CN110301649A - A kind of collagen tripeptide compound and preparation method thereof with anti-aging effects - Google Patents
A kind of collagen tripeptide compound and preparation method thereof with anti-aging effects Download PDFInfo
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- CN110301649A CN110301649A CN201910732155.7A CN201910732155A CN110301649A CN 110301649 A CN110301649 A CN 110301649A CN 201910732155 A CN201910732155 A CN 201910732155A CN 110301649 A CN110301649 A CN 110301649A
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- collagen
- collagen tripeptide
- aging effects
- preparation
- tripeptide compound
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Classifications
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- 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
- A23L33/165—Complexes or chelates
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- 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
- A23L33/17—Amino acids, peptides or proteins
- A23L33/18—Peptides; Protein hydrolysates
-
- 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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- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Organic Chemistry (AREA)
- Mycology (AREA)
- Molecular Biology (AREA)
- Polymers & Plastics (AREA)
- Food Science & Technology (AREA)
- Nutrition Science (AREA)
- Wood Science & Technology (AREA)
- Zoology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Biochemistry (AREA)
- Microbiology (AREA)
- General Chemical & Material Sciences (AREA)
- Biotechnology (AREA)
- Inorganic Chemistry (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Peptides Or Proteins (AREA)
Abstract
The invention discloses a kind of collagen tripeptide compound with anti-aging effects.The collagen tripeptide compound preparation method with anti-aging effects that the invention discloses above-mentioned, it include: that chitosan oligosaccharide is dissolved in organic acid soln, it is subsequently added into collagen, resveratrol and transglutaminase preparation are added, water-bath oscillation is cooling, it is adjusted to neutrality, it adds neutral proteinase to be digested, enzyme deactivation, is dried to obtain cladding wall material；Organic acid zinc is added to the water dissolution, adjusts pH, adds collagen tripeptide, adjusts temperature, heat preservation, dialysis concentration is dried to obtain collagen tripeptide chelated zinc；Collagen tripeptide chelated zinc is added in cladding wall material and is uniformly mixed, spray drying obtains the collagen tripeptide compound with anti-aging effects.Collagen tripeptide compound of the gained with anti-aging effects of the invention has excellent antioxygenic property, has the function that anti-aging；And the holding time is long, can prevent its corruption or oxidation deactivation, while still having bioactivity in vivo.
Description
Technical field
The present invention relates to technical field of bioengineering more particularly to a kind of collagen tripeptide compounds with anti-aging effects
And preparation method thereof.
Background technique
Collagen tripeptide is obtained by collagen or gelatin hydrolysis containing there are three the small peptides of amino acid.Hydrolytic sites are usually located at
The clostridiopetidase A of Y-Gly and certain Special Proteins enzymes, as protease N and protease R hydrolyzable collagen are obtained with Gly-X-Y and its widow
Collagen peptide mixer based on aggressiveness.Collagen hydrolysate by it is different degrees of isolate and purify it is available rich in collagen tripeptide
Mixture or a certain monomer tripeptides.Therefore collagen tripeptide can not only refer to this three peptide mixer, can also refer to that some is single
The tripeptides from collagen.
Collagen tripeptide has the characteristic easily absorbed.There are peptide transporter PepT 1 for human body intestinal canal, can unitransport dipeptides
And tripeptides.Therefore collagen tripeptide can directly be absorbed by the body by intestinal absorption or after incomplete digestion again.At present about glue
There are two types of theoretical for the absorpting form of former tripeptides.It can only first be hydrolyzed to be inhaled again after dipeptides and amino acid one is collagen tripeptide
It receives, another kind is collagen tripeptide other than being absorbed with secondary dipeptides and amino acid, can also directly be absorbed.Such as Aito-
Inoue's, research shows that Gly-Pro-Hyp cannot completely penetrate intestinal epithelial cell, it is usually hydrolyzed to by Aminopeptidase N sweet
Propylhomoserin and Pro-Hyp are conveyed into vivo.And Yamamoto by comparing human body to the absorption of collagen tripeptide and collagen polypeptide and
Homaluria discovery, collagen tripeptide Gly-Pro-Hyp, Gly-Pro-Ala, Gly-Ala-Hyp and their secondary dipeptides can be by
It transports into blood plasma and stablizes transport in human body.No matter which kind of mode absorbs, and collagen tripeptide all can be by human body intestines as small-molecular peptides
Road quickly absorbs.Compared to collagen and its biggish hydrolysate of molecular weight, collagen tripeptide absorbs faster because molecular weight is small.
Collagen tripeptide has multiple biological activities, such as promotes skin regeneration, promotes Ca2+, bone growth, it is anti-oxidant, inhibit
Light aging, antiatherosclerosis determine that collagen tripeptide can be used as the raw material or additive of functional food, promote the battalion of food
Support health-care efficacy.Collagen tripeptide can be applied to nutritional supplement, esthetics food, old people food additive, teenager's food additive
Add agent etc..But existing collagen tripeptide, which enters in body, to be often hydrolyzed, it is difficult to keep bioactivity, and collagen tripeptide has
Stronger antioxygenic property, shelf lives are shorter, therefore need to be protected for collagen tripeptide now, it is made to can smoothly enter into human body
It is absorbed and used.
Summary of the invention
The purpose of the present invention is to solve disadvantages existing in the prior art, and the one kind proposed has anti-aging effects
Collagen tripeptide compound and preparation method thereof.
A kind of collagen tripeptide compound preparation method with anti-aging effects, includes the following steps:
S1, chitosan oligosaccharide is dissolved in organic acid soln, is subsequently added into collagen, add resveratrol and turn glutamy
Amine enzyme preparation, water-bath oscillation is cooling, is adjusted to neutrality, adds neutral proteinase and digested, enzyme deactivation is dried to obtain cladding
Wall material；
S2, organic acid zinc is added to the water to dissolution, adjusts pH to 4-5, add collagen tripeptide, adjust temperature, kept the temperature, thoroughly
Analysis concentration, is dried to obtain collagen tripeptide chelated zinc；
S3, it will be uniformly mixed in collagen tripeptide chelated zinc addition cladding wall material, spray drying is obtained with anti-aging effects
Collagen tripeptide compound.
Chitosan oligosaccharide (chito-oligosaccharid) also known as gucosamine oligosaccharides
It (glucosamineoligosaccharide), is the quasi-oligomer generated by chitosan hydrolyzate, oligosaccharides is usually by 2-12
The low polymer for the linear chain or branched chain that a monosaccharide is formed by β -1,4- glucosides key connection.
In S1 of the present invention, using chitosan oligosaccharide and collagen, under transglutamin-ase 9 enzyme effect, glycosylation is carried out,
The molecular moieties of collagen are unfolded, and the epsilon-amino in structure gradually exposes, and grafting degree is gradually increased, while white Chenopodiaceae is added
Reed alcohol is modified, and is grafted on resveratrol on collagen-chitosan oligosaccharide, on the one hand improves oxidation resistance, on the other hand
Collagen relevant enzyme position is covered by resveratrol and chitosan oligosaccharide, avoid wall material under stomach acid condition by
Cause core material to be released to pepsin effect, Partial digestion is then carried out using neutral proteinase, it is small to generate many solubilities
Molecular polypeptide exposes dissociation group, increases electrostatic charge quantity in solution, to reduce the generation of albumen precipitation aggregation, together
When chitosan oligosaccharide and resveratrol in hydroxyl rich in, improve the hydrophily of cladding wall material, and appropriateness digests and will be embedded in glue
The hydrophobic grouping of former active site of protein is exposed, to reach parent/hydrophobic balance, one side protein hydrophobic side chain is stretched in oil
On the other hand Xiang Zhong is in water phase side by hydroxyl stretching, extension, so that interfacial tension is reduced, the amphiphilic energy of enhancing cladding wall material
Power improves its emulsifying capacity.
Zinc plays a significant role in body, can protect skin health, avoids pachylosis, drying, and wound is promoted to be cured
It closes, and the component part as more than 100 kinds of enzymes in human body, the oxidation resistant ability of effective guarantee body prevent aging.But it is inorganic
Zinc can generate zinc chloride in conjunction with gastric acid, and to the irritating effect of gastrointestinal tract, and organic acid zinc still has certain side effect,
Simultaneously because zinc content is higher, the absorption of the nutrients such as energy antagonism calcium iron.
In S2 of the present invention, organic acid zinc is chelated using collagen tripeptide, although Zn content is lower, chelated zinc activity
Height, it is without any side effects to human body, absorption and utilization of the human body to various nutrients can be effectively facilitated, it will not the battalion such as antagonism calcium iron
The absorption of element is supported, while can be further improved the oxidation resistance of collagen tripeptide.
In S3 of the present invention, collagen tripeptide chelated zinc is coated using cladding wall material, cladding wall material emulsifying capacity is strong, has
The partial size for reducing collagen tripeptide compound is helped, zinc surface can be chelated in collagen tripeptide and form the fine and close oxygen barrier barrier of resistance oxygen, effectively
The oxidation resistance of gained collagen tripeptide compound is improved, the holding time is extended；And cladding wall material uses chitosan oligosaccharide, improves it
Anti-microbial property extends the holding time to further extend the service life of cladding wall material.
Preferably, in S1, the mass ratio of chitosan oligosaccharide and collagen is 6-10:1-1.5.
Preferably, in S1, the molecular weight of chitosan oligosaccharide is less than or equal to 10000.
Chitosan oligosaccharide is as a new class of physiological activator, due to free amine group rich in its structure, thus it is molten
Solution property is greatly improved, and is easy to be absorbed and used, and is lower than 10000 with stronger bioactivity, especially molecular weight
Chitosan oligosaccharide more shows unique physiological activity and functional character.
Preferably, in S1, the mass ratio of resveratrol and collagen is 1-3:1.
Preferably, in S1, water-bath vibration temperature is 30-35 DEG C, and water-bath duration of oscillation is 4-8h.
Preferably, in S1, resveratrol and transglutaminase preparation, the Rate activity of transglutaminase preparation are added
For 10-20U/mg, the activity of transglutaminase preparation is 30-50U/g with the mass ratio of collagen.
Preferably, in S1, the Rate activity of neutral proteinase is 3-4U/mg, activity and the collagen of neutral proteinase
Mass ratio is 0.4-0.9U/g, and hydrolysis temperature is 35-40 DEG C, enzymolysis time 30-50min, maintains pH value in enzymolysis process
It is 7.0.
Preferably, in S1, organic acid is at least one of acetic acid, citric acid, malic acid, tartaric acid.
Preferably, in S2, organic acid zinc is at least one of zinc acetate, zinc citrate, zinc malate, zinc tartrate.
Preferably, in S2, the molar ratio of collagen tripeptide and zinc ion is 3-5:1.
Preferably, in S2,65-85 DEG C is adjusted the temperature to, keeps the temperature 180-300min.
Preferably, in S2, nanofiltration is carried out using nanofiltration membrane in concentration process of dialysing, nanofiltration membrane is preferably that molecular cut off is
The nanofiltration membrane of 200-300Da.
Preferably, in S3, the mass ratio of collagen tripeptide chelated zinc and cladding wall material is 1:4-6.
Preferably, dry using cryospray in S3；It can effectively avoid the drop for leading to oxidation resistance in thermal environment
It is low.
A kind of collagen tripeptide compound with anti-aging effects, it is multiple using the above-mentioned collagen tripeptide with anti-aging effects
Object preparation method is closed to be made.
Specific embodiment
Combined with specific embodiments below the present invention is made further to explain.
Embodiment 1
A kind of collagen tripeptide compound preparation method with anti-aging effects, includes the following steps:
S1,60g chitosan oligosaccharide is dissolved in acetum, is subsequently added into 15g collagen, add 15g resveratrol and
75mg Rate activity is the transglutaminase preparation of 10U/mg, and 4h is vibrated in 35 DEG C of water-baths, cooling, is adjusted to neutrality, adds
1.5mg Rate activity is the neutral protease enzymolysis 30min of 4U/mg, and hydrolysis temperature is 40 DEG C, and maintenance pH value is in enzymolysis process
7.0, enzyme deactivation is dried to obtain cladding wall material；
S2,1.83g zinc acetate is added to the water to dissolution, adjusts pH to 4-5, adds 14g collagen tripeptide, adjust the temperature to
65 DEG C, 300min is kept the temperature, uses molecular cut off to carry out dialysis concentration for the nanofiltration membrane of 200Da, is dried to obtain collagen tripeptide chela
Close zinc；Through detecting, Zn content is 10.26% in collagen tripeptide chelated zinc；
S3, it will be uniformly mixed in 10g collagen tripeptide chelated zinc addition 60g cladding wall material, 50 DEG C of spray drying are had
The collagen tripeptide compound of anti-aging effects.
Embodiment 2
A kind of collagen tripeptide compound preparation method with anti-aging effects, includes the following steps:
S1,100g chitosan oligosaccharide is dissolved in citric acid solution, is subsequently added into 10g collagen, add 30g resveratrol
The transglutaminase preparation for being 20U/mg with 15mg Rate activity, 8h is vibrated in 30 DEG C of water-baths, cooling, is adjusted to neutrality, adds
3mg Rate activity is the neutral protease enzymolysis 50min of 3U/mg, and hydrolysis temperature is 35 DEG C, and it is 7.0 that pH value is maintained in enzymolysis process,
Enzyme deactivation is dried to obtain cladding wall material；
S2, bis- citric acid monohydrate zinc of 61.04g is added to the water to dissolution, adjusts pH to 4-5, adds 252g collagen tripeptide,
85 DEG C are adjusted the temperature to, 180min is kept the temperature, uses molecular cut off to carry out dialysis concentration for the nanofiltration membrane of 280Da, be dried to obtain
Collagen tripeptide chelated zinc；Through detecting, Zn content is 17.41% in collagen tripeptide chelated zinc；
S3, it will be uniformly mixed in 100g collagen tripeptide chelated zinc addition 400g cladding wall material, 40 DEG C of spray drying are had
There is the collagen tripeptide compound of anti-aging effects.
Embodiment 3
A kind of collagen tripeptide compound preparation method with anti-aging effects, includes the following steps:
S1,100g chitosan oligosaccharide is dissolved in malic acid solution, is subsequently added into 20g collagen, add 30g resveratrol
The transglutaminase preparation for being 12U/mg with 75mg Rate activity, 5h is vibrated in 34 DEG C of water-baths, cooling, is adjusted to neutrality, adds
2.70g Rate activity is the neutral protease enzymolysis 35min of 3.7U/mg, and hydrolysis temperature is 38 DEG C, and maintenance pH value is in enzymolysis process
7.0, enzyme deactivation is dried to obtain cladding wall material；
S2, the hydration zinc malate of 233.47g tri- is added to the water dissolution, adjusts pH to 4-5, adds 1260g collagen three
Peptide, adjusts the temperature to 70 DEG C, keeps the temperature 260min, and molecular cut off is used to carry out dialysis concentration for the nanofiltration membrane of 250Da, dry
To collagen tripeptide chelated zinc；Through detecting, Zn content is 12.38% in collagen tripeptide chelated zinc；
S3,100g collagen tripeptide chelated zinc is added in the phosphate buffer that pH is 7.4, adjusts collagen tripeptide chelated zinc
Concentration adds 450g cladding wall material and is uniformly mixed, cryospray is dried to obtain the glue with anti-aging effects to 4.5mol/L
Former three peptide complexes.
Embodiment 4
A kind of collagen tripeptide compound preparation method with anti-aging effects, includes the following steps:
S1,9000g chitosan oligosaccharide is dissolved in tartaric acid solution, is subsequently added into 1200g collagen, it is white adds 3000g
Veratryl alcohol and 2.625g Rate activity are the transglutaminase preparation of 16U/mg, and 7h are vibrated in 32 DEG C of water-baths, cooling, are adjusted to
Property, the neutral protease enzymolysis 45min that 0.255g Rate activity is 3.3U/mg is added, hydrolysis temperature is 36 DEG C, in enzymolysis process
Maintaining pH value is 7.0, and enzyme deactivation is dried to obtain cladding wall material；
S2,2494.8g zinc tartrate is added to the water to dissolution, adjusts pH to 4-5, adds 9800g collagen tripeptide, adjusted
Temperature keeps the temperature 220min, uses molecular cut off to carry out dialysis concentration for the nanofiltration membrane of 250Da, be dried to obtain collagen to 80 DEG C
Three peptide chelated zincs；Through detecting, Zn content is 15.63% in collagen tripeptide chelated zinc；
S3,1000g collagen tripeptide chelated zinc is added in the phosphate buffer that pH is 7.4, adjusts collagen tripeptide chelating
Zinc concentration adds 5500g cladding wall material and is uniformly mixed, cryospray is dried to obtain with anti-aging effects to 3.5mol/L
Collagen tripeptide compound.
Embodiment 5
A kind of collagen tripeptide compound preparation method with anti-aging effects, includes the following steps:
S1,8kg chitosan oligosaccharide is dissolved in citric acid solution, is subsequently added into 1.3kg collagen, add the white Chenopodiaceae of 2.6kg
The pure and mild 3.47g Rate activity of reed is the transglutaminase preparation of 15U/mg, and 6h is vibrated in 33 DEG C of water-baths, cooling, is adjusted to neutrality, then
The neutral protease enzymolysis 40min that 0.22g Rate activity is 3.5U/mg is added, hydrolysis temperature is 37 DEG C, and pH is maintained in enzymolysis process
Value is 7.0, carries out enzyme deactivation using ammonium chloride, then separated with cross-flow ultrafiltration film, and freeze-drying obtains cladding wall material；
S2, bis- citric acid monohydrate zinc of 6.1kg is added to the water to dissolution, adjusts pH to 4-5, adds 33.6kg collagen three
Peptide adjusts the temperature to 75 DEG C, keeps the temperature 240min, molecular cut off is used to carry out dialysis concentration, 30 DEG C of heat for the nanofiltration membrane of 350Da
Wind is dried to obtain collagen tripeptide chelated zinc；Through detecting, Zn content is 14.01% in collagen tripeptide chelated zinc；
S3,10kg collagen tripeptide chelated zinc is added in the phosphate buffer that pH is 7.4, adjusts collagen tripeptide chelated zinc
Concentration is added in 50kg cladding wall material and is uniformly mixed, cryospray is dried to obtain the glue with anti-aging effects to 5mol/L
Former three peptide complexes.
Comparative example 1
A kind of collagen tripeptide compound preparation method, includes the following steps:
S1,8kg chitosan oligosaccharide is dissolved in citric acid solution, is subsequently added into 1.3kg collagen, add the work of 3.47g ratio
Power is the transglutaminase preparation of 15U/mg, and 6h is vibrated in 33 DEG C of water-baths, cooling, is adjusted to neutrality, adds 0.22g Rate activity
For the neutral protease enzymolysis 40min of 3.5U/mg, hydrolysis temperature is 37 DEG C, and it is 7.0 that pH value is maintained in enzymolysis process, using chlorine
Change ammonium and carry out enzyme deactivation, then separated with cross-flow ultrafiltration film, freeze-drying obtains cladding wall material；
S2, bis- citric acid monohydrate zinc of 6.1kg is added to the water to dissolution, adjusts pH to 4-5, adds 33.6kg collagen three
Peptide adjusts the temperature to 75 DEG C, keeps the temperature 240min, molecular cut off is used to carry out dialysis concentration, 30 DEG C of heat for the nanofiltration membrane of 350Da
Wind is dried to obtain collagen tripeptide chelated zinc；
S3,10kg collagen tripeptide chelated zinc is added in the phosphate buffer that pH is 7.4, adjusts collagen tripeptide chelated zinc
Concentration is added in 50kg cladding wall material and is uniformly mixed, cryospray is dried to obtain collagen tripeptide compound to 5mol/L.
Comparative example 2
A kind of collagen tripeptide compound preparation method, includes the following steps:
S1,8kg chitosan oligosaccharide is dissolved in citric acid solution, is subsequently added into 1.3kg collagen, add the white Chenopodiaceae of 2.6kg
The pure and mild 3.47g Rate activity of reed is the transglutaminase preparation of 15U/mg, and 6h are vibrated in 33 DEG C of water-baths, cooling, using ammonium chloride into
Row enzyme deactivation, then separated with cross-flow ultrafiltration film, freeze-drying obtains cladding wall material；
S2, bis- citric acid monohydrate zinc of 6.1kg is added to the water to dissolution, adjusts pH to 4-5, adds 33.6kg collagen three
Peptide adjusts the temperature to 75 DEG C, keeps the temperature 240min, molecular cut off is used to carry out dialysis concentration, 30 DEG C of heat for the nanofiltration membrane of 350Da
Wind is dried to obtain collagen tripeptide chelated zinc；
S3,10kg collagen tripeptide chelated zinc is added in the phosphate buffer that pH is 7.4, adjusts collagen tripeptide chelated zinc
Concentration is added in 50kg cladding wall material and is uniformly mixed, cryospray is dried to obtain collagen tripeptide compound to 5mol/L.
Comparative example 3
A kind of collagen tripeptide compound preparation method, includes the following steps:
S1,8kg chitosan oligosaccharide is dissolved in citric acid solution, is subsequently added into 1.3kg collagen, add the white Chenopodiaceae of 2.6kg
The pure and mild 3.47g Rate activity of reed is the transglutaminase preparation of 15U/mg, and 6h are vibrated in 33 DEG C of water-baths, cooling, using ammonium chloride into
Row enzyme deactivation, then separated with cross-flow ultrafiltration film, freeze-drying obtains cladding wall material；
S2, by 10kg collagen tripeptide be added pH be 7.4 phosphate buffer in, adjust collagen tripeptide chelate zinc concentration to
5mol/L is added in 50kg cladding wall material and is uniformly mixed, and cryospray is dried to obtain collagen tripeptide compound.
Comparative example 4
A kind of collagen tripeptide compound preparation method includes the following steps: for bis- citric acid monohydrate zinc of 6.1kg to be added to the water
Dissolution adjusts pH to 4-5, adds 33.6kg collagen tripeptide, adjusts the temperature to 75 DEG C, keeps the temperature 240min, using retention molecule
Amount is that the nanofiltration membrane of 350Da carries out dialysis concentration, and 30 DEG C of heated-air dryings obtain collagen tripeptide compound.
Comparative example 5
Using collagen tripeptide.
Above-mentioned collagen tripeptide is sold using Wuhan Tiantianhao Biology Products Co., Ltd；Above-mentioned collagen uses Wuxi shellfish
Enlightening bioengineering Co., Ltd is sold；Above-mentioned chitosan oligosaccharide is sold using Zhejiang golden shell pharmaceutcal corporation, Ltd.
Since collagen tripeptide compound obtained by the present invention has the function of anti-aging (mainly anti-oxidant), the present invention is used
Oxidation resistance test is proved in antioxidant activity in vitro test and animal body.
Using collagen tripeptide used in collagen tripeptide compound, comparative example 5 obtained by embodiment 5 and comparative example 1-4 be dissolved in from
Making its mass fraction in sub- water is 6mg/mL, then carries out antioxidant activity in vitro test, specific as follows:
1, measurement of the hexichol for bitter taste diazanyl free radical (DPPH) clearance rate:
A certain amount of DPPH is weighed, is configured to the DPPH mother liquor of 2mol/L with chloroform, with anhydrous second when use
Alcohol dilutes 20 times, is configured to 0.1mmol/L.
Each group sample and DPPH solution are uniformly mixed in the ratio of 1:1, as sample sets；Each group sample and dehydrated alcohol
It is uniformly mixed in the ratio of 1:1, as a control group；Distilled water and DPPH are uniformly mixed in the ratio of 1:1, as blank group,
Room temperature is protected from light 30min, and 8000rpm is centrifuged 10min.It takes supernatant to survey light absorption value at 517nm, and calculates its clearance rate.
The calculation formula of DPPH clearance rate:
DPPH clearance rate=1- (ASample-AControl)/ABlank× 100%.
2, the measurement of hydroxyl radical free radical (OH) clearance rate:
The FeSO of 0.4mL each group sample addition 0.4mL 9mmol/L4Solution, salicylic acid-ethyl alcohol of 0.4mL9mmol/L are molten
The PBS buffer salt solution of liquid and 2.4mL 0.1mol/L pH6.6.Sample is replaced as a control group with the distilled water of equivalent.
It is eventually adding 0.4mL8.8mmol/L H2O2Starting reaction, the water-bath 30min at 37 DEG C, 8000rpm are centrifuged 10min.Take supernatant
Liquid surveys absorbance under 510nm wavelength, and calculates its clearance rate.
The calculation formula of OH clearance rate:
OH clearance rate=(AControl-ASample)/AControl× 100%.
3、Fe2+The measurement of reducing power
It takes 0.5mL each group sample respectively, is added 1% potassium ferricyanide solution of 0.5mL, the heating water bath 20min at 50 DEG C,
10% solution of trichloroacetic acid that 0.5mL is added terminates reaction.Supernatant 1mL is taken after 8000rpm centrifugation 10min, 1mL distillation is added
The ferric chloride solution of 150 μ L 0.1% is added in water after mixing, after reacting at room temperature 10min, surveys and inhales at 700nm wavelength
Light value, A700Value is bigger, illustrates that sample reducing power is stronger.
Experimental result is as follows:
| DPPH clearance rate, % | OH clearance rate, % | A700 | |
| Embodiment 5 | 82.07 | 59.74 | 0.75 |
| Comparative example 1 | 80.63 | 55.31 | 0.71 |
| Comparative example 2 | 78.31 | 54.25 | 0.68 |
| Comparative example 3 | 76.14 | 52.39 | 0.66 |
| Comparative example 4 | 73.36 | 47.28 | 0.62 |
| Comparative example 5 | 80.36 | 77.36 | 0.74 |
As seen from the above table: collagen tripeptide used in comparative example 5 has excellent antioxidation in vitro performance, 4 gained glue of comparative example
Former three peptide complexes are collagen tripeptide chelated zinc, compared with comparative example 5, the decline of mass percent shared by collagen tripeptide, and Er Qieyou
It is not involved in the synthesis of enzyme in vitro in zinc, the antioxygenic property of 4 gained collagen tripeptide compound of comparative example is caused to be weaker than comparative example 5
Collagen tripeptide used.
Relative to comparative example 4, gained collagen tripeptide compound is microencapsulation collagen tripeptide, glue for comparative example 2 and comparative example 3
Cyst wall material is resveratrol and chitosan oligosaccharide modified collagen albumen, improves antioxygenic property by resveratrol and chitosan oligosaccharide, and
Collagen tripeptide or collagen tripeptide chelated zinc are coated by wall material, the fine and close oxygen barrier screen of resistance oxygen can be formed in core surfaces
Barrier effectively improves the oxidation resistance of gained collagen tripeptide compound, but compared with comparative example 4, oxidation resistance is still not
Foot.
Comparative example 1 does not use resveratrol modified collagen albumen, but by carrying out partial hydrolysis to collagen in wall material,
By antioxidant groups exposure in part in collagen, to improve oxidation resistance；And embodiment 5 both uses resveratrol and shell
Oligosaccharides modified collagen albumen, digests further through to collagen, and working along both lines makes 5 gained collagen tripeptide compound of embodiment
Antioxygenic property it is close with collagen tripeptide used in comparative example 5.
It is carried out using collagen tripeptide used in collagen tripeptide compound obtained by embodiment 5 and comparative example 1-4, comparative example 5 internal
Oxidation resistance test, specific as follows:
Experimental animal is divided into 9 groups, successively Normal group, model group, positive controls, 5 groups of embodiment, comparative example 1
Group, 2 groups of comparative example, 3 groups of comparative example, 4 groups of comparative example, 5 groups of comparative example；Every group of 10 cleaning grade rats (male, weight 110-
130g)。
Wherein Normal group feeds basal feed, and other groups are fed high lipid food；High lipid food formula: basal feed 79%
(basal feed is prepared according to GB14924.3-2010), lard 10%, yolk powder 10%, cholesterol 1%.
With distilled water stomach-filling, positive controls are filled with Hedan tablet for each group every morning stomach-filling, Normal group and model group
Stomach, 5 groups of embodiment, 1 group of comparative example, 2 groups of comparative example, 3 groups of comparative example, 4 groups of comparative example embodiment 5 and comparative example 1- is respectively adopted
4 gained collagen tripeptide compound water solution (300mg/kg bw) stomach-fillings, 5 groups of comparative example using collagen tripeptide used in comparative example 5
Aqueous solution (300mg/kg bw) stomach-filling.
The equal ad lib of each group rat and drinking-water during experiment.After persistently feeding 6 weeks, fasting 12h takes blood, and 4 DEG C
4000rpm is centrifuged 10min, prepares serum.
The kit of Bioengineering Research Institute is built up to rat blood serum Glutathione Peroxidase (GSH- using Nanjing
Px), malonaldehyde (MDA), superoxide dismutase (SOD) are measured, and result is as follows:
| GSH-Px, U/mL | MDA, nmol/mL | SOD, U/mL | |
| Normal group | 197.87 | 7.91 | 157.80 |
| Model group | 137.07 | 9.33 | 123.91 |
| Positive controls | 239.81 | 5.07 | 157.11 |
| 5 groups of embodiment | 247.28 | 5.51 | 172.54 |
| 1 group of comparative example | 240.11 | 6.24 | 167.28 |
| 2 groups of comparative example | 236.82 | 6.77 | 165.13 |
| 3 groups of comparative example | 230.07 | 7.21 | 162.04 |
| 4 groups of comparative example | 225.43 | 7.84 | 157.39 |
| 5 groups of comparative example | 229.56 | 7.19 | 161.19 |
As seen from the above table: the present invention is using collagen tripeptide chelated zinc as core material, with resveratrol and chitosan oligosaccharide modified collagen egg
White zymolyte is wall material, makes collagen tripeptide, zinc, resveratrol, chitosan oligosaccharide and collagen enzymatic hydrolysis object collective effect, improves it
In the intracorporal antioxygenic property of machine, to have the function that anti-aging；Simultaneously with resveratrol and chitosan oligosaccharide modified collagen albumen
Zymolyte be wall material, both guaranteed its holding time, prevent its corruption or oxidation deactivation, and guarantee its in vivo still have life
Object activity, makes its oxidation resistance not fail.
The foregoing is only a preferred embodiment of the present invention, but scope of protection of the present invention is not limited thereto,
Anyone skilled in the art in the technical scope disclosed by the present invention, according to the technique and scheme of the present invention and its
Inventive concept is subject to equivalent substitution or change, should be covered by the protection scope of the present invention.
Claims (10)
1. a kind of collagen tripeptide compound preparation method with anti-aging effects, which comprises the steps of:
S1, chitosan oligosaccharide is dissolved in organic acid soln, is subsequently added into collagen, add resveratrol and transglutaminase
Preparation, water-bath oscillation is cooling, is adjusted to neutrality, adds neutral proteinase and digested, enzyme deactivation is dried to obtain cladding wall material；
S2, organic acid zinc is added to the water to dissolution, adjusts pH to 4-5, add collagen tripeptide, adjust temperature, heat preservation is dialysed dense
Contracting, is dried to obtain collagen tripeptide chelated zinc；
S3, it will be uniformly mixed in collagen tripeptide chelated zinc addition cladding wall material, spray drying obtains the glue with anti-aging effects
Former three peptide complexes.
2. according to claim 1 with the collagen tripeptide compound preparation method of anti-aging effects, which is characterized in that S1
In, the mass ratio of chitosan oligosaccharide and collagen is 6-10:1-1.5；Preferably, in S1, the quality of resveratrol and collagen
Than for 1-3:1；Preferably, in S1, water-bath vibration temperature is 30-35 DEG C, and water-bath duration of oscillation is 4-8h.
3. the collagen tripeptide compound preparation method according to claim 1 or claim 2 with anti-aging effects, which is characterized in that
In S1, resveratrol and transglutaminase preparation are added, the Rate activity of transglutaminase preparation is 10-20U/mg, is turned
The activity of glutamine enzyme preparation is 30-50U/g with the mass ratio of collagen.
4. according to claim 1 with the collagen tripeptide compound preparation method of anti-aging effects, feature described in -3 any one
It is, in S1, the Rate activity of neutral proteinase is 3-4U/mg, and the activity of neutral proteinase and the mass ratio of collagen are
0.4-0.9U/g, hydrolysis temperature are 35-40 DEG C, enzymolysis time 30-50min, and it is 7.0 that pH value is maintained in enzymolysis process.
5. according to claim 1 with the collagen tripeptide compound preparation method of anti-aging effects, feature described in -4 any one
It is, in S2, the molar ratio of collagen tripeptide and zinc ion is 3-5:1.
6. according to claim 1 with the collagen tripeptide compound preparation method of anti-aging effects, feature described in -5 any one
It is, in S2, adjusts the temperature to 65-85 DEG C, keep the temperature 180-300min.
7. according to claim 1 with the collagen tripeptide compound preparation method of anti-aging effects, feature described in -6 any one
It is, in S2, nanofiltration is carried out using nanofiltration membrane in concentration process of dialysing, nanofiltration membrane is preferably that molecular cut off is 200-300Da
Nanofiltration membrane.
8. according to claim 1 with the collagen tripeptide compound preparation method of anti-aging effects, feature described in -7 any one
It is, in S3, the mass ratio of collagen tripeptide chelated zinc and cladding wall material is 1:4-6.
9. according to claim 1 with the collagen tripeptide compound preparation method of anti-aging effects, feature described in -7 any one
It is, it is dry using cryospray in S3.
10. a kind of collagen tripeptide compound with anti-aging effects, which is characterized in that have using described in claim 1-9
The collagen tripeptide compound preparation method of anti-aging effects is made.
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