CN113491663B - Superoxide dismutase composition and preparation method thereof - Google Patents
Superoxide dismutase composition and preparation method thereof Download PDFInfo
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- CN113491663B CN113491663B CN202110319847.6A CN202110319847A CN113491663B CN 113491663 B CN113491663 B CN 113491663B CN 202110319847 A CN202110319847 A CN 202110319847A CN 113491663 B CN113491663 B CN 113491663B
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- 102000019197 Superoxide Dismutase Human genes 0.000 title claims abstract description 125
- 108010012715 Superoxide dismutase Proteins 0.000 title claims abstract description 125
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- TXFPEBPIARQUIG-UHFFFAOYSA-N 4'-hydroxyacetophenone Chemical compound CC(=O)C1=CC=C(O)C=C1 TXFPEBPIARQUIG-UHFFFAOYSA-N 0.000 claims abstract description 60
- 229920001495 poly(sodium acrylate) polymer Polymers 0.000 claims abstract description 35
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Classifications
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- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
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- A61K8/81—Cosmetics or similar toiletry preparations characterised by the composition containing organic macromolecular compounds obtained by reactions involving only carbon-to-carbon unsaturated bonds
- A61K8/8141—Compositions of homopolymers or copolymers of compounds having one or more unsaturated aliphatic radicals, each having only one carbon-to-carbon double bond, and at least one being terminated by only one carboxyl radical, or of salts, anhydrides, esters, amides, imides or nitriles thereof; Compositions of derivatives of such polymers
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- A61K8/34—Alcohols
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- A61K8/49—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing heterocyclic compounds
- A61K8/494—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing heterocyclic compounds with more than one nitrogen as the only hetero atom
- A61K8/4953—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing heterocyclic compounds with more than one nitrogen as the only hetero atom containing pyrimidine ring derivatives, e.g. minoxidil
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Abstract
The invention relates to the field of skin care products, in particular to a superoxide dismutase composition and a preparation method thereof. The invention discloses a composition of superoxide dismutase, which comprises sodium polyacrylate, lactobacillus fermentation lysate, superoxide dismutase and fullerene. The invention provides a superoxide dismutase composition and a preparation method thereof, wherein the lactobacillus fermentation lysate and the superoxide dismutase have the effects of scavenging free radicals and resisting aging, can improve the capability of SOD for improving skin problems, and have the effects of whitening, lightening spots, resisting aging, controlling oil and homogenizing skin color; meanwhile, fullerene, tetrahydropyrimidine carboxylic acid and p-hydroxyacetophenone are selected to play a role in protecting SOD, so that the enzyme activity value of the SOD is improved; 1, 2-hexanediol and methyl propylene glycol are used as solvents, so that the solubility of the solvent can be improved, the effect of protecting SOD can be achieved, and the enzyme activity value of the SOD can be improved.
Description
Technical Field
The invention relates to the field of skin care products, in particular to a superoxide dismutase composition and a preparation method thereof.
Background
Superoxide dismutase (SOD) is an important component of the antioxidant enzyme system in biological systems and is widely distributed in microorganisms, plants and animals. SOD is an antioxidant metalloenzyme existing in organisms, can catalyze superoxide anion free radical disproportionation to generate oxygen and hydrogen peroxide, plays a vital role in organism oxidation and antioxidant balance, and is indispensible from the occurrence and development of a plurality of diseases.
Research proves that SOD can play the following roles when being added into cosmetics: the skin-blacking agent has obvious sun-screening effect, and the main reason for blackening the skin by illumination is oxygen free radical damage, so that the SOD can effectively prevent the skin from being damaged by ionizing radiation (particularly ultraviolet rays), thereby playing a role in sun-screening; SOD is antioxidant enzyme, and can effectively prevent skin aging, remove freckle and resist wrinkle; has obvious anti-inflammatory effect and certain effect on preventing and treating skin diseases; SOD has certain effect of preventing and treating formation of speckle. However, superoxide dismutase in a powder state has relatively rapid enzyme activity reduction in the normal temperature preservation process.
Therefore, the preparation of the SOD composition with higher enzyme activity and the preparation method thereof are very urgent. Based on the research, the invention provides a superoxide dismutase composition and a preparation method thereof, and the superoxide dismutase composition is prepared into a solution, so that the enzyme activity of SOD can be effectively improved, and a series of skin problems can be effectively improved by applying the composition to skin care products, so that the effects of whitening, lightening spots, resisting aging, controlling oil and homogenizing skin color are achieved.
Disclosure of Invention
In order to solve the above problems, a first aspect of the present invention provides a composition of superoxide dismutase, comprising sodium polyacrylate, lactobacillus fermentation lysate, superoxide dismutase and fullerene.
As a preferable technical scheme, the superoxide dismutase composition comprises the following components in parts by weight: 0.005-8 parts of sodium polyacrylate, 0.01-8 parts of lactobacillus fermentation lysate, 0.05-10 parts of superoxide dismutase and 0.01-1 part of fullerene.
As a preferable technical scheme, the superoxide dismutase composition comprises the following components in parts by weight: 0.01-5 parts of sodium polyacrylate, 0.02-5 parts of lactobacillus fermentation lysate, 0.1-5 parts of superoxide dismutase and 0.3-0.8 part of fullerene.
As a preferable technical scheme, the superoxide dismutase composition also comprises the following components in parts by weight: 0.008-9 parts of tetrahydropyrimidine carboxylic acid.
As a preferable technical scheme, the superoxide dismutase composition comprises the following components in parts by weight: 0.01-5 parts of tetrahydropyrimidine carboxylic acid.
As a preferable technical scheme, the superoxide dismutase composition also comprises the following components in parts by weight: 0.05-1 part of p-hydroxyacetophenone.
As a preferable technical scheme, the superoxide dismutase composition comprises the following components in parts by weight: 0.1-0.6 part of p-hydroxyacetophenone.
As a preferable technical scheme, the superoxide dismutase composition also comprises the following components in parts by weight: 0.05-10 parts of 1, 2-hexanediol and 0.05-8 parts of methyl propylene glycol.
As a preferable technical scheme, the superoxide dismutase composition comprises the following components in parts by weight: 0.1-5 parts of 1, 2-hexanediol and 0.1-5 parts of methyl propylene glycol.
The second aspect of the present invention provides a method for producing the superoxide dismutase composition, comprising: mixing, heating and dissolving p-hydroxyacetophenone, 1, 2-hexanediol and methyl propylene glycol to obtain a mixed solution; completely and uniformly swelling sodium polyacrylate with hot water, and then adding the mixed solution to uniformly mix; cooling to below 40 ℃; adding superoxide dismutase, tetrahydropyrimidine carboxylic acid and fullerene, and stirring uniformly.
Has the beneficial effects that; the invention provides a superoxide dismutase composition and a preparation method thereof, wherein the lactobacillus fermentation lysate and the superoxide dismutase have the effects of scavenging free radicals and resisting aging, can improve the capability of SOD for improving skin problems, and have the effects of whitening, lightening spots, resisting aging, controlling oil and homogenizing skin color; meanwhile, fullerene, tetrahydropyrimidine carboxylic acid and p-hydroxyacetophenone are selected to play a role in protecting SOD, so that the enzyme activity value of the SOD is improved; 1, 2-hexanediol and methyl propylene glycol are used as solvents, so that the solubility of the solvent can be improved, the effect of protecting SOD can be achieved, and the enzyme activity value of the SOD can be improved; the addition of the fullerene improves the effects of resisting ultraviolet, repairing damaged cells, inhibiting inflammatory factor expression and promoting cell metabolism of the superoxide dismutase composition.
Detailed Description
The disclosure of the present invention will be further understood in conjunction with the following detailed description of the preferred embodiments of the invention, including examples. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. If the definition of a particular term disclosed in the prior art does not conform to any definition provided in this application, the definition of that term provided in this application controls.
As used herein, unless the context clearly indicates otherwise, the absence of a limitation to a plurality of features is also intended to include the plurality of features. It will be further understood that the terms "made of …" and "comprising," "including," "having," "including," and/or "containing," as used herein, are synonymous with "including," "having," "containing," and/or "containing," and when used in this specification, mean the stated composition, step, method, article, or apparatus, but do not preclude the presence or addition of one or more other compositions, steps, methods, articles, or apparatus. Furthermore, when describing embodiments of the present application, the use of "preferred," "more preferred," etc. refers to embodiments of the present invention that may provide certain benefits in certain circumstances. However, other embodiments may be preferred under the same or other circumstances. In addition, the recitation of one or more preferred embodiments does not imply that other embodiments are not useful, nor is it intended to exclude other embodiments from the scope of the invention.
In order to solve the above problems, a first aspect of the present invention provides a composition of superoxide dismutase, comprising sodium polyacrylate, lactobacillus fermentation lysate, superoxide dismutase and fullerene.
Sodium polyacrylate is a novel functional polymer material and an important chemical product, the solid product is white (or pale yellow) block or powder, and the liquid product is colorless (or pale yellow) viscous liquid. Is prepared from acrylic acid and its esters through aqueous solution polymerization. No smell, and is dissolved in aqueous sodium hydroxide solution, and precipitation occurs in aqueous solution such as calcium hydroxide and magnesium hydroxide. Is often used as a water treatment agent, brine refining and latex thickening, and can also be used for food tackifying and emulsifying.
Sodium polyacrylate in this application may be commercially available, for example, as model C sold by Zhengzhou Kangyuan chemical products Co.
The lactobacillus fermentation lysate is obtained by the strains through probiotic evaluation screening, fermentation enrichment culture and complete wall breaking process treatment, and is mainly a series of substances with biological activity generated or released by lactobacillus cell lysis. The product has effects of scavenging free radicals, resisting aging, reducing melanin deposition, brightening skin, promoting fibroblast proliferation, and repairing skin, and can be used for skin care, whitening and moisturizing products, sensitive skin and pregnant women, and is suitable for use in essence, facial mask, facial cream, muscle base solution, etc.
The lactic acid bacteria fermentation lysate in the present application may be commercially available, for example, from the company of Biotechnology, ind. Texas.
Superoxide dismutase (SOD) is an important component of the antioxidant enzyme system in biological systems and is widely distributed in microorganisms, plants and animals. SOD is an antioxidant metalloenzyme existing in organisms, can catalyze superoxide anion free radical disproportionation to generate oxygen and hydrogen peroxide, plays a vital role in organism oxidation and antioxidant balance, and is indispensible from the occurrence and development of a plurality of diseases. Research proves that SOD can play the following roles when being added into cosmetics: the skin-blacking agent has obvious sun-screening effect, and the main reason for blackening the skin by illumination is oxygen free radical damage, so that the SOD can effectively prevent the skin from being damaged by ionizing radiation (particularly ultraviolet rays), thereby playing a role in sun-screening; SOD is antioxidant enzyme, and can effectively prevent skin aging, remove freckle and resist wrinkle; has obvious anti-inflammatory effect and certain effect on preventing and treating skin diseases; SOD has certain effect of preventing and treating formation of speckle.
The superoxide dismutase in the present application may be commercially available, for example, from Hebei Tuohai Biotechnology Co., ltd. Under the product number 10.
In some preferred embodiments, the superoxide dismutase composition comprises the following components in parts by weight: 0.005-8 parts of sodium polyacrylate, 0.01-8 parts of lactobacillus fermentation lysate, 0.05-10 parts of superoxide dismutase and 0.01-1 part of fullerene.
In some preferred embodiments, the superoxide dismutase composition comprises the following components in parts by weight: 0.01-5 parts of sodium polyacrylate, 0.02-5 parts of lactobacillus fermentation lysate, 0.1-5 parts of superoxide dismutase and 0.3-0.8 part of fullerene.
In some preferred embodiments, the superoxide dismutase composition further comprises the following components in parts by weight: 0.008-9 parts of tetrahydropyrimidine carboxylic acid.
Tetrahydropyrimidine carboxylic acid is also known as ectoine. The product can be used for skin care, and has effects of keeping moisture and repairing. The method can ensure that the high halophilic bacteria cannot be dehydrated and dead under a high-salt environment, is an important substance for maintaining osmotic pressure balance, has strong water molecule complexing capability due to the unique molecular structure, can complex four or five water molecules by one molecule of ectoin, and can structure free water in cells. The ectoine can continuously improve the moisture retention capacity of the skin, the moisture retention capacity of the skin can not be immediately reduced after the ectoine is stopped, the moisture retention capacity of the skin is higher than that of a control group after the ectoine is stopped for one week, and the ectoine is a very excellent humectant. Meanwhile, the ectoin can effectively improve the immune protection capability of skin cells, increase the cell repair capability and enable the skin to effectively resist invasion of microorganisms and allergens. Experiments show that: the ectoin can damage ultraviolet rays to skin and repair cell DNA damage caused by ultraviolet rays. Since it is possible to relieve various stresses on the skin, such as ultraviolet ray damage, skin aging due to dry environment, skin deterioration due to surfactant, etc., escidodine is attracting attention as a cell repair additive to be added to cosmetics for the purpose of combating skin aging.
In some preferred embodiments, the superoxide dismutase composition comprises the following components in parts by weight: 0.01-5 parts of tetrahydropyrimidine carboxylic acid.
In some preferred embodiments, the superoxide dismutase composition further comprises the following components in parts by weight: 0.05-1 part of p-hydroxyacetophenone.
The p-hydroxyacetophenone is mainly used as an antioxidant in cosmetics and skin care products, is safer, can be used safely, has no influence on pregnant women generally, and has no poxiness on the p-hydroxyacetophenone. The p-hydroxyacetophenone is widely applied to the fields of medicine, cosmetics and industry, and is a natural plant extract which naturally exists in the roots of stems and leaves of the plants of the Compositae, such as artemisia scoparia, artemisia capillaris, the plants of the Asclepiadaceae, the radix ginseng tylophora, and the like. Multifunctional cosmetic composition with antioxidant and soothing effects; in cosmetics, the product can be used as a substitute of traditional preservatives, has the effects of preserving and enhancing the efficacy of various preservatives, and has excellent stability in high and low pH and temperature ranges. Can be used in various formulas, and has good compatibility with various cosmetic ingredients and packaging materials.
In some preferred embodiments, the superoxide dismutase composition comprises the following components in parts by weight: 0.1-0.6 part of p-hydroxyacetophenone.
In some preferred embodiments, the superoxide dismutase composition further comprises the following components in parts by weight: 0.05-10 parts of 1, 2-hexanediol and 0.05-8 parts of methyl propylene glycol.
The 1, 2-hexanediol has the main function of a humectant in cosmetics and skin care products, has a risk coefficient of 1, is safer, can be used safely, has no influence on pregnant women generally, and has no acne-causing property. The 1, 2-hexanediol is a humectant, belongs to polyol, has antibacterial effect, and has antiseptic effect only when the addition amount is about 5%, but is not equivalent to antiseptic.
Methyl propylene glycol is an organic compound with a molecular formula of C 4 H 10 O 2 Is colorless transparent low-viscosity liquid. Used in polyester resin, paint and other industry, and also used as additive and insecticide for plasticsInsect and bactericide. Used as solvents, moisturizers, emulsifiers, and skin care agents in detergents. Can be used as solvent in skin care product and cosmetic.
In some preferred embodiments, the superoxide dismutase composition comprises the following components in parts by weight: 0.1-5 parts of 1, 2-hexanediol and 0.1-5 parts of methyl propylene glycol.
The second aspect of the present invention provides a method for producing the superoxide dismutase composition, comprising: mixing, heating and dissolving p-hydroxyacetophenone, 1, 2-hexanediol and methyl propylene glycol to obtain a mixed solution; completely and uniformly swelling sodium polyacrylate with hot water, and then adding the mixed solution to uniformly mix; cooling to below 40 ℃; adding superoxide dismutase, lactobacillus fermentation lysate, tetrahydropyrimidine carboxylic acid and fullerene, and stirring.
In the method for producing the superoxide dismutase composition, the temperature at which the p-hydroxyacetophenone, 1, 2-hexanediol and methylpropanediol are mixed and heated is not particularly limited, and those skilled in the art can routinely select them.
In the preparation method of the superoxide dismutase composition, the addition amount of hot water and the temperature of hot water are not particularly limited, and those skilled in the art can make routine selections.
In one embodiment, in the preparation method of the superoxide dismutase composition, 100 parts by weight of hot water is added for preparation, and the temperature of the hot water is higher than 40 ℃.
Wherein, when the composition of the superoxide dismutase does not contain the tetrahydro-methyl pyrimidine carboxylic acid, the p-hydroxy acetophenone, the 1, 2-hexanediol or the methyl propylene glycol, the added weight part of the tetrahydro-methyl pyrimidine carboxylic acid, the p-hydroxy acetophenone, the 1, 2-hexanediol or the methyl propylene glycol is 0 in the preparation method of the composition of the superoxide dismutase.
Examples
The technical scheme of the present invention is described in detail below by way of examples, but the scope of the present invention is not limited to the examples. The raw materials in the present invention are all commercially available unless otherwise specified.
Sodium polyacrylate in this application is available from Zhengzhou Kangyuan chemical company under the model number C.
Superoxide dismutase in this application is purchased from Hebei Tuo sea biotechnology Co., ltd under the product number 10.
Lactic acid bacteria fermentation lysates in this application were purchased from the company of the Biotechnology, freund, germany.
The fullerenes herein are purchased from the akamatsu funakang biotechnology company.
Example 1
Example 1 provides a superoxide dismutase composition comprising the following components in parts by weight: 2.5 parts of sodium polyacrylate, 2.5 parts of lactobacillus fermentation lysate, 3 parts of superoxide dismutase, 0.5 part of fullerene, 2.2 parts of tetrahydropyrimidine carboxylic acid, 0.3 part of p-hydroxyacetophenone, 2.8 parts of 1, 2-hexanediol and 2 parts of methyl propylene glycol.
The preparation method comprises the following steps: mixing, heating and dissolving 0.3 part of p-hydroxyacetophenone, 2.8 parts of 1, 2-hexanediol and 2 parts of methyl propylene glycol to obtain a mixed solution; taking 2.5 parts of sodium polyacrylate, completely swelling uniformly by using 100 parts of hot water with the weight part of 80 ℃, and then adding the sodium polyacrylate into the mixed solution to uniformly mix; cooling to below 40 ℃; adding 3 parts of superoxide dismutase, 2.5 parts of lactobacillus fermentation lysate, 2.2 parts of tetrahydropyrimidine carboxylic acid and 0.5 part of fullerene, and uniformly stirring.
Example 2
Example 2 provides a superoxide dismutase composition comprising the following components in parts by weight: sodium polyacrylate 0.01 part, lactobacillus fermentation lysate 0.02 part, superoxide dismutase 0.1 part, fullerene 0.3 part, tetrahydropyrimidine carboxylic acid 0.01 part, p-hydroxyacetophenone 0.1 part, 1, 2-hexanediol 0.1 part, and methyl propylene glycol 0.1 part.
The preparation method comprises the following steps: mixing, heating and dissolving 0.1 part of p-hydroxyacetophenone, 0.1 part of 1, 2-hexanediol and 0.1 part of methyl propylene glycol to obtain a mixed solution; taking 0.01 part of sodium polyacrylate, completely swelling uniformly by using 100 parts of hot water with the weight part of 80 ℃, and then adding the sodium polyacrylate into the mixed solution to uniformly mix; cooling to below 40 ℃; then adding 0.1 part of superoxide dismutase, 0.02 part of lactobacillus fermentation lysate, 0.01 part of tetrahydropyrimidine carboxylic acid and 0.3 part of fullerene, and stirring uniformly.
Example 3
Example 3 provides a superoxide dismutase composition comprising the following components in parts by weight: 5 parts of sodium polyacrylate, 5 parts of lactobacillus fermentation lysate, 5 parts of superoxide dismutase, 0.8 part of fullerene, 5 parts of tetrahydropyrimidine carboxylic acid, 0.6 part of p-hydroxyacetophenone, 5 parts of 1, 2-hexanediol and 5 parts of methyl propylene glycol.
The preparation method comprises the following steps: mixing, heating and dissolving 0.6 part of p-hydroxyacetophenone, 5 parts of 1, 2-hexanediol and 5 parts of methyl propylene glycol to obtain a mixed solution; taking 5 parts of sodium polyacrylate, completely swelling uniformly by using 100 parts of hot water with the weight part of 80 ℃, and then adding the sodium polyacrylate into the mixed solution to uniformly mix; cooling to below 40 ℃; and adding 5 parts of superoxide dismutase, 5 parts of lactobacillus fermentation lysate, 5 parts of tetrahydropyrimidine carboxylic acid and 0.8 part of fullerene, and uniformly stirring.
Comparative example 1
Comparative example 1 provides a superoxide dismutase composition comprising the following components in parts by weight: 2.5 parts of sodium polyacrylate, 2.5 parts of lactobacillus fermentation lysate, 3 parts of superoxide dismutase, 0.5 part of fullerene, 2.2 parts of tetrahydropyrimidine carboxylic acid, 0.3 part of p-hydroxyacetophenone, 0 part of 1, 2-hexanediol and 0 part of methyl propylene glycol.
The preparation method comprises the following steps: heating and dissolving 0.3 part of p-hydroxyacetophenone; taking 2.5 parts of sodium polyacrylate, completely swelling uniformly by using 100 parts of hot water with the weight part of 80 ℃, and then adding the sodium polyacrylate into the p-hydroxyacetophenone solution for uniform mixing; cooling to below 40 ℃; adding 3 parts of superoxide dismutase, 2.5 parts of lactobacillus fermentation lysate, 2.2 parts of tetrahydropyrimidine carboxylic acid and 0.5 part of fullerene, and uniformly stirring.
Comparative example 2
Comparative example 2 provides a superoxide dismutase composition comprising the following components in parts by weight: 2.5 parts of sodium polyacrylate, 2.5 parts of lactobacillus fermentation lysate, 3 parts of superoxide dismutase, 0.5 part of fullerene, 2.2 parts of tetrahydropyrimidine carboxylic acid, 0 part of p-hydroxyacetophenone, 2.8 parts of 1, 2-hexanediol and 2 parts of methyl propylene glycol.
The preparation method comprises the following steps: mixing 2.8 parts of 1, 2-hexanediol and 2 parts of methyl propylene glycol, heating and dissolving to obtain a mixed solution; taking 2.5 parts of sodium polyacrylate, completely swelling uniformly by using 100 parts of hot water with the weight part of 80 ℃, and then adding the sodium polyacrylate into the mixed solution to uniformly mix; cooling to below 40 ℃; adding 3 parts of superoxide dismutase, 2.5 parts of lactobacillus fermentation lysate, 2.2 parts of tetrahydropyrimidine carboxylic acid and 0.5 part of fullerene, and uniformly stirring.
Comparative example 3
Comparative example 3 provides a superoxide dismutase composition comprising the following components in parts by weight: 2.5 parts of sodium polyacrylate, 2.5 parts of lactobacillus fermentation lysate, 3 parts of superoxide dismutase, 0.5 part of fullerene, 0 part of tetrahydropyrimidine carboxylic acid, 0.3 part of p-hydroxyacetophenone, 2.8 parts of 1, 2-hexanediol and 2 parts of methyl propylene glycol.
The preparation method comprises the following steps: mixing, heating and dissolving 0.3 part of p-hydroxyacetophenone, 2.8 parts of 1, 2-hexanediol and 2 parts of methyl propylene glycol to obtain a mixed solution; taking 2.5 parts of sodium polyacrylate, completely swelling uniformly by using 100 parts of hot water with the weight part of 80 ℃, and then adding the sodium polyacrylate into the mixed solution to uniformly mix; cooling to below 40 ℃; adding 3 parts of superoxide dismutase, 2.5 parts of lactobacillus fermentation lysate and 0.5 part of fullerene, and stirring uniformly.
Evaluation of Performance
The reagents and equipment used in this test were as follows:
SOD detection buffer solution (Biyun day), WST-8 (Biyun day), xanthine oxidase solution (Biyun day), reaction starting solution (Biyun day), and double distilled water; microplate reader (BioTek Co., U.S.A.).
1. Testing of SOD enzyme activity values at different temperatures
The test result shows that 4 samples are subjected to SOD enzyme activity stability detection, and the above example 1 is subjected to treatment under three different temperature conditions of 45 ℃ and normal temperature and 18 ℃ respectively, and then is compared with untreated SOD powder to judge the heat and cold resistance of SOD in the samples and the stability of formula components.
(one) experimental grouping:
SOD powder group and SOD solution prepared in example 1 are treated at 45 deg.c, normal temperature and 18 deg.c to form one group, enzyme solution blank group 1 and zero setting blank group 2, and three experimental holes are formed.
And (II) preparation of a reagent:
(1) Preparation of WST-8/enzyme working solution
An appropriate amount of WST-8/enzyme working solution was prepared in a volume of 160. Mu.L per reaction, and the sample amounts were as shown in Table 1 below, and vortexed and mixed well after the addition. ( Remarks 1: the experiment needs 18 reaction holes, and the proportion is calculated according to 19 holes )
| Number of samples to be measured | 19 |
| SOD detection buffer solution (mu L) | 2869 |
| WST-8(μL) | 152 |
| Enzyme solution (mu L) | 19 |
| WST-8/enzyme working solution (μL) | 3040 |
TABLE 1 WST-8/enzyme working solution formulation Table
(2) Preparation of reaction initiation working solution (1×)
The sample amounts are shown in Table 2 below, and after the addition, the mixture was vortexed and homogenized. ( Remarks 2: in the experiment, 15 holes are added with reaction starting working solution (1X), and the proportion is calculated according to 16 holes )
| Number of reaction holes | 16 |
| SOD detection buffer solution (mu L) | 487.5 |
| Reaction starting liquid (40×) (μl) | 12.5 |
TABLE 2 preparation of reaction initiation working fluid (1X)
(III) sample measurement
(1) Setting a sample hole and a blank control hole by using a 96-well plate, sequentially adding a sample to be tested and other solutions into each hole according to a table 3, and operating in a low-temperature environment to ensure the consistency of the reaction, reduce the error of an experimental result, and quickly using a gun for uniformly mixing after the sample is added;
| sample of | Blank control group 1 | Blank control group 2 | |
| WST-8/enzyme working solution (μL) | 160 | 160 | 160 |
| SOD detection buffer solution (mu L) | - | 20 | 40 |
| Corresponding to the sample to be measured (mu L) | 20 | - | - |
| Reaction starting liquid (mu L) | 20 | 20 | - |
Table 3 sample assay table
(2) Incubation at 37 ℃ for 30 minutes;
(3) The absorbance was measured at 450nm by a microplate reader.
(IV) SOD enzyme activity detection result
TABLE 4 results of SOD enzyme Activity detection
From table 4, it can be obtained: the enzyme activity value of the SOD solution is higher than that of the SOD powder, and meanwhile, the enzyme activity value of the SOD is greatly different under different temperature conditions, wherein the enzyme activity value of the SOD is the highest under the condition of-18 ℃.
2. Testing of SOD enzyme activity values by different protection solutions
The results of this test were that 6 samples were tested for stability of SOD enzyme activity, and the above examples 1-3 and comparative examples 1-3 were treated at-18℃respectively, and then compared with untreated SOD powder for activity of SOD enzyme, thereby judging the heat and cold resistance of SOD in the samples and the stability of the formulation components.
(one) experimental grouping:
SOD powder groups, SOD solutions prepared in examples 1 to 3 and comparative examples 1 to 3 were treated at-18 ℃ to form one group each, enzyme solution blank group 1 and zero-setting blank group 2, and three experimental holes were formed in each group.
And (II) preparation of a reagent:
(1) Preparation of WST-8/enzyme working solution
An appropriate amount of WST-8/enzyme working solution was prepared in a volume of 160. Mu.L per reaction, and the sample amounts were as shown in Table 5 below, and vortexed and mixed well after the addition. ( Remark 3: 27 reaction holes are needed in the experiment, and the proportion is calculated according to 28 holes )
| Number of samples to be measured | 28 |
| SOD detection buffer solution (mu L) | 4228 |
| WST-8(μL) | 224 |
| Enzyme solution (mu L) | 28 |
| WST-8/enzyme working solution (μL) | 4480 |
TABLE 5 WST-8/enzyme working solution formulation Table
(2) Preparation of reaction initiation working solution (1×)
The amount of the sample was as shown in Table 6 below, and after the addition, the mixture was vortexed and homogenized. ( Remarks 4: in the experiment, 24 holes are added with reaction starting working solution (1X), and the proportion is calculated according to 25 holes )
TABLE 6 preparation of reaction initiation working fluid (1X)
(III) sample measurement
(1) Setting a sample hole and a blank control hole by using a 96-well plate, sequentially adding a sample to be tested and other solutions into each hole according to a table 3, and operating in a low-temperature environment to ensure the consistency of the reaction, reduce the error of an experimental result, and quickly using a gun for uniformly mixing after the sample is added;
(2) Incubation at 37 ℃ for 30 minutes;
(3) The absorbance was measured at 450nm by a microplate reader.
(IV) SOD enzyme activity detection result
TABLE 7 SOD enzyme Activity detection results Table
From table 7, it can be derived that: the protecting solution prepared by the above examples and comparative examples can effectively improve the enzyme activity value of SOD, meanwhile, the example 1 is the optimal example at the temperature of-18 ℃, and compared with the examples, the comparative examples lack one or more components, the SOD enzyme activity value of the comparative examples is reduced, so that the components of the protecting solution in the application are mutually synergistic.
From the above experimental data we can derive: the protective solution in the application can effectively improve the enzyme activity value of SOD, and meanwhile, the enzyme activity value is the largest under the condition of-18 ℃.
Finally, it is pointed out that the above examples are only preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above examples, and any other changes, modifications, substitutions, combinations, and simplifications made without departing from the spirit and principles of the present invention should be equivalent to the above examples, and are included in the scope of the present invention.
Claims (4)
1. The superoxide dismutase composition is characterized by comprising the following components in parts by weight: 2.5 parts of sodium polyacrylate, 2.5 parts of lactobacillus fermentation lysate, 3 parts of superoxide dismutase, 0.5 part of fullerene, 2.2 parts of tetrahydropyrimidine carboxylic acid, 0.3 part of p-hydroxyacetophenone, 2.8 parts of 1, 2-hexanediol and 2 parts of methyl propylene glycol.
2. The superoxide dismutase composition is characterized by comprising the following components in parts by weight: sodium polyacrylate 0.01 part, lactobacillus fermentation lysate 0.02 part, superoxide dismutase 0.1 part, fullerene 0.3 part, tetrahydropyrimidine carboxylic acid 0.01 part, p-hydroxyacetophenone 0.1 part, 1, 2-hexanediol 0.1 part, and methyl propylene glycol 0.1 part.
3. The superoxide dismutase composition is characterized by comprising the following components in parts by weight: 5 parts of sodium polyacrylate, 5 parts of lactobacillus fermentation lysate, 5 parts of superoxide dismutase, 0.8 part of fullerene, 5 parts of tetrahydropyrimidine carboxylic acid, 0.6 part of p-hydroxyacetophenone, 5 parts of 1, 2-hexanediol and 5 parts of methyl propylene glycol.
4. A method for producing the superoxide dismutase composition of any one of claims 1 to 3, comprising: mixing, heating and dissolving p-hydroxyacetophenone, 1, 2-hexanediol and methyl propylene glycol to obtain a mixed solution; completely and uniformly swelling sodium polyacrylate with hot water, and then adding the sodium polyacrylate into the mixed solution to uniformly mix; cooling to below 40 ℃; adding superoxide dismutase, lactobacillus fermentation lysate, tetrahydropyrimidine carboxylic acid and fullerene, and stirring.
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