CN102191306A - Enzymatic preparation method for antihypertensive peptides from jellyfish - Google Patents
Enzymatic preparation method for antihypertensive peptides from jellyfish Download PDFInfo
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Abstract
The invention relates to an enzymatic preparation method for antihypertensive peptides from jellyfish, which comprises: selecting fresh jellyfish or jellyfish processed with vitriol as raw materials, desalting, washing and grinding; regulating the protein concentration of jellyfish pulp to 1 to 3 percent; regulating pH value to 2 to 3.5, controlling the temperature to be 35 to 45 DEG C, adding pepsase, and keeping temperature to perform enzymolysis for 4 to 6 hours, wherein the added amount of the pepsase per gram of protein is 2,000 to 4,000um; regulating the pH value of the pepsase hydrolysate of jellyfish to 5 to 6, keeping the temperature between 50 and 55 DEG C, and adding papain to perform enzymolysis for 1 to 3 hours, wherein the added amount of papain per gram of protein is 2,000 to 4,000mu; and deactivating enzymes; and filtering with a membrane for removing impurities, desalinizing by a nanofiltration membrane, concentrating, spray-drying or freeze-drying to obtain powder of antihypertensive peptides from jellyfish. The antihypertensive peptides from jellyfish, which are prepared by the method, are proved in spontaneously hypertensive (SHR) rat oral taking experiments to have remarkable blood pressure reduction assisting effect. In the invention, the production process is advanced, the operability is high, the enzymolysis of the protein is complete, and the product quality is controllable.
Description
Technical field
The present invention relates to a kind of general preparation method of peptide, especially a kind of by enzymatic hydrolysis acquisition jellyfish collagen peptide liquid, again through the enzymatic-process preparation method that separates, concentrates, drying obtains the jellyfish step-down peptide of jellyfish step-down peptide powder.
Background technology
Jellyfish claims jellyfish again, is the enterozoa of Hai Sheng, and not only kind is many for they, and quantity is big, and it is wide to distribute, and is the important component part of marine zooplankton group.Jellyfish vegetal pole horn of plenty, its body contain inorganic salt such as higher protein and calcium, phosphorus, iodine, iron, have the laudatory title of " aquatic products rarity ".Jellyfish is not only edible, but also is the good medicine of curing the disease simply.Described in the Compendium of Material Medica: jellyfish has functions such as clearing heat and detoxicating, resolving phlegm and softening hard masses, step-down detumescence, and is all effective in cure to diseases such as trachitis, asthma, hypertension, stomach ulcer.Therefore, jellyfish eats the homologous macro-organism as a kind of doctor, and it has important economy and edibleness.
The method for preparing bioactive peptide mainly contains: directly extraction, external degradation and utilize recombinant DNA technology, enzyme process or chemical method composite reactive peptide.Extracorporeal hydrolysis protein prepares bioactive peptide three operational paths: acid hydrolysis, basic hydrolysis and enzymatic hydrolysis.The former two's method is simple, but has the deleterious effect of nutrition and toxicity aspect, therefore is greatly limited in Application in Food Processing.Proteinic enzymatic hydrolysis generally can not cause the loss of nutrition aspect, also can not produce the problem on the toxicity; Enzyme effect simultaneously has specificity, can carry out under the condition of gentleness, and energy consumption is also low, so enzymatic hydrolysis is often used in food-processing.
And existing report of technology for preparing the jellyfish bioactive peptide with enzymatic hydrolysis jellyfish protein, for example: by the applicant application and on May 6th, 2009 disclosed CN101422195A Chinese invention patent Shen Qing Publication specification sheets disclosed " a kind of jellyfish active peptide yoghourt and preparation method thereof ", in on May 6th, the 2009 disclosed CN101422266A Chinese invention patent Shen Qing Publication specification sheets disclosed " a kind of jelly-fish nutrient liquid and preparation method thereof ", in on March 17th, the 2010 disclosed CN101669659A Chinese invention patent Shen Qing Publication specification sheets disclosed " a kind of jellyfish water chestnut drink and preparation method thereof ", in on October 20th, the 2010 disclosed CN101862375A Chinese invention patent Shen Qing Publication specification sheets disclosed " a kind of compound jellyfish soluble granules for reducing blood pressure and preparation method thereof ", in the above-mentioned open file to the jellyfish raw material through desalination, clean, broken, behind the thin up or adopt Sumizyme MP or adopt trypsinase to carry out enzymolysis and obtain jellyfish polypeptide liquid, then be used to prepare other functional products.In the disclosed technology, what jellyfish was carried out that enzymolysis all adopts is single enzyme.And in proteolysis, the selection of enzyme is a gordian technique, and the difference of used enzyme and separation purification method can obtain different products, and the activity of the peptide product of different enzymolysis process gained is different.At present, do not see the report that adopts dual-enzyme hydrolysis method to prepare jellyfish step-down peptide as yet.
Summary of the invention
In order to overcome deficiency of the prior art, the invention provides a kind of production technique advanced person, controllable product quality, protein digestion fully, the enzymatic-process preparation method of the active high jellyfish step-down peptide of the jellyfish peptide of gained.
The technical solution adopted for the present invention to solve the technical problems is: a kind of enzymatic-process preparation method of jellyfish step-down peptide is characterized in that: through the following step
(a) processing of raw material is chosen fresh jellyfish or alum system jellyfish is a raw material, and clear water soaks desalination to salinity＜5%, cleans the back and adopts colloidal mill to carry out milled processed, and gained jellyfish slurry particle fineness is controlled at 2～5um;
(b) protein content of the first step enzymolysis determination step (a) gained jellyfish slurry, adding water, to adjust proteinic mass concentration be 1%～3%; Adjusting the pH value is 2～3.5, and controlled temperature is 35～40 ℃, adds stomach en-, and pepsic consumption is the stomach en-that every gram protein adds 2000～4000u, and insulation enzymolysis 4～6h gets jellyfish pepsin hydrolysis liquid;
(c) the second step enzymolysis is 5～6 with the jellyfish pepsin hydrolysis liquid adjustment pH value that step (b) obtains, controlled temperature is 50～55 ℃, adds papoid, and the papoid consumption is the papoid that every gram protein adds 2000～4000u, enzymolysis 1～3h gets the two enzyme enzymolysis solutions of jellyfish;
(d) enzyme-deactivating is heated to 90～100 ℃ with the two enzyme enzymolysis solutions of jellyfish that step (c) obtains, and keeps 10～20 minutes, carries out enzyme-deactivating, the two enzyme enzymolysis solutions of the enzyme jellyfish of must going out;
(e) impurity elimination, desalination are removed impurity with the two enzyme enzymolysis solutions of the enzyme jellyfish of going out that step (d) obtains through tubular membrane, adopt the nanofiltration membrane desalination of 150～300 trapped molecular weights then, and concentrate, get jellyfish step-down peptide concentrated solution, the soluble solid content of this jellyfish step-down peptide concentrated solution is 15%～20%;
(f) the dry jellyfish step-down peptide concentrated solution that step (e) is obtained carries out spraying drying or lyophilize, obtains jellyfish step-down peptide powder.
The two enzyme enzymolysis solutions of described jellyfish after enzyme liquid is gone out are removed the tubular membrane of impurity, and its work aperture is 0.1～0.2 μ m, and working temperature is controlled at 30～40 ℃, advances film pressure and is controlled at 2.5～3bar, and the membrane pressure-controlling is at 0.8～1.0bar.
The described spraying drying that jellyfish step-down peptide concentrated solution is carried out, its inlet temperature is controlled at 185 ℃～190 ℃, and air outlet temperature is controlled at 75～80 ℃, and spray pressure is 0.4MPa.
The described lyophilize that jellyfish step-down peptide concentrated solution is carried out, its vacuum degree control are at 50～133Pa, and temperature of heating plate: 20 ℃ 250min/40 ℃ 400min/30 ℃ up to the freeze-dried material constant weight;
Condenser temperature is controlled at-40 ℃～-50 ℃.
The present invention is to be raw material with bright jellyfish or alum system jellyfish, after desalination, the grinding, obtain jellyfish proteolysis peptide liquid with stomach en-and papoid substep enzymolysis respectively, separate obtaining jellyfish step-down peptide liquid then by membrane filtration removal of impurities, nanofiltration membrane desalination, again through concentrate, spraying drying or lyophilize obtain jellyfish step-down peptide powder.The jellyfish step-down peptide of the present invention's preparation has auxiliary blood pressure reduction effect preferably through oral experimental results show that of SHR rat (original hypertensive rat).The production technique advanced person of this jellyfish step-down peptide, workable, controllable product quality, protein digestion is complete, and the jellyfish step-down peptide powder of gained is white in color or is faint yellow, and is active high, soluble in water, can be used for fields such as food and health care.
Embodiment
Below in conjunction with embodiment the present invention is described in further details.
Embodiment 1
(a) to take by weighing the fresh jellyfish of 500kg be raw material in the processing of raw material, and clear water soaks desalination to salt and is divided into 3%, cleans up the back and adopt colloidal mill to carry out milled processed, and gained jellyfish slurry particle fineness is controlled at 3.5um;
(b) the first step enzymolysis determination step (a) gained jellyfish slurry moisture and protein content, according to measurement result, adding water, to adjust proteinic mass concentration be 2%; Adding the sour pH of adjustment value is 3.26, and controlled temperature is 37 ℃, adds stomach en-, and pepsic consumption is the stomach en-that every gram protein adds 3000u, and insulation enzymolysis 4h gets jellyfish pepsin hydrolysis liquid;
(c) the pH value of the second step enzymolysis jellyfish pepsin hydrolysis liquid that step (b) is obtained is adjusted to 5.02 with alkaline solution, temperature rises to 50 ℃, adds papoid, and the papoid consumption is the papoid that every gram protein adds 4000u, enzymolysis 1h gets the two enzyme enzymolysis solutions of jellyfish;
(d) enzyme-deactivating is heated to 95 ℃ with the two enzyme enzymolysis solutions of jellyfish that step (c) obtains, and keeps 15 minutes, carries out enzyme-deactivating, the two enzyme enzymolysis solutions of the jellyfish of the enzyme that must go out;
(e) impurity elimination, desalination are removed impurity with the two enzyme enzymolysis solutions of the enzyme jellyfish of going out that step (d) obtains through tubular membrane, and wherein tubular membrane work aperture is 0.15 μ m, and working temperature is controlled at 35 ℃, advances film pressure and is controlled at 2.5bar, and the membrane pressure-controlling is at 0.9bar.Adopt the nanofiltration membrane desalination of 250 trapped molecular weights then, concentrate in the time of desalination, resulting concentrated solution can further adopt vacuum concentration, gets jellyfish step-down peptide concentrated solution, and the soluble solid content of this jellyfish step-down peptide concentrated solution is 20%;
(f) the dry jellyfish step-down peptide concentrated solution that step (e) is obtained adopts cryodesiccated mode to carry out drying, cryodesiccated working conditions is: vacuum degree control is at 100Pa, and temperature of heating plate: 20 ℃ 250min/40 ℃ 400min/30 ℃ up to the freeze-dried material constant weight; Condenser temperature is controlled at-45 ℃.Obtain jellyfish step-down peptide powder 3.5kg, measure its IC according to external ACE inhibiting rate
50Be 0.4mg/mL.
Embodiment 2
(a) to take by weighing the alum system jellyfish of getting 500kg be raw material in the processing of raw material, and clear water immersion, rehydration, desalination are to salinity＜5%, and flowing water cleans up the back and adopts colloidal mill to carry out milled processed, and gained jellyfish slurry particle fineness is controlled at 2um;
(b) the first step enzymolysis determination step (a) gained jellyfish slurry moisture and protein content, according to measurement result, adding water, to adjust proteinic mass concentration be 3%; Adding the sour pH of adjustment value is 2, and controlled temperature is 35 ℃, adds stomach en-, and pepsic consumption is the stomach en-that every gram protein adds 4000u, and insulation enzymolysis 5h gets jellyfish pepsin hydrolysis liquid;
(c) pH of the second step enzymolysis jellyfish pepsin hydrolysis liquid that step (b) is obtained is adjusted to 6 with alkaline solution, controlled temperature is 55 ℃, adds papoid, and the papoid consumption is the papoid that every gram protein adds 3500u, enzymolysis 2h gets the two enzyme enzymolysis solutions of jellyfish;
(d) enzyme-deactivating is heated to 100 ℃ with the two enzyme enzymolysis solutions of jellyfish that step (c) obtains, and keeps 10 minutes, carries out enzyme-deactivating, the two enzyme enzymolysis solutions of the jellyfish of the enzyme that must go out;
(e) impurity elimination, desalination are removed impurity with the two enzyme enzymolysis solutions of the enzyme jellyfish of going out that step (d) obtains through tubular membrane, and wherein the work aperture of tubular membrane is 0.2 μ m, and working temperature is controlled at 30 ℃, advances film pressure and is controlled at 3bar, and the membrane pressure-controlling is at 1.0bar.Adopt the nanofiltration membrane desalination of 300 trapped molecular weights then, concentrate in the time of desalination, the concentrated solution that obtains further adopts the vacuum concentration mode to concentrate, and gets jellyfish step-down peptide concentrated solution, and the soluble solid content of this jellyfish step-down peptide concentrated solution is 15%;
(f) the dry jellyfish step-down peptide concentrated solution that step (e) is obtained carries out spraying drying, and its inlet temperature is controlled at 190 ℃ during spraying drying, and air outlet temperature is controlled at 75 ℃, spray pressure is 0.4MPa, obtain jellyfish step-down peptide powder 4.0kg, measure its IC according to external ACE inhibiting rate
50Be 0.5mg/mL.
Embodiment 3
(a) to take by weighing 300kg alum system jellyfish be raw material in the processing of raw material, and clear water immersion, rehydration, desalination are to salinity＜4%, and flowing water is cleaned the back and adopted colloidal mill to carry out milled processed, and gained jellyfish slurry particle fineness is controlled at 5um;
(b) moisture and the protein content of the first step enzymolysis determination step (a) gained jellyfish slurry, adding water, to adjust proteinic mass concentration be 1%; Adding the sour pH of adjustment value is 3, and controlled temperature is 40 ℃, adds stomach en-, and pepsic consumption is the stomach en-that every gram protein adds 2000u, and insulation enzymolysis 6h gets jellyfish pepsin hydrolysis liquid;
(c) the second step enzymolysis is adjusted pH value to 5.5 with the jellyfish pepsin hydrolysis liquid that step (b) obtains with alkaline solution, controlled temperature is 52 ℃, adds papoid, and the papoid consumption is the papoid that every gram protein adds 2000u, enzymolysis 3h gets the two enzyme enzymolysis solutions of jellyfish;
(d) enzyme-deactivating is heated to 90 ℃ with the two enzyme enzymolysis solutions of jellyfish that step (c) obtains, and keeps 20 minutes, carries out enzyme-deactivating, the two enzyme enzymolysis solutions of the enzyme jellyfish of must going out;
(e) impurity elimination, desalination are removed impurity with the two enzyme enzymolysis solutions of the enzyme jellyfish of going out that step (d) obtains through tubular membrane, and the work aperture of its tubular membrane is 0.1 μ m, and working temperature is controlled at 40 ℃, advances film pressure and is controlled at 2.8bar, and the membrane pressure-controlling is at 0.8bar.Adopt the nanofiltration membrane desalination of 150 trapped molecular weights then, concentrate in the time of desalination, the concentrated solution that obtains adopts the vacuum concentration mode further to concentrate, and gets jellyfish step-down peptide concentrated solution, and the soluble solid content of this jellyfish step-down peptide concentrated solution is 18%;
(f) the dry jellyfish step-down peptide concentrated solution that step (e) is obtained carries out lyophilize, and cryodesiccated working conditions is: vacuum degree control is at 133Pa, and temperature of heating plate: 20 ℃ 250min/40 ℃ 400min/30 ℃ up to the freeze-dried material constant weight; Condenser temperature is controlled at-50 ℃, obtains jellyfish step-down peptide powder 2.5kg, measures its IC according to external ACE inhibiting rate
50Be 0.4mg/mL.
Adopt the SHR rat model that prepared jellyfish step-down peptide powder is estimated experimental result such as table 1.
Each administration group of table 1 to the influence of SHR caudal artery systolic pressure (
MmHg)
Annotate: compare * P＜0.05, * * P＜0.01 with the blank group
Compare with the blank group, all obviously reductions (P＜0.05) of 4 week of jellyfish step-down peptide group rat oral gavage back blood pressure level, and also the administration group does not have obvious influence to rat body weight.Jellyfish step-down peptide can obviously reduce the blood pressure of original hypertensive rat.
Claims (4)
1. the enzymatic-process preparation method of a jellyfish step-down peptide is characterized in that: through the following step
(a) processing of raw material is chosen fresh jellyfish or alum system jellyfish is a raw material, and clear water soaks desalination to salinity＜5%, cleans the back and adopts colloidal mill to carry out milled processed, and gained jellyfish slurry particle fineness is controlled at 2～5um;
(b) protein content of the first step enzymolysis determination step (a) gained jellyfish slurry, adding water, to adjust proteinic mass concentration be 1%～3%; Adjusting the pH value is 2～3.5, and controlled temperature is 35～40 ℃, adds stomach en-, and pepsic consumption is the stomach en-that every gram protein adds 2000～4000u, and insulation enzymolysis 4～6h gets jellyfish pepsin hydrolysis liquid;
(c) the second step enzymolysis is 5～6 with the jellyfish pepsin hydrolysis liquid adjustment pH value that step (b) obtains, controlled temperature is 50～55 ℃, adds papoid, and the papoid consumption is the papoid that every gram protein adds 2000～4000u, enzymolysis 1～3h gets the two enzyme enzymolysis solutions of jellyfish;
(d) enzyme-deactivating is heated to 90～100 ℃ with the two enzyme enzymolysis solutions of jellyfish that step (c) obtains, and keeps 10～20 minutes, carries out enzyme-deactivating, the two enzyme enzymolysis solutions of the enzyme jellyfish of must going out;
(e) impurity elimination, desalination are removed impurity with the two enzyme enzymolysis solutions of the enzyme jellyfish of going out that step (d) obtains through tubular membrane, adopt the nanofiltration membrane desalination of 150～300 trapped molecular weights then, and concentrate, get jellyfish step-down peptide concentrated solution, the soluble solid content of this jellyfish step-down peptide concentrated solution is 15%～20%;
(f) the dry jellyfish step-down peptide concentrated solution that step (e) is obtained carries out spraying drying or lyophilize, obtains jellyfish step-down peptide powder.
2. the enzymatic-process preparation method of a kind of jellyfish step-down peptide according to claim 1, it is characterized in that: the two enzyme enzymolysis solutions of described jellyfish after enzyme liquid is gone out are removed the tubular membrane of impurity, its work aperture is 0.1～0.2 μ m, working temperature is controlled at 30～40 ℃, advance film pressure and be controlled at 2.5～3bar, the membrane pressure-controlling is at 0.8～1.0bar.
3. the enzymatic-process preparation method of a kind of jellyfish step-down peptide according to claim 1 and 2, it is characterized in that: the described spraying drying that jellyfish step-down peptide concentrated solution is carried out, its inlet temperature is controlled at 185 ℃～190 ℃, and air outlet temperature is controlled at 75～80 ℃, and spray pressure is 0.4MPa.
4. the enzymatic-process preparation method of a kind of jellyfish step-down peptide according to claim 1 and 2, it is characterized in that: the described lyophilize that jellyfish step-down peptide concentrated solution is carried out, its vacuum degree control is at 50～133Pa, and temperature of heating plate: 20 ℃ 250min/40 ℃ 400min/30 ℃ up to the freeze-dried material constant weight; Condenser temperature is controlled at-40 ℃～-50 ℃.
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| CN104498571A (en) * | 2014-12-05 | 2015-04-08 | 中南林业科技大学 | Intermittent preparation method of camellia meal protein antihypertensive peptide |
| CN104524541A (en) * | 2014-11-27 | 2015-04-22 | 山东好当家海洋发展股份有限公司 | Jellyfish protein effervescent tablet and preparation method thereof |
| CN104774255A (en) * | 2015-04-14 | 2015-07-15 | 山东省科学院生物研究所 | Jellyfis-derived bioactive peptide and preparation method thereof |
| CN105037486A (en) * | 2015-08-07 | 2015-11-11 | 国家海洋局第三海洋研究所 | Method for preparing jellyfish active glycoprotein through multi-stage film separation technology |
| CN105494839A (en) * | 2015-12-26 | 2016-04-20 | 山东好当家海洋发展股份有限公司 | Rhopilema esculentum polypeptide composite tablet candy and preparation method thereof |
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| CN109548950A (en) * | 2018-12-20 | 2019-04-02 | 水母娘娘海洋生物科技有限公司 | The polypeptide and its preparation process in a kind of aurelia source and application |
| CN113812583A (en) * | 2021-08-09 | 2021-12-21 | 庄臣酿酒（福建）有限公司 | Preparation method of jellyfish extract |
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| CN102409072B (en) * | 2011-11-29 | 2014-07-23 | 山东好当家海洋发展股份有限公司 | Method for preparing composite blood pressure lowering peptide by using marine organism |
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| CN103340448A (en) * | 2013-06-09 | 2013-10-09 | 中国海洋大学 | Jellyfish crystal jelly and processing method thereof |
| CN104524541A (en) * | 2014-11-27 | 2015-04-22 | 山东好当家海洋发展股份有限公司 | Jellyfish protein effervescent tablet and preparation method thereof |
| CN104498571B (en) * | 2014-12-05 | 2017-10-20 | 中南林业科技大学 | A kind of Extracted From Oil-tea-cake protein blood pressure-reducing peptide batch (-type) preparation method |
| CN104498571A (en) * | 2014-12-05 | 2015-04-08 | 中南林业科技大学 | Intermittent preparation method of camellia meal protein antihypertensive peptide |
| CN104774255A (en) * | 2015-04-14 | 2015-07-15 | 山东省科学院生物研究所 | Jellyfis-derived bioactive peptide and preparation method thereof |
| CN104774255B (en) * | 2015-04-14 | 2018-01-02 | 山东省科学院生物研究所 | The biologically active peptide in jellyfish source and preparation method thereof |
| CN105037486A (en) * | 2015-08-07 | 2015-11-11 | 国家海洋局第三海洋研究所 | Method for preparing jellyfish active glycoprotein through multi-stage film separation technology |
| CN105494839A (en) * | 2015-12-26 | 2016-04-20 | 山东好当家海洋发展股份有限公司 | Rhopilema esculentum polypeptide composite tablet candy and preparation method thereof |
| CN106591409A (en) * | 2016-12-30 | 2017-04-26 | 浙江海洋大学 | Squid ink oligopeptide with lipid metabolism regulation function |
| CN109548950A (en) * | 2018-12-20 | 2019-04-02 | 水母娘娘海洋生物科技有限公司 | The polypeptide and its preparation process in a kind of aurelia source and application |
| CN113812583A (en) * | 2021-08-09 | 2021-12-21 | 庄臣酿酒（福建）有限公司 | Preparation method of jellyfish extract |
| CN113812583B (en) * | 2021-08-09 | 2023-12-22 | 庄臣酿酒（福建）有限公司 | Preparation method of jellyfish extract |
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