CN108265097A - A kind of extracting method of sturgeon egg protein peptide and application - Google Patents
A kind of extracting method of sturgeon egg protein peptide and application Download PDFInfo
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- CN108265097A CN108265097A CN201810184883.4A CN201810184883A CN108265097A CN 108265097 A CN108265097 A CN 108265097A CN 201810184883 A CN201810184883 A CN 201810184883A CN 108265097 A CN108265097 A CN 108265097A
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P21/00—Preparation of peptides or proteins
- C12P21/06—Preparation of peptides or proteins produced by the hydrolysis of a peptide bond, e.g. hydrolysate products
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- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
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- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
- A61Q19/08—Anti-ageing preparations
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Abstract
Extracting method and application the present invention provides a kind of sturgeon egg protein peptide, method include handling sturgeon ovum, the sturgeon ovum that obtains that treated;Alcohol is added in sturgeon ovum after treatment, impregnates, obtains degreasing sturgeon ovum;Degreasing sturgeon ovum is ground, adds in distilled water, adds protease, enzymolysis, filtering obtain enzymolysis liquid;Enzymolysis liquid is centrifuged, takes supernatant;And be freeze-dried supernatant, obtain sturgeon egg protein peptide.The sturgeon egg protein peptide extracted by the method for the present invention, protein content have antioxidation up to more than 90%.
Description
Technical field
The present invention relates to edible cosmetic product technical field, more particularly, to a kind of extracting method of sturgeon egg protein peptide
And application.
Background technology
Sturgeon ovum, also known as caviar mean fish-egg in Persian, and strictly speaking, only sturgeon ovum just can be described as roe
Sauce, wherein best to originate in the caviar quality in the Caspian Sea for bordering on Iran and Russia.Sturgeon ovum is rich in Multiple components, crude protein
Matter >=20%, crude fat >=16%, crude fibre≤5%, coarse ash≤16%, lysine >=2.0%, rich in mineral such as Fe, Zn
Matter, therefore sturgeon ovum has the characteristics that high protein, low sugar, the amino acid classes in sturgeon egg protein are complete, and proportioning is reasonable, not only
Skin nutrition can effectively be moistened, make skin exquisiteness and it is bright and clean the effect of, can also play the role of rejuvenescent, be a kind of
The untapped high-quality resource for being.
At present, is also rested on for the exploitation of sturgeon ovum the crude product stage, i.e., it is such as current merely through simple allotment processing
A kind of preparation method of disclosed caviar, in the preparation method, add in seasoning smash to pieces stirring after it can be served.But
For the protein peptides of fish-egg, there is presently no relevant method for extraction and purification.
Invention content
In view of the above problems, the present invention provides a kind of method for extracting sturgeon egg protein peptide, including:
Sturgeon ovum is handled, the sturgeon ovum that obtains that treated;Alcohol is added in sturgeon ovum after the treatment, is soaked
Stain obtains degreasing sturgeon ovum;The degreasing sturgeon ovum is ground, adds in distilled water, adds protease, enzymolysis, filtering obtain
Enzymolysis liquid;The enzymolysis liquid is centrifuged, takes supernatant;And be freeze-dried the supernatant, obtain sturgeon ovum
Protein peptides.
In above method, purity >=95% of the alcohol, the quality of the alcohol is treated the sturgeon ovum
4-8 times of quality.
In above method, the dipping is impregnated for constant temperature.
In above method, the temperature of the constant temperature dipping is 30-50 DEG C, and the time of the constant temperature dipping is 30-
60min。
In above method, the quality of the distilled water is 6-15 times of the quality of the degreasing sturgeon ovum.
In above method, the protease include papain, pepsin, trypsase, alkali protease and
Neutral proteinase it is one or more.
In above method, the addition of the protease is 100-10000U/g.
It in above method, further includes, after the protease is added in and before the enzymolysis, adjusts containing described
The pH of the solution of protease is 2.0-11.0.
In above method, the temperature of the enzymolysis is 30-60 DEG C, and the time of the enzymolysis is 0.5-4 hours.
In above method, the rotating speed of the centrifugation is 8000-10000r/min.
The present invention also provides application of the above-mentioned sturgeon egg protein peptide in cosmetics.
The sturgeon egg protein peptide extracted by the method for the present invention, protein content can be removed effectively up to more than 90%
ABTS (2,2- joins nitrogen-two (3- ethyls-benzothiazole -6- sulfonic acid) di-ammonium salts), DPPH (1,1- diphenyl -2- trinitrobenzens
Hydrazine) etc. free radicals, have antioxidation, can be applied to cosmetic field.
Description of the drawings
Fig. 1 is the influence song for the different protease hydrolyzed times ABTS free radical abilities being removed to obtained sturgeon egg protein peptide
Line.
Specific embodiment
The following examples can make those skilled in the art that the present invention be more fully understood, but not limit in any way
The present invention.
The present invention extracts sturgeon egg protein peptide by following steps, and sturgeon egg protein is measured by taking ABTS free radicals as an example
The antioxidation of peptide:
1st, sturgeon ovum is handled:Sturgeon ovum with clear water is cleaned, removes impurity, moisture control is for use, is handled
Sturgeon ovum afterwards;
2nd, degreasing:The 4-8 times of food for measuring (relative to the quality of treated sturgeon ovum) is added in into treated sturgeon ovum
With alcohol, 30-50 DEG C of constant temperature impregnates 30-60min, filters alcohol with filter paper, cleaned repeatedly to no alcohol taste with clear water, obtain degreasing
Sturgeon ovum;Impregnated at 30-50 DEG C 30-60min can ensure sturgeon ovum can complete degreasing, it is and too high or too low for temperature, then it is unfavorable
In sturgeon ovum degreasing;Dip time is too short simultaneously, and degreasing is not complete enough;
3rd, it digests:Degreasing sturgeon ovum is ground into (grain size is about 1.5-2.5mm), 6-15 times is added in and measures (relative to degreasing sturgeon
The quality of fish-egg) distilled water, add in protease (papain, pepsin, trypsase, alkali protease and neutral egg
It is one or more in white enzyme) to 1g substrates (that is, degreasing sturgeon ovum) enzyme activity reach 100-10000U, enzyme activity is effectively ensured
Property, later with diluted acid dilute alkaline soln (sodium hydrate aqueous solution that the dilute hydrochloric acid and mass fraction that such as mass fraction is 5% are 4%)
Adjust pH to 2.0-11.0 (papain 6.5-7.5, pepsin 1.5-2.5, the trypsase of the solution containing protease
7.5-8.5, alkali protease 9.5-10.5, neutral proteinase 6.5-7.5), in 30-60 DEG C of (50-60 DEG C of papain, stomach
0-40 DEG C of protease 3,45-55 DEG C of trypsase, 35-45 DEG C of alkali protease, 45-55 DEG C of neutral proteinase) under digest 0.5-4
Hour, it is filtered with filter paper, obtains enzymolysis liquid;
In this step, it can ensure that the activity of protease is higher by adjusting pH, can preferably digest degreasing sturgeon
Ovum;Meanwhile the 0.5-4 hours abundant enzymolysis for further ensuring degreasing sturgeon ovum are digested at suitable temperature (30-60 DEG C),
If 60 DEG C of temperature > or 30 DEG C of <, enzymatic activity is relatively low, if enzymolysis time is too short, cannot effectively digest degreasing sturgeon ovum, if
Enzymolysis time is too long, then may make the degreasing sturgeon ovum of enzymolysis further combined with influencing enzymolysis product.
4th, it centrifuges:Enzymolysis liquid under the rotating speed of 8000-10000r/min is centrifuged, takes supernatant;
5th, it is freeze-dried:Obtained supernatant is carried out by freeze drier (being purchased from Shanghai precision instrumentation company)
Freeze-drying, wherein, pre-freezing temperature be -30~-25 DEG C, temperature program be -20~-15 DEG C (10h), -10~-8 DEG C
(10h), -5~-3 DEG C (10h), 0~3 DEG C (10h), 5~8 DEG C (30h) obtain sturgeon egg protein peptide;
6th, ABTS antiradical efficiencies measure.
Embodiment 1
1st, sturgeon ovum is handled:Sturgeon ovum (20g) with clear water is cleaned, removes impurity, moisture control is for use, obtains
Treated sturgeon ovum;
2nd, degreasing:The edible of 6 times of amounts (relative to the quality of treated sturgeon ovum) is added in into treated sturgeon ovum
Alcohol, 40 DEG C of constant temperature impregnate 40min, filter alcohol with filter paper, cleaned repeatedly to no alcohol taste with clear water, obtain degreasing sturgeon ovum;
3rd, it digests:Degreasing sturgeon ovum is ground, adds in 8 times of amounts distilled water, addition (relative to the quality of degreasing sturgeon ovum)
Papain to 1g substrates (that is, degreasing sturgeon ovum) enzyme activity reaches 300-3000U, adjusts pH to 6.5-7.5;By more than solution
It is divided into five parts, is digested 0.5 hour, 1 hour, 2 hours, 3 hours and 4 hours at a temperature of 60 DEG C respectively, filtered, obtained with filter paper
To enzymolysis liquid;
4th, it centrifuges:Enzymolysis liquid under the rotating speed of 9000r/min is centrifuged respectively, takes supernatant;
5th, it is freeze-dried:Obtained supernatant by freeze drier is freeze-dried respectively, obtains sturgeon ovum egg
White peptide;
6th, ABTS antiradical efficiencies measure:The different enzymolysis times of the papain of measure are to obtained sturgeon ovum
Protein peptides remove the result is shown in Figure 1 that ABTS antiradical efficiencies influence.
Embodiment 2
1st, sturgeon ovum is handled:Sturgeon ovum (20g) with clear water is cleaned, removes impurity, moisture control is for use, obtains
Treated sturgeon ovum;
2nd, degreasing:The edible of 6 times of amounts (relative to the quality of treated sturgeon ovum) is added in into treated sturgeon ovum
Alcohol, 40 DEG C of constant temperature impregnate 40min, filter alcohol with filter paper, cleaned repeatedly to no alcohol taste with clear water, obtain degreasing sturgeon ovum;
3rd, it digests:Degreasing sturgeon ovum is ground, adds in 8 times of amounts distilled water, addition (relative to the quality of degreasing sturgeon ovum)
Pepsin to 1g substrates (that is, degreasing sturgeon ovum) enzyme activity reaches 300-3000U, adjusts pH to 2.0-2.5;By more than solution point
Into five parts, digested 0.5 hour, 1 hour, 2 hours, 3 hours and 4 hours at a temperature of 30 DEG C respectively, filtered, obtained with filter paper
Enzymolysis liquid;
4th, it centrifuges:Enzymolysis liquid under the rotating speed of 9000r/min is centrifuged respectively, takes supernatant;
5th, it is freeze-dried:Obtained supernatant by freeze drier is freeze-dried respectively, obtains sturgeon ovum egg
White peptide;
6th, ABTS antiradical efficiencies measure:The different enzymolysis times of pepsin are measured to obtained sturgeon egg protein
Peptide removes the result is shown in Figure 1 that ABTS antiradical efficiencies influence.
Embodiment 3
1st, sturgeon ovum is handled:Sturgeon ovum (20g) with clear water is cleaned, removes impurity, moisture control is for use, obtains
Treated sturgeon ovum;
2nd, degreasing:The edible of 6 times of amounts (relative to the quality of treated sturgeon ovum) is added in into treated sturgeon ovum
Alcohol, 40 DEG C of constant temperature impregnate 40min, filter alcohol with filter paper, cleaned repeatedly to no alcohol taste with clear water, obtain degreasing sturgeon ovum;
3rd, it digests:Degreasing sturgeon ovum is ground, adds in 8 times of amounts distilled water, addition (relative to the quality of degreasing sturgeon ovum)
Trypsase to 1g substrates (that is, degreasing sturgeon ovum) enzyme activity reaches 300-3000U, adjusts pH to 7.5-8.5;By more than solution point
Into five parts, digested 0.5 hour, 1 hour, 2 hours, 3 hours and 4 hours at a temperature of 50 DEG C respectively, filtered, obtained with filter paper
Enzymolysis liquid;
4th, it centrifuges:Enzymolysis liquid under the rotating speed of 9000r/min is centrifuged respectively, takes supernatant;
5th, it is freeze-dried:Obtained supernatant by freeze drier is freeze-dried respectively, obtains sturgeon ovum egg
White peptide;
6th, ABTS antiradical efficiencies measure:The different enzymolysis times of the trypsase of measure are to obtained sturgeon ovum egg
White peptide removes the result is shown in Figure 1 that ABTS antiradical efficiencies influence.
Embodiment 4
1st, sturgeon ovum is handled:Sturgeon ovum (20g) with clear water is cleaned, removes impurity, moisture control is for use, obtains
Treated sturgeon ovum;
2nd, degreasing:The edible of 6 times of amounts (relative to the quality of treated sturgeon ovum) is added in into treated sturgeon ovum
Alcohol, 40 DEG C of constant temperature impregnate 40min, filter alcohol with filter paper, cleaned repeatedly to no alcohol taste with clear water, obtain degreasing sturgeon ovum;
3rd, it digests:Degreasing sturgeon ovum is ground, adds in 8 times of amounts distilled water, addition (relative to the quality of degreasing sturgeon ovum)
Alkali protease to 1g substrates (that is, degreasing sturgeon ovum) enzyme activity reaches 300-3000U, adjusts pH to 9.5-10.5;It will be above molten
Liquid is divided into five parts, digests 0.5 hour, 1 hour, 2 hours, 3 hours and 4 hours at a temperature of 40 DEG C, is filtered with filter paper respectively,
Obtain enzymolysis liquid;
4th, it centrifuges:Enzymolysis liquid under the rotating speed of 9000r/min is centrifuged respectively, takes supernatant;
5th, it is freeze-dried:Obtained supernatant by freeze drier is freeze-dried respectively, obtains sturgeon ovum egg
White peptide;
6th, ABTS antiradical efficiencies measure:The different enzymolysis times of the alkali protease of measure are to obtained sturgeon ovum
Protein peptides remove the result is shown in Figure 1 that ABTS antiradical efficiencies influence.
Embodiment 5
1st, sturgeon ovum is handled:Sturgeon ovum (20g) with clear water is cleaned, removes impurity, moisture control is for use, obtains
Treated sturgeon ovum;
2nd, degreasing:The edible of 6 times of amounts (relative to the quality of treated sturgeon ovum) is added in into treated sturgeon ovum
Alcohol, 40 DEG C of constant temperature impregnate 40min, filter alcohol with filter paper, cleaned repeatedly to no alcohol taste with clear water, obtain degreasing sturgeon ovum;
3rd, it digests:Degreasing sturgeon ovum is ground, adds in 8 times of amounts distilled water, addition (relative to the quality of degreasing sturgeon ovum)
Neutral proteinase to 1g substrates (that is, degreasing sturgeon ovum) enzyme activity reaches 300-3000U, adjusts pH to 6.5-7.5;By more than solution
It is divided into five parts, is digested 0.5 hour, 1 hour, 2 hours, 3 hours and 4 hours at a temperature of 40 DEG C respectively, filtered, obtained with filter paper
To enzymolysis liquid;
4th, it centrifuges:Enzymolysis liquid under the rotating speed of 9000r/min is centrifuged respectively, takes supernatant;
5th, it is freeze-dried:Obtained supernatant by freeze drier is freeze-dried respectively, obtains sturgeon ovum egg
White peptide;
6th, ABTS antiradical efficiencies measure:The different enzymolysis times of the neutral proteinase of measure are to obtained sturgeon ovum
Protein peptides remove the result is shown in Figure 1 that ABTS antiradical efficiencies influence.
Embodiment 6
1st, sturgeon ovum is handled:Sturgeon ovum (20g) with clear water is cleaned, removes impurity, moisture control is for use, obtains
Treated sturgeon ovum;
2nd, degreasing:The edible of 4 times of amounts (relative to the quality of treated sturgeon ovum) is added in into treated sturgeon ovum
Alcohol, 30 DEG C of constant temperature impregnate 60min, filter alcohol with filter paper, cleaned repeatedly to no alcohol taste with clear water, obtain degreasing sturgeon ovum;
3rd, it digests:Degreasing sturgeon ovum is ground, adds in 6 times of amounts distilled water, addition (relative to the quality of degreasing sturgeon ovum)
Papain, neutral proteinase and trypsase to 1g substrates (that is, degreasing sturgeon ovum) enzyme activity reach 100-300U, adjust pH
To 6-8;It is digested 4 hours at a temperature of 50 DEG C, is filtered with filter paper, obtain enzymolysis liquid;
4th, it centrifuges:Enzymolysis liquid under the rotating speed of 8000r/min is centrifuged, takes supernatant;
5th, it is freeze-dried:Obtained supernatant by freeze drier is freeze-dried respectively, obtains sturgeon ovum egg
White peptide.
Embodiment 7
1st, sturgeon ovum is handled:Sturgeon ovum (20g) with clear water is cleaned, removes impurity, moisture control is for use, obtains
Treated sturgeon ovum;
2nd, degreasing:The edible of 8 times of amounts (relative to the quality of treated sturgeon ovum) is added in into treated sturgeon ovum
Alcohol, 50 DEG C of constant temperature impregnate 30min, filter alcohol with filter paper, cleaned repeatedly to no alcohol taste with clear water, obtain degreasing sturgeon ovum;
3rd, it digests:Degreasing sturgeon ovum is ground, 15 times is added in and measures (relative to the quality of degreasing sturgeon ovum) distilled water, add
Enter pepsin and alkali protease to 1g substrates (that is, degreasing sturgeon ovum) enzyme activity and reach 3000-10000U, adjust pH to 2-6;
It is digested 4 hours at a temperature of 35 DEG C, is filtered with filter paper, obtain enzymolysis liquid;
4th, it centrifuges:Enzymolysis liquid under the rotating speed of 10000r/min is centrifuged, takes supernatant;
5th, it is freeze-dried:Obtained supernatant by freeze drier is freeze-dried respectively, obtains sturgeon ovum egg
White peptide.
ABTS antiradical efficiency assay methods are as follows:
1) preparation of ABTS working solutions:Weigh ABTS 0.096g, potassium peroxydisulfate (K2S2O8) 0.033 g, it is settled to respectively
Then 25mL mixes ABTS solution with potassium persulfate solution, stored protected from light 12-16h, the ABTS+ ultimately produced in equal volume
Storing solution, that is, ABTS working solutions.
2) ABTS working solutions are diluted to absorbance A at λ=734nm in the range of 0.68-0.72 with ethyl alcohol.
3) 3mL ABTS working solutions are taken, add in 1.5mL ethyl alcohol, absorbance value A is measured at λ=734nm0, with distilled water
For blank control.
4) 3mL ABTS working solutions are taken, 1.5mL dilutions is added in or undiluted sturgeon egg protein peptide solution (is not freeze-dried
When supernatant), absorbance value A is measured at λ=734nm1, using distilled water as blank control.
5) clearance rate calculates:Clearance rate=(A0-A1)/A0× 100%.
With reference to Fig. 1, Fig. 1 shows different protease hydrolyzed times in embodiment 1 to embodiment 5 to obtained sturgeon ovum egg
White peptide removes the influence curve of ABTS free radical abilities.As shown in Figure 1, in the range of 0.5-4 hours, with enzymolysis time
Increase, enzymolysis is more abundant, and the ability that obtained sturgeon egg protein peptide removes ABTS free radicals is stronger, and further may be used by Fig. 1
Know, the ability that ABTS free radicals are removed by the sturgeon egg protein peptide that the enzymolysis of trypsase obtains is most strong.
In addition, with the protein content in the sturgeon egg protein peptide in Kjeldahl nitrogen determination embodiment 6 to embodiment 7, survey
Test result see the table below 1:
Table 1
Embodiment | Protein content |
6 | 91.3% |
7 | 91.7% |
By upper table 1 it is found that the protein content in the sturgeon egg protein peptide extracted by the method for the present invention is more than 90%,
Up to 91.7%, show that this method can effectively extract sturgeon egg protein peptide.
To sum up, the sturgeon egg protein peptide extracted by the method for the present invention, protein content, can be effective more than 90%
Ground is removed but is not limited to ABTS (2,2- connection nitrogen-two (3- ethyls-benzothiazole -6- sulfonic acid) di-ammonium salts) free radical, has antioxygen
Therefore the effect of change, has beauty face-whitening-nourishing, anti-aging function, can be applied to cosmetic field.
It will be understood by those skilled in the art that above example is only exemplary embodiment, in the spirit without departing substantially from the present invention
In the case of range, a variety of variations can be carried out, replaced and changed.
Claims (11)
1. a kind of method for extracting sturgeon egg protein peptide, including:
Sturgeon ovum is handled, the sturgeon ovum that obtains that treated;
Alcohol is added in sturgeon ovum after the treatment, impregnates, obtains degreasing sturgeon ovum;
The degreasing sturgeon ovum is ground, adds in distilled water, adds protease, enzymolysis, filtering obtain enzymolysis liquid;
The enzymolysis liquid is centrifuged, takes supernatant;And
The supernatant is freeze-dried, obtains sturgeon egg protein peptide.
2. according to the method described in claim 1, wherein, purity >=95% of the alcohol, the quality of the alcohol is described
4-8 times of the quality of treated sturgeon ovum.
3. according to the method described in claim 1, wherein, the dipping is impregnated for constant temperature.
4. according to the method described in claim 3, wherein, the temperature of the constant temperature dipping is 30-50 DEG C, what the constant temperature impregnated
Time is 30-60min.
5. according to the method described in claim 1, wherein, the quality of the distilled water is the 6- of the quality of the degreasing sturgeon ovum
15 times.
6. according to the method described in claim 1, wherein, the protease includes papain, pepsin, tryptose
Enzyme, alkali protease and neutral proteinase it is one or more.
7. according to the method described in claim 1, wherein, the addition of the protease is 100-10000U/g.
8. according to the method described in claim 1, further include, after the protease is added in and before the enzymolysis,
The pH for adjusting the solution containing the protease is 2.0-11.0.
9. according to the method described in claim 1, wherein, the temperature of the enzymolysis is 30-60 DEG C, and the time of the enzymolysis is
0.5-4 hours.
10. according to the method described in claim 1, wherein, the rotating speed of the centrifugation is 8000-10000r/min.
11. application of the sturgeon egg protein peptide according to any one of claim 1 to 10 in cosmetics.
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