole plate after the model is established, and treating 30 zebra fish in each hole (experimental group). The xanthium and ephedra original prescription is respectively dissolved in water and is given with the concentration of 6.25 mu L/mL, 12.5 mu L/mL, 25 mu L/mL, 50 mu L/mL and 100 mu L/mL, the xanthium and ephedra flavor is given with the concentration of 1.5625 mu L/mL, 3.125 mu L/mL, 6.25 mu L/mL, 12.5 mu L/mL and 50 mu L/mL, the positive control drug simvastatin is given with the concentration of 40ng/mL, and meanwhile, a model control group is set, and the volume of each hole (experimental group) is 3 mL. After each experimental group is treated for 48 hours, Triglyceride (TG) and cholesterol (TC) kits are used for detecting the content of TG and TC in the zebra fish body, and the influence of the two traditional Chinese medicine decoctions on TG and TC of the zebra fish model with hyperlipidemia is respectively evaluated according to the statistical result of the content of TG and TC. Statistical treatment results are expressed as mean ± SE, and statistical analysis using analysis of variance and Dunnett's T-test, p <0.05 indicated significant differences.
5. Results of the experiment
TG：
The positive control medicine simvastatin with the concentration of 40ng/mL for the whole content of TG in the zebra fish (0.144mmol/gprot) is compared with the p of the model control group (0.226mmol/gprot) to be less than 0.001, which shows that 40ng/mL simvastatin can reduce the whole triglyceride level of the zebra fish.
The integral TG content of the zebra fish in the concentration groups of 6.25 muL/mL, 12.5 muL/mL, 25 muL/mL, 50 muL/mL and 100 muL/mL of the xanthina is respectively 0.221mmol/gprot, 0.230mmol/gprot, 0.150mmol/gprot, 0.142mmol/gprot and 0.194mmol/gprot, and compared with a model control group, the integral TG content of the zebra fish in the concentration groups of 6.25 muL/mL and 12.5 muL/mL is more than 0.05, and the integral TG content of the zebra fish in the concentration groups of 25 muL/mL, 50 muL/mL and 100 muL/mL is less than 0.001, which indicates that the xanthina can reduce the integral triglyceride level of the zebra fish under the concentration conditions of 25 muL/mL, 50 muL/mL and 100 muL/mL.
The sum of the optical densities of blood lipids of tail blood vessels of zebra fish in the concentration groups of 1.5625 muL/mL, 3.125 muL/mL, 6.25 muL/mL, 12.5 muL/mL and 25 muL/mL is respectively 0.245mmol/gprot, 0.219mmol/gprot, 0.246mmol/gprot, 0.248mmol/gprot and 0.239mmol/gprot, and compared with a model control group, the p of each concentration group is greater than 0.05, which indicates that the effect of the xanthium sibiricum taste on reducing the integral triglyceride level of the zebra fish under the condition of the experimental concentration is not obvious.
TC：
Comparing the overall TC content (0.155mmol/gprot) of the zebra fish with the positive control medicament simvastatin of 40ng/mL concentration with the model control group (0.285mmol/gprot), p is less than 0.001, which shows that 40ng/mL simvastatin can obviously reduce the overall cholesterol level of the zebra fish.
The overall TC content of the zebra fish in the concentration groups of 6.25 muL/mL, 12.5 muL/mL, 25 muL/mL, 50 muL/mL and 100 muL/mL of the xanthina is respectively 0.262mmol/gprot, 0.206mmol/gprot, 0.196mmol/gprot, 0.212mmol/gprot and 0.228mmol/gprot, and compared with a model control group, the concentration groups of 6.25 muL/mL p >0.05, 12.5 muL/mL, 25 muL/mL, 50 muL/mL and 100 muL/mL p <0.01& p <0.05 suggest that the xanthina can reduce the overall cholesterol level of the zebra fish under the concentration conditions of 12.5 muL/mL, 25 muL/mL, 50 muL/mL and 100 muL/mL.
The overall TC content of the zebra fish in the concentration groups of 1.5625 muL/mL, 3.125 muL/mL, 6.25 muL/mL, 12.5 muL/mL and 25 muL/mL is respectively 0.297mmol/gprot, 0.291mmol/gprot, 0.299mmol/gprot, 0.271mmol/gprot and 0.303mmol/gprot, compared with the model control group, the p of each concentration group is more than 0.05, which indicates that the effect of the sibra fish flavour on reducing the overall cholesterol level of the zebra fish under the experimental concentration condition is not obvious.
See table 3 for details.
TABLE 3 Zebra fish TG and TC analysis results after treatment of Chinese medicinal decoction (n ═ 5)
Note: p <0.05, p <0.01, p <0.001, compared to model controls
Example 4: effects on fat catabolism
1. Concentration group
2. Basis of concentration determination
According to concentration groping experiments, the MTC of the xanthium sibiricum prescription to the zebra fish is 100 mu L/mL. Effect on fat catabolism experimental concentrations were set as: 6.25 μ L/mL (1/16MTC), 12.5 μ L/mL (1/8MTC), 25 μ L/mL (1/4MTC), 50 μ L/mL (1/2MTC), and 100 μ L/ML (MTC).
According to concentration groping experiments, the MTC of the xanthium sibiricum flavored to the zebra fish is 25 muL/mL. Effect on fat catabolism experimental concentrations were set as: 1.5625. mu.L/mL (1/16MTC), 3.125. mu.L/mL (1/8MTC), 6.25. mu.L/mL (1/4MTC), 12.5. mu.L/mL (1/2MTC) and 50. mu.L/ML (MTC).
3. Experimental methods
And randomly selecting 360 black pigment allele mutant type semitransparent Albino strain zebra fishes 5 days (5dpf) after fertilization in a six-well plate, and treating 30 zebra fishes in each well (experimental group). The xanthium sibiricum original prescription is respectively dissolved in water and is given with the concentration of 6.25 mu L/mL, 12.5 mu L/mL, 25 mu L/mL, 50 mu L/mL and 100 mu L/mL, the xanthium sibiricum is added with 1.5625 mu L/mL, 3.125 mu L/mL, 6.25 mu L/mL, 12.5 mu L/mL and 50 mu L/mL, the concentration of the positive control drug resveratrol is 14.4 mu g/mL, a normal control group is simultaneously arranged, and the volume of each hole (experimental group) is 3 mL. After each experimental group is treated for 24 hours, dissolving the materials in water, and giving nile red for fat staining overnight; after dyeing is finished, 10 zebra fish are randomly taken from each experimental group, photographed under a fluorescence microscope and stored, the dyeing intensity (S) of the zebra fish yolk sac fat is statistically analyzed by NIS-Elements D3.10 advanced image processing software, and the influence of the two traditional Chinese medicine decoctions on the normal zebra fish fat catabolism is respectively evaluated according to the statistical analysis result of the dyeing intensity of the yolk sac fat. Statistical treatment results are expressed as mean ± SE. The fat catabolism calculation formula is as follows:
statistical analysis using analysis of variance and Dunnett's T-test indicated significant differences with p < 0.05.
4. Results of the experiment
The staining intensity of the fat of the yolk sac of the zebra fish in the normal control group is 10034181 pixels, the staining intensity of the fat of the yolk sac of the zebra fish in the positive control drug resveratrol group with the concentration of 14.4 mu g/mL is 5017235 pixels, compared with the normal control group, the staining intensity of the fat of the yolk sac of the zebra fish is less than 0.001, the promotion effect of the staining intensity of the fat of the zebra fish on the catabolism of the zebra fish is 50%, and the promotion effect of the resveratrol on the fat catabolism of the zebra fish is prompted.
The fat staining intensity of the yolk sac of the zebra fish in the concentration groups of 6.25 muL/mL, 12.5 muL/mL, 25 muL/mL, 50 muL/mL and 100 muL/mL of the xanthina original prescription is 8781500 pixels, 7988119 pixels, 8201628 pixels, 7928936 pixels and 5997518 pixels respectively, the fat catabolism promotion effect is 12%, 20%, 18%, 21% and 40% respectively, and compared with a normal control group, the concentration groups of 6.25 muL/mL, 12.5 muL/mL, 25 muL/mL and 50 muL/mL p are all greater than 0.05 and the concentration group of 100 muL/mL p is less than 0.01, which indicates that the xanthina original prescription has promotion effect on the fat catabolism of the zebra fish under the concentration condition of 100 muL/mL.
The dyeing intensity of the fat of the yolk sac of the zebra fish in the concentration groups of 1.5625 muL/mL, 3.125 muL/mL, 6.25 muL/mL, 12.5 muL/mL and 25 muL/mL is 9777147 pixels, 9206031 pixels, 7895828 pixels, 8348262 pixels and 8971160 pixels respectively, the fat catabolism promotion effect is 3%, 8%, 21%, 17% and 11% respectively, and the p of each concentration group is more than 0.05 compared with that of a normal control group, which indicates that the siberian fish has no obvious fat catabolism promotion effect under the concentration condition of the experiment. See table 4, fig. 4 and fig. 5 for details.
TABLE 4 fat catabolism results of treated zebrafish (n ═ 10)
Note: p <0.01, p <0.001, compared to model control group
Example 5
Raw formula of cang ma: wherein 20g of rhizoma atractylodis and 5g of ephedra are decocted with water to obtain 200mL of brown extracting solution.
Example 6
The difference from the example 5 is that the formula of the xanthium and ephedra is as follows: wherein, the atractylodes rhizome is 5g, the ephedra herb is 20g, and 200mL of brown extracting solution is obtained by water decoction.
Example 7
The difference from the example 5 is that the formula of the xanthium and ephedra is as follows: wherein 10g of rhizoma atractylodis and 15g of ephedra are decocted with water to obtain 200mL of brown extracting solution.
Example 8
The difference from the example 5 is that the formula of the xanthium and ephedra is as follows: wherein, the atractylodes rhizome 15g and the ephedra 8g are decocted with water to obtain 200mL of brown extracting solution.
Comparative example 1
The difference from the example 5 is that the formula of the xanthium and ephedra is as follows: wherein, 25g of rhizoma atractylodis and 3g of ephedra are decocted with water to obtain 200mL of brown extracting solution.
Comparative example 2
The difference from the example 5 is that the formula of the xanthium and ephedra is as follows: wherein, the atractylodes rhizome is 5g, the ephedra herb is 25g, and 200mL of brown extracting solution is obtained by water decoction.
The test methods of examples 2 to 4 were repeated at a concentration of 50. mu.L/mL of the Cannabis origin, and the hypolipidemic effects of the Cannabis origins of examples 5-8 and comparative examples 1-2 were examined, and the examination results are shown in Table 5.
TABLE 5
According to the tests, the Cannabis sativa L.prescription provided by the embodiment of the application has the effect of reducing blood fat of zebra fish induced by yolk powder, can obviously reduce the whole triglyceride and cholesterol level of the zebra fish, and has a promotion effect on fat catabolism of the zebra fish. The fructus xanthil and fructus cannabis seasoning has no effect of reducing blood fat of zebra fish induced by yolk powder, and can not reduce the whole triglyceride and cholesterol level of the zebra fish.
From the above description, it can be seen that the above-described embodiments of the present invention achieve the following technical effects:
when the composition of the rhizoma atractylodis and the ephedra is applied to preparing the lipid-lowering medicine, tests prove that the prepared lipid-lowering medicine has the outstanding effects of lowering triglyceride and cholesterol and promoting fat metabolism, and the scope of indications of the rhizoma atractylodis and the ephedra is expanded.
The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention, and various modifications and changes may be made by those skilled in the art. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.
Claims (4)
1. The application of the xanthium and ephedra composition in preparation of a lipid-lowering medicine is characterized in that the xanthium and ephedra composition is composed of 10-15 parts by weight of rhizoma atractylodis and 8-12 parts by weight of ephedra herb, the lipid-lowering medicine is an oral preparation, the oral preparation is a decoction or an oral liquid, each 100mL of the decoction or the oral liquid contains 25-100 μ L of the xanthium and ephedra aqueous extract, and the xanthium and ephedra aqueous extract is 200mL of the aqueous extract obtained by extracting the xanthium and ephedra composition with water.
2. The use as claimed in claim 1, wherein the decoction or the oral liquid contains 100 μ L of the aqueous extract of Siberian cocklebur fruit per 100 mL.
3. The use of claim 1, wherein said lipid-lowering agent further comprises a pharmaceutically acceptable carrier.
4. The use according to claim 1, wherein said use comprises a process for the preparation of said lipid lowering drug, said process comprising:
carrying out water extraction on the xanthium and hemp composition to obtain an extracting solution;
the lipid-lowering medicine is prepared by adopting the extracting solution as a raw material, wherein the step of preparing the lipid-lowering medicine by adopting the extracting solution as a raw material comprises the following steps:
concentrating the extracting solution to obtain a concentrate;
mixing the concentrate with a pharmaceutically acceptable carrier to obtain the lipid-lowering medicine.
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| CN110898093A (en) | 2020-03-24 |
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