CN102286601B - Method for preparing peanut antioxidant peptide by fermentation - Google Patents
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- CN102286601B CN102286601B CN201110229979.6A CN201110229979A CN102286601B CN 102286601 B CN102286601 B CN 102286601B CN 201110229979 A CN201110229979 A CN 201110229979A CN 102286601 B CN102286601 B CN 102286601B
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Abstract
The invention discloses a method for preparing peanut antioxidant peptide by fermentation, and the method comprises the following steps: adding distilled water to low-denatured degreased peanut protein powder, ultrasonically dispersing and dissolving, adjusting the pH value, sterilizing and cooling; inoculating with strain seed culture liquid, and fermenting and culturing; and carrying out ultrasonic-assisted enzymolysis, inactivating enzyme, centrifuging, and freeze-drying supernatant to obtain the peanut antioxidant peptide. The peanut antioxidant peptide prepared by the method disclosed by the invention has the antioxidant activities of scavenging hydroxyl free radical, superoxide anion free radical and DPPH (1,1-diphenyl-2-picrylhydrazyl) free radical, inhibiting hydroxyl-free-radical-mediated deoxyribose injury, inhibiting peroxidation of lipidosome, resisting oxidation of linolic acid system, resisting grease peroxidation capability and the like, and has iron reduction capability, molybdenum reduction capability, iron ion chelating capability and copper ion chelating capability; and the method is simple to operate, has mild reaction conditions and is suitable for industrial production.
Description
Technical field
The present invention relates to a kind of method of preparing peanut antioxidant peptide by fermentation, belong to food protein manufacture field.
Background technology
Along with the continuous progress of free radical life science, the research of oxidative stress damage and anti-oxidation protection effect has been subject to people's growing interest.Under serious oxidative stress status, when the concentration of active oxygen exceedes cytophylaxis ability, oxidative damage just can change the structure and function of cell, affects body health.Equally, food is the system of series of complex, and too much free radical exists the stability that also can have a strong impact on system.The development of modern food industry shows, the method that has an anti-oxidation active substance by interpolation can increase the stability of food system.Along with the development of trophology and biotechnology, it is found that the peptide class between between protein and amino acid, due to its special constructional feature, edible safety is higher, anti-oxidant activity is more remarkable, also can belong to antioxidant, as the Material Source of removing free radical in human body.
At present, enzymolysis process is prepared anti-oxidation peptide and is mainly contained two kinds of Technologies: protease hydrolyzed method and fermentation method.Protease hydrolyzed method reaction conditions gentleness, can obtain the anti-oxidation peptide product of certain molecular weight scope, the main method of using is protease hydrolyzed method now, but due to the restriction of the Substratspezifitaet of enzyme and the space conformation of enzyme, make protease hydrolyzed method can only utilize protein isolate as substrate, increased running cost.
Fermentation method utilizes microorganism growth when breeding can produce the principle of proteolytic enzyme, directly microorganism and raw material is mixed and is fermented, and has saved the step of extraction protein isolate.In addition, microorganism is except producing proteolytic enzyme, can also produce cellulase, phytase etc., can decompose cell walls, be conducive to albumen and be combined with the avtive spot of proteolytic enzyme, improve hydrolysis efficiency, increase the anti-oxidant activity of enzymolysis product, reduce production costs, but fermentation method there is report in the preparation of animal serum protein antioxidant peptide, plant, particularly in peanut antioxidant peptide extraction, have not yet to see report.
Summary of the invention
For addressing the above problem, the invention provides a kind of easy and simple to handle, reaction conditions is gentle, can be continuously produced the fermentation preparation of the peanut antioxidant peptide with better anti-oxidant activity.
Peanut protein content is 25～36%, and its biological value is 58, and protein is tired as 1:7, containing 18 seed amino acids, comprises 8 kinds of indispensable amino acids, and research shows, some small peptides of containing in peanut protein peptide chain have anti-oxidant activity.Because subtilis can produce proteolytic enzyme, aspergillus niger can produce proteolytic enzyme, cellulase and phytase, aspergillus oryzae can produce proteolytic enzyme and cellulase, the present invention intends adopting the mixed strains of subtilis, aspergillus niger and aspergillus oryzae as fermented bacterium, by solid fermentation or liquid fermentation process, prepare peanut antioxidant peptide, to improve the anti-oxidant activity of enzymolysis efficiency and anti-oxidation peptide, for the exploitation of peanut antioxidant peptide provides new way.
For achieving the above object, the present invention adopts following steps:
Low sex change defatted peanut protein powder adds distilled water, stirs, and ultrasonic wave dispersing and dissolving, regulates pH value, and sterilizing is cooling; Access bacterial classification seed culture fluid, fermentation culture; Ultrasound-assisted enzymolysis, the enzyme that goes out, centrifugal, supernatant liquor lyophilize obtains peanut antioxidant peptide.
Specifically, described method is as follows:
(1) in low sex change defatted peanut protein powder, add distilled water, glass stick stirs, and ultrasonic wave dispersing and dissolving regulates pH value, and sterilizing is cooling;
(2) access bacterial classification seed culture fluid, solid fermentation or liquid fermentation and culture, obtain fermented product;
(3) fermented product is through ultrasound-assisted enzymolysis, the enzyme that goes out, and centrifugal, supernatant liquor lyophilize obtains peanut antioxidant peptide.
Preferably, the described low sex change defatted peanut protein powder of step (1) adds distilled water 6.5ml/g raw material (solid fermentation), 19.5ml/g raw material (liquid fermenting), ultrasonic wave dispersing and dissolving condition: ultrasonic power is 300w, ultrasonic frequency is 28kHz, and temperature is 25 ℃, and the time is 15min, the sodium hydroxide solution that is 1.0mol/L by concentration regulates pH value to 6.5 ~ 7.5,121 ℃ of sterilizing 15min in high-pressure sterilizing pot.
Preferably, the described bacterial classification seed culture fluid of step (2) is subtilis, the mixed seeds nutrient solution of aspergillus niger and aspergillus oryzae, access amount is 22.5%(solid fermentation), 6.5%(liquid fermenting), subtilis, the cell age of aspergillus niger and aspergillus oryzae is respectively 46h, 70h and 38h, in mixed seeds nutrient solution, their ratio is subtilis: aspergillus niger: aspergillus oryzae=1.8:1.4:1.8, solid fermentation condition is: 31.5 ℃ of temperature, standing cultivation 34h in incubator, liquid fermentation condition is: 34.5 ℃ of temperature, with 65r/min rotating speed, shaking flask shaking culture 36h.
Preferably, the described fermented product of step (3) through the method for ultrasound-assisted enzymolysis is: solid fermentation thing adds ultrasound-assisted enzymolysis after the distilled water of 10 ~ 15ml/g raw material, and the direct ultrasound-assisted enzymolysis of liquid fermentate, the condition of ultrasound-assisted enzymolysis: ultrasonic power is 180w, ultrasonic frequency is 28kHz, temperature is 48.5 ℃, time is 22.5min, the condition of enzyme of going out is the enzyme 18min that goes out in 100 ℃ of waters bath with thermostatic control, and centrifugal condition is 12 ℃, the centrifugal 12min of 6500r/min rotating speed.
Beneficial effect of the present invention is: peanut antioxidant peptide product prepared by the present invention has removing hydroxy radical qiao, ultra-oxygen anion free radical, DPPH free radical, to the restraining effect of hydroxy radical qiao mediation ribodesose damage, iron reducing power, molybdenum reducing power, iron ion sequestering power, cupric ion sequestering power, suppress liposome peroxidation activity, anti-linoleic acid system oxygenizement, the antioxidation activity in vitro such as anti-oil peroxidation ability, can be used as functional food for removing interior free yl, prevention and assisting therapy modern civilization diseases, also can be used as natural antioxidants be used in food storage and fresh-keeping in, extend Food Shelf-life, the method reaction conditions gentleness of preparing peanut antioxidant peptide by fermentation, environmental friendliness, serialization is produced in enormous quantities, for the exploitation of peanut antioxidant peptide provides new approach.
Embodiment
embodiment 1
Low sex change defatted peanut protein powder adds distilled water 6.5ml/g raw material, glass stick stirs, take ultrasonic power as 300w, ultrasonic frequency is 28kHz, and temperature is the condition of 25 ℃, ultrasonic dispersing and dissolving 15min, obtain peanut protein powder mashed prod, the sodium hydroxide solution that is 1.0mol/L by concentration regulates pH value to 6.86, in high-pressure sterilizing pot, 121 ℃ of sterilizing 15min, are cooled to room temperature; Get subtilis, aspergillus niger and aspergillus oryzae that cell age is respectively 46h, 70h and 38h, in peanut protein powder mashed prod, access with subtilis: aspergillus niger: the seed culture fluid that aspergillus oryzae=1.8:1.4:1.8 ratio is mixed, access amount is 22.5%, in incubator, with 31.5 ℃ of temperature, standing solid fermentation is cultivated 34h, obtains solid fermentation thing; In solid fermentation thing, adding the distilled water of 12.5ml/g raw material, is 180w at ultrasonic power, and ultrasonic frequency is 28kHz, and temperature is under the condition of 48.5 ℃, ultrasound-assisted enzymolysis 22.5min; The enzyme 18min that goes out in 100 ℃ of waters bath with thermostatic control, cool to room temperature, under 12 ℃ of conditions, the centrifugal 12min of 6500r/min rotating speed, gets supernatant liquor lyophilize and obtains peanut antioxidant peptide.The anti-oxidant activity of this anti-oxidation peptide is as follows: the IC that removes hydroxy radical qiao
50value is 0.35mg/mL, removes the IC of ultra-oxygen anion free radical
50value is 1.47mg/mL, removes the IC of DPPH free radical
50value is 1.39mg/mL, to the inhibiting IC of hydroxy radical qiao mediation ribodesose damage
50value is 0.53mg/mL, and the light absorption value of iron reducing power is that the concentration of 0.5 o'clock required peanut antioxidant peptide is 4.14mg/mL, and the light absorption value of molybdenum reducing power is that the concentration of 0.5 o'clock required peanut antioxidant peptide is 1.26mg/mL, the IC of iron ion chelating ability
50value is 1.71mg/mL, the IC of cupric ion chelating ability
50value is 1.48mg/mL, suppresses the IC of liposome peroxidation activity
50value is 1.82mg/mL, the IC of anti-linoleic acid system oxygenizement
50value is 2.06mg/mL, the IC of anti-oil peroxidation ability
50value is 2.13mg/mL.
embodiment 2
Low sex change defatted peanut protein powder adds distilled water 6.5ml/g raw material, glass stick stirs, take ultrasonic power as 300w, ultrasonic frequency is 28kHz, and temperature is the condition of 25 ℃, ultrasonic dispersing and dissolving 15min, obtain peanut protein powder mashed prod, the sodium hydroxide solution that is 1.0mol/L by concentration regulates pH value to 6.62, in high-pressure sterilizing pot, 121 ℃ of sterilizing 15min, are cooled to room temperature; Get subtilis, aspergillus niger and aspergillus oryzae that cell age is respectively 46h, 70h and 38h, in peanut protein powder mashed prod, access with subtilis: aspergillus niger: the seed culture fluid that aspergillus oryzae=1.8:1.4:1.8 ratio is mixed, access amount is 22.5%, in incubator, with 31.5 ℃ of temperature, standing solid fermentation is cultivated 34h, obtains solid fermentation thing; In solid fermentation thing, adding the distilled water of 14.5ml/g raw material, is 180w at ultrasonic power, and ultrasonic frequency is 28kHz, and temperature is under the condition of 48.5 ℃, ultrasound-assisted enzymolysis 22.5min; The enzyme 18min that goes out in 100 ℃ of waters bath with thermostatic control, cool to room temperature, under 12 ℃ of conditions, the centrifugal 12min of 6500r/min rotating speed, gets supernatant liquor lyophilize and obtains peanut antioxidant peptide.The anti-oxidant activity of this anti-oxidation peptide is as follows: the IC that removes hydroxy radical qiao
50value is 0.57mg/mL, removes the IC of ultra-oxygen anion free radical
50value is 0.98mg/mL, removes the IC of DPPH free radical
50value is 1.02mg/mL, to the inhibiting IC of hydroxy radical qiao mediation ribodesose damage
50value is 0.42mg/mL, and the light absorption value of iron reducing power is that the concentration of 0.5 o'clock required peanut antioxidant peptide is 2.46mg/mL, and the light absorption value of molybdenum reducing power is that the concentration of 0.5 o'clock required peanut antioxidant peptide is 0.99mg/mL, the IC of iron ion chelating ability
50value is 1.24mg/mL, the IC of cupric ion chelating ability
50value is 1.11mg/mL, suppresses the IC of liposome peroxidation activity
50value is 1.43mg/mL, the IC of anti-linoleic acid system oxygenizement
50value is 1.08mg/mL, the IC of anti-oil peroxidation ability
50value is 1.29mg/mL.
embodiment 3
Low sex change defatted peanut protein powder adds distilled water 19.5ml/g raw material, glass stick stirs, take ultrasonic power as 300w, ultrasonic frequency is 28kHz, and temperature is the condition of 25 ℃, ultrasonic dispersing and dissolving 15min, obtain peanut protein powder suspension, the sodium hydroxide solution that is 1.0mol/L by concentration regulates pH value to 7.28, in high-pressure sterilizing pot, 121 ℃ of sterilizing 15min, are cooled to room temperature; Get subtilis, aspergillus niger and aspergillus oryzae that cell age is respectively 46h, 70h and 38h, in peanut protein powder mashed prod, access with subtilis: aspergillus niger: the seed culture fluid that aspergillus oryzae=1.8:1.4:1.8 ratio is mixed, access amount is 6.5%, in shaking table, with 34.5 ℃ of temperature, shaking flask liquid fermentation and culture 36h, obtains liquid fermentate; Getting liquid fermentate, is 180w at ultrasonic power, and ultrasonic frequency is 28kHz, and temperature is under the condition of 48.5 ℃, ultrasound-assisted enzymolysis 22.5min; The enzyme 18min that goes out in 100 ℃ of waters bath with thermostatic control, cool to room temperature, under 12 ℃ of conditions, the centrifugal 12min of 6500r/min rotating speed, gets supernatant liquor lyophilize and obtains peanut antioxidant peptide.The anti-oxidant activity of this anti-oxidation peptide is as follows: the IC that removes hydroxy radical qiao
50value is 0.17mg/mL, removes the IC of ultra-oxygen anion free radical
50value is 0.49mg/mL, removes the IC of DPPH free radical
50value is 0.61mg/mL, to the inhibiting IC of hydroxy radical qiao mediation ribodesose damage
50value is 0.33mg/mL, and the light absorption value of iron reducing power is that the concentration of 0.5 o'clock required peanut antioxidant peptide is 1.09mg/mL, and the light absorption value of molybdenum reducing power is that the concentration of 0.5 o'clock required peanut antioxidant peptide is 0.65mg/mL, the IC of iron ion chelating ability
50value is 0.58mg/mL, the IC of cupric ion chelating ability
50value is 0.43mg/mL, suppresses the IC of liposome peroxidation activity
50value is 0.41mg/mL, the IC of anti-linoleic acid system oxygenizement
50value is 0.75mg/mL, the IC of anti-oil peroxidation ability
50value is 0.28mg/mL.
Claims (1)
1. a method for preparing peanut antioxidant peptide by fermentation, comprises the following steps:
(1) in low sex change defatted peanut protein powder, add distilled water, glass stick stirs, and ultrasonic wave dispersing and dissolving regulates pH value, and sterilizing is cooling;
(2) access bacterial classification seed culture fluid, solid fermentation or liquid fermentation and culture, obtain fermented product;
(3) fermented product is through ultrasound-assisted enzymolysis, the enzyme that goes out, and centrifugal, supernatant liquor lyophilize obtains peanut antioxidant peptide;
The described low sex change defatted peanut protein powder of step (1) adds distilled water, solid fermentation during 6.5ml/g raw material, liquid fermenting during 19.5ml/g raw material, ultrasonic wave dispersing and dissolving condition: ultrasonic power is 300w, ultrasonic frequency is 28kHz, and temperature is 25 ℃, and the time is 15min, the sodium hydroxide solution that is 1.0mol/L by concentration regulates pH value to 6.5 ~ 7.5,121 ℃ of sterilizing 15min in high-pressure sterilizing pot;
The described bacterial classification seed culture fluid of step (2) is subtilis, the mixed seeds nutrient solution of aspergillus niger and aspergillus oryzae, solid fermentation when access amount is 22.5%, liquid fermenting in the time of 6.5%, subtilis, the cell age of aspergillus niger and aspergillus oryzae is respectively 46h, 70h and 38h, in mixed seeds nutrient solution, their ratio is subtilis: aspergillus niger: aspergillus oryzae=1.8:1.4:1.8, solid fermentation condition is: 31.5 ℃ of temperature, standing cultivation 34h in incubator, liquid fermentation condition is: 34.5 ℃ of temperature, with 65r/min rotating speed, shaking flask shaking culture 36h,
The described fermented product of step (3) through the method for ultrasound-assisted enzymolysis is: solid fermentation thing adds ultrasound-assisted enzymolysis after the distilled water of 10 ~ 15ml/g raw material, and the direct ultrasound-assisted enzymolysis of liquid fermentate, the condition of ultrasound-assisted enzymolysis: ultrasonic power is 180w, ultrasonic frequency is 28kHz, temperature is 48.5 ℃, and the time is 22.5min, and the condition of the enzyme that goes out is the enzyme 18min that goes out in 100 ℃ of waters bath with thermostatic control, centrifugal condition is 12 ℃, the centrifugal 12min of 6500r/min rotating speed.
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| CN102649974B (en) * | 2012-05-14 | 2014-03-19 | 上海海洋大学 | Method for preparing antioxidant peptide through ultrasonic-microwave synergetic enzymatic hydrolysis |
| CN103053787A (en) * | 2013-02-05 | 2013-04-24 | 杨庆利 | Method for preparing peanut peptide by microbial solid fermentation of peanut meal |
| CN104388502B (en) * | 2014-10-19 | 2019-09-10 | 沈阳农业大学 | A kind of method that mixed bacteria solid state fermentation prepares rice bran active peptide |
| CN105132509B (en) * | 2015-09-25 | 2021-05-14 | 华南理工大学 | Method for extracting strong antioxidant protein polypeptide from peanut meal |
| CN105361168B (en) * | 2015-12-02 | 2017-12-19 | 绥化学院 | A kind of composite type antioxidant peptide and preparation method |
| CN106967768A (en) * | 2017-04-24 | 2017-07-21 | 江苏大学 | It is a kind of to improve the method that liquid state fermentation prepares oligopeptide yield |
| CN106967776A (en) * | 2017-04-24 | 2017-07-21 | 江苏大学 | It is a kind of to improve the method that solid state fermentation prepares oligopeptide yield |
| CN107184464B (en) * | 2017-05-25 | 2018-01-23 | 上海悦目化妆品有限公司 | A kind of Compound Black plant fermentation composition with white-skinned face function and preparation method thereof and the application in facial mask class product |
| CN108185267A (en) * | 2017-12-28 | 2018-06-22 | 广州市科能化妆品科研有限公司 | A kind of anti-oxidation peptide composition and its preparation method and application |
| CN116656763B (en) * | 2023-07-25 | 2023-10-13 | 山东山歌食品科技股份有限公司 | Preparation method of peanut antioxidant peptide |
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