Anti-cancer composition for inhibiting growth of cancer cells and application of anti-cancer composition

Abstract

The invention discloses an anti-cancer composition for inhibiting growth of cancer cells and application of the anti-cancer composition. The anticancer composition for inhibiting the growth of cancer cells is prepared from an angelica keiskei polypeptide micro-ecological extract, an anthraquinone compound, a semen raphani extract, curcumin, resveratrol, fucoidin and lycopene. According to the application, the angelica keiskei polypeptide micro-ecological extract, the anthraquinone compound, the semen raphani extract, curcumin, resveratrol, fucoidin and lycopene are creatively combined for use, so that the composition has an inhibiting effect on various cancer cells such as lung cancer, gastric cancer, liver cancer, breast cancer, colon cancer, cervical cancer, bile duct cancer, esophageal cancer, floor carcinoma of mouth and nasopharynx cancer; and a new solution is provided for the treatment of the cancers.

Description

Technical Field

[0001] The present application relates to the technical field of cancer treatment, and in particular to an anticancer composition for inhibiting the growth of cancer cells and its application. Background Art

[0002] Cancer is one of the diseases that has the greatest impact on human health and life. The cause of cancer is usually due to a series of abnormal gene changes caused by the long-term impact of various chemical, physical, viral and other single or multiple factors on the body, which in turn leads to malignant cell proliferation.

[0003] At present, the main methods for treating cancer include traditional surgery, chemotherapy, radiotherapy, etc. With the research and development of traditional Chinese medicine, various Chinese herbal medicines, Chinese herbal extracts and their combinations have been found to have different anti-cancer functions. However, the anti-cancer effect of Chinese herbal medicines or Chinese herbal extracts is greatly affected by the quality of the Chinese herbal medicines themselves; therefore, further in-depth research on the active ingredients in Chinese herbal medicines and the development of new anti-cancer drugs or drug compositions based on the effective active ingredients of Chinese herbal medicines are the focus and difficulty of the research field of traditional Chinese medicine. This is also an effective way for traditional Chinese medicine to be more widely recognized and used worldwide. Summary of the invention

[0004] The purpose of the present application is to provide a new anticancer composition for inhibiting the growth of cancer cells and its application.

[0005] This application adopts the following technical solutions:

[0006] One aspect of the present application discloses an anticancer composition for inhibiting the growth of cancer cells, which is composed of a microecological extract of a tomorrow leaf polypeptide, anthraquinone compounds, a radish seed extract, curcumin, resveratrol, fucoidan and lycopene.

[0007] Preferably, the Angelica keiskei polypeptide microecological extract accounts for 70-88% of the total weight, the anthraquinone compounds account for 3.9-8.3% of the total weight, the radish seed extract accounts for 7-14% of the total weight, the curcumin accounts for 0.5-2% of the total weight, the resveratrol accounts for 0.05-4.5% of the total weight, the fucoidan accounts for 0.5-1% of the total weight, and the lycopene accounts for 0.05-0.2% of the total weight.

[0008] Preferably, the Angelica keiskei polypeptide microecological extract is prepared by the following method:

[0009] 20-35 parts by weight of Angelica keiskei polypeptide, 15-30 parts by weight of pig blood peptide, 2-5 parts by weight of soybean lecithin, 6-12 parts by weight of complex vitamins, 6-12 parts by weight of complex minerals, 4-6 parts by weight of gentiooligosaccharides and 6-12 parts by weight of mannitol are uniformly mixed and crushed and sieved to obtain a mixed raw material;

[0010] Add 0.1-1 parts by weight of Bifidobacterium lactis and 0.1-1 parts by weight of Bacillus to the mixed raw materials, and incubate at a constant temperature of 30-35° C. for 2-4 hours to obtain a microecological fermentation liquid;

[0011] The microecological fermentation liquid is filtered to remove the residue and collect the filtrate; the filtrate is subjected to 400-500w power ultrasound for 10-30min, and then centrifuged at 6000-8000rpm for 10-30min, and the supernatant is sterilized and matured to obtain the tomorrow leaf polypeptide microecological extract of the present application.

[0012] Preferably, the aging treatment comprises placing the sterilized supernatant in a sterilized aging barrel for sealed aging and storage, the sealed aging and storage temperature is 30-35° C., and the time is 1-3 weeks.

[0013] Preferably, the anthraquinone compound is a natural hydroxyanthraquinone compound.

[0014] Preferably, the natural hydroxyanthraquinone compound is rhubarb and / or rhubarb methyl ether.

[0015] Preferably, rhamnosin and / or rhamnosin methyl ether are extracted from Polygonum cuspidatum.

[0016] Preferably, the radish seed extract is an ethanol extract of radish seeds.

[0017] Preferably, the preparation method of the ethanol extract of radish seeds comprises adding 10-30 times the weight of ethanol to radish seed powder, and extracting with the assistance of ultrasound and heating; then, removing the residue, collecting the filtrate, and removing the solvent in the filtrate to obtain the ethanol extract of radish seeds.

[0018] Preferably, the particle size of the radish seed powder is less than 0.3 mm.

[0019] Preferably, the ethanol is 95% ethanol.

[0020] Preferably, in the preparation method of the radish seed extract, the ultrasonic power is 300-400w, the heating temperature is 40-60°C, and the extraction time is 20-60min.

[0021] Another aspect of the present application discloses the use of the anticancer composition of the present application in the preparation of a drug for treating cancer.

[0022] Preferably, the anticancer composition of the present application is applicable to at least one of lung cancer, gastric cancer, liver cancer, breast cancer, colon cancer, cervical cancer, bile duct cancer, esophageal cancer, floor of mouth cancer and nasopharyngeal cancer.

[0023] The beneficial effects of this application are:

[0024] The anti-cancer composition of the present application creatively combines the Angelica keiskei polypeptide microecological extract, anthraquinone compounds, radish seed extract, curcumin, resveratrol, fucoidan and lycopene, and can inhibit a variety of cancer cells such as lung cancer, gastric cancer, liver cancer, breast cancer, colon cancer, cervical cancer, bile duct cancer, esophageal cancer, floor of mouth cancer and nasopharyngeal cancer, providing a new solution for the treatment of these cancers. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figures 1 to 3 These are the test results of the proliferation inhibition activity of the anticancer composition in the examples of the present application and its control on different cancer cells. DETAILED DESCRIPTION

[0026] The present application is further described in detail below through specific examples. The following examples are only used to further illustrate the present application and should not be construed as limiting the present application.

[0027] Unless otherwise specified, the materials and reagents used in the following examples can be obtained from commercial sources, and the experimental methods used in the following examples are conventional methods unless otherwise specified.

[0028] Example

[0029] The anticancer composition for inhibiting the growth of cancer cells in this example is composed of Angelica keiskei polypeptide microecological extract, anthraquinone compounds, radish seed extract, curcumin, resveratrol, fucoidan and lycopene, and its specific preparation method is as follows:

[0030] 1. Preparation of Angelica keiskei polypeptide microecological extract

[0031] 1) Select raw materials (by weight, 1 part represents 10g)

[0032] Fresh Angelica keiskei (32 parts of Angelica keiskei polypeptides extracted), 20 parts of pig blood peptides, 2 parts of soy lecithin, 6 parts of multivitamins, 6 parts of multiminerals, 6 parts of gentiooligosaccharides and 6 parts of mannitol (all commercially available).

[0033] 2) Preparation process

[0034] Extraction of Angelica keiskei polypeptide: fresh Angelica keiskei is used as raw material for squeezing, pure water is added to the obtained juice and stirred evenly to obtain Angelica keiskei solution, which is placed in a 60°C water bath for 20 minutes; after the solution temperature drops to room temperature, trypsin and subtilisin are added for enzymolysis at pH 7.5, the enzymolysis time is 1 hour, the enzymolysis temperature is 37°C, centrifugation is performed, and the supernatant is taken; papain and serine protease are added for enzymolysis at pH 8.5, the enzymolysis time is 1 hour, the enzymolysis temperature is 42°C, centrifugation is performed, the supernatant is taken, and the pH is adjusted to 5.5 for enzymolysis for 1 hour, the enzymolysis temperature is 55°C, and after enzyme inactivation treatment, it is concentrated and dried to obtain Angelica keiskei polypeptide powder, which is sieved through a 50-mesh sieve for standby use.

[0035] Preparation of Angelica keiskei polypeptide microecological extract: Mix the other raw materials except Angelica keiskei polypeptide according to the above weight parts, grind them through a 100-mesh sieve to obtain a mixed raw material; mix and stir with the Angelica keiskei polypeptide, and then add 0.1 weight parts of Bifidobacterium lactis and 0.1 weight parts of Bacillus, and mix them; incubate at a constant temperature of 35°C for 2h to obtain a microecological fermentation liquid; filter the microecological fermentation liquid to remove the residue, and collect the filtrate; subject the filtrate to 450W power ultrasound for 20min, centrifuge at 7000rpm for 20min, take the supernatant for sterilization, and place the sterilized supernatant in a sterilized maturation barrel and seal and mature it at 35°C for 2 weeks to obtain the Angelica keiskei polypeptide microecological extract.

[0036] At the same time, a mixed raw material without adding Angelica keiskei polypeptide was designed, that is, except for the absence of Angelica keiskei polypeptide, other components were exactly the same, the same Bifidobacterium lactis and Bacillus were added, and the mixture was processed under the same conditions to obtain a control extract.

[0037] 2. Preparation of Anthraquinone Compounds

[0038] The anthraquinone compound used in this example is a natural hydroxyanthraquinone compound, specifically rhubarb extracted from Polygonum cuspidatum. The specific preparation method includes:

[0039] 2 kg of Polygonum cuspidatum was crushed, passed through a 20-mesh sieve, extracted three times with 95% ethanol and refluxed in a water bath, the filtrates were combined, concentrated to 300 mL, 200 mL of 3 mol / L sulfuric acid was added, 1.5 L of water was added after reflux for 1 hour, the mixture was allowed to stand overnight, filtered and dried to obtain a pink powder; the pink powder was put into a Soxhlet extractor, extracted with ether until the ether became colorless, the ether layer was extracted with 5% sodium bicarbonate, the aqueous phase was discarded, and then extracted with 5% sodium carbonate, the combined extracts were adjusted to pH 4 with 5 mol / L hydrochloric acid, filtered, dried, and crystallized with acetone to obtain an orange-yellow crystalline compound, i.e., rhein.

[0040] 3. Preparation of Radish Seed Extract

[0041] In this example, 95% ethanol was used to extract the radish seed extract, and the specific preparation method includes:

[0042] 2 kg of dried radish seeds were crushed, passed through a 60-mesh sieve, 20 times the weight of 95% ethanol was added, heated to 50°C under 350W ultrasonic conditions, extracted for 30 min, filtered, the filtrate was collected, the residue was extracted twice, the three filtrates were combined, concentrated under reduced pressure to 200 mL, frozen at -80°C, and freeze-dried to obtain a powder, i.e., radish seed extract.

[0043] 4. Preparation of Composition

[0044] Take 70-88g of Angelica keiskei polypeptide microecological extract, 3.9-8.3g of anthraquinone compounds, 7-14g of radish seed extract, 0.5-2g of curcumin (Meryer M81102), 0.05-4.5g of resveratrol (McLean MKL-R817262), 0.5-1g of fucoidan (McLean MKL-F889413), and 0.05-0.2g of lycopene (Meryer M74381) to prepare the composition of this example. Specifically prepare the composition of each test group and the composition of the control group according to the formula in Table 1.

[0045] Table 1 Anticancer composition formula

[0046]

[0047]

[0048] In Table 1, the control extract is a microecological extract prepared by the same method as the Angelica keiskei polypeptide microecological extract, but does not contain Angelica keiskei polypeptide and uses the same other raw materials.

[0049] According to the formulations in Table 1, Test 1 to Test 7 and Control 1 to Control 7, a total of 14 compositions, were prepared for testing.

[0050] 5. Anticancer activity test

[0051] The anticancer activities of the above 14 compositions were characterized by their cytotoxicity to cancer cells. The specific cancer cells used in this experiment included: lung cancer cells A549 (Biyuntian), gastric cancer cells GTL-16 (Biyuntian), liver cancer cells HLE (Biyuntian), breast cancer cells HCC1937 (Biyuntian), colon cancer cells HCT116 (Biyuntian), cervical cancer cells HeLa (Biyuntian), bile duct cancer cells RBE (Biyuntian), esophageal cancer cells TE-1 (Biyuntian), floor of mouth cancer cells HTX2346C (Hodi Huatuo Biotechnology), and nasopharyngeal cancer cells IM-H136 (IMMOCELL). Human normal liver cells QSG 7701 (Biyuntian) were used as the control group.

[0052] The proliferation inhibition activity of 14 compositions on 10 types of cancer cells and normal human liver cells was determined by standard MTT colorimetric method, specifically including:

[0053] Cells in the logarithmic growth phase were taken and digested with 0.25% trypsin to prepare a single cell suspension. The cells were counted using a cell counter and diluted to a concentration of 5 × 10 4 / mL single cell suspension; then, 100 μL of each single cell suspension was taken and added to two 96-well plates, and 20 μL of the above 14 compositions were added respectively, and a negative control of adding 20 μL of water was set up. After 72 hours, 20 μL of 5 mg / mL MTT was added to each well, and cultured for another 4 hours, the supernatant was discarded, 150 μL DMSO was added to each well, and oscillated for 15 minutes. The absorbance at 490 nm was detected by a microplate reader Multiskan SkyHigh, and the cell proliferation inhibition rate was calculated according to the following formula.

[0054] Cell growth inhibition rate = (1-cancer cell absorbance value / normal cell absorbance value) × 100%

[0055] The ratio of the absorbance value of the negative control of cancer cells with added water to the absorbance value of the negative control of normal cells with added water was used as a correction coefficient and multiplied by the above-mentioned inhibition rate of cell growth to obtain the final cancer cell inhibition rate result. The results of the proliferation inhibition activity of the 14 compositions on 10 types of cancer cells and normal human liver cells are shown in Tables 2, 3 and Figures 1 to 3 shown.

[0056] Table 2 Test results of proliferation inhibition activity in the experimental group

[0057] cell Test 1 Test 2 Test 3 Test 4 Test 5 Test 6 Test 7 Lung cancer 66％ 78％ 69％ 71％ 70％ 77％ 76％ Gastric cancer 62％ 71％ 66％ 67％ 65％ 72％ 70％ Liver cancer 56％ 67％ 58％ 62％ 61％ 66％ 66％ Breast cancer 39％ 48％ 39％ 40％ 42％ 47％ 45％ Colon cancer 48％ 59％ 51％ 54％ 50％ 59％ 58％ Cervical cancer 35％ 49％ 40％ 41％ 40％ 47％ 46％ Cholangiocarcinoma 47％ 59％ 51％ 52％ 51％ 58％ 57％ Esophageal cancer 46％ 58％ 49％ 52％ 51％ 58％ 57％ Floor of mouth cancer 42％ 48％ 39％ 40％ 42％ 49％ 48％ Nasopharyngeal carcinoma 44％ 57％ 49％ 50％ 49％ 56％ 53％ Normal cells 0％ 0％ 0％ 0％ 0％ 0％ 0％

[0058] Table 3 Results of proliferation inhibition activity test in control group

[0059] cell Comparison 1 Comparison 2 Comparison 3 Comparison 4 Comparison 5 Comparison 6 Comparison 7 water Lung cancer 10％ 21％ 27％ 28％ 28％ 24％ 25％ 0％ Gastric cancer 8％ 17％ 21％ 20％ 19％ 15％ 14％ 0％ Liver cancer 5％ 21％ 26％ 25％ 23％ 21％ 19％ 0％ Breast cancer 2％ 8％ 10％ 11％ 10％ 9％ 7％ 0％ Colon cancer 5％ 11％ 15％ 18％ 16％ 13％ 11％ 0％ Cervical cancer 2％ 7％ 9％ 9％ 8％ 7％ 7％ 0％ Cholangiocarcinoma 3％ 10％ 15％ 16％ 18％ 16％ 14％ 0％ Esophageal cancer 4％ 15％ 17％ 17％ 16％ 14％ 16％ 0％ Floor of mouth cancer 3％ 9％ 8％ 10％ 9％ 8％ 7％ 0％ Nasopharyngeal carcinoma 7％ 10％ 11％ 9％ 9％ 10％ 9％ 0％ Normal cells 0％ 0％ 0％ 0％ 0％ 0％ 0％ 0％

[0060] Table 2, Table 3, Figures 1 to 3 The results showed that the seven compositions of Experiment 1 to Experiment 7 had good growth inhibition effects on lung cancer cells A549, gastric cancer cells GTL-16, liver cancer cells HLE, breast cancer cells HCC1937, colon cancer cells HCT 116, cervical cancer cells HeLa, bile duct cancer cells RBE, esophageal cancer cells TE-1, floor of mouth cancer cells HTX2346C and nasopharyngeal cancer cells IM-H136, and had almost no effect on human normal liver cells QSG 7701. In contrast, the seven compositions of Control 1 to Control 7 had significantly weakened growth inhibition effects on each cancer cell.

[0061] Morphological observation of cancer cells: Lung cancer cells A549, gastric cancer cells GTL-16, liver cancer cells HLE and human normal liver cells QSG 7701 were inoculated into a new 96-well plate. After culturing for 1 hour, the 14 compositions were added to the cultured cells respectively, and 0.1% DMSO was set as a negative control. The culture was continued for 72 hours, and the cells were collected, washed with pre-cooled phosphate buffer, stained with DAPI, and observed under a fluorescence microscope.

[0062] The results showed that the lung cancer cells A549, gastric cancer cells GTL-16, liver cancer cells HLE and human normal liver cells QSG 7701 in the negative control group all grew well. At the same time, the human normal liver cells QSG 7701 added with 14 kinds of compositions also grew well; in the lung cancer cells A549, gastric cancer cells GTL-16 and liver cancer cells HLE added with experiments 1 to 7, different degrees of nuclear rupture and chromosome condensation were observed, and apoptotic bodies were produced; relatively speaking, although a small amount of nuclear rupture occurred in controls 1 to 7, the degree was significantly lower than that in experiments 1 to 7.

[0063] Furthermore, flow cytometry was used to detect the apoptosis rates of lung cancer cells A549, gastric cancer cells GTL-16, liver cancer cells HLE, and human normal liver cells QSG 7701. The results were highly inhibitory to the inhibition rate results in Table 2, that is, experiments 1 to 7 could better promote the apoptosis of lung cancer cells A549, gastric cancer cells GTL-16 and liver cancer cells HLE, but had almost no effect on human normal liver cells QSG7701; relatively speaking, the apoptosis effects of controls 1 to 7 were significantly reduced.

[0064] The above experiments show that the anticancer composition composed of 70-88g of Angelica keiskei polypeptide microecological extract, 3.9-8.3g of anthraquinone compounds, 7-14g of radish seed extract, 0.5-2g of curcumin, 0.05-4.5g of resveratrol, 0.5-1g of fucoidan, and 0.05-0.2g of lycopene can effectively inhibit the growth of lung cancer cells, gastric cancer cells, liver cancer cells, breast cancer cells, colon cancer cells, cervical cancer cells, bile duct cancer cells, esophageal cancer cells, floor of mouth cancer cells and nasopharyngeal cancer cells, and promote their apoptosis; and it has almost no effect on normal human cells. Therefore, the anticancer composition of this experiment can be used to prepare drugs for treating lung cancer, gastric cancer, liver cancer, breast cancer, colon cancer, cervical cancer, bile duct cancer, esophageal cancer, floor of mouth cancer or nasopharyngeal cancer.

[0065] The above contents are further detailed descriptions of the present application in combination with specific implementation methods, and it cannot be determined that the specific implementation of the present application is limited to these descriptions. For ordinary technicians in the technical field to which the present application belongs, several simple deductions or substitutions can be made without departing from the concept of the present application.

Claims

1. An anticancer composition for inhibiting the growth of cancer cells, characterized in that: It is composed of tomorrow leaf polypeptide microecological extract, anthraquinone compounds, radish seed extract, curcumin, resveratrol, fucoidan and lycopene.

2. The anticancer composition according to claim 1, characterized in that: The Angelica keiskei polypeptide microecological extract accounts for 70-88% of the total weight, the anthraquinone compounds account for 3.9-8.3% of the total weight, the radish seed extract accounts for 7-14% of the total weight, the curcumin accounts for 0.5-2% of the total weight, the resveratrol accounts for 0.05-4.5% of the total weight, the fucoidan accounts for 0.5-1% of the total weight, and the lycopene accounts for 0.05-0.2% of the total weight.

3. The anticancer composition according to claim 1, characterized in that: The Angelica keiskei polypeptide microecological extract is prepared by the following method: 20-35 parts by weight of Angelica keiskei polypeptide, 15-30 parts by weight of pig blood peptide, 2-5 parts by weight of soybean lecithin, 6-12 parts by weight of complex vitamins, 6-12 parts by weight of complex minerals, 4-6 parts by weight of gentiooligosaccharides and 6-12 parts by weight of mannitol are uniformly mixed and crushed and sieved to obtain a mixed raw material; Adding 0.1-1 parts by weight of Bifidobacterium lactis and 0.1-1 parts by weight of Bacillus to the mixed raw materials, incubating at a constant temperature of 30-35° C. for 2-4 hours to obtain a microecological fermentation liquid; The microecological fermentation liquid is filtered to remove the residue and collect the filtrate; the filtrate is subjected to ultrasound at a power of 400-500w for 10-30min, and then centrifuged at 6000-8000rpm for 10-30min, and the supernatant is sterilized and matured to obtain the Angelica keiskei polypeptide microecological extract.

4. The anticancer composition according to claim 3, characterized in that: The aging treatment includes placing the sterilized supernatant in a sterilized aging barrel for sealed aging and storage, the sealed aging and storage temperature is 30-35° C., and the time is 1-3 weeks.

5. The anticancer composition according to any one of claims 1 to 4, characterized in that: The anthraquinone compounds are natural hydroxyanthraquinone compounds.

6. The anticancer composition according to claim 5, characterized in that: The natural hydroxyanthraquinone compound is rhamnosine and / or rhamnosine methyl ether.

7. The anticancer composition according to claim 6, characterized in that: The emodin and / or emodin methyl ether are extracted from Polygonum cuspidatum.

8. The anticancer composition according to any one of claims 1 to 4, characterized in that: The radish seed extract is an ethanol extract of radish seeds; Preferably, the preparation method of the ethanol extract of radish seeds comprises adding 10-30 times the weight of ethanol to radish seed powder, and extracting with ultrasound and heating assistance; then, removing the residue, collecting the filtrate, and removing the solvent in the filtrate to obtain the ethanol extract of radish seeds; Preferably, the particle size of the radish seed powder is less than 0.3 mm; Preferably, the ethanol is 95% ethanol; Preferably, the power of the ultrasound is 300-400w, the heating temperature is 40-60°C, and the extraction time is 20-60min.

9. Use of the anticancer composition according to any one of claims 1 to 8 in the preparation of a drug for treating cancer.

10. The use according to claim 9, characterized in that: The cancer includes at least one of lung cancer, stomach cancer, liver cancer, breast cancer, colon cancer, cervical cancer, bile duct cancer, esophageal cancer, floor of mouth cancer and nasopharyngeal cancer.

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