CN117462658A - Marine low molecular weight polypeptide composition and preparation method and application thereof - Google Patents
Marine low molecular weight polypeptide composition and preparation method and application thereof Download PDFInfo
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Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/41—Porphyrin- or corrin-ring-containing peptides
-
- 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)
- A61K36/52—Juglandaceae (Walnut family)
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/04—Peptides having up to 20 amino acids in a fully defined sequence; Derivatives thereof
- A61K38/10—Peptides having 12 to 20 amino acids
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- A61K38/1767—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from invertebrates
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/0087—Galenical forms not covered by A61K9/02 - A61K9/7023
- A61K9/0095—Drinks; Beverages; Syrups; Compositions for reconstitution thereof, e.g. powders or tablets to be dispersed in a glass of water; Veterinary drenches
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
Abstract
The invention provides a marine low molecular weight polypeptide composition, a preparation method and application thereof, and relates to the technical field of natural medicines, wherein the composition mainly comprises the following raw materials in parts by weight: 20-30 parts of white ginseng polypeptide, 15-20 parts of cyclocarya paliurus leaf extract, 10-15 parts of conch polypeptide, 5-10 parts of oyster shell polypeptide and 5-10 parts of spirulina phycocyanin; the composition can be used for weak positive people in coarse screening of cancers, and experiments prove that the weak positive can be converted into negative by using the product, so that the effect of preventing cancers is achieved.
Description
[ field of technology ]
The invention relates to the technical field of natural medicines, in particular to a marine low molecular weight polypeptide composition, a preparation method and application thereof.
[ background Art ]
The diversity of marine organisms, the uniqueness of the marine environment and the novelty of the marine bioactive substance structure make the ocean a resource repository for innovative and functional/health food. Since the 70 s of the last century, tens of thousands of novel active substances have been isolated from marine organisms, including peptides, proteins, polysaccharides, alkaloids, terpenes, macrocyclic polyesters and the like. Among these, peptides are the most abundant class of compounds. It has been demonstrated that various marine peptides have physiological activities such as anti-tumor, anti-AIDS, antifungal, antiviral, and immunomodulating.
China has become a well-known "cancer kingdom" based on the global latest cancer burden data of 2020 issued by the world health organization international cancer research Institute (IARC) recently. 457 cases of new cancer in China in 2020, 248 ten thousand men, 209 ten thousand women, 300 ten thousand cases of cancer death in China in 2020, 182 ten thousand men and 118 ten thousand women. The world is first of new cases and deaths in China. The high mortality rate of cancer brings great loss, pain and medical burden to individuals and families, and more seriously consumes social resources. At present, the treatment cost for cancer patients in China is up to 1000 hundred million yuan each year, and the treatment cost accounts for 20% of the total national sanitary cost.
Early diagnosis and early treatment of tumors are key to improving tumor cure rate. The existing diagnostic means commonly used in clinic include chest radiography, B ultrasonic, CT, nuclear magnetic resonance and the like, which are often accompanied by the serious pain of patients such as puncture, blood drawing and the like and even the possibility of cross infection, and are expensive, more importantly, the tumors which can be detected by the means are generally in middle and late stages, and the cure rate is greatly reduced. Abnormal nucleotide metabolism in cancer cells produces a monohydroxyphenol-type metabolite in which the content of p-hydroxyphenylalanine is far higher than that of normal persons, and this substance can be discharged through urine. Therefore, the detection of the p-hydroxyphenylalanine in urine is a noninvasive, painless, rapid and convenient method for screening malignant tumors, and has important significance in early detection, early diagnosis, early treatment and prognosis judgment of malignant tumors. Studies have shown that the content of para-hydroxyphenylalanine and its metabolic derivatives in malignant tumors is significantly higher than in healthy and benign tumor patients; the sensitivity of the detection of the p-hydroxyphenylalanine and the metabolic derivatives thereof in diagnosing malignant tumors is 89.8%, the negative predictive value is 91.4%, the detection of the p-hydroxyphenylalanine has important value in screening malignant tumors (especially untreated malignant tumors), whether a human body has cancer can be deduced, the tumors can be found early, the lives are saved, the additional expense is not needed, and the fear and the pain of patients are avoided.
Based on the above, the application provides a composition which is researched and developed according to the modern medicine theory so as to achieve the effect of preventing the disease before the disease and reduce the occurrence of cancer diseases.
[ invention ]
In view of the above, the invention aims to provide a marine low molecular weight polypeptide composition, and a preparation method and application thereof, which have the advantages of simple process, easy operation and the like, and the marine low molecular weight polypeptide composition has positive significance for preventing cancers.
In order to solve the technical problems, the invention adopts the following technical scheme:
the marine low molecular weight polypeptide composition is characterized by mainly comprising the following raw materials in parts by weight: 20-30 parts of white ginseng polypeptide, 15-20 parts of cyclocarya paliurus leaf extract, 10-15 parts of conch polypeptide, 5-10 parts of oyster shell polypeptide and 5-10 parts of spirulina phycocyanin.
In the invention, further, the composition mainly comprises the following raw materials in parts by weight: 25 parts of white ginseng polypeptide, 17 parts of cyclocarya paliurus leaf extract, 13 parts of pearl snail polypeptide, 8 parts of oyster shell polypeptide and 8 parts of spirulina phycocyanin.
In the invention, the white ginseng polypeptide is 28064n, an island white ginseng polypeptide.
Specifically, the 280647-palea white ginseng polypeptide is obtained by the following method: pulverizing 28064Mr. Zhou island Ginseng radix alba, adding into NaCl solution with 15 times of weight and 10% concentration, maintaining temperature in 40deg.C water bath for 4 hr, filtering, retaining filtrate, adding filter residue with 8 times of weight, preserving heat in acetic acid solution with concentration of 0.5mol/L in water bath at 5deg.C for 6h, filtering, mixing the obtained filtrate with the filtrate to obtain protein extractive solution of radix Ginseng Indici of Paederia on the sea/island of 280647, regulating pH value of protein extract of the white ginseng of the Zhou island to 6.5, adding 10mg of trypsin per gram of the white ginseng of the Zhou island to 280642, carrying out enzymolysis reaction for 4 hours in a water bath at 60 ℃, taking out the white ginseng, carrying out enzyme deactivation treatment for 15 minutes in the water bath at 90 ℃, carrying out centrifugal separation to obtain supernatant, carrying out ultrafiltration through an ultrafiltration membrane with molecular cut-off of 3kDa and 1kDa, collecting solution with molecular weight of 1-3kDa, and carrying out freeze drying to obtain polypeptide of the white ginseng of the Zhou island of the\28064continent island; and determining the structure and isoelectric point of the polypeptide; the N-terminal sequence of the polypeptide of the white ginseng of the intercontinental island is shown as SEQ ID NO.1, specifically GGVGTDRIDTNMFEIQNTR, the molecular weight of the polypeptide is 2.12kDa, and the isoelectric point of the polypeptide is 8.90.
In the invention, the conch polypeptide is 280643 or a Severe island conch polypeptide.
Specifically, the 280647-plocalyx concha polypeptide is obtained by the following method: crushing the intercontinental island conch, adding deionized water with the weight of 12 times of the crushed intercontinental island conch, stirring uniformly to obtain 280642-day conch protein suspension slurry, regulating the pH value of the white ginseng suspension slurry to 7, adding 20mg of neutral protease per gram of 280642-day conch protein, carrying out enzymolysis reaction for 4 hours in a water bath with the temperature of 50 ℃, taking out, carrying out enzyme deactivation treatment for 15 minutes in the water bath with the temperature of 90 ℃, carrying out centrifugal separation to obtain supernatant, carrying out ultrafiltration by an ultrafiltration membrane with the molecular cut-off of 3kDa, collecting a solution with the molecular weight of not more than 3kDa, and carrying out freeze drying to obtain 280643-day conch polypeptide, and measuring the structure and isoelectric point of the polypeptide; the N-terminal sequence of the polypeptide of the intercontinental island conch is shown as SEQ ID NO.2, specifically GGVGTDRIDTNMFEIQNTR, the molecular weight is 3.0kDa, and the isoelectric point is 8.60.
In the invention, further, the oyster-in-the-river polypeptide is obtained by the following method: crushing oyster in the near river, adding deionized water 10 times the oyster weight, stirring uniformly, preserving heat in water bath at 80 ℃ for 20min, and cooling to obtain oyster pulp in the near river; adding 0.05mol/L hydrochloric acid to regulate the pH value of the oyster shell slurry to 5, adding 5mg of trypsin per gram of oyster shell, performing enzymolysis in a water bath at 40 ℃ for 3 hours, taking out, performing enzyme deactivation in the water bath at 90 ℃ for 15 minutes, performing centrifugal separation to obtain supernatant, performing ultrafiltration by an ultrafiltration membrane with molecular cut-off of 3kDa, collecting a solution with molecular weight not more than 3kDa, performing freeze drying to obtain oyster shell polypeptide, and determining the polypeptide structure and isoelectric point; the N-terminal sequence of the oyster in the river is shown as SEQ ID NO.3, specifically GEAPLKKLLGDHNAGLSKAGGLIPDTTDAVYIPRA, the molecular weight of the oyster in the river is 3.56kDa, and the isoelectric point of the oyster in the river is 8.85.
In the invention, further, the spirulina phycocyanin is high-purity spirulina phycocyanin, and the preparation method is as follows: the phycocyanin extract was further purified by gel chromatography column, and the adsorption column was rinsed with 0.2mol/L phosphate buffer solution having pH of 6.8. The sample solution was also assayed for phycocyanin purity a 620/a280=3.2-3.5. Finally, the obtained solution is subjected to nanofiltration, concentration and desalination, and spray drying is carried out, so that the spirulina phycocyanin with high purity can be obtained.
In the invention, the cyclocarya paliurus leaf extract is prepared by crushing cyclocarya paliurus leaves, extracting with hot water at 60-80 ℃ for three times, concentrating the obtained extract under reduced pressure at a feed-liquid ratio of 1:5-20, and freeze-drying under vacuum.
The invention also provides a preparation method of the composition, which comprises the following steps: adding white ginseng polypeptide powder, pearl snail polypeptide, oyster shell polypeptide, cyclocarya paliurus leaf extract and spirulina phycocyanin into 1000 parts by weight of water, stirring until the white ginseng polypeptide powder, the pearl snail polypeptide, the oyster shell polypeptide, the cyclocarya paliurus leaf extract and the spirulina phycocyanin are completely dissolved, and heating to 90 ℃ to obtain the composition.
The composition of the invention can be prepared into drink by adding auxiliary materials accepted in the preparation technology, wherein the auxiliary materials comprise honey, pectin and/or xylitol.
The composition is used for preparing a drink, and the specific method is as follows: adding cyclocarya paliurus leaf extract, spirulina phycocyanin, xylitol and pectin into water, stirring and dissolving completely, adding 280648/and Concha Ostreae polypeptide powder, stirring for 30min, heating to 80deg.C, maintaining the temperature for 10min, and packaging to obtain the beverage. The drink can be used for preventing tumors, wherein the tumors are lung cancer or colorectal cancer.
In summary, due to the adoption of the technical scheme, the beneficial effects of the invention are as follows:
the invention provides a clear-structured/280648-structured white ginseng polypeptide powder, a 280642-structured Zhoula pearl snail polypeptide, a oyster in the near river and the like for preparing a composition drink for preventing cancers, wherein the composition can be used for preventing cancers, and in the detection of the parahydroxyphenylalanine by using a national second-class medical instrument urine, the negative is normal, the positive is cancer, the weak positive is a certain amount of cancerous cells existing in the body, and the weak positive patient can be changed into the negative after using the composition, so the preparation method of the composition is simple, the components are safe, no side effect is caused, and the application prospect is very wide.
[ description of the drawings ]
FIG. 1 is a drawing showing the establishment of a mouse model;
in the figure, A is a schematic drawing of the collection of mouse tissue and blood at the end of each cycle; b is a cancer cell amplification process diagram of performing pathological section verification on the collected tissues; c is a comparison of normal mouse intestinal epithelium and tumor growth.
[ detailed description ] of the invention
The following examples will assist those skilled in the art in a more complete understanding of the invention, but are not intended to limit the invention in any way.
Example 1
The embodiment provides a marine low molecular weight polypeptide composition, which mainly comprises the following raw materials in parts by weight: 20 parts of an intercontinental white ginseng polypeptide, 15 parts of a cyclocarya paliurus leaf extract, 10 parts of an intercontinental pearl snail polypeptide, 5 parts of a oyster polypeptide and 5 parts of spirulina phycocyanin.
The 28064seven-leg white ginseng polypeptide is obtained by the following method: pulverizing 28064Mr. Zhou island Ginseng radix alba, adding into NaCl solution with 15 times of weight and 10% concentration, maintaining temperature in 40deg.C water bath for 4 hr, filtering, retaining filtrate, adding filter residue with 8 times of weight, preserving heat in acetic acid solution with concentration of 0.5mol/L in water bath at 5deg.C for 6h, filtering, mixing the obtained filtrate with the filtrate to obtain protein extractive solution of radix Ginseng Indici of Paederia on the sea/island of 280647, regulating pH value of protein extract of the white ginseng of the Zhou island to 6.5, adding 10mg of trypsin per gram of the white ginseng of the Zhou island to 280642, carrying out enzymolysis reaction for 4 hours in a water bath at 60 ℃, taking out the white ginseng, carrying out enzyme deactivation treatment for 15 minutes in the water bath at 90 ℃, carrying out centrifugal separation to obtain supernatant, carrying out ultrafiltration through an ultrafiltration membrane with molecular cut-off of 3kDa and 1kDa, collecting solution with molecular weight of 1kDa, and carrying out freeze-drying to obtain polypeptide of the white ginseng of the Zhou island of the\280643; and determining the structure and isoelectric point of the polypeptide; the N-terminal sequence of the 28064m white ginseng polypeptide is GGVGTDRIDTNMFEIQNTR, the molecular weight is 2.12kDa, and the isoelectric point is 8.90.
The 280642-site conch polypeptide is obtained by the following method: crushing the intercontinental island conch, adding deionized water with the weight of 12 times of the crushed intercontinental island conch, stirring uniformly to obtain 280642-day conch protein suspension slurry, regulating the pH value of the white ginseng suspension slurry to 7, adding 20mg of neutral protease per gram of 280642-day conch protein, carrying out enzymolysis reaction for 4 hours in a water bath with the temperature of 50 ℃, taking out, carrying out enzyme deactivation treatment for 15 minutes in the water bath with the temperature of 90 ℃, carrying out centrifugal separation to obtain supernatant, carrying out ultrafiltration by an ultrafiltration membrane with the molecular cut-off of 3kDa, collecting a solution with the molecular weight of not more than 3kDa, and carrying out freeze drying to obtain 280643-day conch polypeptide, and measuring the structure and isoelectric point of the polypeptide; the N-terminal sequence of the 28064m-Zhouabain conch polypeptide is GGVGTDRIDTNMFEIQNTR, the molecular weight is 3.0kDa, and the isoelectric point is 8.60.
The oyster polypeptide is obtained by the following method: crushing oyster in the near river, adding deionized water 10 times the oyster weight, stirring uniformly, preserving heat in water bath at 80 ℃ for 20min, and cooling to obtain oyster pulp in the near river; adding 0.05mol/L hydrochloric acid to regulate the pH value of the oyster shell slurry to 5, adding 5mg of trypsin per gram of oyster shell, performing enzymolysis in a water bath at 40 ℃ for 3 hours, taking out, performing enzyme deactivation in the water bath at 90 ℃ for 15 minutes, performing centrifugal separation to obtain supernatant, performing ultrafiltration by an ultrafiltration membrane with molecular cut-off of 3kDa, collecting a solution with molecular weight not more than 3kDa, performing freeze drying to obtain oyster shell polypeptide, and determining the polypeptide structure and isoelectric point; the N-terminal sequence of the oyster in the river is GEAPLKKLLGDHNAGLSKAGGLIPDTTDAVYIPRA, the molecular weight of the oyster in the river is 3.56kDa, and the isoelectric point of the oyster in the river is 8.85.
The spirulina phycocyanin is high-purity spirulina phycocyanin, and the preparation method comprises the following steps: the phycocyanin extract was further purified by gel chromatography column, and the adsorption column was rinsed with 0.2mol/L phosphate buffer solution having pH of 6.8. The sample solution was simultaneously assayed for phycocyanin purity a 620/a280=3.2. Finally, the obtained solution is filtered, concentrated and desalted by sodium, and spray-dried to obtain high-purity spirulina phycocyanin;
the cyclocarya paliurus leaf extract is obtained by crushing cyclocarya paliurus leaves, extracting with hot water at 60 ℃ for three times, wherein the ratio of feed to liquid is 1:5, concentrating the obtained extract under reduced pressure, and freeze-drying in vacuum.
Example 2
The embodiment provides a marine low molecular weight polypeptide composition, which mainly comprises the following raw materials in parts by weight: 25 parts of an intercontinental white ginseng polypeptide, 17 parts of a cyclocarya paliurus leaf extract, 13 parts of an intercontinental pearl snail polypeptide, 8 parts of a oyster polypeptide and 8 parts of spirulina phycocyanin.
The 28064seven-leg white ginseng polypeptide is obtained by the following method: pulverizing 28064Mr. Zhou island Ginseng radix alba, adding into NaCl solution with 15 times of weight and 10% concentration, maintaining temperature in 40deg.C water bath for 4 hr, filtering, retaining filtrate, adding filter residue with 8 times of weight, preserving heat in acetic acid solution with concentration of 0.5mol/L in water bath at 5deg.C for 6h, filtering, mixing the obtained filtrate with the filtrate to obtain protein extractive solution of radix Ginseng Indici of Paederia on the sea/island of 280647, regulating pH value of protein extract of the white ginseng of the Zhou island to 6.5, adding 10mg of trypsin per gram of the white ginseng of the Zhou island to 280642, carrying out enzymolysis reaction for 4 hours in a water bath at 60 ℃, taking out the white ginseng, carrying out enzyme deactivation treatment for 15 minutes in the water bath at 90 ℃, carrying out centrifugal separation to obtain supernatant, carrying out ultrafiltration through an ultrafiltration membrane with molecular cut-off of 3kDa and 1kDa, collecting solution with molecular weight of 2kDa, and carrying out freeze-drying to obtain polypeptide of the white ginseng of the Zhou island of the\280643; and determining the structure and isoelectric point of the polypeptide; the N-terminal sequence of the 28064m white ginseng polypeptide is GGVGTDRIDTNMFEIQNTR, the molecular weight is 2.12kDa, and the isoelectric point is 8.90.
The 280642-site conch polypeptide is obtained by the following method: crushing the intercontinental island conch, adding deionized water with the weight of 12 times of the crushed intercontinental island conch, stirring uniformly to obtain 280642-day conch protein suspension slurry, regulating the pH value of the white ginseng suspension slurry to 7, adding 20mg of neutral protease per gram of 280642-day conch protein, carrying out enzymolysis reaction for 4 hours in a water bath with the temperature of 50 ℃, taking out, carrying out enzyme deactivation treatment for 15 minutes in the water bath with the temperature of 90 ℃, carrying out centrifugal separation to obtain supernatant, carrying out ultrafiltration by an ultrafiltration membrane with the molecular cut-off of 3kDa, collecting a solution with the molecular weight of not more than 3kDa, and carrying out freeze drying to obtain 280643-day conch polypeptide, and measuring the structure and isoelectric point of the polypeptide; the N-terminal sequence of the 28064m-Zhouabain conch polypeptide is GGVGTDRIDTNMFEIQNTR, the molecular weight is 3.0kDa, and the isoelectric point is 8.60.
The oyster polypeptide is obtained by the following method: crushing oyster in the near river, adding deionized water 10 times the oyster weight, stirring uniformly, preserving heat in water bath at 80 ℃ for 20min, and cooling to obtain oyster pulp in the near river; adding 0.05mol/L hydrochloric acid to regulate the pH value of the oyster shell slurry to 5, adding 5mg of trypsin per gram of oyster shell, performing enzymolysis in a water bath at 40 ℃ for 3 hours, taking out, performing enzyme deactivation in the water bath at 90 ℃ for 15 minutes, performing centrifugal separation to obtain supernatant, performing ultrafiltration by an ultrafiltration membrane with molecular cut-off of 3kDa, collecting a solution with molecular weight not more than 3kDa, performing freeze drying to obtain oyster shell polypeptide, and determining the polypeptide structure and isoelectric point; the N-terminal sequence of the oyster in the river is GEAPLKKLLGDHNAGLSKAGGLIPDTTDAVYIPRA, the molecular weight of the oyster in the river is 3.56kDa, and the isoelectric point of the oyster in the river is 8.85.
The spirulina phycocyanin is high-purity spirulina phycocyanin, and the preparation method comprises the following steps: the phycocyanin extract was further purified by gel chromatography column, and the adsorption column was rinsed with 0.2mol/L phosphate buffer solution having pH of 6.8. The sample solution was simultaneously assayed for phycocyanin purity a 620/a280=3.3. Finally, the obtained solution is filtered, concentrated and desalted by sodium, and spray-dried to obtain high-purity spirulina phycocyanin;
the cyclocarya paliurus leaf extract is obtained by crushing cyclocarya paliurus leaves, extracting with hot water at 70 ℃ for three times, wherein the ratio of feed to liquid is 1:12, concentrating the obtained extract under reduced pressure, and freeze-drying in vacuum.
Example 3
This example provides a method of preparing a composition having the same compositional components as described in example 2;
the preparation method of the composition comprises the following steps:
adding white ginseng polypeptide powder, pearl snail polypeptide, oyster shell polypeptide, cyclocarya paliurus leaf extract and spirulina phycocyanin into 1000 parts by weight of water, stirring until the white ginseng polypeptide powder, the pearl snail polypeptide, the oyster shell polypeptide, the cyclocarya paliurus leaf extract and the spirulina phycocyanin are completely dissolved, and heating to 90 ℃ to obtain the composition.
Example 4
The embodiment provides a drink, which comprises the following raw materials in parts by weight: 25 parts of an intercontinental white ginseng polypeptide, 17 parts of a cyclocarya paliurus leaf extract, 13 parts of an intercontinental pearl snail polypeptide, 8 parts of a oyster polypeptide, 8 parts of spirulina phycocyanin, 50 parts of xylitol, 0.5 part of pectin and 1000 parts of water;
the raw materials are obtained in the same manner as in example 2, and the preparation method of the drink comprises the following steps: adding cyclocarya paliurus leaf extract, spirulina phycocyanin, xylitol and pectin into water, stirring and dissolving completely, adding 280648/and Concha Ostreae polypeptide powder, stirring for 30min, heating to 80deg.C, maintaining the temperature for 10min, and packaging to obtain the beverage.
Test examples
The applicant conducted the following tests to verify the practical value of the present application:
1.1 construction of a model of the occurrence of mouse lung cancer
Grouping and treating experimental animals: SPF class C57BL/6 mice of 8 weeks of age, each half male and female, 180 in total. After 2 weeks of post-purchase adaptation, mice were anesthetized with the gas anesthetic isoflurane, infected with air by inhalation tracheal instillation, and randomly divided into 3 groups: (1) 60 CTPE group mice were instilled with 50. Mu.L CTPE (VDMSO: V corn oil=1:4) solution, 1 mg/mouse, 1 time per week, and stopped after 4 weeks; (2) 60 mice in a solvent control (VC) group and an experimental group are instilled with a mixed liquid gas tube of equal volume of DMSO and corn oil; (3) normal Control (NC) group of 60 mice were not treated at all. Mice were kept under the same conditions, were free to eat, were observed for mental state and eating and were recorded.
1.2 staged viewing
Batch planing mice 3 months, 6 months, 9 months and 12 months after the first contamination, weighing the weight before planing, and anaesthetizing with 1% sodium pentobarbital, wherein 15 mice are planing each group; removing the plane, exposing the main vein of the mouse abdomen, taking blood, placing in a blood collection tube containing EDTA anticoagulant, centrifuging at 4deg.C and 1500r/min for 5min, collecting upper plasma, and storing at-80deg.C; the lung neoplasia conditions of the mice in each group are observed, the sizes of the mice are measured by a vernier caliper, and the macroscopic neoplasia number of the lungs of the mice in each stage is counted: tumor number = total number of lung tumors per group of mice/total number of mice in the group; the tumor formation rate of each stage of the mice is calculated by the following formula: lung neoplasia rate (%) = number of mice observed to develop tumors per group x 100% of total mice in the group.
1.3 mouse base case
After CTPE trachea instillation, the mice are in listlessness, appetite is slightly reduced, food intake is reduced, activity is reduced, after a period of time, the mice are relieved, and the mental state, the food intake, the activity and the like of the mice are restored to be normal; throughout the experiment, the differences in body weight of the mice in each group were not statistically significant (P > 0.05).
1.4 Coke tar pitch smoke extract induces the occurrence of pulmonary tumors in mice
Mice were dissected in batches 3 months and 6 months after the end of the first contamination, and the lung tumor formation of the mice was observed. Macroscopic tumor bodies are observed in the lungs of mice 6 months after CTPE infection, mostly have a plurality of tumor bodies and a few tumor bodies have single tumor bodies, and the tumor bodies are round, grey-white and semitransparent and are clear in boundary with surrounding tissues. No macroscopic tumor mass was observed at each stage after contamination in the normal control group and the solvent control group. No macroscopic tumor was observed in CTPE group at 3 months, whereas the tumor formation rate at 6 months after contamination was 26.67%, tumor sizes were unequal, and diameters were less than 4mm. Pathological results show that after CTPE contamination, mucus secretion and massive inflammatory cell infiltration are observed in early lung tissues, whereas inflammatory changes are significantly attenuated in late stages; a large amount of papillary hyperplasia was observed at 6 months. Lung cancer types are mainly lung adenocarcinoma, lung squamous carcinoma and lung adenosquamous carcinoma; in the whole experimental process, 14 cases of adenocarcinoma, 6 cases of squamous carcinoma and 6 cases of adenosquamous carcinoma are found in the CTPE group, wherein the lung adenocarcinoma accounts for 53.85 percent, the lung squamous carcinoma accounts for 23.08 percent and the lung adenosquamous carcinoma accounts for 23.07 percent.
1.5 testing of the fluid drink of the present application for negative-turning Activity on mice Pre-colonization with Lung tumors (Weak Positive)
The mice 1 month after the end of the contamination were used to set up test groups of 8 mice, each half of the male and female mice were filled with 0.5mL of the drink described in example 4 each day, and urine was collected on day 0, day 10, day 20, and day 30 and day 40 after the filling. By using the p-hydroxyphenylalanine kit for detection, the detection result shows that on the 0 th day, all the tested groups are weak positive, and gradually turn into negative along with the progress of gastric lavage, and the specific table is shown below.
2.1 establishing an AOM/DSS mouse colorectal cancer model
BALB/c mice were selected for culture, the mice were divided into normal (PBS) and colorectal cancer (AOM/DSS) groups, and after single dose (12.5 mg/kg) of AOM was intraperitoneally injected, 1 week later, fed with 2.5% DSS in water for 7 days, and then changed to normal drinking for 14 days, 1 cycle, 3 cycles altogether, and different cycles of urine and tissue were collected from the mice. Urine collected with a sterile tube was frozen at-20℃and after fixing organs and tumor tissues with formalin solution, the tissues were immersed in Phosphate Buffer (PBS) overnight, dehydrated and embedded, and then prepared into 5 μm thick sections. And (3) putting the slices into an oven to bake until paraffin melts (60 ℃), taking out, taking the slices according to the traditional dyeing steps, dewaxing, dyeing, dehydrating and sealing the slices in sequence, and observing histopathology under a microscope.
2.2 model building
Mice were divided into PBS and AOM groups, and after single dose (12.5 mg/kg) of AOM was intraperitoneally injected, 1 week later on with 2.5% dss in water for 7 days, and then normal water was changed to 1 cycle for 14 days, and total cycles were 2 times, and mouse tissues and blood were collected at the end of each cycle (see fig. 1A). The collected tissues are subjected to pathological section verification, so that the rectum tissues of the mice can be seen to be changed, cancer cells are continuously amplified (see fig. 1B), and the tumor of the intestinal epithelium of the mice can be directly observed to grow from fig. 1C, so that the success of model construction is demonstrated.
2.3 testing of the beverage of the present application for transvaginal Activity in mice with colorectal Pre-tumor (Weak Positive)
The mice in the 1 st period after the end of the contamination were used as the test group, 8 mice were used as the test group, and each half of the mice was filled with 0.5mL of the composition described in example 3 every day, and urine was collected on the 0 th, 10 th, 20 th, and 30 th days and 40 th days after the filling. By using the p-hydroxyphenylalanine kit for detection, the detection result shows that on the 0 th day, all the tested groups are weak positive, and gradually turn into negative along with the progress of gastric lavage, and the specific table is shown below.
The applicant carried out the test by replacing the above composition with the drink obtained in example 4, and could obtain the same test result, the drink being relatively acceptable to the volunteer in terms of taste, and therefore the test was carried out by replacing the drink in the following test.
3.1 selection criteria
The volunteers who are collected to participate in the study, on the basis of the primary screening, any single index in 14 examinations of tumor markers in the hospital is raised to be more than 2 times of the normal value, and no solid tumor is generated as a healthy subject of the study.
3.2 inclusion criteria
(1) A male or female meeting the above criteria; (2) non-basic diseases such as diabetes, AIDS, hypertension, etc.; (3) signing an informed consent form; (4) the detection is a weak positive population.
3.3 sample
In order to ensure consistency, the same batch of testers are adopted, 100ml of the drink described in the embodiment 4 of the application is taken every day, a fresh clean urine sample of the testers in the early morning is collected every 7 days, fresh urine is sucked by a urine suction tube, a drop of urine is added into a test tube, the color of the test tube and a color chart is observed within 10 minutes, and the test results are shown in the following table.
In summary, due to the adoption of the technical scheme, the beneficial effects of the invention are as follows:
the invention provides a clear-structured/280648-structured white ginseng polypeptide powder, a 280642-structured Zhoula pearl snail polypeptide, a oyster in the near river and the like for preparing a composition drink for preventing cancers, wherein the composition can be used for preventing cancers, and in the detection of the parahydroxyphenylalanine by using a national second-class medical instrument urine, the negative is normal, the positive is cancer, the weak positive is a certain amount of cancerous cells existing in the body, and the weak positive patient can be changed into the negative after using the composition, so the preparation method of the composition is simple, the components are safe and have no side effect, and the application prospect is very wide.
While the invention has been described in detail in the foregoing general description and with reference to specific embodiments thereof, it will be apparent to one skilled in the art that modifications and improvements can be made thereto. Accordingly, such modifications or improvements may be made without departing from the spirit of the invention and are intended to be within the scope of the invention as claimed.
Claims (10)
1. The marine low molecular weight polypeptide composition is characterized by mainly comprising the following raw materials in parts by weight: 20-30 parts of white ginseng polypeptide, 15-20 parts of cyclocarya paliurus leaf extract, 10-15 parts of conch polypeptide, 5-10 parts of oyster shell polypeptide and 5-10 parts of spirulina phycocyanin.
2. The composition according to claim 1, characterized in that it consists essentially of the following raw materials in parts by weight: 25 parts of white ginseng polypeptide, 17 parts of cyclocarya paliurus leaf extract, 13 parts of pearl snail polypeptide, 8 parts of oyster shell polypeptide and 8 parts of spirulina phycocyanin.
3. The composition of claim 1, wherein the white ginseng polypeptide is a 28064n-type white ginseng polypeptide, the N-terminal sequence of the 28064n-type white ginseng polypeptide is shown in SEQ ID No.1, the molecular weight is 2.12KDa, and the isoelectric point is 8.90.
4. The composition of claim 1, wherein the conch polypeptide is a 28064n-type conch polypeptide, the N-terminal sequence of the 28064n-type conch polypeptide is shown in SEQ ID No.2, the molecular weight is 3.0KDa, and the isoelectric point is 8.60.
5. The composition of claim 1, wherein the oyster shell polypeptide has an N-terminal sequence shown in SEQ ID NO.3, a molecular weight of 3.56kDa and an isoelectric point of 8.85.
6. A composition according to claim 1, wherein the purity of the spirulina phycocyanin is a620/a280 = 3.2-3.5.
7. The composition of claim 1, wherein the cyclocarya paliurus leaf extract is obtained by pulverizing cyclocarya paliurus leaves, extracting with hot water at 60-80deg.C for three times at a feed liquid ratio of 1:5-20, concentrating under reduced pressure, and vacuum freeze drying.
8. A process for preparing the composition according to claim 1 of any one of claims 1 to 7, characterized in that it comprises: adding radix Ginseng alba polypeptide powder, margarita polypeptide, concha Ostreae polypeptide, cyclocarya paliurus leaf extract and spirulina phycocyanin into water, stirring to dissolve completely, and heating to 90deg.C to obtain the final product.
9. Use of a composition according to any one of claims 1-7 for the preparation of a beverage for the prevention of tumors.
10. Use of a composition according to any one of claims 1-7 for the preparation of a prophylactic tumour medicament.
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