Application of ziziphus jujube neutral polysaccharide

Abstract

The invention discloses application of ziziphus jujube neutral polysaccharide. In-vitro anti-tumor activity determination on the polysaccharide proves that the neutral polysaccharide of the ziziphus jujuba has an excellent inhibition effect on tumor cells, so that the neutral polysaccharide of the ziziphus jujuba can show anti-tumor activity, and the application of the neutral polysaccharide of the ziziphus jujuba in preparation of a raw material for preparing anti-tumor drugs is provided. The tumor cells are Hela human cervical cancer cells and HGC-27 human gastric cancer cells. The polysaccharide provides a good foundation for application in the industries of medicines, health care products and food.

Description

Technical Field

[0001] The invention belongs to the field of anti-tumor drugs and relates to the use of gray jujube neutral polysaccharide. Background Art

[0002] Tumor is one of the malignant diseases that endanger human health. Although many anticancer drugs used clinically have certain inhibitory and killing effects on cancer cells, they also cause certain damage to normal human cells, especially the human immune system, causing a variety of complications. On the other hand, during treatment, tumors gradually develop multidrug resistance to anticancer drugs, which is also an important reason for the poor effect of tumor treatment. The results of polysaccharide research show that many polysaccharides have immunomodulatory effects, and can enhance the anticancer effect when used in combination with radiotherapy and chemotherapy drugs. Plant polysaccharides have no obvious toxic side effects, so they are expected to become one of the ingredients of tumor treatment drugs, which lays the foundation for the high-value utilization of plant polysaccharides.

[0003] Polysaccharides are natural high-molecular carbohydrates composed of at least 10 monosaccharides linked by glycosidic bonds. They are widely found in animals, plants, microorganisms, seaweed and other organisms. Plant polysaccharides have a unique structure-activity relationship due to their complex structure, special structure and biological activity. Studies have shown that polysaccharides have biological activity in anti-tumor, anti-inflammatory, anti-viral, hypoglycemic, anti-aging, and immune promotion. In addition, natural plant polysaccharides are relatively safe for the human body, with high absorption and utilization rates. Therefore, with the development of molecular biology, the various physiological functions of polysaccharides have gradually been valued by people, especially the function of enhancing immunity has been widely recognized.

[0004] Ziziphus Jujuba Mill. is the fruit of the plant of the genus Ziziphus of the family Rhamnaceae. It is native to China and has a history of more than 4,000 years. More than 700 varieties have been found in China, especially the planting area and output of Xinjiang red dates, which currently rank first in the country. Modern research has found that the gray date contains a variety of nutrients and active substances such as carbohydrates, proteins, alkaloids, triterpenes, saponins, flavonoids, cyclic adenosine monophosphate, cyclic guanosine monophosphate, amino acids, vitamins, minerals, etc. Among them, polysaccharides are one of the main components with functional activity in the gray date, and have multiple biological activities such as antioxidant, immunomodulatory, anti-tumor, hypoglycemic, hypolipidemic and cavity flora regulation, which indicates that the gray date polysaccharide has a high application value in the fields of functional food, medical care, etc. However, there is no research report on the extraction and purification of a neutral part polysaccharide from the gray date and its inhibitory effect on Hela human cervical cancer cells and HGC-27 human gastric cancer cells. Therefore, the present invention prepares a neutral part polysaccharide from gray jujube and exhibits strong inhibitory activity against two tumor cells, providing a new way for the efficient utilization and development of Xinjiang gray jujube resources. Summary of the invention

[0005] The purpose of the present invention is to provide a use of neutral polysaccharide of gray jujube. Experiments show that gray jujube polysaccharide has excellent anti-tumor activity and is used as a raw material in the preparation of anti-tumor drugs. The tumors are Hela human cervical cancer cells and HGC-27 human gastric cancer cells. It provides a good basis for the application in the pharmaceutical, health care and food industries.

[0006] The invention discloses use of the neutral polysaccharide of jujube as a raw material in preparing anti-tumor drugs.

[0007] The anti-tumor cells are Hela human cervical cancer cells and HGC-27 human gastric cancer cells.

[0008] The use of the neutral polysaccharide of gray jujube described in the present invention, and the preparation method of the neutral polysaccharide of gray jujube involved are carried out according to the following steps:

[0009] a. Hot water extraction: remove the core of the selected gray jujube medicinal material, freeze-dry and grind to obtain gray jujube powder, add water at a solid-liquid ratio of 1:20, extract twice in water at a temperature of 60-90°C for 2.5-3.5 hours, filter and combine the clear liquid to obtain the total extract;

[0010] b. Concentrating the extract: Concentrate the total extract obtained in step a at 50-60° C. under reduced pressure to a volume of 1 / 4-1 / 6 of the total extract;

[0011] c. Alcohol precipitation treatment: the concentrated solution obtained in step b is precipitated with edible alcohol at a volume ratio of 1:3-5 and left overnight to obtain a crude polysaccharide precipitate of gray jujube;

[0012] d. washing the precipitate obtained in step c with edible alcohol and freeze-drying to obtain crude polysaccharide;

[0013] e. The crude polysaccharide obtained in step d was redissolved in water, and chromatographed on a DEAE-650M ion exchange chromatography column, eluted with purified water and 0.4 mol / L NaCl solution in sequence, and the portion with purified water as the eluent was collected. The purified water was dialyzed with a dialysis bag with a molecular weight cutoff of 1 kDa for 48 h. The portion in the dialysis bag was freeze-dried to obtain the neutral polysaccharide portion of gray jujube. BRIEF DESCRIPTION OF THE DRAWINGS

[0014] Figure 1 This is the elution curve of the jujube polysaccharide from DEAE-650M ion exchange chromatography column;

[0015] Figure 2 The inhibitory effect of the gray jujube polysaccharide of the present invention on Hela human cervical cancer cells;

[0016] Figure 3 The present invention shows the inhibitory effect of the gray jujube polysaccharide on HGC-27 human gastric cancer cells. DETAILED DESCRIPTION

[0017] In order to further understand the present invention, the present invention is further described in detail below in conjunction with the embodiments, and the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments.

[0018] Example 1

[0019] a. Hot water extraction: remove the core of the selected gray jujube medicinal material, freeze-dry and grind to obtain 10g of gray jujube powder, add water at a solid-liquid ratio of 1:20, extract twice in water at a temperature of 85°C for 3 hours, filter and combine the clear liquid to obtain the total extract;

[0020] b. Concentration of the extract: Concentrate the total extract obtained in step a under reduced pressure at 55° C. to a concentrated solution having a volume of 1 / 4 of the total extract;

[0021] c. Alcohol precipitation treatment: the concentrated solution obtained in step b was precipitated with edible alcohol at a volume ratio of 1:4 overnight, and the precipitate was centrifuged to obtain gray jujube polysaccharide precipitate;

[0022] d. Wash the precipitate obtained in step c with edible alcohol and freeze-dry to obtain a crude extract; that is, 0.51 g of crude sample of gray jujube polysaccharide, and the polysaccharide extraction rate is 5.1%; the analysis results show that the polysaccharide content of the extract is 84.6%;

[0023] e. The crude polysaccharide extract obtained in step d was redissolved in water, and chromatographed on a DEAE-650M ion exchange chromatography column, eluted with purified water and 0.4 mol / L NaCl solution in sequence, and the portion with purified water as the eluent was collected. The purified water was dialyzed with a dialysis bag with a molecular weight cutoff of 1 kDa for 48 h. The portion in the dialysis bag was freeze-dried to obtain the neutral polysaccharide portion of gray jujube.

[0024] Example 2

[0025] a. Hot water extraction: remove the core of the selected gray jujube medicinal material, freeze-dry and grind to obtain gray jujube powder, add water at a solid-liquid ratio of 1:20, extract twice in water at a temperature of 60°C for 2.5 hours, filter and combine the clear liquid to obtain the total extract;

[0026] b. Concentrating the extract: Concentrate the total extract obtained in step a under reduced pressure at 50° C. to a concentrated solution having a volume of 1 / 6 of the total extract;

[0027] c. Alcohol precipitation treatment: the concentrated solution obtained in step b was precipitated with edible alcohol at a volume ratio of 1:5 overnight to obtain gray jujube polysaccharide precipitate;

[0028] d. Wash the precipitate obtained in step c with edible alcohol and freeze-dry to obtain a crude extract; that is, 0.40 g of the crude sample of gray jujube polysaccharide, and the polysaccharide extraction rate is 4.0%;

[0029] e. The crude polysaccharide extract obtained in step d was redissolved in water, and chromatographed on a DEAE-650M ion exchange chromatography column, eluted with purified water and 0.4 mol / L NaCl solution in sequence, and the portion with purified water as the eluent was collected. The purified water was dialyzed with a dialysis bag with a molecular weight cutoff of 1 kDa for 48 h. The portion in the dialysis bag was freeze-dried to obtain the neutral polysaccharide portion of gray jujube.

[0030] Example 3

[0031] a. Hot water extraction: remove the core of the selected gray jujube medicinal material, freeze-dry and grind to obtain gray jujube powder, add water at a solid-liquid ratio of 1:20, extract twice in water at a temperature of 90°C for 3.5 hours, filter and combine the clear liquid to obtain the total extract;

[0032] b. Concentration of the extract: Concentrate the total extract obtained in step a under reduced pressure at 60° C. to a concentrated solution having a volume of 1 / 5 of the total extract;

[0033] c. Alcohol precipitation treatment: the concentrated solution obtained in step b was precipitated with edible alcohol at a volume ratio of 1:4 overnight to obtain gray jujube polysaccharide precipitate;

[0034] d. Wash the precipitate obtained in step c with edible alcohol and freeze-dry to obtain a crude extract; that is, 0.45 g of crude sample of gray jujube polysaccharide, and the polysaccharide extraction rate is 4.5%;

[0035] e. The crude polysaccharide extract obtained in step d was redissolved in water, and chromatographed on a DEAE-650M ion exchange chromatography column, eluted with purified water and 0.4 mol / L NaCl solution in sequence, and the portion with purified water as the eluent was collected. The purified water was dialyzed with a dialysis bag with a molecular weight cutoff of 1 kDa for 48 h. The portion in the dialysis bag was freeze-dried to obtain the neutral polysaccharide portion of gray jujube.

[0036] Example 4

[0037] a. Hot water extraction: remove the core of the selected gray jujube medicinal material, freeze-dry and grind to obtain gray jujube powder, add water at a solid-liquid ratio of 1:20, extract twice in water at a temperature of 70°C for 3 hours, filter and combine the clear liquid to obtain the total extract;

[0038] b. Concentration of the extract: Concentrate the total extract obtained in step a under reduced pressure at 60° C. to a concentrated solution having a volume of 1 / 4 of the total extract;

[0039] c. Alcohol precipitation treatment: the concentrated solution obtained in step b was precipitated with edible alcohol at a volume ratio of 1:3 overnight to obtain gray jujube polysaccharide precipitate;

[0040] d. Wash the precipitate obtained in step c with edible alcohol and freeze-dry to obtain a crude extract; that is, 0.41 g of crude sample of gray jujube polysaccharide, and the polysaccharide extraction rate is 4.1%;

[0041] e. The crude polysaccharide extract obtained in step d was redissolved in water, and chromatographed on a DEAE-650M ion exchange chromatography column, eluted with purified water and 0.4 mol / L NaCl solution in sequence, and the portion with purified water as the eluent was collected. The purified water was dialyzed with a dialysis bag with a molecular weight cutoff of 1 kDa for 48 h. The portion in the dialysis bag was freeze-dried to obtain the neutral polysaccharide portion of gray jujube.

[0042] Example 5

[0043] Detection of the effect of polysaccharides from the neutral part of gray dates on the survival rate of Hela human cervical cancer cells:

[0044] 2.5×10 3 Hela human cervical cancer cells were inoculated in 96-well plates (100 μL / well) and cultured for 24 h. Different concentrations of 0.094 mg / mL, 0.189 mg / mL, 0.375 mg / mL, 0.750 mg / mL, 1.5 mg / mL, 3.0 mg / mL and 6.0 mg / mL of neutral polysaccharide of jujube were added to the 96-well plates and treated for 48 h. The cell viability was determined by MTT method. The results are shown in Tables 1 and Figure 2 As shown:

[0045] Table 1 shows the effects of the neutral polysaccharide of gray jujube on the survival rate of tumor cells Hela and HGC-27

[0046]

[0047] from Figure 2 It can be seen that the neutral polysaccharide of gray jujube at a concentration of 0.094-6.0 mg / mL affects the vitality of Hela human cervical cancer cells. The inhibition rate corresponding to the neutral polysaccharide concentration of gray jujube of 0.094-6 mg / mL is 39.01-67.07%. When the concentration of neutral polysaccharide of gray jujube is greater than and equal to 0.375 mg / mL, the inhibition rate exceeds 50%. The inhibition rates of neutral polysaccharides of gray jujube at concentrations of 0.094 mg / mL and 0.189 mg / mL on the vitality of Hela human cervical cancer cells are both less than 50%. This result shows that the neutral polysaccharide part of gray jujube has the effect of inhibiting the growth of tumor cells Hela human cervical cancer cells.

[0048] Example 6

[0049] Detection of the survival rate of HGC-27 human gastric cancer cells by polysaccharides from the neutral part of gray dates:

[0050] 3.0×10 3 HGC-27 human gastric cancer cells were inoculated in 96-well plates (100 μL / well) and cultured for 24 h. Different concentrations of 0.189 mg / mL, 0.375 mg / mL, 0.750 mg / mL, 1.50 mg / mL, 3.0 mg / mL and 6.0 mg / mL of neutral polysaccharide of jujube were added to the 96-well plates and treated for 48 h. Cell viability was determined by MTT method. The results are shown in Tables 1 and Figure 3 As shown:

[0051] from Figure 3 It can be seen that 0.189-6.0 mg / mL of gray jujube neutral polysaccharide has an effect on the vitality of HGC-27 human gastric cancer cells. The inhibition rate corresponding to the gray jujube neutral polysaccharide concentration of 0.189 mg / mL-6 mg / mL is 44.45%-67.30%, among which when the gray jujube neutral polysaccharide concentration is greater than 1.5 mg / mL, the inhibition rate exceeds 50%, and the gray jujube polysaccharide concentrations of 0.189 mg / mL, 0.375 mg / mL, and 0.750 mg / mL have an inhibition rate on the vitality of HGC-27 human gastric cancer cells that is less than 50%. The results show that the gray jujube neutral polysaccharide part has the effect of inhibiting the growth of tumor cells HGC-27 human gastric cancer cells.

Claims

1. Use of neutral polysaccharide of jujube as a raw material in the preparation of anti-tumor drugs.

2. The use according to claim 1, characterized in that: The anti-tumor cells are Hela human cervical cancer cells and HGC-27 human gastric cancer cells.

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