CN101948899B - Method for preparing antihypertensive peptides by using enzymatic degradation on mussel-digested protein - Google Patents
Method for preparing antihypertensive peptides by using enzymatic degradation on mussel-digested protein Download PDFInfo
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Abstract
The invention discloses a method for preparing angiotensin converting enzyme inhibitory peptide (ACEIP) by using enzymatic degradation on mussel-digested protein, which comprises the following steps: 1) mixing defatted dry mussel powder or mussel protein powder serving as raw material with water, then adjusting the pH value to 9.0 to 9.5 so as to obtain a mixed liquor; 2) raising the temperature of the mixed liquor to 60 to 65 DEG C for preheating, then adding alkali protease into the mixed liquor to carry out zymolysing reaction, wherein the zymolysing temperature is 60 to 65 DEG C and the zymolysing time is 180 to 240 minutes; and 3) carrying out enzyme inactivation processing on zymolysing products obtained in step 2 firstly so as to obtain zymolysing liquid; then sequentially carryingout ultra-filtration, freeze drying and chromatography on the obtained zymolysing liquid so as to obtain the ACEIP. The ACEIP prepared by using the method of the invention has the characteristic of strong activity.
Description
Technical field
The invention belongs to technical field of bioengineering, be specifically related to medicine biological technique or functional foodstuff processing technology, especially a kind of enzymolysis mussel protein prepares the method for blood pressure lowering peptide.
Background technology
Angiotensin-converting enzyme (ACE) plays an important role in the blood pressure regulation of human body: the ACE hyperactivity in human circulation or the local organization; The Angiotensin II growing amount is increased; Simultaneously, the amount of bradykinin significantly reduces, and directly causes elevation of blood pressure.Therefore, can play the effect that brings high blood pressure down through the activity that suppresses ACE in the body.
At present; ACE inhibitor has become the hypertensive line medicine of treatment; Compare with the synthetic ACE inhibitor, the ACE that obtains through enzymolysis food endogenous binding protein matter suppresses bioactive peptide and has lot of advantages: the one, and security is high, and having ACE, to suppress active peptide all be to be raw material with food endogenous binding protein matter; Adopt the food grade protease hydrolysis to prepare nontoxicity; The 2nd, have no side effect, the animal experiment of multiple ace inhibitory peptide and Clinical Laboratory result show, do not observe it and have spinoff; The 3rd, preparation condition is gentle, is easy to control, carries out suitability for industrialized production easily, and cost is low, and raw material sources are wide; The 4th, antihypertensive effect is single-minded, and research shows that food source property ace inhibitory peptide only has hypotensive effect to the hyperpietic, and normal arterial pressure person's blood pressure is not had influence; The 5th, be easy to digest and assimilate; Therefore, select the outer protein hydrolysate of suitable proteasome, just might prepare and have ACE and suppress active bioactive peptide, thereby research and develop out bioactive peptide food with hypotensive activity according to industrially scalable.
There are some researches show that people have successfully obtained ace inhibitory peptide from Sunlover 10, zein, fish protein, clam, oyster, algae, mushroom.But these raw materials or protein contnt are on the low side, cause the yield of blood pressure lowering peptide lower, or are people's fresh and alive food, and cost is higher.
Yu Ya etc. (in " it is active that the preparation of oyster functional small peptide and ACE suppress " [J] of Ya, Yang Ruijin, Wang Zhang etc. Wuxi light industry college journal; 2004; 23 (3): 50-52.) utilize the hydrolysis by novo oyster; And through Sephadex G-15 gel chromatography column separating purification, thereby preparation has ACE to suppress active function small peptide.Zhang Miansong (Zhang Miansong " enzyme process prepares the research of clam ace inhibitory peptide " [D]. Southern Yangtze University's master thesis; 2008) optimized the processing condition that neutral protease and Sumizyme MP complex enzyme hydrolysis clam prepare ace inhibitory peptide, inquired into ultrafiltration simultaneously ACE is suppressed active influence.
And mussel is as the main cultivated shellfish of China, and cultured output is very big.But the mussel major part that China produced is main with marketing fresh, minority processing gelation article and ready-to-eat, and all the other process dry products or as feed.Therefore, compare with other marine food, the processing and utilization rate of mussel is extremely low, thereby has caused the significant wastage of mussel resource, and has limited further developing of China's mussel industry.Therefore, development and use mussel protein resource has great importance and wide prospect.
China is in the blank stage basically for the research of mussel ace inhibitory peptide, does not still report for practicable enzymolysis process and separation and purification process.For ability preparation of industrialization blood pressure lowering peptide, should consider hypotensive activity, also to consider the product yield, the economy of production cost etc.And at present known be the method for feedstock production ace inhibitory peptide and not to be suitable for the mussel be that raw material prepares blood pressure lowering peptide (reason is the active low of gained ace inhibitory peptide) with clam, oyster.
Summary of the invention
The technical problem that the present invention will solve provides the method that a kind of Alcalase of utilization Sumizyme MP prepares the mussel blood pressure lowering peptide, and this method mild condition, simple to operate, raw material sources are abundant, cheap, and the blood pressure lowering peptide activity of preparation is stronger.
In order to solve the problems of the technologies described above, the present invention provides a kind of enzymolysis mussel protein to prepare the method for blood pressure lowering peptide, may further comprise the steps:
1), with the dried mussel powder of degreasing or mussel protein powder as raw material, raw material is mixed according to 1: 15～25 weight ratio with water, regulate pH value to 9.0～9.5 (optimal ph is 9.1～9.3) then; Get mixed solution;
2), mixeding liquid temperature is risen to 60～65 ℃ of preheatings, add Sumizyme MP (Alcalase) then and carry out enzyme digestion reaction, the weight of Sumizyme MP is 1.0%～1.5% of raw material, and hydrolysis temperature is 60～65 ℃, and enzymolysis time is 180～240 minutes;
3), with step 2) enzymolysis product of gained handles through the enzyme that goes out earlier, enzymolysis solution; Said enzymolysis solution gets blood pressure lowering peptide successively through ultrafiltration, lyophilize and chromatography.
The improvement for preparing the method for blood pressure lowering peptide as enzymolysis mussel protein of the present invention: step 2) under whipped state the time of preheating be 20～25 minutes.
The further improvements in methods that prepare blood pressure lowering peptide as enzymolysis mussel protein of the present invention: the enzyme that goes out of step 3) is treated to:
With step 2) enzymolysis product of gained is warming up to 90～100 ℃ and kept 8～10 minutes in this temperature, is cooled to 0～20 ℃ again to stop enzyme reaction, and is centrifugal then, enzymolysis solution.
The further improvements in methods that prepare blood pressure lowering peptide as enzymolysis mussel protein of the present invention:
The ultrafiltration of step 3) is: said enzymolysis solution adopts ultra-filtration membrane to carry out uf processing under the working temperature of the WP of 0.1～0.15MPa and 25～45 ℃, the ultrafiltration enzymolysis solution;
The ultrafiltration enzymolysis solution carries out lyophilize, gets ultrafiltration zymolyte dry powder;
Ultrafiltration zymolyte dry powder is carried out chromatography, get blood pressure lowering peptide.
The further improvements in methods that prepare blood pressure lowering peptide as enzymolysis mussel protein of the present invention: used ultra-filtration membrane molecular weight cut-off is 10K dalton (KDa) in the step 3).
The further improvements in methods that prepare blood pressure lowering peptide as enzymolysis mussel protein of the present invention: the chromatography of step 3) is:
Sephadex G-10 or G-15 gel chromatography column are gone up in ultrafiltration zymolyte dry powder dissolving back, and the highly active small-molecular peptides component of collecting gained gets the mussel blood pressure lowering peptide through lyophilize.
The further improvements in methods that prepare blood pressure lowering peptide as enzymolysis mussel protein of the present invention: the enzyme activity of Sumizyme MP is>=9.4 * 10
4U/g.
The further improvements in methods that prepare blood pressure lowering peptide as enzymolysis mussel protein of the present invention: the preparation method of the dried mussel powder of degreasing is following:
After fresh mussel water cleaned up, byssus was removed in shelling, and the homogenate postlyophilization gets the mussel powder, and the mussel powder is adopted the ethanolic soln degreasing of volumetric concentration 95%, after the vacuum-drying the dried mussel powder of degreasing (albumen weight content >=60%).
The further improvements in methods that prepare blood pressure lowering peptide as enzymolysis mussel protein of the present invention: mussel is Mytilus edulis or Mytilus crassitesta Lischke.
In the present invention, dried mussel powder of degreasing or mussel protein powder (albumen weight content >=70%) can obtain through commercial mode, also can prepare voluntarily and get.
The present invention considers the compositing characteristic of mussel protein, selects Sumizyme MP to use enzyme as the enzymolysis of preparation blood pressure lowering peptide, and experimental result shows that the enzymolysis solution of Sumizyme MP enzymolysis gained has the strongest ACE and suppresses active, and yield is very high.Through the effective ace inhibitory peptide in the enrichment zymolyte of ultrafiltration and gel chromatography.
Enzymolysis mussel protein of the present invention prepares the method for blood pressure lowering peptide, has following advantage:
1. the prepared blood pressure lowering peptide of the present invention is that dried mussel powder of degreasing or mussel protein powder make through enzymic hydrolysis, safely, have no side effect.The gained blood pressure lowering peptide has the active effect of significant inhibition ACE, and the hyperpietic is had hypotensive effect, and the normotensive is had health care and prophylactic effect.
2. the raw material that adopts of the present invention is dried mussel powder of degreasing or mussel protein powder, can be from the low value mussel of ocean, also can from the processing waste of mussel, prepare and get; Therefore it is abundant to have the source, and cheap characteristics can effectively improve the added value of mussel through technical transform of the present invention, and the Sustainable development of mussel industry is had great importance.
3. the present invention is a kind of biological enzyme, is prone to through the monitoring to enzymolysis process ace inhibitory peptide farthest discharged, and improves utilization ratio of raw materials.
4. product of the present invention can be used as medicine, protective foods, food, foodstuff additive etc., and craft science is reasonable, and is simple to operate, has stronger industrial implementation property.
The actual usage of the blood pressure lowering peptide that adopts the inventive method preparation and get and consumption are with present existing ace inhibitory peptide.
Than prior preparation method, enzymolysis, ultrafiltration and three kinds of means of gel chromatography have been merged in the present invention simultaneously, and method is more perfect, and the blood pressure lowering peptide activity that obtains is higher.
Description of drawings
Do further explain below in conjunction with the accompanying drawing specific embodiments of the invention.
Fig. 1 is a Mytilus edulis blood pressure lowering peptide Sephadex G-10 gel chromatography spectrogram (embodiment 1);
Fig. 2 is a Mytilus crassitesta Lischke blood pressure lowering peptide Sephadex G-10 gel chromatography spectrogram (embodiment 2);
Fig. 3 is a Mytilus edulis blood pressure lowering peptide Sephadex G-10 gel chromatography spectrogram (embodiment 3);
Fig. 4 is a Mytilus edulis blood pressure lowering peptide Sephadex G-15 gel chromatography spectrogram (embodiment 4);
Fig. 5 is a Mytilus crassitesta Lischke blood pressure lowering peptide Sephadex G-15 gel chromatography spectrogram (Comparative Examples 1).
Embodiment
Utilize embodiment that the present invention is described further below:
Embodiment 1, a kind of enzymolysis mussel protein prepare the method for blood pressure lowering peptide, carry out following steps successively:
1), the dried mussel powder of preparation degreasing:
After fresh Mytilus edulis water cleaned up, byssus was removed in shelling, hollander homogenate to pasty state, lyophilize (50 ℃, 72 hours), the mussel powder.The mussel powder under 40 ℃ of conditions, is added the ethanolic soln degreasing 4h of volumetric concentration 95%, triplicate in the ratio of 1g: 4ml; 100 mesh sieves are pulverized and crossed to mussel powder after the above-mentioned skimming treatment in 40 ℃ of vacuum-dryings (5 hours),, get the dried mussel powder of degreasing of moisture content≤2.34%.
2), take by weighing the dried mussel powder of above-mentioned degreasing 5.0g (albumen weight content 60.23%) in enzyme reactor, it is even to add the 100ml deionized water and stirring, homogeneous 10min is with the NaOH solution adjusting pH to 9.2 of 0.5mol/L; Get mixed solution.
3), mixeding liquid temperature is risen to 60 ℃ stir preheatings, and in whipped state insulation 20 minutes down; (Alcalase, enzyme activity are 9.4 * 10 to add the 0.075g Sumizyme MP then
4U/g) start enzyme digestion reaction, hydrolysis temperature is 60～65 ℃, continuously stirring, and the temperature and the pH constant (temperature is 60～65 ℃, and pH is 9.2) of maintenance reaction system, enzymolysis time is 240 minutes.Get enzymolysis product.
4), the enzymolysis product of step 3) gained is handled through the enzyme that goes out earlier, enzymolysis solution; Said enzymolysis solution gets blood pressure lowering peptide successively through ultrafiltration, lyophilize and chromatography.Be specially:
1., the enzyme that goes out:
Enzymolysis product is rapidly heated to 95 ℃, and, is cooled to 15 ℃ then in 95 ℃ of water-baths enzyme 10min that goes out; Thereby the termination enzyme reaction, then centrifugal 15min under 5000r/min gets supernatant as enzymolysis solution.
2., ultrafiltration
Enzymolysis solution is carried out micro-filtration with the hollow-fibre membrane of 0.45 μ m earlier; Enzymolysis solution behind the micro-filtration is under the working temperature of the WP of 0.13Mpa and 25 ℃; Carry out ultrafiltration through molecular weight cut-off for the 10KDa ultra-filtration membrane; To remove macro-molecular protein and polysaccharide etc., collected is the ultrafiltration enzymolysis solution through liquid.
3., the ultrafiltration enzymolysis solution is carried out lyophilize (50 ℃, 72 hours), ultrafiltration zymolyte dry powder.
4., chromatography:
Ultrafiltration zymolyte dry powder uses deionized water to be mixed with concentration to be 20mg/kg solution, with SephadexG-10 gel chromatography column on this solution of 3ml, carries out wash-out with the ultrapure water of 30ml/h, collects (10min one pipe, every pipe 5mL) and obtains six components.
Wherein, the 5th peak of collection has the highest ACE and suppresses active (see figure 1), with its collection, lyophilize (50 ℃, 72 hours), gets blood pressure lowering peptide 168mg.
The little Huang of blood pressure lowering peptide color and luster that obtains utilizes vitro detection ACE to suppress active HPLC, records its IC
50Be 0.076mg/mL.
Embodiment 2, a kind of enzymolysis mussel protein prepare the method for blood pressure lowering peptide, carry out following steps successively:
1), the dried mussel powder of preparation degreasing:
Substitute fresh Mytilus edulis with fresh Mytilus crassitesta Lischke, all the other are with embodiment 1.
2), take by weighing the dried mussel powder of degreasing 5.0g (albumen weight content 72.56%) in enzyme reactor, it is even to add the 100ml deionized water and stirring, homogeneous 10min is with the NaOH solution adjusting pH to 9.2 of 0.5mol/L; Get mixed solution.
3), mixeding liquid temperature is risen to 60 ℃ stir preheatings, and in whipped state insulation 20 minutes down; (Alcalase, enzyme activity are 9.4 * 10 to add the 0.075g Sumizyme MP then
4U/g) start enzyme digestion reaction, hydrolysis temperature is 60～65 ℃, and continuously stirring keeps the temperature and the pH of reaction system constant, and enzymolysis time is 240 minutes.Get enzymolysis product.
4), the enzymolysis product of step 3) gained is handled through the enzyme that goes out earlier, enzymolysis solution; Said enzymolysis solution gets blood pressure lowering peptide successively through ultrafiltration, lyophilize and chromatography.Be specially:
1., the enzyme that goes out:
With embodiment 1.
2., ultrafiltration
With embodiment 1.
3., with embodiment 1.
4., chromatography:
Ultrafiltration zymolyte dry powder uses deionized water to be mixed with concentration to be 20mg/kg solution, with SephadexG-10 gel chromatography column on this solution of 3mL, carries out wash-out with the ultrapure water of 30ml/h, collects (10min one pipe, every pipe 5mL) and obtains six components.
Wherein, the 5th peak of collection has the highest ACE and suppresses active (see figure 2), with its collection, lyophilize (50 ℃, 72 hours), gets blood pressure lowering peptide 171mg.
The little Huang of blood pressure lowering peptide color and luster that obtains utilizes vitro detection ACE to suppress active HPLC, records its IC
50Be 0.078mg/mL.
Embodiment 3, a kind of enzymolysis mussel protein prepare the method for blood pressure lowering peptide, carry out following steps successively:
1), the dried mussel powder of preparation degreasing:
With embodiment 1.
2), take by weighing the dried mussel powder of degreasing 200.0g (albumen weight content 60.23%) in enzyme reactor, it is even to add the 5L deionized water and stirring, homogeneous 10min is with the NaOH solution adjusting pH to 9.3 of 0.5mol/L; Get mixed solution.
3), mixeding liquid temperature is risen to 65 ℃ stir preheatings, and in whipped state insulation 20 minutes down; (Alcalase, enzyme activity are 9.4 * 10 to add the 2.0g Sumizyme MP then
4U/g) start enzyme digestion reaction, hydrolysis temperature is 65 ℃, and continuously stirring keeps the temperature and the pH of reaction system constant, and enzymolysis time is 180 minutes.Get enzymolysis product.
4), the enzymolysis product of step 3) gained is handled through the enzyme that goes out earlier, enzymolysis solution; Said enzymolysis solution gets blood pressure lowering peptide successively through ultrafiltration, lyophilize and chromatography.Be specially:
1., the enzyme that goes out:
With embodiment 1.
2., ultrafiltration
With embodiment 1.
3., with embodiment 1.
4., chromatography:
Ultrafiltration zymolyte dry powder uses deionized water to be mixed with concentration to be 20mg/kg solution, with SephadexG-10 gel chromatography column on this solution of 3ml, carries out wash-out with the ultrapure water of 30ml/h, collects (10min one pipe, every pipe 5mL) and obtains six components.
Wherein, the 5th peak of collection has the highest ACE and suppresses active (see figure 3), with its collection, lyophilize (50 ℃, 72 hours), gets blood pressure lowering peptide 6253mg.
The little Huang of blood pressure lowering peptide color and luster that obtains utilizes vitro detection ACE to suppress active HPLC, records its IC
50Be 0.083mg/mL.
Embodiment 4, a kind of enzymolysis mussel protein prepare the method for blood pressure lowering peptide, carry out following steps successively:
1), preparation mussel protein powder:
After fresh Mytilus edulis water cleaned up, byssus was removed in shelling, hollander homogenate to pasty state, lyophilize (50 ℃, 72 hours), the mussel powder.The mussel powder under 40 ℃ of conditions, is added the ethanolic soln degreasing 4h of volumetric concentration 95%, triplicate in the ratio of 1g: 4ml; 100 mesh sieves are pulverized and crossed to mussel powder after the above-mentioned skimming treatment in 40 ℃ of vacuum-dryings (5h),, get the dried mussel powder of degreasing of moisture content≤2.51%.
Take by weighing the dried mussel powder of degreasing 200.0g, utilize the sodium hydroxide solution extraction albumen of 6L pH11.5,35 ℃ of temperature; Extraction time is 2.5 hours; Under 4 ℃ of conditions of 3000r/min centrifugal 20 minutes, supernatant used concentration as the hydrochloric acid of 2N the pH value to be transferred to 3.5, precipitates (50 ℃ of centrifugal postlyophilizations; 72 hours) obtain mussel protein powder 176g, protein contnt is 74.32%.
2), take by weighing mussel protein powder 5.0g (albumen weight content 74.32%) in enzyme reactor, it is even to add the 80ml deionized water and stirring, homogeneous 10min is with the NaOH solution adjusting pH to 9.1 of 0.5mol/L; Get mixed solution.
3), mixeding liquid temperature is risen to 60 ℃ stir preheatings, and in whipped state insulation 23 minutes down; (Alcalase, enzyme activity are 9.4 * 10 to add the 0.065g Sumizyme MP then
4U/g) start enzyme digestion reaction, hydrolysis temperature is 60 ℃, and continuously stirring keeps the temperature and the pH of reaction system constant, and enzymolysis time is 200 minutes.Get enzymolysis product.
4), the enzymolysis product of step 3) gained is handled through the enzyme that goes out earlier, enzymolysis solution; Said enzymolysis solution gets blood pressure lowering peptide successively through ultrafiltration, lyophilize and chromatography.Be specially:
1., the enzyme that goes out:
With embodiment 1.
2., ultrafiltration
With embodiment 1.
3., with embodiment 1.
4., chromatography:
Ultrafiltration zymolyte dry powder uses deionized water to be mixed with concentration to be 20mg/kg solution, with SephadexG-15 gel chromatography column on this solution of 3mL, carries out wash-out with the ultrapure water of 30ml/h, collects (10min one pipe, every pipe 5mL) and obtains five components.The 3rd peak has the highest ACE and suppresses active (see figure 4), and its collection, lyophilize are got blood pressure lowering peptide 181mg.The little Huang of blood pressure lowering peptide color and luster that obtains utilizes vitro detection ACE to suppress active HPLC, records its IC
50Be 0.061mg/mL.
Comparative Examples 1, be used in the method that Ya etc. prepares the oyster functional small peptide and prepare the mussel blood pressure lowering peptide, carry out following steps successively:
1), the dried mussel powder of preparation degreasing:
Get Mytilus crassitesta Lischke dry powder, add sherwood oil and soak, suction filtration, air-dry, get the dried mussel powder of degreasing (moisture content≤4.34%).
2), take by weighing the dried mussel powder of degreasing 5.0g (albumen weight content 71.08%) in enzyme reactor; It is even to add the 50ml deionized water and stirring; Mixeding liquid temperature is risen to 60 ℃ stir preheating, (Alcalase, enzyme activity are 9.4 * 10 to add the 0.9575mg Sumizyme MP then
4U/g) under pH8.5, start enzyme digestion reaction, keep the temperature and the pH of reaction system constant, control hydrolysis degree DH is 20%, stopped reaction.
4), the enzymolysis product of step 3) gained is heated to 90～100 ℃, the enzyme 10min that goes out gets the supernatant spraying drying behind centrifugal (3000r/min) 15min, zymolyte dry powder.
5) zymolyte dry powder uses deionized water to be mixed with concentration to be 20mg/kg solution, and with SephadexG-15 gel chromatography column on this solution of 2mL, (0.1mol/L pH4.0) carries out wash-out, collects to obtain four components to receive damping fluid with acetic acid-acetic acid of 21.3ml/h.
Wherein, the 3rd peak of collection has the highest ACE and suppresses active, with its collection, lyophilize (50 ℃, 72 hours), gets blood pressure lowering peptide 122mg.Utilize vitro detection ACE to suppress active HPLC, record its IC
50Be 1.63mg/mL.
Comparative Examples 2, the method for utilizing the Zhang Miansong enzyme process to prepare the clam ace inhibitory peptide prepare the mussel blood pressure lowering peptide, carry out following steps successively:
1), gets Mytilus crassitesta Lischke degreasing dry powder 5.0g (albumen weight content 71.08%) in enzyme reactor; It is even to add the 142ml deionized water and stirring; Mixeding liquid temperature is risen to 60 ℃ stir preheating, (Alcalase, enzyme activity are 9.4 * 10 to add the 0.76g Sumizyme MP then
4U/g) under pH9.0, start enzyme digestion reaction, keep the temperature and the pH of reaction system constant, control hydrolysis time 34.3min, under identical condition, adding 0.05g trypsin enzyme activity then is 5.6 * 10
4U/g), behind the hydrolysis 12.9min, 100 ℃ of enzyme 10min that go out get supernatant behind centrifugal (4000r/min) 20min, are enzymolysis solution.
2), the enzymolysis solution of above-mentioned steps gained under the working temperature of the WP of 0.13Mpa and 25 ℃, carry out ultrafiltration through molecular weight cut-off for the 3KDa ultra-filtration membrane, collect and see through liquid, lyophilize (50 ℃, 72 hours), blood pressure lowering peptide 247mg.Utilize vitro detection ACE to suppress active HPLC, recording its IC50 is 1.87mg/mL.
At last, it is also to be noted that what more than enumerate only is several specific embodiments of the present invention.Obviously, the invention is not restricted to above embodiment, many distortion can also be arranged.All distortion that those of ordinary skill in the art can directly derive or associate from content disclosed by the invention all should be thought protection scope of the present invention.
Claims (8)
1. an enzymolysis mussel protein prepares the method for blood pressure lowering peptide, it is characterized in that may further comprise the steps:
1), with the dried mussel powder of degreasing or mussel protein powder as raw material, raw material is mixed according to 1: 15～25 weight ratio with water, regulate pH value to 9.0～9.5 then; Get mixed solution;
2), mixeding liquid temperature is risen to 60～65 ℃ of preheatings, add Sumizyme MP then and carry out enzyme digestion reaction, the weight of said Sumizyme MP is 1.0%～1.5% of raw material, and hydrolysis temperature is 60～65 ℃, and enzymolysis time is 180～240 minutes; Said Sumizyme MP is enzyme activity>=9.4 * 10
4The Alcalase Sumizyme MP of U/g;
3), with step 2) enzymolysis product of gained handles through the enzyme that goes out earlier, enzymolysis solution; Said enzymolysis solution gets blood pressure lowering peptide successively through ultrafiltration, lyophilize and chromatography.
2. enzymolysis mussel protein according to claim 1 prepares the method for blood pressure lowering peptide, it is characterized in that: preheating 20～25 minutes under whipped state said step 2).
3. enzymolysis mussel protein according to claim 1 and 2 prepares the method for blood pressure lowering peptide, it is characterized in that: the enzyme that goes out of said step 3) is treated to:
With step 2) enzymolysis product of gained is warming up to 90～100 ℃ and kept 8～10 minutes in this temperature, is cooled to 0～20 ℃ again to stop enzyme reaction, and is centrifugal then, enzymolysis solution.
4. enzymolysis mussel protein according to claim 3 prepares the method for blood pressure lowering peptide, it is characterized in that:
The ultrafiltration of said step 3) is: said enzymolysis solution adopts ultra-filtration membrane to carry out uf processing under the working temperature of the WP of 0.1～0.15MPa and 25～45 ℃, the ultrafiltration enzymolysis solution;
Said ultrafiltration enzymolysis solution carries out lyophilize, gets ultrafiltration zymolyte dry powder;
Said ultrafiltration zymolyte dry powder is carried out chromatography, get blood pressure lowering peptide.
5. enzymolysis mussel protein according to claim 4 prepares the method for blood pressure lowering peptide, it is characterized in that: used ultra-filtration membrane molecular weight cut-off is 10K dalton (KDa) in the said step 3).
6. enzymolysis mussel protein according to claim 5 prepares the method for blood pressure lowering peptide, it is characterized in that: the chromatography of said step 3) is:
Sephadex G-10 or G-15 gel chromatography column are gone up in ultrafiltration zymolyte dry powder dissolving back, and the highly active small-molecular peptides component of collecting gained gets the mussel blood pressure lowering peptide through lyophilize.
7. enzymolysis mussel protein according to claim 6 prepares the method for blood pressure lowering peptide, it is characterized in that: the preparation method of the dried mussel powder of said degreasing is following:
After fresh mussel water cleaned up, byssus was removed in shelling, and the homogenate postlyophilization gets the mussel powder, and the mussel powder is adopted the ethanolic soln degreasing of volumetric concentration 95%, after the vacuum-drying the dried mussel powder of degreasing.
8. enzymolysis mussel protein according to claim 7 prepares the method for blood pressure lowering peptide, it is characterized in that: said mussel is Mytilus edulis or Mytilus crassitesta Lischke.
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| CN106720914A (en) * | 2016-12-01 | 2017-05-31 | 中国科学院烟台海岸带研究所 | The preparation method of scallop edge polypeptide dry powder |
| CN106834399A (en) * | 2016-12-30 | 2017-06-13 | 浙江海洋大学 | A kind of Antihypertensive Peptides from Trachyostracous mussel closed shell flesh |
| CN107365818A (en) * | 2017-07-07 | 2017-11-21 | 浙江海润达科技有限公司 | A kind of preparation method of mussel protein small peptide and its preparation method of respective combination chewable tablets |
| CN107383168A (en) * | 2017-09-11 | 2017-11-24 | 浙江海洋大学 | A kind of Trachyostracous mussel Antihypertensive Peptides |
| CN108396048A (en) * | 2018-02-08 | 2018-08-14 | 舟山海研食品科技有限公司 | The preparation method of oyster active peptides |
| CN110438192A (en) * | 2019-09-19 | 2019-11-12 | 大连工业大学 | A kind of preparation method and applications of sugar-free small molecule Gly-His-Lys |
| CN111955621B (en) * | 2020-09-02 | 2022-12-16 | 青岛隆和生物科技有限公司 | Application of composite polypeptide in preparation of feed for promoting egg laying of chickens |
| CN112625088A (en) * | 2021-03-15 | 2021-04-09 | 烟台市华昕生物医药科技有限公司 | Preparation method and application of mussel ACE inhibitory peptide |
| CN114164247B (en) * | 2021-11-23 | 2022-06-14 | 华南理工大学 | Compound ACE inhibitory peptide and preparation method and application thereof |
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| CN101755998B (en) * | 2009-10-30 | 2012-05-30 | 嵊泗海洋之星生物科技发展有限公司 | Preparation method of mussel active polypeptide |
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