CN114671924A - Periplaneta americana polypeptide extract and application thereof in oral cleaning product - Google Patents
Periplaneta americana polypeptide extract and application thereof in oral cleaning product Download PDFInfo
- Publication number
- CN114671924A CN114671924A CN202210297208.9A CN202210297208A CN114671924A CN 114671924 A CN114671924 A CN 114671924A CN 202210297208 A CN202210297208 A CN 202210297208A CN 114671924 A CN114671924 A CN 114671924A
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- China
- Prior art keywords
- extract
- periplaneta americana
- parts
- purified water
- polypeptide
- Prior art date
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- C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
- C07K1/14—Extraction; Separation; Purification
- C07K1/36—Extraction; Separation; Purification by a combination of two or more processes of different types
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Abstract
The invention relates to the fields of medicines, medical instruments and daily chemical products, in particular to a periplaneta americana polypeptide extract for diminishing inflammation, easing pain and repairing mucosa and application thereof in a mouth cleaning product. The periplaneta americana polypeptide extract is an active polypeptide directly obtained after being treated by a biological activity bionic affinity purification technology on the basis of a periplaneta americana extract crude product, has the advantages of safety, high purity, less impurities, high stability and strong activity compared with the traditional periplaneta americana extract crude product, has an obvious repairing effect on oral mucosa, and provides a new material basis for developing oral medicaments, medical instruments, daily chemical products and the like.
Description
Technical Field
The invention relates to the fields of medicines, medical instruments and daily chemical products, in particular to a periplaneta americana polypeptide extract for diminishing inflammation, easing pain and repairing mucosa and application thereof in a mouth cleaning product.
Background
Periplaneta americana (Periplaneta americana) belongs to the class Insecta, the order Blattaria, the family Blattaceae, and has survived for 3.5 hundred million years on earth, being one of the strongest, oldest, and by far the most successful groups of insects in the world. The earliest existing pharmaceutical monograph in China is recorded in Shen nong Ben Cao Jing, and the Chinese patent publication states that the Chinese patent publication states is salty and cold, mainly causes blood stasis, is hard in cold and heat, breaks accumulation, has sore throat and is cold in the interior, and belongs to a Chinese medicament which can treat diseases and nourish the body. Modern pharmacological studies show that the periplaneta americana has obvious effects of promoting tissue repair and regeneration, resisting tumors and hepatitis (2007, 32:2326-2331, a Chinese traditional medicine).
In the prior art, the periplaneta americana extract oral cleaning product is a mouthwash mainly using the periplaneta americana extract crude product as a raw material, the main effect is to use the anti-inflammatory and detoxifying effects of the periplaneta americana protein, and the product location is mainly to inhibit the growth of bacteria.
Disclosure of Invention
In order to overcome the defects and shortcomings of the prior art, the invention provides a preparation method of the periplaneta americana polypeptide extract and also provides application of the periplaneta americana polypeptide extract extracted by the extraction method.
The specific technical scheme is as follows:
a periplaneta americana polypeptide extract, which is prepared by the following steps:
taking dried protozoon of periplaneta americana, and crushing the protozoon on an ultrafine crusher to obtain nanoscale powder; placing the nano-scale powder in a vacuum extraction tank, adding 10 times volume of 95% ethanol, heating at 60 ℃ under-8 mpa vacuum degree, carrying out countercurrent dynamic extraction to obtain an extract fluid extract, and carrying out vacuum drying to obtain a periplaneta americana extract crude product dry powder;
taking Periplaneta americana extract crude product dry powder, dispersing, stirring and dissolving with purified water in a ratio of 1:100, filtering with a 1um folding filter, and injecting the filtrate into a polyamide membrane reverse osmosis (R/O) device to obtain percolate for later use;
injecting the percolate into a macroporous resin separation column filled with D101, D941, H103, NKA-II and other macroporous resins with different types and different mixture ratios, eluting with a water-alcohol mixed solution (water: ethanol is 100:33), allowing the water-alcohol mixed solution to pass through the macroporous resin separation column at a flow rate of 0.65L/min, and removing specific impurities such as mucoid, mucopolysaccharide, cellulose and the like by utilizing different adsorption effects of the macroporous resins to obtain a primary pure solution (fluid extract) for later use;
extracting the fluid extract, injecting into supercritical carbon dioxide extraction equipment, further removing macromolecular heterocyclic protein impurities in the fluid extract by utilizing the phase change characteristic of liquefied carbon dioxide, and obtaining an extract (thick extract) with higher purity for later use;
taking the thick extract, and adding purified water according to the weight ratio of 1: dispersing, stirring, dissolving at a ratio of 50, injecting into a special hollow fiber ultrafiltration device, and removing macromolecular protein components to obtain ultrafiltration extractive solution;
injecting the ultrafiltration extract into a separation device of a vertical electrophoresis combined system, and performing chromatography to obtain different separation liquids under the action of an electric field by utilizing the principle that retention times of different molecular weights in the components are different, so as to achieve the purpose of removing macromolecular peptides or long-chain peptides and obtain a chromatography liquid of an active polypeptide component, namely an extract of the polypeptide component of the periplaneta americana;
taking the periplaneta americana polypeptide component extracting solution, drying for 12 hours at the temperature of minus 40 +/-5 ℃ and under the condition of minus 10 +/-2 mPa by adopting ultralow temperature vacuum drying equipment to obtain the periplaneta americana polypeptide extract, and storing for later use.
The periplaneta americana polypeptide extract extracted by the method is placed in sodium alginate for dissolution and storage, so that the periplaneta americana polypeptide extract can be stably stored.
The periplaneta americana polypeptide extract can be applied to a mouth cleaning product.
Preferably, the mouth cleaning product can be a medicine, a medical appliance or a daily chemical product.
Preferably, the mouth wash product is in the form of a spray, a mouthwash, a tablet, a gel, a film.
Preferably, the oral cleaning product comprises 0.1-6 parts of periplaneta americana polypeptide extract, 0.5-0.8 part of rabdosia rubescens extract, 0.5-0.8 part of radix tetrastigme extract, 0.5-0.8 part of anoectochilus formosanus extract, 0.3-0.6 part of eugenol, 0.3-0.6 part of cymene and 0.3-0.6 part of asiaticoside.
Preferably, the oral cleaning product spray comprises 3-6 parts of periplaneta americana polypeptide extract, 0.5-0.8 part of rabdosia rubescens extract, 0.5-0.8 part of radix tetrastigme extract, 0.5-0.8 part of anoectochilus formosanus extract, 0.3-0.6 part of eugenol, 0.3-0.6 part of cymene, 0.3-0.6 part of asiaticoside, 11-13 parts of polyoxyethylene 40 hydrogenated castor oil, 25-35 parts of propylene glycol, 85-95 parts of glycerol, 0.08-0.12 part of dipotassium glycyrrhizinate, 0.15-0.2 part of sucralose, 0.08-0.12 part of ixiong sugar, 0.25-0.3 part of freshener, 0.8-0.12 part of methylparaben, 0.4-0.6 part of propylparaben, 1-3 parts of essence and 900-1000 parts of purified water.
Preferably, the preparation method of the mouth cleaning product spray comprises the following steps:
s1, injecting purified water with the total preparation amount of 80% into a preparation tank, adding glycerol, stirring uniformly, dispersing and dissolving the American cockroach polypeptide extract in purified water with the preparation amount of 5%, adding into the preparation tank, and stirring uniformly;
s2, taking purified water with the total preparation amount of 10%, adding dipotassium glycyrrhizinate, sucralose and Italy euphoria sugar into another suitable container to prepare a sweet solution, pouring the sweet solution into the solution obtained in the step S1, and continuing stirring;
s3, mixing propylene glycol and hydrogenated castor oil, stirring uniformly, adding methyl hydroxybenzoate, propyl hydroxybenzoate and a freshener, dissolving completely, adding essence, stirring uniformly, pouring into the prepared solution obtained in S2, and stirring continuously;
s4, dissolving rabdosia rubescens extract, radix tetrastigme extract, anoectochilus roxburghii extract, eugenol, cymene and asiaticoside with purified water accounting for 5% of the total preparation amount, pouring into a preparation tank, and uniformly stirring to obtain the product;
s5, subpackaging the prepared semi-finished product liquid into suitable containers, capping and sealing, transferring to an outer packaging process, and boxing.
The invention has the advantages that: the invention uses the active periplaneta americana polypeptide extract directly obtained after the treatment of the biological activity bionic affinity purification technology, the preparation steps are simple and convenient, the industrialization is facilitated, and the test result shows that the periplaneta americana polypeptide extract has obvious effect of repairing the oral mucosa, and provides a new material basis for developing oral cleaning medicines, medical instruments, daily chemical products and the like.
Drawings
FIG. 1 is a schematic diagram of the preparation steps of the Periplaneta americana polypeptide extract of the present invention;
FIG. 2 is an image of different groups of cells in the cell scratch test of example 4 (microscope 100X).
Detailed Description
For better understanding of the present invention, the following embodiments are further described, but the present invention is not limited to the following embodiments.
Example 1 preparation of periplaneta americana polypeptide extract:
taking dried protozoon of periplaneta americana, and crushing the protozoon on an ultrafine crusher to obtain nanoscale powder; placing the nano-scale powder in a vacuum extraction tank, adding 10 times volume of 95% ethanol, heating at 60 ℃ under-8 mpa vacuum degree, carrying out countercurrent dynamic extraction to obtain an extract fluid extract, and carrying out vacuum drying to obtain a periplaneta americana extract crude product dry powder;
taking Periplaneta americana extract crude product dry powder, dispersing, stirring and dissolving with purified water in a ratio of 1:100, filtering with a 1um folding filter, and injecting the filtrate into a polyamide membrane reverse osmosis (R/O) device to obtain percolate for later use;
injecting the percolate into a macroporous resin separation column filled with D101, D941, H103, NKA-II and other macroporous resins with different types and different mixture ratios, eluting with a water-alcohol mixed solution (water: ethanol is 100:33), allowing the water-alcohol mixed solution to pass through the macroporous resin separation column at a flow rate of 0.65L/min, and removing specific impurities such as mucoid, mucopolysaccharide, cellulose and the like by utilizing different adsorption effects of the macroporous resins to obtain a primary pure solution (fluid extract) for later use;
extracting the fluid extract, injecting into supercritical carbon dioxide extraction equipment, further removing macromolecular heterocyclic protein impurities in the fluid extract by utilizing the phase change characteristic of liquefied carbon dioxide, and obtaining an extract (thick extract) with higher purity for later use;
taking the thick extract, and adding purified water according to the weight ratio of 1: dispersing, stirring, dissolving at a ratio of 50, injecting into a special hollow fiber ultrafiltration device, and removing macromolecular protein components to obtain ultrafiltration extractive solution;
injecting the ultrafiltration extract into a separation device of a vertical electrophoresis combined system, and performing chromatography to obtain different separation liquids under the action of an electric field by utilizing the principle that retention times of different molecular weights in the components are different, so as to achieve the purpose of removing macromolecular peptides or long-chain peptides and obtain a chromatography liquid of an active polypeptide component, namely an extract of the polypeptide component of the periplaneta americana;
taking the periplaneta americana polypeptide component extracting solution, drying for 12 hours at the temperature of minus 40 +/-5 ℃ and under the condition of minus 10 +/-2 mPa by adopting ultralow temperature vacuum drying equipment to obtain the periplaneta americana polypeptide extract, and storing for later use.
Example 2 periplaneta americana polypeptide safety assay:
2.1 test materials:
2.1.1 test strains:
histidine-deficient strains TA97, TA98, TA100 and TA102 of Salmonella typhimurium were purchased from KRISHGEN BioSystems and CHI SCIENTIFIC. Before testing, four strains were identified in the laboratory and the biological properties of the strains met the test requirements.
2.1.2 metabolite activation System: s9
2.1.3 positives:
2.1.3.1 sodium azide (NaN3) was purchased from the national pharmaceutical group Chemicals Co., Ltd. (batch No.: 20180321).
2.1.3.2 Dixon (Dexon) from AccuStandard (batch No.: 4030165-01).
2.1.3.32-Aminofluorene (2-AF) was purchased from aladdin (batch: H1511011).
2.1.3.41, 8-dihydroxyanthraquinone (Dorbane) was purchased from Sigma-Aldrich (batch: WXBB 0392).
2.1.3.52-Aminoanthracene (2-AA) was purchased from Sigma-Aldrich (batch number: STBD 3302V).
2.1.4 culture conditions: 37 ℃ incubator
2.2 test procedure:
2.2.1 preparation of bacteria: before the test, the strain was inoculated into a nutrient broth and incubated at about 37 ℃ in a constant temperature incubator for approximately 16 hours. The strain reaches about 1X 10 per ml9。
2.2.2 test article preparation: sterile ultrapure water is used as a solvent to prepare contamination solutions with different concentrations, namely 50mg/mL, 15.81mg/mL, 5mg/mL and 1.58mg/mL respectively.
2.2.3 grouping: setting 4 different dosage groups, performing metabolic activation and non-metabolic activation treatment, and respectively setting a solvent control group, a positive control group, a sterile control group and a blank control group.
2.2.4 contamination time: 48h
2.2.5 plate incorporation:
non-metabolic activation: 2.0mL of top agar culture medium is subpackaged in test tubes, heat preservation is carried out in a water bath at 45 ℃, then 0.1mL of test strain enrichment liquid, 0.1mL of test object solution and 0.5mL of phosphate buffer solution are sequentially added into each tube, the mixture is fully and uniformly mixed, the mixture is rapidly poured onto a bottom culture medium plate, and a plate is rotated to be uniformly distributed. All dose groups and controls were set up in 3 replicate dishes. Placing the plate horizontally for condensation and solidification, and culturing in a 37 deg.C incubator for 48 h. After the incubation was completed, the growth of background colonies was observed and the number of colonies varied back per dish was recorded.
Metabolic activation system: the phosphate buffer solution (0.5 mL) in the nonmetabolic activation test was replaced with the S9 mixed solution, and the other operations were the same as in the nonmetabolic activation test.
2.3 test results:
bacterial regression individual values, mean and standard deviation:
TABLE 2-1 statistical value table of colony counts tested (Mean + -SD)
TABLE 2-2 statistical value table of colony counts tested (Mean + -SD)
Tables 2-3 colony counts
Tables 2-4 colony counts for the first test
2.4 conclusion:
no obvious bacterial toxicity is generated in each dose group compared with a solvent control group whether the metabolism is activated or not, and the number of reversion colonies of each dose of all the tested strains is 2 times lower than that of the solvent control group.
According to the judgment standard of technical Specification for safety of cosmetics (2015), the test sample has no mutagenicity to the test strain under the test condition.
Example 3 mucosal irritancy of periplaneta americana polypeptide extract:
3.1 test substance:
the sample was diluted to a 5% aqueous solution.
3.2 Experimental animals and Environment:
3.2.1 general grade animal house: license number for experimental animals: SYXK (Yue) 2017-.
3.2.2 animal species strains: common grade New Zealand rabbits; animal certification number: 44007600008193.
3.2.3 animal sources: SCXK (Guangdong) 2019-0023 of Dongxin laboratory animal farm in Guangzhou city Huadu district.
3.2.4 Environment: temperature (T): 18-25 ℃, relative humidity (% RH): 40-70 percent.
3.3 test procedure:
both eyes of the experimental animals were routinely examined 24h prior to the experiment.
During the test, the lower eyelid of the eye on one side of the experimental animal is slightly pulled open, 0.1mL of the test object is sucked and dripped into the conjunctival sac, the eye is passively closed for 1s, and the eye is not washed within 24 hours after the test object is dripped. The other eye was left untreated as a self control.
Eyes were examined and scored at 1h, 24h, 48h, 72h, 4d, 7d, 14d and 21d after the test was completed, e.g., no irritation occurred within 72h, or eyes at 7d or 14d recovered completely, and the test was terminated prematurely. The cornea was examined with 2% sodium fluorescein if necessary.
3.4 evaluation of results: and calculating stimulation response integral according to scores of cornea, iris or conjunctiva of the tested animals at the observation time points of 24h, 48h and 72h, and grading the eye irritation response according to the evaluation of the stimulation response integral and the recovery time according to the table 3-1.
3-1 grading of products by eye-irritating reaction
Note: when the integral of cornea, iris and conjunctiva is 0, it is judged to be nonirritating.
3.5 test results
3-2 eye irritation test results
According to the judgment standard of the acute eye irritation/corrosion test of chapter six of the 2015 technical safety standard of cosmetics, under the test condition, the result of the acute eye irritation test of the sample periplaneta americana polypeptide extract to the new zealand rabbit is non-irritant.
Example 4 periplaneta americana polypeptide Human Gingival Fibroblast (HGF) scratch test:
4.1 Experimental materials:
4.1.1 cell lines: human Gingival Fibroblast (HGF), purchased from north nai hitching, algebra: 6
4.1.2 complete Medium: DMEM medium containing 10% Fetal Bovine Serum (FBS) (FBS from Gibco, Lot:42Q1095K)
4.1.3 maintenance Medium: DMEM medium with 1% FBS (FBS from Gibco, Lot:42Q1095K)
4.1.4 test conditions: temperature 37 + -0.5 deg.C, humidity > 90%, and concentration of 5% CO2
4.1.5 solutions and controls:
negative control group (NC): maintenance medium
Positive control group (PC): maintenance medium containing 1ng/mL EGF
Test substance (TA): 0.2153g of sample is weighed, 2.15mL of maintenance medium is added to prepare a stock solution of 100mg/mL, and then the stock solution is diluted to the concentration to be measured by using the maintenance medium in turn.
4.2 test procedure:
4.2.1 cell Activity assay
4.2.1.1 cells were routinely cultured. Preparing cell suspension with the density of 5-7 multiplied by 104/mL, inoculating the cell suspension into a 96-well cell culture plate, culturing for 18-24h, wherein each well is 100 mu L.
4.2.1.2 discard the stock culture from the wells, add 100. mu.L of test samples of different concentrations to each well, return to the incubator and incubate for 48. + -. 1 h.
4.2.1.3 the plates were removed and 20. mu.L of MTT solution was added to each well and the incubator was incubated for 3-4 h. Removing liquid in the wells, adding 100 μ L DMSO into each well, placing in an oscillator, oscillating for 10-15min, and measuring absorbance at 570nm wavelength of a microplate reader.
4.2.1.4 data analysis: cell activity relative cell activity (viatility) was calculated for each group, taking the cell activity of the negative control group as 100%.
4.2.2 cell scratch test
4.2.2.1 cells were cultured routinely, cell density was adjusted to 3.0-5.0X 105 cells/mL, using 24-well cell culture plates with 2-well inserts, 70. mu.L of each well was inoculated, returned to the incubator and cultured for 18-24h until cells fused.
4.2.2.2 carefully remove the culture insert from the plate with forceps, a flat cell-free area is formed between the 2 wells of the culture insert. The cell layer was washed 3 times with PBS to remove exfoliated cells.
4.2.2.3 Add 1mL of culture medium containing different concentrations of test sample to each well while taking an image under the mirror, noted 0h, and mark the location of the acquisition.
4.2.2.4 pictures were taken every 24h, at the same marked positions, noted 24h and 48 h.
4.2.2.5 data analysis: and analyzing the area of the scratch region of each image by using IPP image analysis software, and calculating the relative area by taking 0h as 100%.
4.3 test results:
4.3.1 cell Activity assay
TABLE 4-1 relative Activity of the groups of cells (Mean. + -. SD)
4.3.2 cell scratch test
TABLE 4-2 relative scratch area size (%)
Under the test condition of 4.3.2, at 24h, the relative scratch area of the positive control group is obviously reduced (P <0.05), and the relative scratch area of the American cockroach polypeptide extract sample is obviously reduced (P <0.05) under the concentrations of 4.0mg/mL, 1.0mg/mL and 0.25 mg/mL. At 48h, compared with the negative control group, the relative scratch area of the positive control group is obviously reduced (P <0.05), and the relative scratch area of the American cockroach polypeptide extract is obviously reduced (P <0.05) under the concentration of 4.0mg/mL, 1.0mg/mL and 0.25 mg/mL. The sample periplaneta americana polypeptide extract is prompted to have a certain effect of promoting and repairing human gingival fibroblast.
Example 5 oral spray formulation one:
3 parts of periplaneta americana active polypeptide extract, 0.5 part of rabdosia rubescens extract, 0.5 part of tetrastigma hemsleyanum extract, 0.5 part of anoectochilus roxburghii extract, 0.3 part of eugenol, 0.3 part of cymene, 0.3 part of asiaticoside, 11 parts of polyoxyethylene 40 hydrogenated castor oil, 25 parts of propylene glycol, 85 parts of glycerol, 0.08 part of dipotassium glycyrrhizinate, 0.15 part of sucralose, 0.08 part of ixiong sugar, 0.25 part of freshener, 0.8 part of methylparaben, 0.4 part of propylparaben, 1 part of essence and 870.84 parts of purified water.
Example 6 oral spray formulation two:
3 parts of periplaneta americana active polypeptide extract, 0.65 part of rabdosia rubescens extract, 0.65 part of tetrastigma hemsleyanum extract, 0.65 part of anoectochilus roxburghii extract, 0.45 part of eugenol, 0.45 part of cymene, 0.45 part of asiaticoside, 12 parts of polyoxyethylene 40 hydrogenated castor oil, 30 parts of propylene glycol, 90 parts of glycerol, 0.1 part of dipotassium glycyrrhizinate, 0.175 part of sucralose, 0.1 part of ixiong sugar, 0.275 part of freshener, 0.1 part of methyl paraben, 0.5 part of propyl paraben, 2 parts of essence and 855.45 parts of purified water.
Example 7 oral spray formulation three:
6 parts of periplaneta americana active polypeptide extract, 0.8 part of rabdosia rubescens extract, 0.8 part of tetrastigma hemsleyanum extract, 0.8 part of anoectochilus roxburghii extract, 0.6 part of eugenol, 0.6 part of cymene, 0.6 part of asiaticoside, 13 parts of polyoxyethylene 40 hydrogenated castor oil, 35 parts of propylene glycol, 95 parts of glycerol, 0.12 part of dipotassium glycyrrhizinate, 0.2 part of sucralose, 0.12 part of idaxin sugar, 0.3 part of freshener, 0.12 part of methyl paraben, 0.6 part of propyl paraben, 3 parts of essence and 842.34 parts of purified water.
Example 8 oral spray preparation method:
the preparation method of the mouth cleaning spray comprises the following steps: s1, injecting purified water with the total preparation amount of 80% into a preparation tank, adding glycerol, stirring uniformly, dispersing and dissolving the American cockroach polypeptide extract in purified water with the preparation amount of 5%, adding into the preparation tank, and stirring uniformly;
s2, taking purified water with the total preparation amount of 10%, adding dipotassium glycyrrhizinate, sucralose and Italy euphoria sugar into another suitable container to prepare a sweet solution, pouring the sweet solution into the solution obtained in the step S1, and continuing stirring;
s3, mixing propylene glycol and hydrogenated castor oil, stirring uniformly, adding methyl hydroxybenzoate, propyl hydroxybenzoate and a freshener, dissolving completely, adding essence, stirring uniformly, pouring into the prepared solution obtained in S2, and stirring continuously;
s4, dissolving the rabdosia rubescens extract, the radix tetrastigme extract, the anoectochilus roxburghii extract, the eugenol, the cymene and the asiaticoside with purified water accounting for 5 percent of the total preparation amount of a proper amount of water, pouring the mixture into a preparation tank, and uniformly stirring the mixture to obtain the traditional Chinese medicine preparation;
s5, subpackaging the prepared semi-finished product liquid into suitable containers, capping and sealing, transferring to an outer packaging process, and boxing.
TABLE 6-1 oral spray product test results
Example 9 mouthwash composition formula one:
0.1 part of periplaneta americana active polypeptide extract, 0.5 part of rabdosia rubescens extract, 0.5 part of tetrastigma hemsleyanum extract, 0.5 part of anoectochilus roxburghii extract, 0.3 part of eugenol, 0.3 part of cymene, 0.3 part of asiaticoside, 11 parts of polyoxyethylene 40 hydrogenated castor oil, 25 parts of propylene glycol, 85 parts of glycerol, 0.08 part of dipotassium glycyrrhizinate, 0.15 part of sucralose, 0.08 part of ixiong sugar, 0.25 part of freshener, 0.8 part of methyl paraben, 0.4 part of propyl paraben, 1 part of essence and 873.74 parts of purified water.
Example 10 mouthwash composition formula ii:
1.1 parts of periplaneta americana active polypeptide extract, 0.65 part of rabdosia rubescens extract, 0.65 part of tetrastigma hemsleyanum extract, 0.65 part of anoectochilus roxburghii extract, 0.45 part of eugenol, 0.45 part of cymene, 0.45 part of asiaticoside, 12 parts of polyoxyethylene 40 hydrogenated castor oil, 30 parts of propylene glycol, 90 parts of glycerol, 0.1 part of dipotassium glycyrrhizinate, 0.175 part of sucralose, 0.1 part of ixiong sugar, 0.275 part of freshener, 0.1 part of methyl paraben, 0.5 part of propyl paraben, 2 parts of essence and 860.35 parts of purified water.
Example 11 mouthwash composition formula three:
2 parts of periplaneta americana active polypeptide extract, 0.8 part of rabdosia rubescens extract, 0.8 part of tetrastigma hemsleyanum extract, 0.8 part of anoectochilus roxburghii extract, 0.6 part of eugenol, 0.6 part of cymene, 0.6 part of asiaticoside, 13 parts of polyoxyethylene 40 hydrogenated castor oil, 35 parts of propylene glycol, 95 parts of glycerol, 0.12 part of dipotassium glycyrrhizinate, 0.2 part of sucralose, 0.12 part of ixiong sugar, 0.3 part of freshener, 0.12 part of methylparaben, 0.6 part of propylparaben, 3 parts of essence and 846.34 parts of purified water.
Example 12 method of preparing a mouthrinse comprising a mouthwash:
the preparation method of the mouth cleaning liquid containing cleaner comprises the following steps: s1, injecting purified water with the total preparation amount of 80% into a preparation tank, adding glycerol, stirring uniformly, dispersing and dissolving the American cockroach polypeptide extract in purified water with the preparation amount of 5%, adding into the preparation tank, and stirring uniformly;
s2, taking purified water with the total preparation amount of 10%, adding dipotassium glycyrrhizinate, sucralose and Italy euphoria sugar into another suitable container to prepare a sweet solution, pouring the sweet solution into the solution obtained in the step S1, and continuing stirring;
s3, mixing propylene glycol and hydrogenated castor oil, stirring uniformly, adding methyl hydroxybenzoate, propyl hydroxybenzoate and a freshener, dissolving completely, adding essence, stirring uniformly, pouring into the prepared solution obtained in S2, and stirring continuously;
s4, dissolving rabdosia rubescens extract, radix tetrastigme extract, anoectochilus roxburghii extract, eugenol, cymene and asiaticoside with purified water accounting for 5% of the total preparation amount, pouring into a preparation tank, and uniformly stirring to obtain the product;
s5, subpackaging the prepared semi-finished product liquid into suitable containers, capping and sealing, transferring to an outer packaging process, and boxing.
Claims (10)
1. A periplaneta americana polypeptide extract is characterized in that the preparation method comprises the following steps:
taking dried protozoon of periplaneta americana, and crushing the protozoon on an ultrafine crusher to obtain nanoscale powder; placing the nano-scale powder in a vacuum extraction tank, adding 10 times volume of 95% ethanol, heating at-8 mpa vacuum degree for 60 ℃, performing countercurrent dynamic extraction to obtain an extract fluid extract, and performing vacuum drying to obtain crude periplaneta americana extract dry powder;
taking Periplaneta americana extract crude product dry powder, dispersing, stirring and dissolving with purified water in a ratio of 1:100, filtering with a 1um folding filter, and injecting the filtrate into a polyamide membrane reverse osmosis (R/O) device to obtain percolate for later use;
injecting the percolate into macroporous resin separation columns filled with D101, D941, H103, NKA-II and other macroporous resin separation columns with different types and different mixture ratios, eluting with water-alcohol mixed solution (water: ethanol is 100:33), and allowing the water-alcohol mixed solution to pass through at a flow rate of 0.65L/min to obtain fluid extract for later use;
taking the fluid extract, injecting into supercritical carbon dioxide extraction equipment to obtain a thick extract for later use;
taking the thick extract, and adding purified water according to the weight ratio of 1: dispersing, stirring, dissolving at a ratio of 50, injecting into a special hollow fiber ultrafiltration device, and removing macromolecular protein components to obtain ultrafiltration extractive solution;
injecting the ultrafiltration extract into a separation device of a vertical electrophoresis combined system to obtain a chromatography liquid of active polypeptide components, namely the periplaneta americana polypeptide component extract;
and (3) taking the periplaneta americana polypeptide component extracting solution, drying for 12 hours at the temperature of minus 40 +/-5 ℃ and under the condition of minus 10 +/-2 mPa by adopting ultralow-temperature vacuum drying equipment, and storing the periplaneta americana polypeptide extract for later use.
2. The periplaneta americana polypeptide extract as claimed in claim 1, wherein the periplaneta americana polypeptide extract is dissolved and preserved in sodium alginate.
3. Use of the polypeptide extract of periplaneta americana according to claim 1 or claim 2 in a mouth-cleansing product.
4. Use according to claim 3, wherein the oral cleaning product is a pharmaceutical product, a medical device or a daily chemical product.
5. A use according to claim 4 wherein the oral cleaning product is in the form of a spray, mouthwash, tablet, gel, film.
6. The application of claim 5, wherein the oral cleaning product comprises 0.1-6 parts of periplaneta americana polypeptide extract, 0.5-0.8 part of rabdosia rubescens extract, 0.5-0.8 part of radix tetrastigme extract, 0.5-0.8 part of anoectochilus roxburghii extract, 0.3-0.6 part of eugenol, 0.3-0.6 part of cymene and 0.3-0.6 part of asiaticoside.
7. The application of claim 6, wherein the oral cleaning product spray comprises 3-6 parts of periplaneta americana polypeptide extract, 0.5-0.8 part of rabdosia rubescens extract, 0.5-0.8 part of radix tetrastigme extract, 0.5-0.8 part of anoectochilus roxburghii extract, 0.3-0.6 part of eugenol, 0.3-0.6 part of cymene, 0.3-0.6 part of asiaticoside, 11-13 parts of polyoxyethylene 40 hydrogenated castor oil, 25-35 parts of propylene glycol, 85-95 parts of glycerol, 0.08-0.12 part of dipotassium glycyrrhizinate, 0.15-0.2 part of sucralose, 0.08-0.12 part of ixionsin, 0.25-0.3 part of freshener, 0.8-0.12 part of methyl paraben, 0.4-0.6 part of propyl paraben, 1-3 parts of essence and 900-1000 parts of purified water.
8. The use of claim 7, wherein the oral cleaning product spray is prepared by:
s1, injecting purified water with the total preparation amount of 80% into a preparation tank, adding glycerol, stirring uniformly, dispersing and dissolving the American cockroach polypeptide extract in purified water with the preparation amount of 5%, adding into the preparation tank, and stirring uniformly;
s2, taking purified water with the total preparation amount of 10%, adding dipotassium glycyrrhizinate, sucralose and Italy euphoria sugar into another suitable container to prepare a sweet solution, pouring the sweet solution into the solution obtained in the step S1, and continuing stirring;
s3, mixing propylene glycol and hydrogenated castor oil, stirring uniformly, adding methyl hydroxybenzoate, propyl hydroxybenzoate and a freshener, dissolving completely, adding essence, stirring uniformly, pouring into the prepared solution obtained in S2, and stirring continuously;
s4, dissolving rabdosia rubescens extract, radix tetrastigme extract, anoectochilus roxburghii extract, eugenol, cymene and asiaticoside with purified water accounting for 5% of the total preparation amount, pouring into a preparation tank, and uniformly stirring to obtain the product;
s5, subpackaging the prepared semi-finished product liquid into suitable containers, capping and sealing, transferring to an outer packaging process, and boxing.
9. The use of claim 6, wherein the components of the liquid mouthwash product comprise 0.1-2 parts of periplaneta americana polypeptide extract, 0.5-0.8 part of rabdosia rubescens extract, 0.5-0.8 part of radix tetrastigme extract, 0.5-0.8 part of anoectochilus roxburghii extract, 0.3-0.6 part of eugenol, 0.3-0.6 part of cymene, 0.3-0.6 part of asiaticoside, 11-13 parts of polyoxyethylene 40 hydrogenated castor oil, 25-35 parts of propylene glycol, 85-95 parts of glycerol, 0.08-0.12 part of dipotassium glycyrrhizinate, 0.15-0.2 part of sucralose, 0.08-0.12 part of isoeuphorbia hirsuta, 0.25-0.3 part of freshener, 0.8-0.12 part of methylparaben, 0.4-0.6 part of propylparaben, 1-3 parts of essence, and 1000 parts of purified water.
10. A use according to claim 9, wherein the mouthwash product is prepared by a method comprising:
s1, injecting purified water with the total preparation amount of 80% into a preparation tank, adding glycerol, stirring uniformly, dispersing and dissolving the American cockroach polypeptide extract in purified water with the preparation amount of 5%, adding into the preparation tank, and stirring uniformly;
s2, taking purified water with the total preparation amount of 10%, adding dipotassium glycyrrhizinate, sucralose and Italy euphoria sugar into another suitable container to prepare a sweet solution, pouring the sweet solution into the solution obtained in the step S1, and continuing stirring;
s3, mixing propylene glycol and hydrogenated castor oil, stirring uniformly, adding methyl hydroxybenzoate, propyl hydroxybenzoate and a freshener, dissolving completely, adding essence, stirring uniformly, pouring into the prepared solution obtained in S2, and stirring continuously;
s4, dissolving rabdosia rubescens extract, radix tetrastigme extract, anoectochilus roxburghii extract, eugenol, cymene and asiaticoside with purified water accounting for 5% of the total preparation amount, pouring into a preparation tank, and uniformly stirring to obtain the product;
s5, subpackaging the prepared semi-finished product liquid into suitable containers, capping and sealing, transferring to an outer packaging process, and boxing.
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