CN108403775B - Gorgon fruit shell extract for inhibiting cancer cell proliferation as well as preparation method and application thereof - Google Patents
Gorgon fruit shell extract for inhibiting cancer cell proliferation as well as preparation method and application thereof Download PDFInfo
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- CN108403775B CN108403775B CN201810439331.3A CN201810439331A CN108403775B CN 108403775 B CN108403775 B CN 108403775B CN 201810439331 A CN201810439331 A CN 201810439331A CN 108403775 B CN108403775 B CN 108403775B
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Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/62—Nymphaeaceae (Water-lily family)
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/105—Plant extracts, their artificial duplicates or their derivatives
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/30—Extraction of the material
- A61K2236/33—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones
- A61K2236/333—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones using mixed solvents, e.g. 70% EtOH
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/50—Methods involving additional extraction steps
- A61K2236/51—Concentration or drying of the extract, e.g. Lyophilisation, freeze-drying or spray-drying
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/50—Methods involving additional extraction steps
- A61K2236/55—Liquid-liquid separation; Phase separation
Abstract
The invention belongs to the technical field of natural product extraction, and relates to a gorgon fruit shell extract for inhibiting cancer cell proliferation and a preparation method and application thereof. The preparation method comprises the following steps: i) ultrasonic extracting the gordon euryale seed shell powder by an ultrasonic extractor to obtain an extracting solution; ii) purifying the extracting solution obtained in the step i) by silica gel column chromatography, heating, concentrating and freeze-drying to obtain the gordon euryale seed shell extract. The gordon euryale seed shell extract has obvious effect of inhibiting tumor cell proliferation, and has good application prospect in the aspect of developing natural cancer prevention and anticancer health-care food.
Description
Technical Field
The invention belongs to the technical field of natural product extraction, and particularly relates to a gorgon fruit shell extract for inhibiting cancer cell proliferation as well as a preparation method and application thereof.
Background
Gorgon fruit is a traditional food and also a good traditional Chinese medicine, and is recorded in compendium of materia medica: qian Shi quenches thirst and tonifies the kidney, and is used for treating urinary incontinence, spermatorrhea and leukorrhagia. It is listed in the list of 'medicinal and edible' plants published by Ministry of health of China.
At present, the research on the extracts and the biological activity of the gorgon fruit shell is less. Through reference of documents, chinese patent application publication No. CN105147781A discloses "an inflammation-inhibiting and allergy-resisting preparation containing gorgon fruit shell extract as main ingredient and its application", which relates to a preparation obtained by adding gorgon fruit shell extract with different concentrations into honeycomb water extract/vitamin C, and having an effect of relieving skin inflammation, especially allergic dermatitis. Chinese patent application with publication number CN104489147A discloses a gordon euryale seed tea and a preparation method thereof, which relates to a gordon euryale seed tea for reducing blood fat and blood sugar, which is obtained by stir-frying gordon euryale seed shells containing water after being crushed through gas explosion. The large amount of gorgon fruit shells generated in the industrial production is mostly directly discarded. At present, the extraction preparation of the effective components of the gordon euryale seed shell and the research on the biological activity are almost blank at home and abroad. The problem to be solved urgently is to develop and utilize the gorgon fruit shell resource, extract the functional components thereof, research the biological activity and apply the components.
Disclosure of Invention
The invention aims to provide a gorgon fruit shell extract and a preparation method and application thereof. The gorgon fruit shell extract is a gorgon fruit shell flavone extract, has an obvious effect of inhibiting tumor cell proliferation, and has a good application prospect in the aspect of developing natural cancer prevention and anticancer health-care foods.
In order to achieve the above object, a first aspect of the present invention provides a method for preparing a gorgon euryale seed shell extract for inhibiting cancer cell proliferation, comprising the steps of:
i) ultrasonic extracting the gordon euryale seed shell powder by an ultrasonic extractor to obtain an extracting solution;
ii) purifying the extracting solution obtained in the step i) by silica gel column chromatography, heating, concentrating and freeze-drying to obtain the gordon euryale seed shell extract.
According to a preferred embodiment of the invention, the method comprises the following steps:
1) drying, pulverizing and sieving semen euryales shell to obtain semen euryales shell powder;
2) mixing the gordon euryale seed shell powder with an ethanol solution, then carrying out ultrasonic extraction, and carrying out suction filtration and heating concentration to obtain an extracting solution;
3) adsorbing and eluting the extracting solution obtained in the step 2) through silica gel column chromatography, collecting eluent, heating, concentrating and freeze-drying the eluent to obtain the gordon euryale seed shell extract.
The gorgon fruit powder is crushed to facilitate subsequent extraction, and the sieving mesh number can be 30-50 meshes, for example.
In the present invention, the heating concentration in step 2) and step 3) is preferably performed under reduced pressure, and the temperature is preferably 40 to 50 ℃.
According to the present invention, preferably, the conditions of the ultrasonic extraction include: the ultrasonic power is 100-300w, the ultrasonic extraction time is 10-30min, the ultrasonic extraction temperature is 40-60 ℃, and the ultrasonic extraction is carried out twice.
According to the present invention, preferably, in the step 2), the ethanol solution is an ethanol aqueous solution with a volume fraction of 30% -90%; the weight ratio of the gordon euryale seed shell powder to the ethanol solution is 1: 5-30.
According to the present invention, preferably, in step 3), the adsorption conditions include: regulating the concentration of the extractive solution to 3.0-12.0mg/mL, the adsorption flow rate to 0.6-1.2 times of column volume/h, and the adsorption volume to 2-6 times of column volume. The elution conditions include: after adsorption equilibrium, 50-80% ethanol solution is used as eluent for elution, the elution flow rate is 0.8-2mL/min, and the volume of the eluent is 3-6 times of the column volume.
The concentration of the extracting solution is the concentration of soluble solids in the gorgon fruit shell extracting solution.
The second aspect of the present invention provides the gorgon euryale seed shell extract for inhibiting cancer cell proliferation prepared according to the above preparation method.
The third aspect of the invention provides an application of the gorgon fruit shell extract in preparing a product for inhibiting cancer cell proliferation.
Specifically, the cancer cell is selected from human gastric cancer SGC7901 cell, human liver cancer HepG2 cell and human cervical cancer Hela cell.
The product for inhibiting the proliferation of the cancer cells comprises a preparation and a medicine for inhibiting the proliferation of the cancer cells, and also comprises an anticancer and cancer-preventing health-care functional food.
Compared with the prior art, the invention has the following beneficial effects:
(1) the invention adopts the ultrasonic extraction method, can quickly destroy the cell wall of gordon euryale seed shell tissue, is more beneficial to the extraction of flavone components, greatly shortens the low-temperature extraction time and improves the yield.
(2) The preparation process is simple, the ultrasonic extraction is carried out, the silica gel column chromatography purification is adopted, the biological activity of the extract is kept as far as possible, the highest purity of the gordon euryale seed shell flavone can reach 42.5 percent through the enrichment of the silica gel column, the gordon euryale seed shell flavone has an obvious anti-tumor effect, and the purity of the gordon euryale seed shell flavone obtained by a conventional hot extraction method is only about 20 percent. The method also reduces the use of a large amount of organic reagents, and can be used for developing natural anticancer food.
(3) The extraction and concentration temperature of the gorgon fruit shell extract is controlled below 55 ℃, and the activity of effective components is more favorably maintained than high-temperature extraction.
Additional features and advantages of the invention will be set forth in the detailed description which follows.
Detailed Description
Preferred embodiments of the present invention will be described in more detail below. While the following describes preferred embodiments of the present invention, it should be understood that the present invention may be embodied in various forms and should not be limited by the embodiments set forth herein.
Example 1
Separation and preparation of gordon euryale seed shell extract
1. Preparing a gorgon euryale seed shell extract 1:
(1) extraction: mixing dried, crushed and 40-mesh-screened gordon euryale seed shells with a 50% ethanol solution according to a ratio of 1:20, placing the mixture in an ultrasonic extractor for ultrasonic extraction, wherein the ultrasonic extraction power is 100W, the ultrasonic extraction time is 20min, the extraction temperature is 50 ℃, extracting twice, combining filtrates of the two times, concentrating under reduced pressure at 45 ℃ until no ethanol exists to obtain a gordon euryale seed shell extracting solution, and taking a part of the mixture to freeze-dry to obtain an ultrasonic crude extract 1;
(2) adsorption: adjusting the concentration of the gordon euryale seed shell extracting solution to be 5.0mg/mL, and carrying out silica gel column chromatography; the adsorption flow rate is 0.8 times of the column volume/h, and the adsorption volume is 2.5 times of the column volume;
(3) and (3) elution: eluting with 70% ethanol solution as eluent at an elution flow rate of 1.0mL/min, wherein the volume of the eluent is 4 column volumes, and collecting the eluent;
(4) concentration: concentrating the eluate, concentrating under reduced pressure at 50 deg.C, and freeze drying to obtain semen euryales shell extract 1.
2. Preparing a gorgon euryale seed shell extract 2:
(1) extraction: mixing dried, crushed and 40-mesh-screened gordon euryale seed shells and 80% ethanol solution according to a ratio of 1:10, placing the mixture in an ultrasonic extractor for extraction, wherein the ultrasonic extraction power is 300W, the ultrasonic extraction time is 10min, the extraction temperature is 45 ℃, extracting twice, combining filtrates of the two times, concentrating under reduced pressure at 45 ℃ until no ethanol exists to obtain gordon euryale seed shell flavone extracting solution, and taking a part of the gordon euryale seed shells and freeze-drying to obtain an ultrasonic crude extract 2;
(2) adsorption: adjusting the concentration of the gordon euryale seed shell extracting solution to be 10.0mg/mL, and carrying out silica gel column chromatography; the adsorption flow rate is 1 time of column volume/h, and the adsorption volume is 5 times of column volume;
(3) and (3) elution: eluting with 60% ethanol solution as eluent at an elution flow rate of 1.5mL/min, wherein the volume of the eluent is 5 column volumes, and collecting the eluent;
(4) concentration: concentrating the eluate, concentrating under reduced pressure at 45 deg.C, and freeze drying to obtain semen euryales shell flavone extract 2.
3. Drawing of rutin standard curve
Accurately preparing 0.30mg/mL rutin solution in a 50mL volumetric flask. Accurately sucking the rutin standard solution 1.0mL, 2.0mL, 3.0mL, 4.0mL, 5.0mL, 6.0mL, 7.0mL, and 8.0mL, respectively placing into 25mL volumetric flasks, adding anhydrous ethanol to 12.5mL, and respectively adding 5% NaNO20.7ml of water solution is shaken up and then kept stand for 5 min; adding 10% Al (NO)3)30.7mL of aqueous solution, shaking up and standing for 6 min; respectively adding 5mL of L moL/L NaOH aqueous solution, adding absolute ethyl alcohol to 25mL, standing for 10min, measuring the absorbance value at 510nm by using an ultraviolet-visible spectrophotometer, and calculating a standard curve regression equation.
4. Determination of flavone content in sample
Accurately sucking 1.0mL of sample into a 25mL volumetric flask, adding 11.5mL of absolute ethyl alcohol, and shaking up; adding 5% NaNO respectively20.7mL of aqueous solution, shaking and standing for 5min, adding 10% Al (NO)3)30.7mL of aqueous solution, shaking up, standing for 6min, adding 5mL of 1mol/L NaOH aqueous solution, fixing the volume to the scale with absolute ethyl alcohol, shaking up, standing for 10min, making blank with absolute ethyl alcohol, and measuring with an ultraviolet-visible spectrophotometer at 510nmAnd measuring the absorbance value. Substituting into standard curve equation to calculate flavone concentration.
The content of flavone in the gordon euryale seed shell extract 1 extracted by the process is 42.5 percent. The content of flavone in the gorgon fruit shell extract 2 is 39.4%.
Comparative example 1
Mixing dried, crushed and 40-mesh-screened gorgon fruit shell in the preparation of the gorgon fruit shell extract 1 with 50% ethanol solution according to a ratio of 1:20, performing reflux extraction for 1 hour for two times, combining filtrates, concentrating under pressure at 45 ℃ until no ethanol exists, and performing freeze drying to obtain an ethanol reflux crude extract 1.
Mixing dried, crushed and 40-mesh-screened gorgon fruit shell in the preparation of the gorgon fruit shell extract 2 with 80% ethanol solution according to a ratio of 1:10, performing reflux extraction for 1h, performing reflux extraction twice, combining filtrates, performing pressure concentration at 45 ℃ until no ethanol exists, and performing freeze drying to obtain an ethanol reflux crude extract 2.
Test example 1
The extraction rate and purity of the crude extract obtained by ultrasonic extraction and conventional reflux extraction were determined as shown in table 1 below.
TABLE 1 comparison of the yield and purity of the crude extract of Gorgon fruit shell obtained by ultrasonic extraction and reflux extraction
| Sample (I) | Yield (%) | Purity (%) |
| Ultrasonic crude extract 1 | 8.43±0.86 | 21.32±0.96 |
| Ultrasonic crude extract 2 | 7.82±0.97 | 22.89±1.37 |
| Ethanol reflux crude extract 1 | 6.19±0.28 | 19.19±0.81 |
| Ethanol reflux crude extract 2 | 6.57±0.57 | 18.03±1.23 |
As can be seen from Table 1, the ultrasonic extraction method of the present invention can improve the yield, and in addition, the total time of the conventional reflux extraction time is generally more than 2h, while the ultrasonic extraction method of the present invention can be controlled within 1h, and the extraction time can be obviously shortened.
Test example 2
The CCK-8 method is used for detecting the influence of the gorgon fruit shell extract on the proliferation of different tumor cells.
Experimental materials:
sample preparation: the gorgon euryale seed shell extracts of this experiment were gorgon euryale seed shell extract 1 and gorgon euryale seed shell extract 2 prepared in example 1.
Reagent: human liver cancer HepG2 cell, human stomach cancer SGC7901 cell, human cervical carcinoma Hela cell, DMSO, Baiostoma Biotech; DMEM high sugar, F12 medium, PBS, Hyclone; pancreatin, hangzhou geno; 5-fluorouracil, sigma; fetal bovine serum, dual antibiotics, Hangzhou ilex bioengineering Co., Ltd; CCK-8 reagent, Japan Dojingzhan.
Equipment: t25 flasks, Coming, USA; 96-well plates, corning, usa; centrifuge tubes, Extragene, inc; a vertical pressure steam sterilizer, Shanghai Bocheng industries, Ltd; CJ-1F type medical purification workbench, Suzhou city Jinyan purification equipment Co., Ltd; model BX51 electron microscope, Olympus, japan; SCHP-80CO2 incubator, constant, Henshou constant instruments Limited manufacturing company; AXTD5A desk-top low speed centrifuge, salt city safety experiment instruments ltd; microplate reader, Thermo corporation, usa.
The test method comprises the following steps:
1. cell resuscitation
The cells frozen in the refrigerator at-80 ℃ are taken out, put into a water bath kettle at 37 ℃ for quick thawing, and the cells in the tube are gently blown by a pipette gun to suspend the cells. Preparing a required culture medium in a clean bench, centrifuging all cell suspensions in a centrifuge tube at 1000rpm for 3min, and discarding the supernatant. The cells were floated by gently pipetting in 1mL of complete medium, and a quantity of complete medium was added to the flask in T25, into which the cell suspension was pipetted. Turning the bottle cover of the culture bottle for a half circle after screwing the bottle cover, and putting the bottle cover at 37 ℃ and 5% CO2Culturing in an incubator. The following day the medium was changed.
2. Cell passage
When the degree of cell fusion reaches 90% or more, cell growth is affected by contact between cells, nutrient deficiency of the medium, and harmful metabolites, and thus, cell passaging is required. And (3) discarding the culture medium in the culture bottle, adding 2mL of PBS cleaning solution for cleaning, discarding the PBS cleaning solution, and cleaning again. And adding pancreatin for digestion for 1-3 min, and removing the pancreatin when the bottle wall is fogged. The digestion was stopped by adding 1mL of medium and the cells attached to the wall of the flask were blown down with a pipette and centrifuged in a centrifuge tube under the following conditions: 1000rpm, 3 min. Sucking out supernatant, adding a small amount of complete culture medium, gently blowing and beating to mix cells uniformly, transferring the cells into a proper amount of T25 culture bottles added with 4mL of culture medium according to the cell concentration, blowing and beating by a liquid transfer gun, and gently shaking the bottles to shake uniformly.
3. Cell plating
Adding 2mL of culture medium into human gastric cancer cells or liver cancer cells in logarithmic growth phase, blowing and beating uniformly, estimating and reading the cell suspension concentration by using a blood counting chamber, adding a proper amount of culture medium to dilute to the final concentration, and mixing uniformly. 100 μ L of the cell suspension was aspirated by pipette, and the cells were injected into each well and then blown out uniformly.
4. CCK-8 method for detecting cell viability
0.02g of the sample was weighed, added to 200. mu.L of DMSO solution for solubilization, and then 1800. mu.L of distilled water was added. The sample solution was prepared at 2000. mu.g/mL, sterilized by filtration through a 0.22 μm filter, and then diluted to 50. mu.g/mL, 100. mu.g/mL, 200. mu.g/mL, 400. mu.g/mL, 800. mu.g/mL for use. 5-FU uracil was used as a positive control.
Cell plating was performed the day before dosing and the density was adjusted to 4X 10 with complete medium4cells/mL, 100. mu.L of cell suspension was added per well in a 96-well plate. Duplicate wells were designed for each drug concentration and incubated for 24 hours. Adding medicine with corresponding concentration, using complete culture solution instead of sample solution as control group, taking another 5 wells, adding 200 μ L of complete culture solution as blank group, culturing for 24 hr, discarding sample solution in well plate, adding 200 μ L of complete culture medium, adding 10 μ L of CCK-8 reagent in dark place, adding 5% CO at 37 deg.C2Incubate for 3 hours and read the microplate reader at a wavelength of 450 nm. The cell proliferation inhibition rate was calculated as follows:
cell inhibition rate (%) [1- (OD experimental group-OD blank)/(OD control group-OD blank) ] × 100
Statistical analysis of the data was performed using Microsoft Excel2007, SPSS 4.0.
Test results
1. Effect of gorgon euryale seed shell extract on HepG2 tumor cell proliferation:
TABLE 2 proliferation inhibition of HepG2 by Gorgon fruit shell extract
TABLE 3 IC50 values for inhibition of HepG2 proliferation by Gorgon fruit shell extract
| Sample (I) | IC50(μg/mL) | R2 |
| Gorgon fruit shell extract 1 | 97.51 | 0.8411 |
| Gorgon fruit shell extract 2 | 102.34 | 0.7931 |
| 5-FU uracils | 18.42 | 0.9098 |
As can be seen from tables 2 and 3, both the Gorgon fruit shell extract 1 and the Gorgon fruit shell extract 2 have a certain inhibition effect on the proliferation of HepG2 cells, and the inhibition effect is weaker than that of the positive control 5-FU. The IC50 values of the gorgon fruit shell extract 1 and the gorgon fruit shell extract 2 are 97.51 mug/mL and 102.34 mug/mL respectively, which shows that the gorgon fruit shell extract has better effect of inhibiting the proliferation of liver cancer cells.
2. Influence of the gorgon fruit shell extract on proliferation of human gastric cancer SGC7901 tumor cells:
TABLE 4 proliferation inhibitory Effect of Gorgon fruit Shell extract on SGC7901
TABLE 5 IC50 value for inhibition of SGC7901 proliferation by Gorgon fruit Shell extract
| Sample name | IC50 | R2 |
| Gorgon fruit shell extract 1 | 177.86 | 0.9346 |
| Gorgon fruit shell extract 2 | 253..98 | 0.9648 |
| 5-FU uracils | 208.52 | 0.9922 |
As can be seen from tables 4 and 5, the Gorgon fruit shell extract 1 and the Gorgon fruit shell extract 2 both have certain inhibitory effects on the in vitro proliferation of human gastric cancer cell SGC 7901. The Gordon euryale seed shell extract 1 has an IC50 value of less than 5-FU for the inhibition rate of SGC7901 proliferation, is comparable to a positive control, and has a good effect of inhibiting SGC 7901.
3. Influence of the euryale ferox shell extract on proliferation of human cervical cancer Hela tumor cells:
TABLE 6 proliferation inhibiting effect of euryale ferox shell extract on human cervical carcinoma Hela cells
TABLE 7 IC50 value of Gorgon fruit shell extract for inhibiting human cervical carcinoma Hela cell proliferation
| Sample (I)Name (R) | IC50 | R2 |
| Gorgon fruit shell extract 1 | 237.86 | 0.9346 |
| Gorgon fruit shell extract 2 | 268.98 | 0.9648 |
| 5-FU uracils | 132.52 | 0.9922 |
As can be seen from tables 6 and 7, the gorgon fruit shell extract 1 and the gorgon fruit shell extract 2 both have certain inhibition effect on the proliferation of human cervical cancer Hela cells, and the inhibition effect is weaker than that of the positive control 5-FU uracil. The IC50 values of the gorgon fruit shell extract 1 and the gorgon fruit shell extract 2 are 237.86 mug/mL and 268.98 mug/mL respectively, which indicates that the gorgon fruit shell extract has certain effect of inhibiting the proliferation of liver cancer cells.
Having described embodiments of the present invention, the foregoing description is intended to be exemplary, not exhaustive, and not limited to the embodiments disclosed. Many modifications and variations will be apparent to those of ordinary skill in the art without departing from the scope and spirit of the described embodiments.
Claims (6)
1. The application of the gorgon fruit shell extract in preparing products for inhibiting cancer cell proliferation is characterized in that the preparation method of the gorgon fruit shell extract comprises the following steps:
1) drying, pulverizing and sieving semen euryales shell to obtain semen euryales shell powder;
2) mixing the gordon euryale seed shell powder with an ethanol solution, then carrying out ultrasonic extraction, and carrying out suction filtration and heating concentration to obtain an extracting solution; the ultrasonic extraction temperature is 40-60 ℃;
3) adsorbing and eluting the extracting solution obtained in the step 2) through silica gel column chromatography, collecting eluent, heating, concentrating and freeze-drying the eluent to obtain the gordon euryale seed shell extract;
the heating and concentration in the step 2) and the step 3) are carried out under reduced pressure, and the temperature is 40-50 ℃.
2. The use according to claim 1, wherein the conditions of ultrasound extraction comprise: the ultrasonic power is 100-300w, the ultrasonic extraction time is 10-30min, and the ultrasonic extraction is carried out twice.
3. The use according to claim 1, wherein in step 2), the ethanol solution is 30-90% ethanol water solution by volume fraction; the weight ratio of the gordon euryale seed shell powder to the ethanol solution is 1: 5-30.
4. Use according to claim 1, characterized in that, in step 3),
the adsorption conditions include: regulating the concentration of the extractive solution to 3.0-12.0mg/mL, the adsorption flow rate to 0.6-1.2 times of column volume/h, and the adsorption volume to 2-6 times of column volume.
5. Use according to claim 1, characterized in that, in step 3),
the elution conditions include: after adsorption equilibrium, 50-80% ethanol solution is used as eluent for elution, the elution flow rate is 0.8-2mL/min, and the volume of the eluent is 3-6 times of the column volume.
6. The use of claim 1, wherein the cancer cell is selected from the group consisting of human gastric cancer SGC7901 cell, human liver cancer HepG2 cell, human cervical cancer Hela cell.
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