CN113230316A - Application of peony and licorice decoction preparation in preventing and treating tumor diseases - Google Patents
Application of peony and licorice decoction preparation in preventing and treating tumor diseases Download PDFInfo
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- 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 belongs to the field of traditional Chinese medicines, and relates to a new application of a peony and licorice decoction preparation in preventing and treating tumor diseases. The invention provides a new application of the peony and licorice decoction as a unique or main effective component for resisting tumors, which can inhibit the growth and migration of cancer cells, induce the apoptosis of the cancer cells, has an anti-tumor effect, expands the brand-new medical effect of the peony and licorice decoction, widens the clinical application range of the peony and licorice decoction, and is beneficial to the development of new drugs in the field of traditional Chinese medicines.
Description
Technical Field
The invention relates to a new application of a traditional Chinese medicine composition, in particular to a new application of a peony and licorice decoction preparation as a sole or main effective component in preventing and treating tumor diseases.
Background
In recent years, cancer gradually invades into our daily lives, the incidence rate of the cancer presents a remarkable rising trend and has extremely high mortality rate, and the cancer becomes one of serious diseases endangering human life and health. With the progress of scientific research, people find that a plurality of traditional Chinese medicine compositions have potential good anti-tumor effects.
China is a hometown of natural Chinese herbal medicines and has tens of thousands of medicinal materials and thousands of Chinese patent medicines. The famous prescription preparation explicitly recorded in the ancient classical medical book contains medical experience accumulated by Chinese people in life practice for more than five thousand years, is the essence of Chinese traditional medicine culture, and has very important clinical research value.
The peony and licorice decoction is one of ancient and famous classical prescriptions in China and is originated from the treatise on febrile disease by the holy Zhang Zhongjing in medicine. Although the prescription only contains two medicines of white paeony root and liquorice (roasted), the effective components of the prescription are very complex, and the prescription has obvious effects of invigorating and nourishing yin, softening liver and relaxing muscles and tendons, relieving spasm and relieving pain and the like. In the current clinical medication process, researches on the peony and licorice decoction preparation mainly focus on analgesic and anti-inflammatory effects, but researches on the overall anti-tumor effect of the decoction are not reported.
Disclosure of Invention
Based on the current application situation of the peony and licorice decoction, the inventor further researches the peony and licorice decoction to find the antitumor effect of the peony and licorice decoction. Therefore, the invention provides a new application of the peony and licorice decoction preparation in preventing and treating tumor diseases.
The peony and licorice decoction is prepared by decocting and extracting radix paeoniae alba and honey-fried licorice root in water according to the weight ratio of 1: 1; or filtering the extractive solution, collecting filtrate, and lyophilizing to obtain lyophilized powder.
The raw material medicines of the white paeony root and the honey-fried licorice root related by the invention both accord with the record of Chinese pharmacopoeia (2020 edition). In the formula, the white paeony root is bitter and sour in property, enters liver and spleen channels, nourishes yin, benefits blood, relieves spasm and relieves pain, and is a monarch drug; prepared licorice root, radix Glycyrrhizae Praeparata, warm in nature and sweet in taste, enters heart, lung, stomach and spleen meridians and can tonify the middle-jiao and Qi.
The tumor includes human esophagus cancer, colon cancer, cervical cancer and gastric cancer.
To fully verify the antitumor effect of the peony and licorice decoction, the following studies were performed: firstly, the evaluation of the in vitro anti-tumor activity of the peony and licorice decoction is carried out, and the peony and licorice decoction is preliminarily proved to have a certain anti-tumor effect.
Secondly, cell monoclonal formation and cell scratch experiments are carried out, and the peony and licorice decoction is clear to obviously inhibit the growth and migration of cancer cells.
Thirdly, cell apoptosis experiments are carried out to prove that the peony and licorice decoction can induce cancer cells to generate concentration-dependent apoptosis, thereby playing a direct anti-tumor role.
Based on the detailed research, the invention provides a new application of the peony and licorice decoction as a unique or main effective component in preventing and treating tumor diseases, and the composition of the peony and licorice decoction comprises pharmaceutically acceptable auxiliary components.
The preparation formulation of the peony and licorice decoction provided by the invention can be freeze-dried powder injection, decoction, granules, tablets, pills, capsules, suspending agents, injection, controlled release or sustained release agents and the like.
Compared with the prior art, the invention has the advantages that:
the peony and licorice decoction is used for preparing the antitumor drug, so that the growth and migration of cancer cells can be inhibited, the cancer cells can be induced to undergo apoptosis, the direct antitumor effect is exerted, the new medical effect of the peony and licorice decoction is expanded, the clinical application range of the peony and licorice decoction is expanded, and a new research idea and direction are provided for the research of the current antitumor drug. Has good guiding and researching significance for developing innovative drugs with independent intellectual property rights.
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FIG. 1 shows the effect of Paeoniae radix and Glycyrrhizae radix decoction on the growth of TE-1 cells;
FIG. 2 shows the effect of Paeoniae radix and Glycyrrhizae radix decoction on TE-1 cell migration ability;
FIG. 3 shows the effect of Paeoniae radix and Glycyrrhizae radix decoction on the apoptosis of TE-1 cells.
DETAILED DESCRIPTION OF EMBODIMENT (S) OF INVENTION
The present invention will be described in further detail with reference to specific examples for better illustrating the objects, technical solutions and advantages of the present invention, but the scope of the present invention is not limited thereto.
The experimental materials used in the following examples are not specifically described, and the biochemical reagents are conventional in the art, and can be prepared according to the conventional methods in the art or commercially available.
The source of the medicinal materials is as follows: radix Paeoniae alba is purchased from Anhui; licorice is purchased from inner Mongolia.
Preparing the medicine: the raw materials and the weight ratio are white paeony root: and (3) adding water into the honey-fried licorice root which is 1:1, decocting and extracting, filtering an extracting solution, taking a filtrate, and performing a freeze dryer to obtain freeze-dried powder. Dissolving and diluting the freeze-dried powder to a certain concentration gradient by using dimethyl sulfoxide (DMSO).
Example 1 evaluation of antitumor Activity of Paeoniae radix and Glycyrrhizae radix decoction against five common human tumor cells (TE-1, Eca-109, SW-620, Hela, MGC-803)
The experimental method comprises the following steps:
adopting SRB method, digesting, centrifuging and resuspending and counting tumor cells in logarithmic growth phase, inoculating 10000 cells/well into 96-well plate according to 1000-2And cultured in a cell culture box at 37 ℃ for 24 hours. After the cells are completely attached to the wall, discarding the old culture medium, setting a negative control group which is not added with drugs and only adds the culture medium, diluting the drugs to the required series concentration by using the complete culture medium, adding 150 mu L of drug-containing culture medium with the corresponding drug concentration into each hole of a 96-hole plate, and setting 3 groups of multiple holes for each concentration. Placing the well plate with the added medicine in 5% CO by volume2And culturing in a 37 ℃ cell culture box for 48 or 72 hours. And taking out the 96-well plate after 48 or 72 hours, discarding the old culture medium containing the medicine, adding 150 mu L of trichloroacetic acid solution with the mass percentage content of 10% into each well, and placing the mixture into a refrigerator at 4 ℃ for fixing for two hours. After the fixation is completed, the solution is taken out, discarded, 300 mu L of ultrapure water is added into each hole, washed three times, and dried for one hour or even longer at room temperature. After the mixture is dried, 200 mu L of SRB solution with the mass percentage content of 0.4% is added into each hole for dyeing, the plate is shaken for 20min, the dyeing solution is discarded after the time, the mixture is washed for three times by acetic acid with the mass percentage content of 1%, and the mixture is dried thoroughly at room temperature. After drying, 200. mu.L of 10mM Tris solution was added to each well, and the plate was shaken for 20min to dissolve it sufficiently. In the experiment, an enzyme-labeling instrument is used for reading the absorbance value of each hole at 562nm, the inhibition rate of the drug on cell proliferation is calculated, and IC is calculated according to the inhibition rate50The value is obtained. The experiments were performed in triplicate and the mean and standard deviation of triplicates were obtained. The results are shown in Table 1.
TABLE 1 evaluation results of antitumor and antitumor activities of Paeoniae radix and Glycyrrhizae radix decoction
And (4) analyzing results:
as can be seen from the table above, the peony and licorice decoction has good anti-tumor activity on the common human tumor cells in 5, presents obvious time dependence, and can be applied to the anti-proliferative activity research of human esophageal cancer, colon cancer, cervical cancer and gastric cancer cells.
EXAMPLE 2 cell monocloning experiments
The experimental method comprises the following steps:
subjecting the logarithmic growth phase of TE-1 cells to a cell growth reaction at a rate of 1X 104One for each well, inoculated into a 6-well plate, shaken up and put into a cell incubator. After 24h, the supernatant was discarded and a blank and culture medium containing different mass percent concentrations of drug (i.e., 0, 2%, 4%, 6%) was added along the walls of the wells. The 6-well plate was placed in the cell incubator again and the medium was changed every 2-3 days. When the cells showed macroscopic colonies, the supernatant was discarded, washed 3 times with PBS, and then fixed with paraformaldehyde. After 20min, abandoning the supernatant, washing with PBS for 3 times, dyeing with crystal violet dye solution for 30min, slowly washing off the dye solution, standing and airing, and calculating the corresponding inhibition rate of clone formation.
The percent (%) inhibition of clone formation was [ (% clone number in control group-clone number in experimental group)/clone number in control group ]. times.100%
And (4) analyzing results:
in order to investigate whether the peony and licorice decoction can influence the in-vitro clone forming capability of TE-1 cells, a cell monoclonal experiment is adopted for verification. As shown in FIG. 1, compared with the NC group, the number of cell clonogenic decreases significantly with the increase of the concentration of the test drug, and when the concentration of the test drug reaches 6%, clonogenic inhibition is almost complete, indicating that peony-licorice decoction can inhibit the clonogenic formation of TE-1 cells in vitro. The data in the figure are the average values from three independent experiments, wherein P < 0.05 and P < 0.01.
Example 3 cell scratch test
The experimental method comprises the following steps:
inoculating TE-1 cells into a 6-well plate, shaking uniformly, and placing into a cell culture box. After 24h, the tip of the tip was scribed vertically with a 100. mu.L lance tip (the tip was not tilted). After washing with 1mL PBS, the blank and the culture medium (i.e. 0, 2%, 3%, 4%, 6%) containing different mass percent drug were added to the 6-well plate along the wall, and cultured in a 37 ℃ cell incubator. Samples were taken at 0, 12, and 24h, respectively, and photographed under a microscope for recording.
Cell mobility (%) - (scratch width of 0 h-scratch width after incubation for several hours)/scratch width of 0h × 100%
And (4) analyzing results:
in order to investigate whether the peony and licorice decoction can influence the in vitro invasion and migration capacity of TE-1 cells, a cell scratch experiment is adopted for verification. As a result, as shown in FIG. 2, the scratch healing tendency of the NC group was very significant as the cell culture time was increased. When the medicine concentration is 2%, the inhibition effect on the healing of cell scratches is weak. But when the concentration of the medicine reaches 4% -6%, the medicine has obvious inhibition effect on healing of cell scratches, which shows that the peony and licorice decoction can inhibit the invasion and metastasis capability of cancer cells. The data in the figure are the average values from three independent experiments, wherein P < 0.05 and P < 0.01.
Example 4 apoptosis assay
The experimental method comprises the following steps:
TE-1 cells were stained at 1X 10 using Annexin V-FITC/PI double staining5One well per well, inoculated into a 6-well plate, shaken well, and placed into a cell incubator. After 24h, the supernatant was discarded and the blank and the culture medium containing different mass percent concentrations of drug (i.e., 0, 24%, 48%) were added to the corresponding wells along the side walls. The 6-well plate was placed in the cell incubator again. After 24h, the growth state of the cells was observed, and the suspended cells in each well and the cells obtained by digestion were combined in the same EP tube, centrifuged at 1000rpm for 5min, and the cells were collected. The cell sediment is blown off evenly by using Binding Buffer, 2 mu L Annexin V-FITC and 2 mu L PI staining solution are added into each EP tube, vortex and mix evenly, the mixture is placed in an ice box and react for 5-15min in a dark place, and the detection is carried out by using a flow cytometer.
And (4) analyzing results:
the peony and licorice soup (0, 24 percent and 48 percent) with different mass percentage concentrations are respectively used for acting on TE-1 cells for 24 hours, and the influence of the peony and licorice soup on the TE-1 cell apoptosis is detected by a flow cytometer. As shown in FIG. 3, the ratio of TE-1 apoptosis was gradually increased with the increase of the concentration of the test drug compared to NC group, indicating that Paeoniae and Glycyrrhizae radix decoction can induce TE-1 apoptosis to exert in vitro anti-tumor effect and have concentration dependence. The data in the figure are the average values from three independent experiments, wherein P < 0.05 and P < 0.01.
Claims (5)
1. Use of a Chinese medicinal composition prepared from radix Paeoniae alba and radix Glycyrrhizae Preparata in preparing medicine for treating tumor diseases; the traditional Chinese medicine composition is prepared from the following raw material medicines in parts by weight: white peony root: honey-fried licorice root =1: 1.
2. The use of the composition of claim 1, wherein the composition is a decoction of radix Paeoniae alba and radix Glycyrrhizae Preparata.
3. The use of the composition of claim 1 or 2, wherein the tumor is human esophageal cancer, colon cancer, cervical cancer or gastric cancer.
4. The use of the composition as claimed in claim 1 or 2, wherein the composition is prepared from radix Paeoniae alba and radix Glycyrrhizae Preparata as the sole active ingredient or in combination with pharmaceutically acceptable auxiliary ingredients.
5. The use of the traditional Chinese medicine composition prepared from radix paeoniae alba and radix glycyrrhizae preparata according to claim 4, wherein the dosage form comprises freeze-dried powder injection, decoction, granules, tablets, pills, capsules, suspending agents, injections, controlled release or sustained release agents.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN115444825A (en) * | 2022-06-07 | 2022-12-09 | 神威药业集团有限公司 | Preparation method of peony and liquorice soup freeze-dried powder |
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2021
- 2021-06-04 CN CN202110623260.4A patent/CN113230316A/en active Pending
Non-Patent Citations (3)
| Title |
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| 梁岚等: "甘草甜素对人食管癌细胞ECA109的影响及机制", 《中国老年学杂志》 * |
| 王付: "《经方妙用治百病》", 31 July 2008, 人民军医出版社 * |
| 许树才: "芍药甘草汤对晚期癌症患者临床疗效观察", 《辽宁中医药大学学报》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN115444825A (en) * | 2022-06-07 | 2022-12-09 | 神威药业集团有限公司 | Preparation method of peony and liquorice soup freeze-dried powder |
| CN115444825B (en) * | 2022-06-07 | 2024-01-26 | 神威药业集团有限公司 | Preparation method of radix paeoniae alba and liquorice soup freeze-dried powder |
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