CN110423791A - A kind of reproducibility polypeptide and its preparation method and application - Google Patents
A kind of reproducibility polypeptide and its preparation method and application Download PDFInfo
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- CN110423791A CN110423791A CN201910732432.4A CN201910732432A CN110423791A CN 110423791 A CN110423791 A CN 110423791A CN 201910732432 A CN201910732432 A CN 201910732432A CN 110423791 A CN110423791 A CN 110423791A
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- polypeptide
- selenium
- reproducibility
- active nano
- food
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- 108090000765 processed proteins & peptides Proteins 0.000 title claims abstract description 79
- 229920001184 polypeptide Polymers 0.000 title claims abstract description 77
- 102000004196 processed proteins & peptides Human genes 0.000 title claims abstract description 77
- 238000002360 preparation method Methods 0.000 title claims abstract description 19
- BUGBHKTXTAQXES-UHFFFAOYSA-N Selenium Chemical compound [Se] BUGBHKTXTAQXES-UHFFFAOYSA-N 0.000 claims abstract description 93
- 229940091258 selenium supplement Drugs 0.000 claims abstract description 88
- 239000011669 selenium Substances 0.000 claims abstract description 82
- 229910052711 selenium Inorganic materials 0.000 claims abstract description 82
- 235000018102 proteins Nutrition 0.000 claims abstract description 19
- 102000004169 proteins and genes Human genes 0.000 claims abstract description 19
- 108090000623 proteins and genes Proteins 0.000 claims abstract description 19
- 239000011781 sodium selenite Substances 0.000 claims abstract description 15
- 108091005804 Peptidases Proteins 0.000 claims abstract description 13
- 239000004365 Protease Substances 0.000 claims abstract description 13
- BVTBRVFYZUCAKH-UHFFFAOYSA-L disodium selenite Chemical compound [Na+].[Na+].[O-][Se]([O-])=O BVTBRVFYZUCAKH-UHFFFAOYSA-L 0.000 claims abstract description 13
- 235000019419 proteases Nutrition 0.000 claims abstract description 13
- 239000011655 sodium selenate Substances 0.000 claims abstract description 13
- 229960001471 sodium selenite Drugs 0.000 claims abstract description 13
- 235000015921 sodium selenite Nutrition 0.000 claims abstract description 13
- 230000007071 enzymatic hydrolysis Effects 0.000 claims abstract description 12
- 238000006047 enzymatic hydrolysis reaction Methods 0.000 claims abstract description 12
- 239000000843 powder Substances 0.000 claims abstract description 12
- PMYDPQQPEAYXKD-UHFFFAOYSA-N 3-hydroxy-n-naphthalen-2-ylnaphthalene-2-carboxamide Chemical compound C1=CC=CC2=CC(NC(=O)C3=CC4=CC=CC=C4C=C3O)=CC=C21 PMYDPQQPEAYXKD-UHFFFAOYSA-N 0.000 claims abstract description 11
- 229960001881 sodium selenate Drugs 0.000 claims abstract description 11
- 235000018716 sodium selenate Nutrition 0.000 claims abstract description 11
- 235000017060 Arachis glabrata Nutrition 0.000 claims abstract description 10
- 235000010777 Arachis hypogaea Nutrition 0.000 claims abstract description 10
- 235000018262 Arachis monticola Nutrition 0.000 claims abstract description 10
- 235000020232 peanut Nutrition 0.000 claims abstract description 10
- QCVGEOXPDFCNHA-UHFFFAOYSA-N 5,5-dimethyl-2,4-dioxo-1,3-oxazolidine-3-carboxamide Chemical compound CC1(C)OC(=O)N(C(N)=O)C1=O QCVGEOXPDFCNHA-UHFFFAOYSA-N 0.000 claims abstract description 7
- 102000002322 Egg Proteins Human genes 0.000 claims abstract description 7
- 108010000912 Egg Proteins Proteins 0.000 claims abstract description 7
- 235000014103 egg white Nutrition 0.000 claims abstract description 7
- 210000000969 egg white Anatomy 0.000 claims abstract description 7
- 102000005927 Cysteine Proteases Human genes 0.000 claims abstract description 5
- 108010005843 Cysteine Proteases Proteins 0.000 claims abstract description 5
- 102000005158 Subtilisins Human genes 0.000 claims abstract description 5
- 108010056079 Subtilisins Proteins 0.000 claims abstract description 5
- 239000003513 alkali Substances 0.000 claims abstract description 5
- 241001553178 Arachis glabrata Species 0.000 claims abstract 2
- 102100037486 Reverse transcriptase/ribonuclease H Human genes 0.000 claims abstract 2
- 238000006243 chemical reaction Methods 0.000 claims description 12
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 11
- 239000002994 raw material Substances 0.000 claims description 10
- 230000002779 inactivation Effects 0.000 claims description 9
- 238000000034 method Methods 0.000 claims description 8
- 238000001816 cooling Methods 0.000 claims description 7
- 239000006228 supernatant Substances 0.000 claims description 7
- 102000004190 Enzymes Human genes 0.000 claims description 6
- 108090000790 Enzymes Proteins 0.000 claims description 6
- 235000013305 food Nutrition 0.000 claims description 4
- 101000693530 Staphylococcus aureus Staphylokinase Proteins 0.000 claims 1
- 238000005119 centrifugation Methods 0.000 claims 1
- 238000010521 absorption reaction Methods 0.000 abstract description 7
- 230000000694 effects Effects 0.000 abstract description 6
- 241001465754 Metazoa Species 0.000 abstract description 5
- 230000006870 function Effects 0.000 abstract description 3
- 230000003078 antioxidant effect Effects 0.000 abstract description 2
- 235000013402 health food Nutrition 0.000 abstract description 2
- 230000007935 neutral effect Effects 0.000 abstract description 2
- 230000001151 other effect Effects 0.000 abstract description 2
- 230000008827 biological function Effects 0.000 abstract 1
- 238000004519 manufacturing process Methods 0.000 abstract 1
- 239000000243 solution Substances 0.000 description 20
- 102000035195 Peptidases Human genes 0.000 description 11
- 239000000523 sample Substances 0.000 description 11
- 239000007788 liquid Substances 0.000 description 10
- 244000105624 Arachis hypogaea Species 0.000 description 8
- 238000006722 reduction reaction Methods 0.000 description 8
- 238000005259 measurement Methods 0.000 description 7
- 230000009467 reduction Effects 0.000 description 7
- 239000012488 sample solution Substances 0.000 description 7
- YXFVVABEGXRONW-UHFFFAOYSA-N Toluene Chemical compound CC1=CC=CC=C1 YXFVVABEGXRONW-UHFFFAOYSA-N 0.000 description 6
- 238000002835 absorbance Methods 0.000 description 6
- HHEAADYXPMHMCT-UHFFFAOYSA-N dpph Chemical compound [O-][N+](=O)C1=CC([N+](=O)[O-])=CC([N+]([O-])=O)=C1[N]N(C=1C=CC=CC=1)C1=CC=CC=C1 HHEAADYXPMHMCT-UHFFFAOYSA-N 0.000 description 5
- 235000012054 meals Nutrition 0.000 description 5
- 102000009027 Albumins Human genes 0.000 description 4
- 108010088751 Albumins Proteins 0.000 description 4
- 230000002776 aggregation Effects 0.000 description 4
- 238000004220 aggregation Methods 0.000 description 4
- 238000004458 analytical method Methods 0.000 description 4
- 239000008367 deionised water Substances 0.000 description 4
- 229910021641 deionized water Inorganic materials 0.000 description 4
- 238000002156 mixing Methods 0.000 description 4
- IOLCXVTUBQKXJR-UHFFFAOYSA-M potassium bromide Chemical compound [K+].[Br-] IOLCXVTUBQKXJR-UHFFFAOYSA-M 0.000 description 4
- 238000012360 testing method Methods 0.000 description 4
- 238000004566 IR spectroscopy Methods 0.000 description 3
- ATJFFYVFTNAWJD-UHFFFAOYSA-N Tin Chemical compound [Sn] ATJFFYVFTNAWJD-UHFFFAOYSA-N 0.000 description 3
- 230000005540 biological transmission Effects 0.000 description 3
- 235000013339 cereals Nutrition 0.000 description 3
- 238000009826 distribution Methods 0.000 description 3
- 235000013601 eggs Nutrition 0.000 description 3
- 239000008187 granular material Substances 0.000 description 3
- 239000002105 nanoparticle Substances 0.000 description 3
- 239000002245 particle Substances 0.000 description 3
- -1 potassium ferricyanide Chemical compound 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- 238000005303 weighing Methods 0.000 description 3
- 240000001548 Camellia japonica Species 0.000 description 2
- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 description 2
- 229910003424 Na2SeO3 Inorganic materials 0.000 description 2
- 240000007594 Oryza sativa Species 0.000 description 2
- 235000007164 Oryza sativa Nutrition 0.000 description 2
- 238000004364 calculation method Methods 0.000 description 2
- 239000003638 chemical reducing agent Substances 0.000 description 2
- 235000018597 common camellia Nutrition 0.000 description 2
- 239000000706 filtrate Substances 0.000 description 2
- 235000013372 meat Nutrition 0.000 description 2
- 230000010355 oscillation Effects 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 235000009566 rice Nutrition 0.000 description 2
- 150000003839 salts Chemical class 0.000 description 2
- 229940065287 selenium compound Drugs 0.000 description 2
- 150000003343 selenium compounds Chemical class 0.000 description 2
- 238000001228 spectrum Methods 0.000 description 2
- 208000024172 Cardiovascular disease Diseases 0.000 description 1
- 229920001661 Chitosan Polymers 0.000 description 1
- RYGMFSIKBFXOCR-UHFFFAOYSA-N Copper Chemical compound [Cu] RYGMFSIKBFXOCR-UHFFFAOYSA-N 0.000 description 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Natural products CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- 241000282412 Homo Species 0.000 description 1
- 208000019926 Keshan disease Diseases 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 206010039921 Selenium deficiency Diseases 0.000 description 1
- 238000005411 Van der Waals force Methods 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 239000000853 adhesive Substances 0.000 description 1
- 230000001070 adhesive effect Effects 0.000 description 1
- 230000000840 anti-viral effect Effects 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 238000005253 cladding Methods 0.000 description 1
- 239000011248 coating agent Substances 0.000 description 1
- 238000000576 coating method Methods 0.000 description 1
- 229910052802 copper Inorganic materials 0.000 description 1
- 239000010949 copper Substances 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 235000015872 dietary supplement Nutrition 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 239000002270 dispersing agent Substances 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 230000004064 dysfunction Effects 0.000 description 1
- 235000019441 ethanol Nutrition 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 230000007760 free radical scavenging Effects 0.000 description 1
- 230000005714 functional activity Effects 0.000 description 1
- 125000000524 functional group Chemical group 0.000 description 1
- 238000000227 grinding Methods 0.000 description 1
- 230000036737 immune function Effects 0.000 description 1
- 230000002163 immunogen Effects 0.000 description 1
- 238000002329 infrared spectrum Methods 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 238000002347 injection Methods 0.000 description 1
- 239000007924 injection Substances 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- 229910052742 iron Inorganic materials 0.000 description 1
- FBAFATDZDUQKNH-UHFFFAOYSA-M iron chloride Chemical compound [Cl-].[Fe] FBAFATDZDUQKNH-UHFFFAOYSA-M 0.000 description 1
- 229910052751 metal Inorganic materials 0.000 description 1
- 239000002184 metal Substances 0.000 description 1
- 239000011812 mixed powder Substances 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 235000016709 nutrition Nutrition 0.000 description 1
- 230000035764 nutrition Effects 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- 230000003647 oxidation Effects 0.000 description 1
- 238000007254 oxidation reaction Methods 0.000 description 1
- 239000008188 pellet Substances 0.000 description 1
- 239000008363 phosphate buffer Substances 0.000 description 1
- 230000017854 proteolysis Effects 0.000 description 1
- 150000003254 radicals Chemical class 0.000 description 1
- PRKRTJSKTGAVMH-UHFFFAOYSA-N selenic acid;sodium Chemical compound [Na].O[Se](O)(=O)=O PRKRTJSKTGAVMH-UHFFFAOYSA-N 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 238000005507 spraying Methods 0.000 description 1
- 230000000087 stabilizing effect Effects 0.000 description 1
- 239000013589 supplement Substances 0.000 description 1
- 230000001360 synchronised effect Effects 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- YNJBWRMUSHSURL-UHFFFAOYSA-N trichloroacetic acid Chemical compound OC(=O)C(Cl)(Cl)Cl YNJBWRMUSHSURL-UHFFFAOYSA-N 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K20/00—Accessory food factors for animal feeding-stuffs
- A23K20/20—Inorganic substances, e.g. oligoelements
- A23K20/30—Oligoelements
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/16—Inorganic salts, minerals or trace elements
-
- C—CHEMISTRY; METALLURGY
- C01—INORGANIC CHEMISTRY
- C01B—NON-METALLIC ELEMENTS; COMPOUNDS THEREOF; METALLOIDS OR COMPOUNDS THEREOF NOT COVERED BY SUBCLASS C01C
- C01B19/00—Selenium; Tellurium; Compounds thereof
- C01B19/02—Elemental selenium or tellurium
-
- 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
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
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- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Organic Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Polymers & Plastics (AREA)
- Inorganic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Food Science & Technology (AREA)
- Biochemistry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Microbiology (AREA)
- Animal Husbandry (AREA)
- Biotechnology (AREA)
- Molecular Biology (AREA)
- Mycology (AREA)
- Nutrition Science (AREA)
- Coloring Foods And Improving Nutritive Qualities (AREA)
Abstract
The invention discloses a kind of reproducibility polypeptides and its preparation method and application, the reproducibility polypeptide is that egg white powder and/or tuna albumen and/or peanut cake protein cysteine proteinase and/or ALcalase alkali protease and/or neutral protein are carried out to enzymatic hydrolysis to be made, and has reproducibility.By acting on the reproducibility polypeptide and sodium selenite or sodium selenate, the active nano selenium with biological function can be made, realize body to the two-component absorption of active nano selenium, difunctional effect and other effects.Nanometer selenium partial size prepared by the present invention is smaller, be conducive to absorption and utilization of the body to nanometer selenium, also there is antioxidant activity, be expected to become a kind of novel, safe and efficient, prominent function selenium supplement, be widely used in animal productiong, Feed Manufacturing and application or human health food field.
Description
Technical field
The invention belongs to biochemical industry or field of biological pharmacy, are related to a kind of reproducibility polypeptide, and in particular to a kind of
The preparation method and applications of reproducibility polypeptide.
Background technique
Selenium is one of microelement necessary to humans and animals.The oxidation resistance of selenium and body, antitumaous effect, it is antiviral,
Immune function etc. has important relationship.Numerous studies and clinical test show that selenium deficiency can make some important organs in human body
Dysfunction occurs, so as to cause a variety of diseases, such as tumour, cardiovascular disease and Keshan disease etc..Currently, supplement selenium element
Mainly sodium selenite (Na2SeO3) or sodium selenate (Na2SeO4) etc. inorganic salts, absorption and residual toxicity of the body to inorganic salts
It is not easy to control, there is the risk for causing selenosis.
Relative to inorganic selenium, nanometer selenium has safer more efficiently absorption rate.Pass through inorganic selenium reduction preparation
Elemental selenium is prone to assemble, and forms the insoluble state of aggregation elemental selenium of grey or black, and state of aggregation elemental selenium is living without biology
Property, also it is unfavorable for cell and is absorbed and utilized.It recent studies have shown that, use the macromoleculars such as protein or chitosan as point of elemental selenium
Powder or embedding medium can prepare nano granules of selenium, and can prevent elemental selenium from assembling, but the macromolecular nano granules of selenium formed
Partial size compares larger, is unfavorable for being absorbed and utilized for body, nutrition and functional all relatively simple.
Summary of the invention
In view of the above-mentioned problems, the activity for the polypeptide cladding for being easy to absorb is received the object of the present invention is to provide a kind of partial size is small
Rice selenium new product.
To achieve the above object, the technical scheme adopted by the invention is as follows: a kind of reproducibility polypeptide, the reproducibility egg
White polypeptide carries out enzymatic hydrolysis by food-borne protein raw materials and is made；The food-borne protein sources are egg white powder, tuna egg
At least one of white, peanut cake protein；The enzymatic hydrolysis with enzyme be cysteine proteinase, ALcalase alkali protease, in
At least one of property protease.
Present invention firstly discovers that by using cysteine proteinase and/or ALcalase alkali protease and/or neutrality
Protease can get the albumen with reproducibility to egg white powder and/or tuna albumen and/or peanut meal proteolysis
Polypeptide.The protein raw materials and protease that the present invention uses are not replaceable, do not find other protein raw materials or other protease temporarily
Using can produce the polypeptide with reproducibility.
As one embodiment of the present invention, the preparation method of the reproducibility polypeptide the following steps are included:
S1) pretreatment: taking the food-borne protein raw materials, and it is 1~20 times of food-borne protein raw materials that quality, which is added,
Water mixes.
S2 it) digests: adjusting pH to 4.5~9.5, it is the described of food-borne protein raw materials 0.1%~10.0% that quality, which is added,
Protease, 25~65 DEG C of 5~500min of enzymatic hydrolysis；
S3 it) is centrifuged: being placed in 90 DEG C of inactivation 20min in water-bath, be centrifuged after cooling, take supernatant to get the reduction is arrived
Property polypeptide.
Preferably, S3) inactivation condition is 90 DEG C of water-bath 20min, centrifugal condition 4000g, 20min, 20 DEG C in step.
The reproducibility polypeptide of the invention can be used for preparing active nano selenium.
Reproducibility polypeptide of the present invention has both reduction and package action, adds reducing agent without additional,
Active nano selenium can efficiently be prepared.
A kind of preferred embodiment of active nano selenium, the reduction are used to prepare as reproducibility polypeptide of the present invention
Property the polypeptide method for preparing active nano selenium, comprising the following steps: taking concentration is the reproducibility of 1.0~500mg/mL
Polypeptide, with 0.001~10M, the sodium selenite or sodium selenate that pH is 3.0~8.0 are mixed, and 4~100 DEG C are reacted 10 minutes extremely
10 hours to get arrive the solution containing active nano selenium.
Reproducibility polypeptide of the present invention is both used as natural reducing agent, plays reduction, and be as simple substance
The dispersing agent and covering of selenium, promote sodium selenite (Na2SeO3) or sodium selenate (Na2SeO4) high price selenium (IV, VI) in molecule
The reduction of being reduced property polypeptide generates active nano selenium, and the high price selenium in sodium selenite or selenic acid sodium molecule, which is reduced, generates zeroth order list
While matter selenium, the peptide molecule in solution is by intermolecular or interatomic interactions such as Van der Waals forces, in elemental selenium
Surrounding forms polypeptide clad, prevents the aggregation of elemental selenium, and forms the active nano selenium that partial size is less than 300nm, is conducive to inhale
It receives and plays stabilizing active effect.
Active nano selenium prepared according to the methods of the invention is also claimed in the present invention.
Active nano selenium of the present invention can be used as application of the selenium supplement in food.
Active nano selenium of the present invention can be used as application of the selenium supplement in feed.
The active nano selenium partial size of this patent preparation is small, is the elemental selenium of a kind of high security, high activity, can be used as function
Property ingredient or nutritional supplement be widely used in the fields such as animal productiong, animal feed, medicine, special doctor's Food and hygienical food.
Present invention firstly discovers that utilizing cysteine proteinase and/or ALcalase alkali protease and/or neutral protein
Enzyme digests egg white powder and/or tuna albumen and/or peanut cake protein, and can get has stronger reproducibility
Polypeptide,；The polypeptide and sodium selenite or sodium selenate are acted on, the reduction reaction that can promote selenium generates simple substance
Selenium, it is synchronous with the coating function of polypeptide carry out, can effectively prevent the aggregation solid state of elemental selenium, be formed by nano granules of selenium compared with
It is small, it is conducive to body and absorbs；In addition, imparting the more biological functional activities of nanometer selenium due to being loaded with polypeptide fractions, machine is realized
Body is to the two-component absorption of active nano selenium, difunctional effect and other effects.Active nano selenium partial size prepared by the present invention is smaller, also
With antioxidant activity, be conducive to absorption and utilization of the body to selenium, be expected to become a kind of novel, safe and efficient selenium supplement
Agent is widely used in animal productiong, feed or human health food field.
Detailed description of the invention
Fig. 1 is the infrared spectrogram of tuna polypeptide and polypeptide nano selenium.
Fig. 2 compares for total reducing power of tuna polypeptide and polypeptide nano selenium.
Fig. 3 is that the infrared spectrogram of albumin polypeptide, polypeptide nano selenium and sodium selenite compares.
Fig. 4 is influence of the albumin polypeptide to DPPH clearance rate and reducing power.
Fig. 5 is total reducing power of peanut camellia meal protein polypeptide.
Fig. 6 is influence of the peanut camellia meal protein polypeptide nanometer selenium to the clearance rate of DPPH.
Fig. 7 is electron microscope SEM figure, and A is albumin polypeptide；B is polypeptide nano selenium.
Fig. 8 is that the electron microscope SEM of Peanut Polypeptide schemes.
Fig. 9 is that the transmission electron microscope TEM of active nano selenium of the present invention schemes.
Specific embodiment
To better illustrate the object, technical solutions and advantages of the present invention, below in conjunction with the drawings and specific embodiments pair
The present invention is described further.
Embodiment 1
The preparation of polypeptide: acquisition tuna is cleaned and is made into meat gruel, mixes by solid-liquid mass ratio 1:5 and water.It adjusts
2.0% protease, 50 DEG C of enzymatic hydrolysis 30min are added in pH to 7；90 DEG C of water-bath are placed in, 20min inactivation is centrifuged after cooling, takes
Clear liquid to get arrive the reproducibility polypeptide.
The preparation of active nano selenium: taking concentration is the reproducibility polypeptide 80mL of 5.0mg/mL, and 40mL0.005M,
The sodium selenite or sodium selenate that pH is 6.5 are uniformly mixed, and 60 DEG C of reaction 30min are to get to the solution containing active nano selenium.
Embodiment 2
The preparation of polypeptide: acquisition tuna is cleaned and is made into meat gruel, mixes by solid-to-liquid ratio 1:20 and water.Adjust pH
To 9.5,10% protease, 25 DEG C of enzymatic hydrolysis 500min are added；90 DEG C of water-bath are placed in, 20min inactivation is centrifuged after cooling, takes
Clear liquid to get arrive the reproducibility polypeptide.
The preparation of active nano selenium: taking concentration is the reproducibility polypeptide 100mL of 500mg/mL, and 0.1mL10M,
The sodium selenite or sodium selenate that pH is 7.0 are uniformly mixed, and 4 DEG C of reaction 600min are to get to the solution containing active nano selenium.
Embodiment 3
The preparation of polypeptide: weighing egg white solution, mixes by solid-to-liquid ratio 1:10 and water.PH to 6 is adjusted, 3% albumen is added
Enzyme, 37 DEG C of enzymatic hydrolysis 350min；90 DEG C of water-bath are placed in, 20min inactivation is centrifuged after cooling, takes supernatant to get the reduction is arrived
Property polypeptide.
The preparation of active nano selenium: taking concentration is the reproducibility polypeptide 100mL of 100mg/mL, and 10mL0.1M,
The sodium selenite or sodium selenate that pH is 7.0 are uniformly mixed, and 50 DEG C of reaction 200min are to get to the solution containing active nano selenium.
Embodiment 4
The preparation of polypeptide: weighing egg albumen powder, mixes by solid-to-liquid ratio 1:1 and water.PH to 5 is adjusted, 3% egg is added
White enzyme, 50 DEG C of enzymatic hydrolysis 120min；90 DEG C of water-bath are placed in, 20min inactivation is centrifuged after cooling, and supernatant is taken to go back to get to described
Immunogenic peptide polypeptide.
The preparation of active nano selenium: taking concentration is the reproducibility polypeptide 100mL of 350mg/mL, with 1mL 5M, pH
It is uniformly mixed for 6.5 sodium selenite or sodium selenate, 70 DEG C of reaction 60min are to get to the solution containing active nano selenium.
Embodiment 5
The preparation of polypeptide: weighing peanut meal albumen powder, mixes by solid-to-liquid ratio 1:1 and water.PH to 8.5 is adjusted, is added
3% protease, 50 DEG C of enzymatic hydrolysis 180min；90 DEG C of water-bath are placed in, 20min inactivation is centrifuged after cooling, takes supernatant to get arriving
The reproducibility polypeptide.
The preparation of active nano selenium: taking concentration is the reproducibility polypeptide 100mL of 250mg/mL, with 10mL
0.05M, pH be 7.0 sodium selenite or sodium selenate be uniformly mixed 87 DEG C of reaction 100min to get arrive contain active nano selenium
Solution.
6 active nano selenium functional evaluation of embodiment
(1) experimental method:
I. physical and chemical property determining
(1) measuring method of selenium conversion ratio
It takes polypeptide-selenium solution after 10K Minitype centrifugal screen pipe 5000g is centrifuged 30min, takes lower layer's filtrate and DAN solution
Oscillation mixing, is in mass ratio filtrate: mixed liquor is moved to clean separatory funnel after being stored at room temperature 2h by DAN solution=1:10
In, 5mL toluene is added, funnel is sufficiently placed in iron stand after oscillation and stands 30s, takes supernatant liquid after layering.It is sky with toluene
It is white, absorbance is measured at 380nm.According to the absorbance of gained sample solution, standard curve is corresponded to, the selenium that polypeptide can be obtained is dense
Degree, and then the selenium conversion ratio of sample solution is calculated, the calculation method of selenium conversion ratio is as follows:
(2) polypeptide and the analysis of the infrared spectrum measurement of polypeptide nano selenium structure
Tabletting: pellet technique is used.The range of test is 4000~400cm-1, the analysis rate of spectrum is 4cm-1.It takes
Pure potassium bromide and sample mixed grinding are analyzed in right amount to powdered, are dispersed in sample powder in potassium bromide powder, are taken suitable
Amount mixed-powder is laid on dedicated mold, and using hydraulic press tabletting, pressure is 30Mpa or so, and the tabletting time is 1 minute.
Tabletting require thickness be suitable for, while it is noted that sample granularity and hardness, operate rapidly to prevent the sample moisture absorption, avoid damage to press
Tongue surface smoothness.
Detection: blank control is done with air, measures CO2Infrared spectroscopy after measure sample, and subtract blank test into
Row amendment；Analysis: the difference of functional group is embodied in the difference of position and strong and weak variation in bands of a spectrum, analyzes polypeptide with this, polypeptide-is received
The difference of structure between rice selenium.
(3) polypeptide nano selenium scanning electron microscope (SEM) measures
Polypeptide powder and four kinds of selenium compounds are pre-processed, the double-sided adhesive and tinfoil of suitable size processed are cut, by tinfoil
Stick on sample stage, then draw a small amount of sample solution with capillary and drip on tinfoil, carried out in ion injection instrument metal spraying it
Afterwards, sem analysis is carried out to polypeptide powder and four kinds of selenium compounds respectively, observes the configuration of surface of sample, voltage 15.00kV.
(4) transmission electron microscope (TEM) measures
Solution is made in sample, ultrasonic 10min pipettes a drop sample liquid on copper mesh with 50 μ L pipettors, is placed in 60 DEG C of bakings
Case drying can be placed in TEM and be measured.
Ii. the measurement of total reducing power
Sample solution is prepared with deionized water.Take 2mL sample solution and 2mL 0.2mol/L phosphate buffer (pH6.6),
The potassium ferricyanide of 2mL 1% mixes in test tube.The solution mixed is placed in 50 DEG C of water-bath 20min, is added 2mL's 10%
Solution of trichloroacetic acid, 3000r/min is centrifuged 10min after mixing.2mL supernatant is taken, 2mL deionized water and 0.4mL is added
0.1% iron chloride stands 10min after mixing, measures the absorbance value at 700nm.
Iii. to the measurement of free radical scavenging activity:
Sample solution is prepared with deionized water；Ethyl alcohol compound concentration with 95% is 2 × 10-4The DPPH solution of M.Take 2mL
2mL DPPH solution is added, after mixing avoid light place 30min in sample solution.(blank control group replaces sample with deionized water
Product liquid.) absorbance value at the ultraviolet 517nm of measurement.The clearance rate calculation method of free radical is as follows:
In formula: A0--- the absorbance value of blank group, AS--- the absorbance value of sample sets.
(2) experimental result:
Polypeptide and active nano selenium obtained in embodiment 1 is measured.Pass through Malvern nano particle size point
It is 22nm that analyzer, which measures nanometer selenium average grain diameter, and particle diameter distribution PDI is 0.152, and the conversion ratio that high price selenium becomes elemental selenium is
77.53%.Fig. 1 is the infrared spectroscopy of tuna polypeptide nano selenium, the measurement result of total reducing power such as Fig. 2.Wherein: polypeptide walks
It is rapid a) in polypeptide, polypeptide selenium be step b) in active nano selenium, Fig. 9 be active nano selenium of the present invention transmission electron microscope
TEM figure.
Polypeptide and active nano selenium obtained in embodiment 3 is measured.Pass through Malvern nano particle size point
It is 233nm that analyzer, which measures nanometer selenium average grain diameter, and particle diameter distribution PDI is 0.091, and the conversion ratio that high price selenium becomes elemental selenium is
94.40%.Fig. 3, Fig. 4 are respectively the infrared spectroscopy and reducing power measurement result of albumin polypeptide nanometer selenium, and Fig. 7 is electronic display
Micro mirror SEM figure.Wherein: polypeptide, that is, polypeptide, polypeptide selenium, that is, active nano selenium.
Polypeptide and active nano selenium obtained in embodiment 5 is measured.Pass through Malvern nano particle size point
It is 130nm that analyzer, which measures nanometer selenium average grain diameter, and particle diameter distribution PDI is 0.075, and the conversion ratio that high price selenium becomes elemental selenium is
97.21%.Fig. 5, Fig. 6 are respectively reducing power and DPPH clearance rate measurement result, and Fig. 8 is peanut meal polypeptide electron microscope SEM
Figure.
Finally it should be noted that the above embodiments are merely illustrative of the technical solutions of the present invention, rather than the present invention is protected
The limitation of range is protected, although explaining in detail referring to preferred embodiment to the present invention, those skilled in the art are answered
Work as understanding, it can be with modification or equivalent replacement of the technical solution of the present invention are made, without departing from the reality of technical solution of the present invention
Matter and range.
Claims (8)
1. a kind of reproducibility polypeptide, which is characterized in that the reproducibility polypeptide is carried out by food-borne protein raw materials
Enzymatic hydrolysis is made；The food-borne protein sources are at least one of egg white powder, tuna albumen, peanut cake protein；Institute
It is at least one of cysteine proteinase, ALcalase alkali protease, neutral proteinase that enzymatic hydrolysis, which is stated, with enzyme.
2. the preparation method of reproducibility polypeptide as described in claim 1, which comprises the following steps:
S1) pretreatment: taking the food-borne protein raw materials, and the water that quality is 1~20 times of food-borne protein raw materials is added, mixes
It is even.
S2 it) digests: adjusting pH to 4.5~9.5, the albumen that quality is food-borne protein raw materials 0.1%~10.0% is added
Enzyme, 25~65 DEG C of 5~500min of enzymatic hydrolysis；
S3 it) is centrifuged: being placed in water-bath inactivation, be centrifuged after cooling, take supernatant to get the reproducibility polypeptide is arrived.
3. method according to claim 2, which is characterized in that S3) inactivation condition is 90 DEG C of water-bath 20min, centrifugation in step
Condition is 4000g, 20min, 20 DEG C.
4. reproducibility polypeptide as described in claim 1 is preparing the application in active nano selenium.
5. a kind of method for preparing active nano selenium, which comprises the following steps: taking concentration is 1.0~500mg/mL
The reproducibility polypeptide, the sodium selenite or sodium selenate that and 0.001~10M, pH are 3.0~8.0 be uniformly mixed, 4~
100 DEG C of 10~600min of reaction to get arrive the solution containing active nano selenium.
6. the active nano selenium of method preparation as claimed in claim 5.
7. application of the active nano selenium as claimed in claim 6 as selenium supplement in food.
8. application of the active nano selenium as claimed in claim 6 as selenium supplement in feed.
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| CN112063675A (en) * | 2020-08-04 | 2020-12-11 | 广州大学 | High-hydrolysis-degree polypeptide nano-selenium particle and preparation method and application thereof |
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