CN102363800A - Method for preparing antioxidant oligopeptide by performing ultrasonic enzymolysis on swim bladders - Google Patents
Method for preparing antioxidant oligopeptide by performing ultrasonic enzymolysis on swim bladders Download PDFInfo
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- CN102363800A CN102363800A CN2011103637107A CN201110363710A CN102363800A CN 102363800 A CN102363800 A CN 102363800A CN 2011103637107 A CN2011103637107 A CN 2011103637107A CN 201110363710 A CN201110363710 A CN 201110363710A CN 102363800 A CN102363800 A CN 102363800A
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Abstract
The invention provides a method for preparing antioxidant oligopeptide by performing ultrasonic enzymolysis on swim bladders. The method comprises the following steps of: crushing the swim bladders into swim bladder powder; adding a phosphate buffer solution with the pH value of 6.0 and trypsin into the swim bladder powder and stirring uniformly to obtain a swim bladder solution; performing enzymolysis on the swim bladder solution in water bath; performing ultrasonic treatment on the swim bladder solution for enzymolysis; after ultrasonic treatment, continuously performing enzymolysis in constant-temperature water bath; after the enzymolysis is finished, deactivating enzyme, cooling and filtering to obtain supernate; performing ultrafiltration on the supernate by using a 10,000-Da ultrafiltration membrane; and freeze-drying the solution obtained after ultrafiltration to obtain the swim bladder antioxidant oligopeptide. The antioxidant oligopeptide prepared by the method has a good effect of eliminating free radicals of a human body, can prevent ageing, inflammation and heart cerebrovascular diseases caused by arteriosclerosis, and can serve as a nutritional additive to be applied to food.
Description
Technical field
The present invention relates to the method that a kind of air bladder enzymolysis prepares anti-oxidant oligopeptides, particularly a kind of UW enzymolysis air bladder prepares the method for anti-oxidant oligopeptides.
Background technology
Air bladder has another name called fish bubble, fish maw, the fish sperm, pressure born of the same parents, is the density controller official that the fish body weight is wanted; Live in the teleosts in seawater upper strata and middle level; Great majority all have fish glue, and the volume of air bladder accounts for about 5% of health, and its shape has oval, taper shape, heart-shaped, shape of a hoof or the like.The main nutrient composition of air bladder is various trace elements such as senior collagen protein, mucopolysaccharide, multivitamin and calcium, zinc, iron, selenium, and wherein protein contnt is generally more than 80%, and lipid content is lower, is ideal high protein and low fat food.Contain multiple inorganic elements in the air bladder, especially trace elements zn, Cu, Fe, Se content are very abundant, and these trace elements play an important role to keeping fit.
Nutrient composition content in the air bladder changes because of kind is different, and maximum with the lipid content variation of different air bladders, the variation of protein contnt is minimum.Amino acid whose analytical results shows that 6 seed amino acids such as glycocoll, L-Ala, L-glutamic acid, proline(Pro), l-arginine and aspartic acid have constituted the advantage amino acid of fish maw protein matter.Except glycocoll, L-glutamic acid, proline(Pro) and the content difference maximum of aspartic acid 4 seed amino acids in different air bladders, most of content of amino acids change little.The modern Chinese medicine pharmacy thinks, the air bladder sweet property of distinguishing the flavor of is flat, the function of nourishing YIN and benefiting blood is arranged, stop blooding, invigorating the kidney and stopping nocturnal emission." it is former that book on Chinese herbal medicine is asked " carries: air bladder can " be supported smart arteries and veins, decide hand war, controlling nocturnal emission with astringent drugs." air bladder also has the good health care effect, can promote seminal vesicle secretion fructose, for sperm provides energy; Can promote gastrointestinal to digest and assimilate, appetite stimulator is prevented and treated apocleisis, maldigestion, abdominal distension, constipation; Can increase the toughness and the elastic force of muscle tissue, physical strength reinforcing, Ginseng Extract; The ability skin care makes skin moist; Can strengthen mentality and nervous function, promote to grow, improve thinking and intelligence, keep body of gland and normally secrete; And can prevent that intelligence from subtracting late, nerve conduction is slow, and is slow in reacting; Postpartum lactescence is not enough, senile dementia, forgetful, insomnia etc.; Can improve Abwehrkraft des Koepers and immunizing power, antifatigue suppresses tumor growth.
The present processed and applied of air bladder mainly contains the processing of air bladder dry products, and its method is in fish belly, to take air bladder, cleans up, and with alum solution dipping back stripping, liming soaks degrease, cleans the draining after drying then.The air bladder added value that this working method is produced is low, and tangible region characteristics are arranged, and mainly concentrates on the coastland, and rare interiorly edible, this has restricted the sale of air bladder.
Application number 200810091845.0; Applying date 2008.3.30, publication number CN101545001A, open day 2009.9.30; Artificial section China of shaking of application has applied for a patent, has introduced a kind of preparation method of swimbladder oligopeptide solution: 1. sharp fish maw dry product is carried out cleaning and crushing and handle; 2. after soaking 5-50min with the tap water of 5 times of weights of sharp fish maw, ultrasonication 10-80min; 3. by solid-to-liquid ratio 1: 10-1: 60 mend tap water, smash to pieces and pull an oar and handle 5min with high-speed tissue mashing machine; 4. the slurries with preparation carry out the constant temperature enzymolysis, and enzyme amount ratio is 1: 1-1: 7, and be under the 0.1-0.9% condition at temperature 40-60 ℃, pH value 5.0-8.0, enzyme amount, enzymolysis 3-9h, enzymolysis finish at the 80-100 ℃ of enzyme processing 8-15min that goes out; 5. high speed centrifugation enzymolysis solution is got bottling behind the supernatant liquid filtering, is sealed, and the rotating speed of supercentrifuge is 4000-20, and 000rpm, centrifugation time are 5-15min; 6. with autoclave sterilizer bottle-packaging solution is carried out sterilising treatment, sterilising temp 100-135 ℃, sterilization time 20-50min promptly is prepared into swimbladder oligopeptide solution after the cooling.What this method was prepared is only to be used for replenishing amino acid whose common oligopeptides.And the technology of using the air bladder enzymolysis to prepare anti-oxidation peptide is not at present also appeared in the newspapers, and this patent is mainly produced the oligopeptides with oxidation-resistance through the enzymatic hydrolysis condition of control air bladder.
Summary of the invention
The present invention provides a kind of UW enzymolysis air bladder to prepare the method for anti-oxidant oligopeptides, preparation be anti-oxidant oligopeptides, not only can be used for replenishing amino acid, and can also remove radical.
The technical scheme that the present invention adopted is that a kind of UW enzymolysis air bladder prepares the method for anti-oxidant oligopeptides, comprises following operation steps:
Step 1,
It is the air bladder powder that air bladder is pulverized;
Step 2,
Adding pH value is 6.0 phosphate buffered saline buffer and trypsinase in the air bladder powder, stirs, and obtains air bladder solution; Wherein, the air bladder powder quality is 6% of a phosphate buffered saline buffer quality, and the trypsinase quality is 1.6～2.0% of an air bladder powder quality;
Step 3,
Earlier with air bladder solution enzymolysis 5～10min in 50 ℃ of waters bath with thermostatic control; Again air bladder solution supersound process is carried out enzymolysis, control ultrasonic power 110～220W, ultrasonic time 1～9min; 30～65 ℃ of ultrasonic temperature, the supersound process continued is enzymolysis 30～90min in 50 ℃ water bath with thermostatic control; After enzymolysis finished, the enzyme that goes out, cooling, filtration obtained supernatant;
Step 4,
With the ultrafiltration of 10000Da ultra-filtration membrane, the solution after the ultrafiltration obtains the anti-oxidant oligopeptides of air bladder through lyophilize with supernatant.
Wherein, phosphate buffered saline buffer is that the biphosphate sodium water solution of 0.2mol/L and Sodium phosphate, dibasic aqueous solution that volumetric molar concentration is 0.2mol/L form by volumetric molar concentration, and the volume ratio of the biphosphate sodium water solution and the Sodium phosphate, dibasic aqueous solution is 87.7: 12.3.
The invention has the beneficial effects as follows that using ultrasound ripple enzymolysis fish maw protein matter has improved enzymolysis efficiency; Improve the burst size of peptide in the enzymolysis process, shortened the needed time in the enzymolysis process, simple to operate; With short production cycle, and the anti-oxidant oligopeptides of the present invention preparation has good scavenging(action) to the radical of human body, can play preventing aging; Inflammation, and arteriosclerosis and the cardiovascular and cerebrovascular diseases that causes also can be used as a kind of nutritional additive and are applied in the food.
Embodiment
Embodiment 1
Air bladder is cut into small pieces, puts into kibbler and be ground into 60 purpose air bladder powder; Take by weighing 24g air bladder powder; In the air bladder powder, add 400ml (400g) pH value and be 6.0 phosphate buffered saline buffer (wherein; Phosphate buffered saline buffer is that the biphosphate sodium water solution of 0.2mol/L and Sodium phosphate, dibasic aqueous solution that the 49.2ml volumetric molar concentration is 0.2mol/L form by the 350.8ml volumetric molar concentration) and 0.384g trypsinase; Stir, obtain air bladder solution; With air bladder solution enzymolysis 5min in 50 ℃ of waters bath with thermostatic control, again air bladder solution supersound process is carried out enzymolysis, control ultrasonic power 220W, ultrasonic time 5min, 40 ℃ of ultrasonic temperature, the supersound process continued is enzymolysis 45min in 50 ℃ water bath with thermostatic control; Enzymolysis is elevated to 100 ℃ of enzymes that go out with bath temperature after finishing, and the enzyme postcooling that goes out filters and obtains supernatant to room temperature, and the protein contnt that uses micro-triumphant formula nitriding to record supernatant is 6.02mg/ml; At last with supernatant with the ultrafiltration of 10000Da ultra-filtration membrane, solution after the ultrafiltration is through at-20 ℃, lyophilize obtains the anti-oxidant oligopeptides of air bladder under the 0.43bar condition.
Above-mentioned trypsinase can biological ltd of section be produced by the Wuxi is outstanding.
The ability that records product removing DPPH radical is 74.11%, and the ability of removing hydroxy radical qiao is 76.58%, and the ability of removing ABTS solution is 77.54%.
Embodiment 2
Air bladder is cut into small pieces, puts into kibbler and be ground into 60 purpose air bladder powder; Take by weighing 24g air bladder powder; The pH value that in the air bladder powder, adds 400ml (400g) be 6.0 phosphate buffered saline buffer (wherein; Phosphate buffered saline buffer is that the biphosphate sodium water solution of 0.2mol/L and Sodium phosphate, dibasic aqueous solution that the 49.2ml volumetric molar concentration is 0.2mol/L form by the 350.8ml volumetric molar concentration) and 0.384g trypsinase; Stir, obtain air bladder solution; With air bladder solution enzymolysis 10min in 50 ℃ of waters bath with thermostatic control, again air bladder solution supersound process is carried out enzymolysis, control ultrasonic power 220W, ultrasonic time 9min, 30 ℃ of ultrasonic temperature, the supersound process continued is enzymolysis 45min in 50 ℃ water bath with thermostatic control; Enzymolysis is elevated to 100 ℃ of enzymes that go out with bath temperature after finishing, and the enzyme postcooling that goes out filters and obtains supernatant to room temperature, and the protein contnt that uses micro-triumphant formula nitriding to record supernatant is 6.13mg/ml; At last with supernatant with the ultrafiltration of 10000Da ultra-filtration membrane, solution after the ultrafiltration is through at-20 ℃, lyophilize obtains the anti-oxidant oligopeptides of air bladder under the 0.43bar condition.
Above-mentioned trypsinase can biological ltd of section be produced by the Wuxi is outstanding.
The ability that records product removing DPPH radical is 71.13%, and the ability of removing hydroxy radical qiao is 72.36%, and the ability of removing ABTS solution is 72.61%.
Embodiment 3
Air bladder is cut into small pieces, puts into kibbler and be ground into 60 purpose air bladder powder; Take by weighing 24g air bladder powder; The pH value that in the air bladder powder, adds 400ml (400g) be 6.0 phosphate buffered saline buffer (wherein; Phosphate buffered saline buffer is that the biphosphate sodium water solution of 0.2mol/L and Sodium phosphate, dibasic aqueous solution that the 49.2ml volumetric molar concentration is 0.2mol/L form by the 350.8ml volumetric molar concentration) and 0.48g trypsinase; Stir, obtain air bladder solution; With air bladder solution enzymolysis 10min in 50 ℃ of waters bath with thermostatic control, again air bladder solution supersound process is carried out enzymolysis, control ultrasonic power 220W, ultrasonic time 7min, 45 ℃ of ultrasonic temperature, the supersound process continued is enzymolysis 30min in 50 ℃ water bath with thermostatic control; Enzymolysis is elevated to 100 ℃ of enzymes that go out with bath temperature after finishing, and the enzyme postcooling that goes out filters and obtains supernatant to room temperature, and the protein contnt that uses micro-triumphant formula nitriding to record supernatant is 6.0mg/ml; At last with supernatant with the ultrafiltration of 10000Da ultra-filtration membrane, solution after the ultrafiltration is through at-20 ℃, lyophilize obtains the anti-oxidant oligopeptides of air bladder under the 0.43bar condition.
Above-mentioned trypsinase can biological ltd of section be produced by the Wuxi is outstanding.
The ability that records product removing DPPH radical is 68.36%, and the ability of removing hydroxy radical qiao is 67.92%, and the ability of removing ABTS solution is 69.01%.
Embodiment 4
Air bladder is cut into small pieces, puts into kibbler and be ground into 60 purpose air bladder powder; Take by weighing 24g air bladder powder; The pH value that in the air bladder powder, adds 400ml (400g) be 6.0 phosphate buffered saline buffer (wherein; Phosphate buffered saline buffer is that the biphosphate sodium water solution of 0.2mol/L and Sodium phosphate, dibasic aqueous solution that the 49.2ml volumetric molar concentration is 0.2mol/L form by the 350.8ml volumetric molar concentration) and 0.384g trypsinase; Stir, obtain air bladder solution; With air bladder solution enzymolysis 8min in 50 ℃ of waters bath with thermostatic control, again air bladder solution supersound process is carried out enzymolysis, control ultrasonic power 110W, ultrasonic time 1min, 65 ℃ of ultrasonic temperature, the supersound process continued is enzymolysis 90min in 50 ℃ water bath with thermostatic control; Enzymolysis is elevated to 100 ℃ of enzymes that go out with bath temperature after finishing, and the enzyme postcooling that goes out filters and obtains supernatant to room temperature, and the protein contnt that uses micro-triumphant formula nitriding to record supernatant is 5.96mg/ml; At last with supernatant with the ultrafiltration of 10000Da ultra-filtration membrane, solution after the ultrafiltration is through at-20 ℃, lyophilize obtains the anti-oxidant oligopeptides of air bladder under the 0.43bar condition.
Above-mentioned trypsinase can biological ltd of section be produced by the Wuxi is outstanding.
The ability that records product removing DPPH radical is 64.23%, and the ability of removing hydroxy radical qiao is 64.52%, and the ability of removing ABTS solution is 65.03%.
The present invention with ultrasonic applications in enzymolysis; Make full use of cavitation effect of ultrasonic waves and made the particulate high vibration in the solution, produced fierce collision solution temperature is raise, played good stirring action; Quickened chemical reaction; Improve enzymolysis efficiency, improved the burst size of peptide in the enzymolysis process, shortened the needed time of enzymolysis process.
Claims (2)
1. a UW enzymolysis air bladder prepares the method for anti-oxidant oligopeptides, it is characterized in that, comprises following operation steps:
Step 1,
It is the air bladder powder that air bladder is pulverized;
Step 2,
Adding pH value is 6.0 phosphate buffered saline buffer and trypsinase in the air bladder powder, stirs, and obtains air bladder solution; Wherein, the air bladder powder quality is 6% of a phosphate buffered saline buffer quality, and the trypsinase quality is 1.6～2.0% of an air bladder powder quality;
Step 3,
Earlier with air bladder solution enzymolysis 5～10min in 50 ℃ of waters bath with thermostatic control; Again air bladder solution supersound process is carried out enzymolysis, control ultrasonic power 110～220W, ultrasonic time 1～9min; 30～65 ℃ of ultrasonic temperature, the supersound process continued is enzymolysis 30～90min in 50 ℃ water bath with thermostatic control; After enzymolysis finished, the enzyme that goes out, cooling, filtration obtained supernatant;
Step 4,
With the ultrafiltration of 10000Da ultra-filtration membrane, the solution after the ultrafiltration obtains the anti-oxidant oligopeptides of air bladder through lyophilize with supernatant.
2. according to the method for the anti-oxidant oligopeptides of the said preparation of claim 1; It is characterized in that: said phosphate buffered saline buffer is that the biphosphate sodium water solution of 0.2mol/L and Sodium phosphate, dibasic aqueous solution that volumetric molar concentration is 0.2mol/L form by volumetric molar concentration, and the volume ratio of the biphosphate sodium water solution and the Sodium phosphate, dibasic aqueous solution is 87.7: 12.3.
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| CN103451258A (en) * | 2013-06-19 | 2013-12-18 | 浙江海洋学院 | Preparation technology of croaker fish bladder collagen with high anti-fatigue activity |
| CN104286784A (en) * | 2014-10-28 | 2015-01-21 | 威海市桢昊生物技术有限公司 | Preparation method for fragrance flavor seafood seasoning |
| CN104313096A (en) * | 2014-10-23 | 2015-01-28 | 哈尔滨派特纳生物科技开发有限公司 | Method for preparing beluga swimming bladder protein peptide |
| CN104313098A (en) * | 2014-10-28 | 2015-01-28 | 威海市桢昊生物技术有限公司 | Method for preparing swimming bladder collagen powder |
| CN105254737A (en) * | 2015-11-10 | 2016-01-20 | 浙江海洋学院 | Swim bladder collagen anti-oxidation and anti-fatigue protein peptide |
| CN105410945A (en) * | 2015-11-10 | 2016-03-23 | 浙江海洋学院 | Application of collagen peptide of slim bladders |
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| CN103451258A (en) * | 2013-06-19 | 2013-12-18 | 浙江海洋学院 | Preparation technology of croaker fish bladder collagen with high anti-fatigue activity |
| CN103451258B (en) * | 2013-06-19 | 2015-04-22 | 浙江海洋学院 | Preparation technology of croaker fish bladder collagen with high anti-fatigue activity |
| CN104313096A (en) * | 2014-10-23 | 2015-01-28 | 哈尔滨派特纳生物科技开发有限公司 | Method for preparing beluga swimming bladder protein peptide |
| CN104286784A (en) * | 2014-10-28 | 2015-01-21 | 威海市桢昊生物技术有限公司 | Preparation method for fragrance flavor seafood seasoning |
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| CN105254737A (en) * | 2015-11-10 | 2016-01-20 | 浙江海洋学院 | Swim bladder collagen anti-oxidation and anti-fatigue protein peptide |
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| CN107118256A (en) * | 2017-06-29 | 2017-09-01 | 江苏省农业科学院 | The preparation method of anti-oxidant silver carp liver protein |
| CN109567072A (en) * | 2019-01-02 | 2019-04-05 | 广州南颐贸易有限公司 | A kind of instant colored glue processing method |
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