CN114404465A

CN114404465A - A black ginseng paste for treating tumor and its preparation method

Info

Publication number: CN114404465A
Application number: CN202210012489.9A
Authority: CN
Inventors: 解慧勇
Original assignee: Individual
Current assignee: Individual
Priority date: 2022-01-06
Filing date: 2022-01-06
Publication date: 2022-04-29

Abstract

The invention belongs to the technical field of tumor drugs, and discloses a black ginseng paste for treating tumors and a preparation method thereof. The invention effectively improves the content of rare saponins Rg3 and Rg5, enhances intestinal absorption, improves the absorptivity of rare saponins, realizes the improvement of the anticancer effect to the maximum value, and avoids the generation of toxic and side effects. The invention can generate more effective substances by utilizing a special process; concentrating at low temperature, and retaining more content components of rare saponins Rg3 and Rg5 with high concentration of effective components; the application range is wide; the extraction efficiency is high; can effectively improve the absorption rate of human body.

Description

A black ginseng paste for treating tumor and its preparation method

Technical Field

The invention belongs to the technical field of tumor medicines, and particularly relates to black ginseng paste for treating tumors and a preparation method thereof.

Background

Currently, in 2018, 9, the world health organization international cancer research institute published a content entitled "global cancer report" that relates to global cancer epidemics and deaths due to this disease. Reports indicate that by the end of the year it is expected that about 1810 new cancer cases, 960 ten thousand cancer death cases will be found. There was a significant rise compared to 1410 ten thousand new cases and 820 ten cancer death cases in 2012. The report shows that worldwide, the cumulative risk of developing cancer before age 75 is 21.4%, the risk of dying from cancer is 17.7%, and the second leading killer to cardiovascular disease, threatening human health.

Tumor infiltration and metastasis are the leading cause of death in cancer. The lack of depth in its mechanism has hindered effective measures to prevent cancer invasion and metastasis. But recently have made encouraging progress in the study of the molecular mechanisms of tumors and are leading to new strategies for prevention and treatment.

The main traditional therapeutic methods for malignant tumor include surgery, radiation, and chemical drug therapy, and the fourth and fifth therapeutic modes of biological therapy and cancer thermotherapy are now considered. In all cases, even if a malignant tumor is extensively resected at an early stage, it cannot be said that the cancer is completely cleared, and extremely complicated interrelations exist among the host, the tumor, and the treatment. There are three possible outcomes of cancer patients after treatment: firstly, permanent cure is carried out; ② the cancer tumor continues to develop irrepressibly and leads to death, the treatment can not eliminate cancer; after a period of obvious remission, the tumor recurs, metastasizes or presents a new focus, which indicates that the defense function of the organism is low and the tolerance of the anticancer drug occurs. The use of plants for the treatment of tumours has a long history and one can find a number of effective anti-cancer components from plants, such as the flavonoid-frataxin, although it is already fully synthetically available, but the new skeleton is derived from the natural product rohitukine. Rohitukine was isolated in india in the early 90 s of the 20 th century and has anti-inflammatory and immunomodulatory activity. Through the structure-activity relationship research, the fraveline which has tyrosine kinase activity and growth inhibition effect on a series of breast cancer and lung cancer cells is finally found from more than 100 analogs. Data are related to the fact that the proportion of the drug actually derived from synthesis is reported to be 37%. Considering the kind of disease, 68.3% of anti-infective drugs (antibacterial, antifungal, antiparasitic, antiviral) are derived from natural products, while 79.8% in the antitumor field are derived from natural products.

The ginseng element is called 'Baicaozhiwang' and has been used as a medicinal plant in Asian countries such as China, Korea, Japan for thousands of years, and has pharmacological effects of tonifying, regulating immunity, resisting aging and the like. With the development of modern separation and purification technology, ginsenoside, polysaccharide, terpenoid, flavone, alkynol, organic acid, vitamin, amino acid, oligopeptide, saccharide and inorganic salt are separated and purified in ginseng. The biological activity research of various components shows that the pharmacological action of the ginseng is mainly derived from ginsenoside. At present, more than 60 ginsenosides have been isolated from panax genus of ginseng family, and each ginsenoside has its unique physiological activity. The research shows that: ginsenoside has effects in inhibiting tumor cell growth, enhancing immunity, relieving fatigue, delaying aging, regulating central nervous system, and improving cardiovascular and cerebrovascular insufficiency.

The black ginseng is a product of ginseng which is artificially processed, and the active ingredients of the black ginseng are increased. The black ginseng extract (black ginseng paste obtained by normal temperature extraction and low temperature vacuum concentration) comprehensively utilizes the effective components of the black ginseng extract, and has remarkable effect particularly in the deep research of anti-tumor. However, in the traditional preparation method, due to overhigh temperature, rare saponin components in the prepared paste are greatly damaged, and the absorption rate and the efficacy of a human body are influenced.

Through the above analysis, the problems and defects of the prior art are as follows:

in the traditional preparation method, due to overhigh temperature, rare saponin components in the prepared paste are greatly damaged, and the absorption rate and the efficacy of a human body are influenced.

The significance of solving the problems and the defects is as follows: the rare ginsenoside is secondary saponin, can be generated only in the processing process of ginseng, is very sensitive to temperature, humidity and pH value, the yield of the rare ginsenoside can be ensured only by strictly controlling the temperature, humidity and pH value parameters of different processing stages, and the low-temperature extraction and concentration are kept in the process of processing the black ginseng paste to avoid the hydrolysis of the rare ginsenoside.

Disclosure of Invention

Aiming at the problems in the prior art, the invention provides a black ginseng paste for treating tumors and a preparation method thereof.

The invention is realized in such a way that the preparation method of the black ginseng paste for treating the tumor comprises the following steps:

step one, crushing the processed black ginseng;

step two, extracting by adopting a low-temperature high-pressure extraction mode, and avoiding the rare saponin from being hydrolyzed due to overhigh temperature;

filtering the total extract to obtain a black ginseng liquid, putting the black ginseng liquid into a storage tank, and centrifuging;

and step four, carrying out low-temperature vacuum concentration on the black ginseng liquid, and packaging in a glass bottle.

Further, the preparation method of black ginseng in the first step comprises:

cleaning fresh Ginseng radix, sorting, placing into a fermentation box, fermenting under the conditions of controlling temperature, humidity and pH value to promote the production of rare ginsenoside, so that common ginsenoside is converted into rare ginsenoside with conversion rate of above 70%.

Fermenting the fresh ginseng in a fermentation box for 5-10 days, and then drying at 50 ℃.

The fresh ginseng is fresh ginseng for more than five years in a long term.

Further, in the first step, the black ginseng is ground and crushed on a milling machine to 20-90 meshes.

Further, in the second step, when a low-temperature high-pressure extraction mode is adopted for extraction, the adopted extraction mode specifically comprises:

soaking in 10 times of distilled water or 50% ethanol at 30 deg.C for 8 hr, and extracting at 40 deg.C or below with ultrahigh pressure cold isostatic pressing equipment for 6 hr.

Further, in the third step, the total extract is filtered by a 10 micron filter element during filtering.

Further, in the fourth step, when the black ginseng liquid is subjected to low-temperature vacuum concentration, the filtrate is subjected to vacuum concentration at the temperature of 60 ℃ or below, and the paste is concentrated to the density of 1.2 and stops.

Another object of the present invention is to provide a black ginseng paste prepared by the method for preparing black ginseng paste for treating tumors.

The invention also aims to provide the application of the black ginseng paste in preparing the medicines for improving the immunity of the organism, resisting fatigue, delaying senility, regulating the central nervous system and improving the blood supply insufficiency of the heart and the brain.

By combining all the technical schemes, the invention has the advantages and positive effects that:

the invention effectively improves the content of rare saponins Rg3 and Rg5, enhances intestinal absorption, improves the absorptivity of rare saponins, realizes the improvement of the anticancer effect to the maximum value, and avoids the generation of toxic and side effects.

The invention can generate more effective substances by utilizing a special process; concentrating at low temperature, and retaining more content components of rare saponins Rg3 and Rg5 with high concentration of effective components; the application range is wide; the extraction efficiency is high; can effectively improve the absorption rate of human body.

Drawings

In order to more clearly illustrate the technical solutions of the embodiments of the present application, the drawings needed to be used in the embodiments of the present application will be briefly described below, and it is obvious that the drawings described below are only some embodiments of the present application, and it is obvious for those skilled in the art that other drawings can be obtained from the drawings without creative efforts.

Fig. 1 is a flow chart of a preparation method of black ginseng cream for treating tumors according to an embodiment of the present invention.

FIG. 2 is a schematic diagram illustrating the effect of the black ginseng digestion lyophilized powder on the proliferation capacity of HepG2 cells.

FIG. 3 is a schematic diagram showing the effect of black ginseng paste on the proliferation potency of HepG2 cells according to the present invention.

FIG. 4 is a schematic diagram illustrating the effect of the digestion lyophilized powder of black ginseng on the proliferation capacity of HCT-116 cells according to the embodiment of the present invention.

FIG. 5 is a schematic diagram showing the effect of black ginseng paste on the proliferation ability of HCT-116 cells according to an embodiment of the present invention.

FIG. 6 is a schematic diagram showing the effect of black ginseng paste on INS-1 cell proliferation potency according to an embodiment of the present invention.

Fig. 7 is a schematic diagram illustrating the effect of black ginseng cream on the proliferation capacity of a549 cells according to an embodiment of the present invention.

FIG. 8 is a schematic diagram showing the effect of black ginseng paste on the proliferation potency of Hela cells according to an embodiment of the present invention.

FIG. 9(a) is a morphological diagram of cells cultured for 24h in A375 according to the present invention.

FIG. 9(b) is a morphological diagram of cells cultured for 48h A375 according to the present invention.

FIG. 9(c) is a 24h cell morphology map of cisplatin administration as provided by the example of the invention.

FIG. 9(d) is a cell morphology map of 48h cisplatin administration provided by the example of the present invention.

FIG. 9(e) is a cell morphology diagram of black ginseng paste administered for 24h according to an embodiment of the present invention.

FIG. 9(f) is a cell morphology diagram of 48h after administration of black ginseng paste according to an embodiment of the present invention.

Detailed Description

In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is further described in detail with reference to the following embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.

Aiming at the problems in the prior art, the invention provides black ginseng paste for treating tumors and a preparation method thereof, and the invention is described in detail below with reference to the accompanying drawings.

As shown in fig. 1, the preparation method of black ginseng paste for treating tumor provided by the embodiment of the present invention includes:

s101, crushing the processed black ginseng;

s102, extracting by adopting a low-temperature high-pressure extraction mode, and avoiding the rare saponin from being hydrolyzed due to overhigh temperature;

s103, filtering the total extract to obtain a black ginseng liquid, putting the black ginseng liquid into a storage tank, and standing for centrifugation;

and S104, carrying out low-temperature vacuum concentration on the black ginseng liquid, and packaging in a glass bottle.

The invention is further described with reference to specific examples.

Selecting fresh ginseng with more than five years of growth period, cleaning, sorting, then placing into a fermentation box, fermenting for 5-10 days under the conditions of controlling temperature, humidity and pH value, then drying at about 50 ℃ to promote the generation of a large amount of rare ginsenoside, so that the common ginsenoside is converted into the rare ginsenoside, the conversion rate is more than 70%, and the black ginseng is prepared, thereby greatly improving the effective components.

Grinding 100 kg of the prepared black ginseng on a milling machine to 20-90 meshes, preferably 50 meshes;

the preparation method comprises grinding black ginseng sample on a milling machine, sieving with 50 mesh sieve to obtain fine powder, soaking with 10 times of distilled water or 50% ethanol at 30 deg.C for 8 hr, extracting at 40 deg.C under low temperature and high pressure by using ultrahigh pressure cold isostatic pressing equipment to avoid hydrolysis of rare saponin due to overhigh temperature, and extracting for 6 hr;

filtering the total extractive solution with 10 μm filter core to obtain black Ginseng radix solution, placing in storage tank, and centrifuging;

pumping the black ginseng liquid into a liquid storage tank at a low temperature of below 60 ℃, performing low-temperature vacuum concentration, stopping concentrating the paste until the density is 1.2, and packaging in a glass bottle.

The efficacy of the black ginseng paste prepared according to the present invention will be further described with reference to specific experiments.

1. Effect of black ginseng cream on proliferation of HepG2 cells.

HepG2 cells treated by the black ginseng paste for 48 hours have lower inhibition rate than that treated by the black ginseng digestion freeze-dried powder for 24 hours, but the inhibition rate is increased to 26.85 percent when the increase concentration is 2000 mug/mL, and particularly shown in figure 1 and figure 2.

The experimental results show that the black ginseng paste has a certain inhibition effect on gastric tumors after oral administration.

2. Effect of black ginseng extract on proliferation of HCT-116 cells.

The inhibition rate of the black ginseng paste reaches 31.21% at maximum when the drug concentration is 2000 mug/mL for 24 hours, the inhibition rate of the black ginseng paste reaches 23.92% and 39.04% at 500 mug/mL and 1000 mug/mL for 48 hours, and the inhibition rate reaches 64.21% at 2000 mug/mL, particularly shown in figures 3 and 4(a) to 4 (c).

3. Effect of black ginseng paste on the proliferation of INS-1 cells.

The inhibition rate of the black ginseng extract on INS-1 cells is only 7.19% when the black ginseng extract is treated for 24 hours and the drug concentration is 1000 mug/mL, and the inhibition rate of the stock solution is found to be 58.37% at the maximum when the black ginseng extract inhibits the proliferation of INS-1 cells and 62.27% at the maximum when the concentration reaches 2000 mug/mL along with the increase of the administration concentration when the black ginseng extract inhibits the proliferation of INS-1 cells after being treated for 48 hours. See fig. 5 and fig. 6(a) to 6(b) in detail.

4. The effect of black ginseng extract on the proliferation of a549 cells.

As shown in figure 7, the inhibition rate of the black ginseng extract on A549 cells reaches 37.50% when the black ginseng extract is treated for 24 hours and the concentration is 1000 mug/mL; the black ginseng paste is treated for 48 hours, when the concentration of the medicine is 1000 mug/mL, the inhibition rate of the black ginseng paste on A549 cells reaches 41.17%, and when the concentration of the medicine is 2000 mug/mL, the inhibition rate of the black ginseng paste on the A549 cells reaches 72.60%. See in particular fig. 7(a) to 7 (c).

5. The effect of black ginseng paste on the proliferation of Hela cells.

The black ginseng paste is processed for 24 hours, the concentration is 1000 mug/mL, the inhibition rate to cervical cancer (Hela) cells reaches 34.53%, the concentration is 2000 mug/mL, and the inhibition rate to cervical cancer (Hela) cells reaches 74.18%; when the black ginseng extract is treated for 48 hours and the concentration is 1000 mug/mL, the inhibition rate of the black ginseng extract on Hela cells is 43.70%. At a concentration of 2000. mu.g/mL, the inhibition rate of HeLa cells by black ginseng paste was 89.38%, as shown in FIGS. 8(a) to 8 (d).

The experimental results show that the medicine effect is better than 24h in 48h and shows dose dependence, so that the black ginseng ointment has a certain inhibition effect on the cervical cancer Hela.

6. Effects of black ginseng cream and cisplatin on a375 cell morphology.

From FIGS. 9(a) to 9(f), it can be seen that the cell morphology of A375 cells cultured for 24 hours and 48 hours showed the characteristics of atrophy and decrease compared with the cell morphology without the drug, indicating that cisplatin and black ginseng paste can induce apoptosis and inhibit proliferation of cells.

The black ginseng has obvious effect of inhibiting cancer cells and has no toxic or side effect.

The results of the screening research project on the anti-tumor effect of the black ginseng are proved by the results of the report of the Jilin university Life sciences institute:

1. research on influence of black ginseng paste on cells of three digestive systems of liver cancer, colorectal cancer and pancreatic cancer

The black ginseng paste has an inhibitory effect on HepG2 cells, HCT-116 cells and pancreatic cancer. After the action of 1000 mu g/mL of the medicine for 24 hours, the inhibition rates of the medicine on HepG2 cells and HCT-116 cells are 47 percent and 11 percent respectively, and the inhibition rate of the medicine on pancreatic cancer (INS-1) after 48 hours is 5.8 percent respectively.

2. Research on influence of black ginseng paste on lung cancer and cervical cancer

The black ginseng cream can inhibit the proliferation of A549 cells and Hela cells, has higher inhibition rate on the proliferation of the two cells, particularly has the drug concentration of 1000 mu g/mL, has the inhibition rates of 37.50 percent and 34.53 percent on the two cells respectively when being treated for 24 hours, has the inhibition rates of 41.17 percent and 43.70 percent on the two cells respectively when being treated for 48 hours, has the inhibition rates of 2000 mu g/mL, has the inhibition rates of 70.50 percent and 74.18 percent on the two cells respectively when being treated for 24 hours, and has the inhibition rates of 72.60 percent and 89.38 percent on the two cells respectively when being treated for 48 hours.

3. Research on influence of black ginseng paste on A375 cell apoptosis and cycle by using flow cytometry

The black ginseng paste acts on A375 cells, and after the cells are cultured for 24 hours and 48 hours, the cells of the black ginseng paste have the characteristics of shrinking and diminishing compared with the cells of the black ginseng paste without the drug, which shows that the black ginseng paste can promote the apoptosis of the cells.

The apoptosis experiment result shows that after the black ginseng paste is used for treating the A375 cells, the proliferation of the cells is inhibited, and the inhibition rate is increased along with the increase of time. After the medicine acts for 24 hours, the inhibition rate of the black ginseng paste medicine on cells can reach 22.18% when the medicine concentration is 1 mg/mL; after the medicine acts for 48 hours, the inhibition rate of the black ginseng paste medicine concentration of 1mg/mL on cells can reach 44%. The black ginseng extract can inhibit the proliferation of liver cancer cells, which is the same as the MTT result.

The research shows that the black ginseng paste has obvious inhibition effect on liver cancer, colorectal cancer, lung cancer, cervical cancer and pancreatic cancer.

The above description is only for the purpose of illustrating the present invention and the appended claims are not to be construed as limiting the scope of the invention, which is intended to cover all modifications, equivalents and improvements that are within the spirit and scope of the invention as defined by the appended claims.

Claims

1. The preparation method of the black ginseng paste for treating the tumors is characterized by comprising the following steps of:

step one, crushing the processed black ginseng;

2. The method of preparing black ginseng paste for treating tumor according to claim 1, wherein the method of preparing black ginseng in step one comprises:

cleaning fresh Ginseng radix, sorting, placing into a fermentation box, fermenting under the conditions of controlling temperature, humidity and pH value to promote the production of rare ginsenoside.

3. The method of preparing black ginseng paste for treating tumor according to claim 2, wherein the fresh ginseng is fermented in a fermentation tank for 5-10 days and then dried at 50 ℃.

4. The method of preparing black ginseng paste for treating tumor according to claim 2, wherein the fresh ginseng is fresh ginseng having a growth period of more than five years.

5. The method of preparing black ginseng paste for treating tumor according to claim 1, wherein the black ginseng is ground and pulverized on a milling machine to 20-90 mesh in the first step.

6. The method for preparing black ginseng paste for treating tumor according to claim 1, wherein when the low-temperature high-pressure extraction is adopted in the second step, the adopted extraction method specifically comprises:

soaking in 10 times of distilled water or 50% ethanol at 30 deg.C for 8 hr, and extracting at 40 deg.C for 6 hr by using ultrahigh pressure cold static pressure equipment.

7. The method for preparing black ginseng paste for treating tumor according to claim 1, wherein the total extract in step three is filtered by a 10 μm filter.

8. The method for preparing black ginseng paste for treating tumor according to claim 1, wherein the concentration of the paste to a density of 1.2 is stopped by vacuum concentrating the filtrate at a temperature of 60 ℃ or less while vacuum concentrating the black ginseng liquid at a low temperature in the fourth step.

9. A black ginseng paste prepared by the method for preparing black ginseng paste for treating tumors according to any one of claims 1 to 8.

10. The use of the black ginseng paste of claim 9 in preparing a medicament for enhancing immunity, resisting fatigue, delaying aging, regulating the central nervous system, and improving cardiovascular and cerebrovascular insufficiency.