CN104862364B - Bee pollen form cole Oligopeptide Compositions and preparation method thereof - Google Patents
Bee pollen form cole Oligopeptide Compositions and preparation method thereof Download PDFInfo
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Abstract
The present invention relates to a kind of method that Oligopeptide Compositions are extracted from bee pollen form cole and the Oligopeptide Compositions extracted by this method.Bee pollen form cole Oligopeptide Compositions bioavilability provided by the invention is high, safer in immunology.Invention additionally provides the food containing bee pollen form cole Oligopeptide Compositions, health products or cosmetic composition.
Description
Technical field
The present invention relates to biological technical fields.It is obtained specifically, being extracted the present invention provides one kind from bee pollen form cole
The method of Oligopeptide Compositions and the composition extracted by this method.The invention further relates to extracted from bee pollen form cole
To application of the composition containing oligopeptides in health products and cosmetics.
Background technology
Rape, alias rape flower or rape, Latin literary fame Brassica rapa L, original producton location Europe with one band of the Central Asia,
Belong to annual herb plant, Cruciferae in botany.Rape is now wider in Chinese cultivated, for the agricultural of large area plantation
Industrial crops.Rape Main Cultivation (kind) type includes turnip type rape (Brassica rapa (campestris) at present
L.), mustard type rape (Brassica juncea L.), cabbage type rape (Brassica napus L.).
Bee Pollen (bee pollen) refers to the pollen that honeybee gathers out of flowering nectariferous plant flower pesticide, by honeybee to its
Inside adds in nectar and saliva, is mixed into irregular oblateness, carries the embedding dough for holding trace under the arm of honeybee hind leg above.Melissa
Powder usually contains 200 multiple nutritional components, wherein containing more than 20 kinds of amino acid, more than 100 kinds of enzyme and coenzyme, 10 multivitamins,
More than 30 kinds of macroelement and trace element, 2% or so nucleic acid and a variety of organic acids, unrighted acid, flavonoid
Object and hormone etc..Bee Pollen is described as " nutrition library of concentration " " the miniature natural medicament of concentration " and " complete nutrition by people as a result,
Food " etc..Modern medicine proves that Bee Pollen has effects that following：(1) functions of intestines and stomach is adjusted：There are many sterilization component, energy for pollen
Kill Escherichia coli etc.；(2) human body comprehensive immune function is improved；(3) brain cardiovascular disease is prevented：Flavonoid in pollen
Object can effectively remove deposition fatty on vascular wall, so as to act to soften blood vessel and reducing blood lipid；(4) anti-aging, beauty is made
With：Belonging to the cosmetics of trophism, the ingredients such as VE, superoxide dismutase (SOD), selenium in pollen can moisten nutrition skin,
Recover the elastic and bright and clean of skin；(5) nervous system is adjusted, promotes sleep etc..
Oligopeptides refers to that the smaller polypeptide of amino acid number, such as amino acid are less than the polypeptide of 10, particularly containing 1,2
Or the polypeptide of 3 amino acid.In human body, oligopeptides can be rapidly absorbed into the circulatory system and be utilized by tissue with higher efficiency.
Protein content is quite abundant in bee pollen form cole, accounts for the 7%~35% of its dry weight, and wherein water-solubility protein is not
To 30%, and protein exists with macromolecular state, and bioavilability is low, and easily causes the anaphylaxis of part population anti-
It should.In nutrition and cosmetic industry, it is common that the active ingredient substance of processing acquisition higher purity is extracted to bee pollen form cole
Means.It has been reported that extracting method be mainly mechanical means, microwave method, fermentation process and enzyme solution.Using existing
Protein usually also retains almost complete macromolecular form in the cole melissa powder preparation that method obtains.Complete protein molecular
Unwanted immunological regulation phenomenon is brought sometimes.
CN102746413B reports a kind of bee pollen polysaccharide leach extraction method, wherein by Bee Pollen mechanical crushing, adds in fine
The plain enzyme of dimension, forms melissa amidin in the pasty state after broken wall, is then extracted with 400W ultrasonic powers.
CN101401821B reports the mechanically method with cellulase and Protease Treatment Bee Pollen, to obtain
A kind of bee pollen peptide slice.
At " the Bee Pollen peptide separation purify and immunocompetence research " of Sun Peilong etc., Food Science, the 3rd phase in 2008,
In Vol.29, the means such as Superose-12 gel chromatographies, DEAE Sepharose F.F. ion-exchange chromatographies are employed to locate
Bee pollen form cole is managed, obtains albumen therein, and it was found that the albumen plays an important role of to influence immunity.
This field still needs with safer in immune angle, i.e., contained by the molecular weight of protein or polypeptide it is lower
Cole melissa powder product, such as protein therein or polypeptide are most of to be less than five amino acid containing length, are preferably
The small-molecular peptides of one or two amino acid.
The content of the invention
It extracts from bee pollen form cole to obtain the method for Oligopeptide Compositions the present invention provides a kind of, wherein locating Bee Pollen in advance
It is hydrolyzed again using combined-enzyme method after reason, obtains the composition containing small molecule oligopeptides, which is characterized in that the molecular weight of the oligopeptides
Below 500D is mainly distributed on, preferably in below 300D.In one aspect of the invention, the composition middle-molecular-weihydroxyethyl is
The Gaussian Profile peak area percent of the oligopeptides of below 500D is more than 60%, and preferably greater than 70%.In another side of the present invention
Face, the Oligopeptide Compositions middle-molecular-weihydroxyethyl are that the Gaussian Profile peak area percent of the oligopeptides of below 300D is more than 50%.That is,
The Gaussian Profile peak area percent of peptide containing 1 or 2 or 3 amino acid in the Oligopeptide Compositions is more than 50%.
In the present invention, using first being pre-processed to bee pollen form cole, then bee pollen form cole is hydrolyzed with combined-enzyme method
Method.Wherein, it is alkali process pretreatment to be carried out to bee pollen form cole, i.e., by bee pollen form cole in the solution of about pH9-11
The regular hour is managed, such as 2-3 more than hour.In the one aspect of the present invention, bee pollen form cole is pre-processed
It is that bee pollen form cole is made solution, and its pH is adjusted to about 9-11 (such as with NaOH or Na2CO3Or NaHCO3Or its arbitrary group
The bases such as close and adjust pH), it is more than hour that 2 are kept under the pH, for example, about 3-8 hour.Preferably, the alkali process exists
It is carried out at about 30-60 DEG C, for example, about 40-50 DEG C.
In the present invention, bee pollen form cole, the enzyme that wherein combined-enzyme method uses are hydrolyzed with combined-enzyme method again after the pre-treatment
It is the combination of 3 kinds or 3 kinds or more hydrolases.It must include alkali protease, neutral proteinase in the combination of the hydrolase
And pepsin.
Bee Pollen has complicated structure and ingredient, the saliva including nectar and honeybee etc..Have outside Bee Pollen hard
Outer wall, inside be the complicated inclusion containing various nutriments and reproduction cell, complicated chemical component causes
Many technical problems are run into using the method for enzyme hydrolysis, such as in the prior art using the combined treatment side of single enzyme or some enzymes
Method extends enzymolysis time anyway and increases the amount of enzyme, cannot all make in protein ingredient containing one or more (2-3) ammonia
The ratio of the polypeptide of base acid increases.
Then the pretreatment that alkali process is first carried out to bee pollen form cole that the present invention uses carries out combined-enzyme method hydrolysis and comes again
The method of Oligopeptide Compositions is obtained, the Oligopeptide Compositions that its middle-molecular-weihydroxyethyl is mainly distributed below 500D have been prepared, it is described
Composition middle-molecular-weihydroxyethyl is that the Gaussian Profile peak area percent of the oligopeptides of below 500D is more than 70%, and the oligopeptides combines
Object middle-molecular-weihydroxyethyl is that the Gaussian Profile peak area percent of the oligopeptides of below 300D is more than 50%.It did not report in the prior art
The composition that above-mentioned molecular weight is mainly distributed on the oligopeptides of below 500D is obtained from bee pollen form cole.
In an additional aspect of the present invention, when bee pollen form cole is hydrolyzed using the combined-enzyme method, enzyme is added in
Order be alkali protease, pepsin and neutral proteinase.In the method for the invention, according to the suitable of the addition enzyme
Sequence, the technique that can improve the method for the present invention are much less the time of processing step and operation and save raw material and intermediate wood
Material.
A kind of method for extracting to obtain Oligopeptide Compositions from bee pollen form cole provided by the invention comprises the following steps：
A) bee pollen form cole is ground into 40-150 mesh fine powders；
B) fine powder and deionized water are pressed 1:20-1:40 mixing；
C) mixed liquor is adjusted to pH9-11 with alkali, 40-50 DEG C, when keeping 3-8 small；
D) pretreated mixed liquor is warming up to 50-55 DEG C, by 0.1 weight %-5 weight % of bee pollen form cole dry weight
500,000 u/g alkali proteases are added in, adjust pH to 8-9, keep the temperature 2-4h；
E) pH to 3-4 is adjusted, 50,000 u/g pepsins, 50-60 are added in by 1 weight %-3 weight % of bee pollen form cole dry weight
DEG C, keep 2-4h；
F) pH to 7-8 is adjusted, 150,000 u/g neutral proteinases are added in by 1 weight %-3 weight % of bee pollen form cole dry weight,
40-50 DEG C, keep 2-4h；
G) coarse filtration；
H) activated carbon of 1-3 weight %, 50-55 DEG C of decoloration 0.5-1h are added；
I) clear, pale yellowish liquid is filtered to obtain with diatomite filter；
J) concentrate, be spray-dried or be freeze-dried to obtain yellow to brown ceramic powder.
In the one aspect of this method, pass through above-mentioned steps a)-j) the molecule measurer of oligopeptides in obtained composition
There is following Gaussian Profile：
In the above method of the present invention, preferred mesh number is 50-150 mesh in step a), and 1 is preferably in a proportion of in step b):
20-1:Preferable temperature is about 90 DEG C in 30, step c)；Time is about 10min, and the amount that enzyme is preferably added in step d) is seed oil dish
The 1-1.2 weight % of melissa powder and fine powder dry weight；Preferable temperature is about 50 DEG C；It is preferred that pH8.2-8.5；It holds time as 3-3.2h.
The amount that enzyme is preferably added in step e) is the 1-3 weight % of Brassica campestris L pollen fine powder dry weight, is most preferably 1 weight of fructification dry weight
Measure %；Preferably ph 3.6, preferable temperature are 55 DEG C, time 3-3.2h.The amount that enzyme is preferably added in step f) is bee pollen form cole
The 1-1.2 weight % of fine powder dry weight；Preferable temperature is about 50 DEG C；It is preferred that pH7.3；It holds time as 3-3.2h.It is excellent in step g)
Select 95 DEG C of temperature, time 15min.The amount and hydrolysis time using enzyme used in optimal technical scheme in the method for the present invention
The oligopeptides extract that can be more effectively needed, i.e. molecular weight are mainly distributed on the oligopeptides of below 500D, the present invention's
On one side, the Gaussian Profile peak area percent of the oligopeptides of 300D is more than 50% in the composition.
The present invention provides the composition extracted by the above method from bee pollen form cole, wherein locating in advance to Bee Pollen
It is hydrolyzed again using combined-enzyme method after reason, obtains the composition containing oligopeptides, which is characterized in that the molecular weight of the oligopeptides mainly divides
Cloth is in below 500D, preferably in below 300D.In one aspect of the invention, the composition middle-molecular-weihydroxyethyl is below 500D
Oligopeptides Gaussian Profile peak area percent be more than 60%, preferably greater than 70%.In an additional aspect of the present invention, the widow
Peptide combinations middle-molecular-weihydroxyethyl is that the Gaussian Profile peak area percent of the oligopeptides of below 300D is more than 50%.That is, the oligopeptides
The Gaussian Profile peak area percent of peptide containing 1 or 2 or 3 amino acid in composition is more than 50%.
In an additional aspect of the present invention, the molecular weight of the oligopeptides of the composition small molecular is with following Gauss point
Cloth：
Also containing carbohydrate, vitamin and mineral etc. in the composition provided by the invention extracted from bee pollen form cole
Nutritional ingredient.The method of the present invention can also effectively retain more than bee pollen form cole its in extract in addition to oligopeptides
Its active material.
Bee pollen form cole provided by the invention can be widely used in food, health products, cosmetic field.
The present invention provides preparing food from the Oligopeptide Compositions that bee pollen form cole extracts as elucidated before, protecting
Application in strong product, cosmetics.
In one aspect of the invention, a kind of cosmetic composition is provided, is contained as elucidated before from rape bee
The Oligopeptide Compositions that pollen extracts.
In one aspect of the invention, a kind of Halth-care composition is provided, is contained as elucidated before from rape bee
The Oligopeptide Compositions that pollen extracts.
In one aspect of the invention, a kind of food compositions are provided, are contained as elucidated before from cole melissa
The Oligopeptide Compositions that powder extracts.
Food provided by the invention, health products, the dosage form of cosmetic composition are included for liquid or powder, i.e., oral liquid,
Capsule, tablet, ball etc..
The present invention can be used for food, health products, the additive of cosmetics that can simultaneously carry out industrialized production.
Specific embodiment
With reference to specific embodiment, the invention will be further described, but the protection model of the present invention should not be limited with this
It encloses.
Embodiment 1
1. the preparation of bee pollen form cole powder
Bee pollen form cole is ground into 100 mesh fine powders with crusher.
2. the pretreatment of bee pollen form cole
Brassica campestris L pollen fine powder and deionized water are pressed 1:20 mixing adjust mixed liquor to pH9-11, at 45 DEG C with NaOH
It is lower holding 6-8 it is small when.
3. the hydrolysis of bee pollen form cole
50-55 DEG C is warming up to after heat preservation, adds in the 500000 u/g alkalescence of 1 weight % of Brassica campestris L pollen fine powder dry weight
Protease adjusts pH to 8.5,50 DEG C of heat preservation 3h with 10%NaOH solution.After heat preservation, Brassica campestris L pollen fine powder dry weight is added in
1 weight % 50,000 u/g pepsins, then with 10%HCl solution tune pH to 3.5,50 DEG C of holding 3h.After heat preservation, add in
The 150000 u/g neutral proteinases of 1 weight % of Brassica campestris L pollen fine powder dry weight, with 10%NaOH solution adjust pH to 7.3,50 DEG C
Keep the temperature 3h.After heat preservation, 95 DEG C are warming up to, keeps 20min, inactivation treatment is carried out to enzyme.
After inactivation, with coarse sand cloth filtering reacting liquid.Then in filtrate liquid feeding body 3% powdered active carbon, 50 DEG C are stirred
Decoloration 0.5h is mixed, is then filtered with diatomite filter, obtains clear, pale yellowish liquid.With 0.25-0.5 μm of nuclepore membrane filter
Then filtering carries out ultrafiltration by the film for 10000 dalton of molecular weight that throttles, collects permeate.
Concentration, freeze-drying obtain yellow to brown ceramic powder.
4. the measurement pair Brassica campestris L pollen extract oligopeptides
According to the JY/T 024- of the molecular weight distribution of common high efficient liquid phase analysis method detection small-molecular peptides in the industry
1996, QB/T2879-2007 standards measure the peptide molecular weight distribution of extract, and it is as follows to obtain its Gaussian Profile result：
Embodiment 2
1. the preparation of bee pollen form cole powder
Bee pollen form cole is ground into 100 mesh fine powders with crusher.
2. the hydrolysis of bee pollen form cole
Brassica campestris L pollen fine powder and deionized water are pressed 1:20 mixing, are warming up to 50-55 DEG C, add in Brassica campestris L pollen fine powder
The 500000 u/g alkali proteases of 1 weight % of dry weight adjust pH to 8.5,50 DEG C of heat preservation 6h with 10%NaOH solution.Heat preservation knot
Shu Hou adds in the 50000 u/g pepsins of 1 weight % of Brassica campestris L pollen fine powder dry weight, then with 10%HCl solution tune pH extremely
3.5,50 DEG C of holding 3h.After heat preservation, the 150000 u/g neutral proteinases of 1 weight % of Brassica campestris L pollen fine powder dry weight are added in,
PH to 7.3,50 DEG C of heat preservation 3h are adjusted with 10%NaOH solution.After heat preservation, 95 DEG C are warming up to, keeps 20min, enzyme is carried out
Inactivation treatment.
After inactivation, with coarse sand cloth filtering reacting liquid.Then in filtrate liquid feeding body 3% powdered active carbon, 50 DEG C are stirred
Decoloration 0.5h is mixed, is then filtered with diatomite filter, obtains clear, pale yellowish liquid.With 0.25-0.5 μm of nuclepore membrane filter
Then filtering carries out ultrafiltration by the film for 10000 dalton of molecular weight that throttles, collects permeate.
Concentration, freeze-drying obtain yellow to brown ceramic powder.
3. the measurement pair Brassica campestris L pollen extract oligopeptides
The powder that will be prepared, according to the molecular weight of common high efficient liquid phase analysis method detection small-molecular peptides in the industry
JY/T 024-1996, the QB/T2879-2007 standards of distribution measure the peptide molecular weight distribution of extract, obtain its Gauss
Distribution results are as follows：
Rich in protein in bee pollen form cole, and protein exists with macromolecular state, bioavilability is low, and
Easily cause the allergic reaction of part population.The present invention is pre-processed by alkaline process, and with biological complex enzyme mild hydrolysis
Method, obtain bee pollen form cole Oligopeptide Compositions, macro-molecular protein made to be degraded to the higher oligopeptides of bioavilability, is not broken
The original activity of bad Bee Pollen, it is safer in immune angle, while retain other nutritional ingredients such as polysaccharide, minerals, dimension life
Element etc., and the smell and mouthfeel of product are improved, add its application.
Unless otherwise noted, practice of the invention will use the conventional skill of biotechnology and organic chemistry, inorganic chemistry etc.
Art, it is clear that in addition to being particularly described in above description and embodiment, can also realize the present invention otherwise.It is other this
Technical solution and improvement in invention scope will be obvious to those skilled in the art in the invention.According to the present invention
Introduction, many change and variations are feasible, therefore are within the scope of the present invention.
Claims (5)
1. a kind of extraction from bee pollen form cole obtains the method for Oligopeptide Compositions, wherein bee pollen form cole is pre-processed,
Then hydrolyze to obtain Oligopeptide Compositions with combined-enzyme method, in the Oligopeptide Compositions molecular weight of oligopeptides be mainly distributed on 500D with
Under, wherein the Gaussian Profile peak area percent that the Oligopeptide Compositions middle-molecular-weihydroxyethyl is the oligopeptides of below 500D is more than 60%,
Enzyme includes alkali protease, pepsin and neutral proteinase and adds in the suitable of enzyme used by the combined-enzyme method hydrolysis
Sequence is alkali protease, pepsin and neutral proteinase,
It the described method comprises the following steps：
A) bee pollen form cole is ground into 40-150 mesh fine powders；
B) fine powder is mixed with deionized water by 1: 20-1: 40；
C) mixed liquor is adjusted to pH9-11 with alkali, 40-50 DEG C, when keeping 3-8 small；
D) pretreated mixed liquor is warming up to 50-55 DEG C, is added in by 0.1 weight %-5 weight % of bee pollen form cole dry weight
500000 u/g alkali proteases adjust pH to 8-9, keep the temperature 2-4h；
E) pH to 3-4 is adjusted, 50,000 u/g pepsins is added in by 1 weight %-3 weight % of bee pollen form cole dry weight, 50-60 DEG C, protects
Hold 2-4h；
F) pH to 7-8 is adjusted, 150,000 u/g neutral proteinases, 40-50 are added in by 1 weight %-3 weight % of bee pollen form cole dry weight
DEG C, keep 2-4h；
G) coarse filtration；
H) activated carbon of 1-3 weight %, 50-55 DEG C of decoloration 0.5-1h are added；
I) clear, pale yellowish liquid is filtered to obtain with diatomite filter；
J) concentrate, be spray-dried or be freeze-dried to obtain yellow to brown ceramic powder,
The molecular weight of oligopeptides has following Gaussian Profile in obtained composition：
2. the method for claim 1 wherein the pretreatment is to handle bee pollen form cole in the solution of pH9-11.
3. a kind of bee pollen form cole Oligopeptide Compositions being prepared by method of claim 1, which is characterized in that described group
Close the oligopeptides that is hydrolyzed containing bee pollen form cole in object, in the Oligopeptide Compositions molecular weight of peptide be mainly distributed on 500D with
Under, wherein the Gaussian Profile peak area percent that the Oligopeptide Compositions middle-molecular-weihydroxyethyl is the oligopeptides of below 500D is more than 60%,
Wherein the molecular weight of oligopeptides has following Gaussian Profile：
4. a kind of Halth-care composition, the bee pollen form cole Oligopeptide Compositions containing claim 3.
5. a kind of cosmetic composition, the bee pollen form cole Oligopeptide Compositions containing claim 3.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101058600A (en) * | 2007-04-28 | 2007-10-24 | 中国农业科学院蜜蜂研究所 | Oxidation resistance zymolyte rich in pollen active peptide |
| CN103039709A (en) * | 2013-01-11 | 2013-04-17 | 江西旺大动物科技有限公司 | Preparation method of wall-breaking and enzymolysis bee pollen |
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101058600A (en) * | 2007-04-28 | 2007-10-24 | 中国农业科学院蜜蜂研究所 | Oxidation resistance zymolyte rich in pollen active peptide |
| CN103039709A (en) * | 2013-01-11 | 2013-04-17 | 江西旺大动物科技有限公司 | Preparation method of wall-breaking and enzymolysis bee pollen |
Non-Patent Citations (2)
| Title |
|---|
| 油菜蜂花粉ACE抑制物的制备及分离;李天娇;《中国优秀硕士学位论文全文库 工程科技Ⅰ辑》;20111015(第10期);第B016-143页 * |
| 油菜蜂花粉蛋白酶解条件的研究;朱晓丽等;《食品科学》;20080430;第29卷(第4期);摘要，第234页1.2.2和第235页表1 * |
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