CN100558906C - Biocatalysis prepares the method for corn protein peptide in the organic medium - Google Patents
Biocatalysis prepares the method for corn protein peptide in the organic medium Download PDFInfo
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- CN100558906C CN100558906C CNB2006101239142A CN200610123914A CN100558906C CN 100558906 C CN100558906 C CN 100558906C CN B2006101239142 A CNB2006101239142 A CN B2006101239142A CN 200610123914 A CN200610123914 A CN 200610123914A CN 100558906 C CN100558906 C CN 100558906C
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Abstract
The invention discloses a kind of in organic medium biocatalysis prepare the method for corn protein peptide.This method is that the ratio by Zein powder and the consumption of organic solvent is that 0.01～10g: 1ml adds Zein powder in reactor, weight ratio according to Zein powder and water is 20～1: 1 adds entry, in the weight ratio of biological catalyst and zein is that 0.1～10: 1 ratio adds immobilized enzyme, in temperature is after 30～70 ℃, hunting speed are to react 3～48 hours under 100～300rpm, the normal pressure, separates obtaining corn protein peptide.The present invention has overcome that the easy inactivation of enzyme in the prior art, reaction efficiency are low, high temperature goes out the shortcoming that the enzyme energy consumption is big, zymoprotein is easily residual; Have that cost is low, reaction conditions is gentle, the easy isolating advantage from reaction mixture of environmental friendliness, reaction process simple controllable, product.
Description
Technical field
The present invention relates to biocatalysis and biosynthesizing field, be specifically related to a kind of in organic medium biocatalysis prepare the method for corn protein peptide.
Background technology
China's corn annual production has reached more than 1.1 hundred million t, accounts for 22% of Gross World Product, occupies the second place of the world.About 4,000,000 t of corn wet method Starch Production factory annual consumption corn, produce about 210,000 t of Zein powder, these Zein powders contain 40%～60% protein, but because the amino acid imbalance in the Zein powder, especially the necessary amino acid of some organisms such as Methionin, tryptophane etc. lack, make its value lower, limited its application.For a long time, Zein powder can only be sold at a bargain or discharge naturally as feed, and the corn protein that China every year drains with waste liquid is up to 90,000 t, not only contaminate environment, and caused proteinic huge waste, therefore imperative to its research of carrying out deep processing.Corn protein peptide is a kind of hydrolysate of zein, and is little and have very highly active little peptide molecule and form by molecular weight.There are some researches show, corn protein peptide has the inhibition elevation of blood pressure, antifatigue and reduce cholesterol in the blood, reduce absorption of human body alcoholic acid speed, anti-oxidant, radioprotective and the effects such as formation of root system when suppressing plant seed germination, in fields such as preventive medicine, pharmacy industry, food chemical industry, biotechnology, all have important use and be worth and wide application prospect.The hydrolysate of corn protein peptide can have different physico-chemical properties and biological activity according to the difference of its structure, can be used as pharmaceutical preparation auxiliary material with biocompatibility or physiologically active, natural medicinal formulations (as blood pressure lowering peptide), green chemical (as antioxidant, weedicide) etc.
The present domestic main employing enzyme process of preparation of corn protein peptide, generally with alkaline aqueous solution or short chain alcohol one alkaline aqueous solution as reaction medium, enzyme powder or liquid enzymes are as catalyzer.This method steps is various, and percent hydrolysis is low, energy consumption is big, the difficult separation of product; Adopt alcoholic solution in the reaction process,, very easily make enzyme that irreversible inactivation takes place, greatly reduce reaction efficiency although can dissolve substrate preferably; And owing to can only utilize the high temperature enzyme that goes out to come stopped reaction after the reaction, not only increase energy consumption, and it is low to be difficult to leave away enzyme, enzyme utilization ratio of branch behind the enzyme that goes out, and has increased the Financial cost of this method and the difficulty of product separation and purification greatly; In addition, the discharging of a large amount of alkali wasteliquids in reaction back can bring the serious environmental pollution problem.Therefore, research and development corn protein peptide new preparation process has great practice significance.
Summary of the invention
The objective of the invention is in order to solve the deficiency that above-mentioned prior art exists, the objective of the invention is to the problem that exists at prior art processes, the method that provide that a kind of environmental friendliness, production cost are low, reaction process simple controllable, the segregative organic medium biocatalysis of product prepares corn protein peptide.
Purpose of the present invention is achieved through the following technical solutions:
A kind of in organic medium biocatalysis prepare the method for corn protein peptide: the ratio by Zein powder and the consumption of organic solvent in reactor is that 0.01～10g: 1ml adds Zein powder, weight ratio according to Zein powder and water is 20～1: 1 adds entry, in the weight ratio of biological catalyst and zein is that 0.1～10: 1 ratio adds immobilized enzyme, in temperature is after 30～70 ℃, hunting speed are to react 3～48 hours under 100～300rpm, the normal pressure, separates obtaining corn protein peptide.
Described proteolytic enzyme derives from Bacillus licheniformis (Bacillus licheniforms), bacillus amyloliquefaciens (Bacillus amyloliquefaciens), subtilis (Bacillus subtilis), Rhizomucor miehei (Rhizomucor miehei), the brown Mucor of plum (Mucor miehei), papaya latex, Fructus Fici, the dirty or pancreas of animal stomach.
Described organic solvent is selected from a kind of in butane, toluene, ethyl acetate, tetrahydrofuran (THF), the trimethyl carbinol, the pyridine or two kinds.
It is as follows to separate the corn protein peptide method: reaction mixture is removed immobilized enzyme after filtration, and organic solvent is removed in evaporation, the corn protein peptide that resulting throw out can obtain having different molecular weight with the ultra-filtration membrane or the nanofiltration membrane fractional separation of different interceptions.
Principle of the present invention: adopt organic medium, as catalyzer, the reaction that is hydrolyzed of catalysis corn protein peptide obtains corn protein peptide with immobilization proteinase.
Compared with prior art, the present invention has following advantage:
(1) adopts biological catalyst immobilization proteinase efficient, high stability to come the catalysis corn protein peptide synthetic, overcome the easy inactivation of enzyme in the prior art, the shortcoming that reaction efficiency is low; Reaction process simple controllable, product are easily separated;
(2) reaction terminating adopt to filter the method for dezymotizing, and is simple, convenient and rapid, overcome in the prior art high temperature shortcomings such as the enzyme energy consumption is big, zymoprotein is easily residual of going out; And immobilized enzyme can use repeatedly, reduces production costs;
(3) utilized the enzymatic new technology of nonaqueous phase, adopted organic solvent as reaction medium, reaction back solvent can be removed by evaporation, has simplified the product separating step, and the product impurity that obtains is few; The organic solvent of removing through evaporation can recycle and reuse, and has reduced production cost;
(4) reaction conditions gentleness, no alkali waste discharging, environmental friendliness.
Embodiment
For better understanding the present invention, below in conjunction with embodiment the present invention is done detailed description further, but the scope of protection of present invention is not limited to the scope that embodiment represents.
Embodiment 1
The immobilization proteinase and the 6ml butane that 0.5g zein, 25mg water, 50mg are derived from papaya latex are put into tool plug triangular flask, place vibration in 30 ℃, the constant temperature water bath vibrator of 100rpm, behind the reaction 3h, remove by filter immobilization proteinase, organic solvent is removed in evaporation, and resulting throw out can obtain the corn protein peptide of molecular weight at 6000～5000Da with ultra-filtration membrane 6000 films and 5000 film fractional separation.
Embodiment 2
The proteolytic enzyme and the 10ml trimethyl carbinol that 0.5g corn protein peptide, 3mg water, 50mg are derived from Bacillus licheniformis (Bacilluslicheniforms) are put into tool plug triangular flask, place vibration in 60 ℃, the constant temperature water bath vibrator of 200rpm, behind the reaction 12h, remove by filter immobilization proteinase, organic solvent is removed in evaporation, and resulting throw out can obtain the corn protein peptide of molecular weight at 5000～2000Da with ultra-filtration membrane 5000 films and the 2000 film fractional separation of different interceptions.
Embodiment 3
0.5g corn protein peptide, 30mg water, 200mg are derived from bacillus amyloliquefaciens (Bacillusamyloliquefaciens) proteolytic enzyme and 6ml toluene is put into tool plug triangular flask, place vibration in 50 ℃, the constant temperature water bath vibrator of 300rpm, behind the reaction 24h, remove by filter immobilization proteinase, organic solvent is removed in evaporation, and resulting throw out can obtain the corn protein peptide of molecular weight at 2000～1000Da with ultra-filtration membrane 2000 films and 1000 film fractional separation.
Embodiment 4
0.5g corn protein peptide, 50mg water, 600mg are derived from the proteolytic enzyme and 6ml ethyl acetate of subtilis (Bacillus subtilis), place vibration in 70 ℃, the constant temperature water bath vibrator of 250rpm, behind the reaction 36h, remove by filter immobilization proteinase, organic solvent is removed in evaporation, and resulting throw out molecular weight cut-off is that 800 and 1000 nanofiltration membrane fractional separation can obtain the corn protein peptide of molecular weight at 800～1000Da.
Embodiment 5
Put into tool plug triangular flask to what 0.5g corn protein peptide, 100mg water, 1000mg derived from Rhizomucor miehei (Rhizomucormiehei) with the 6ml pyridine, place vibration in 50 ℃, the constant temperature water bath vibrator of 150rpm, behind the reaction 24h, remove by filter immobilization proteinase, organic solvent is removed in evaporation, and resulting throw out molecular weight cut-off is that 700～800 nanofiltration membrane separation can obtain the corn protein peptide of molecular weight at 700～800Da.
Embodiment 6
The proteolytic enzyme and the 50 μ l tetrahydrofuran (THF)s that 0.5g corn protein peptide, 500mg water, 5000mg are derived from the brown Mucor of plum (Mucor miehei) are put into tool plug triangular flask, place vibration in 40 ℃, the constant temperature water bath vibrator of 200rpm, behind the reaction 30h, remove by filter immobilization proteinase, organic solvent is removed in evaporation, and resulting throw out molecular weight cut-off is that 400～500 nanofiltration membrane fractional separation can obtain the corn protein peptide of molecular weight at 400～500Da.
Embodiment 7
The immobilization proteinase and the 10ml butane-tetrahydrofuran (THF) mixed solvent that 0.5g corn protein peptide, 100mg water, 1000mg are derived from animal pancreas are put into tool plug triangular flask, place vibration in 50 ℃, the constant temperature water bath vibrator of 150rpm, behind the reaction 48h, remove by filter immobilization proteinase, organic solvent is removed in evaporation, and resulting throw out molecular weight cut-off is that 150～200 nanofiltration membrane separation can obtain the corn protein peptide of molecular weight at 150～200Da.
Embodiment 8
0.5g zein, 25mg water, 50mg are derived from the dirty immobilization proteinase of animal stomach and 10ml pyridine-trimethyl carbinol is put into tool plug triangular flask, place vibration in 70 ℃, the constant temperature water bath vibrator of 250rpm, behind the reaction 48h, remove by filter immobilization proteinase, organic solvent is removed in evaporation, and resulting throw out molecular weight cut-off is that 80～100 nanofiltration membrane separation can obtain the corn protein peptide of molecular weight at 80～100Da.
Embodiment 9
The immobilization proteinase and the 10ml pyridine-trimethyl carbinol that 0.5g zein, 25mg water, 50mg are derived from Fructus Fici are put into tool plug triangular flask, place vibration in 60 ℃, the constant temperature water bath vibrator of 300rpm, behind the reaction 48h, remove by filter immobilization proteinase, organic solvent is removed in evaporation, and resulting throw out molecular weight cut-off is that 80～100 nanofiltration membrane separation can obtain the corn protein peptide of molecular weight at 80～100Da.
Claims (6)
1, a kind of in organic medium biocatalysis prepare the method for corn protein peptide, it is characterized in that: the ratio by Zein powder and the consumption of organic solvent in reactor is that 0.01～10g: 1ml adds Zein powder, weight ratio according to Zein powder and water is 20～1: 1 adds entry, in the weight ratio of biological catalyst and zein is that 0.1～10: 1 ratio adds immobilization proteinase, in temperature is after 30～70 ℃, hunting speed are to react 3～48 hours under 100～300rpm, the normal pressure, separates obtaining corn protein peptide;
Described immobilization proteinase derives from Bacillus licheniformis (Bacillus licheniforms), bacillus amyloliquefaciens (Bacillus amyloliquefaciens), subtilis (Bacillus subtilis), Rhizomucor miehei (Rhizomucor miehei), the brown Mucor of plum (Mucor miehei), papaya latex, Fructus Fici, the dirty or pancreas of animal stomach;
Described organic solvent is selected from one or both in butane, toluene, ethyl acetate, tetrahydrofuran (THF), the trimethyl carbinol and the pyridine.
2, according to claim 1 in organic medium biocatalysis prepare the method for corn protein peptide, it is characterized in that described immobilization proteinase derives from microorganism, derive from Bacillus licheniformis, bacillus amyloliquefaciens, subtilis, the brown Mucor of Rhizomucor miehei or plum.
3, according to claim 1 in organic medium biocatalysis prepare the method for corn protein peptide, it is characterized in that described immobilization proteinase derives from plant, derives from papaya latex or Fructus Fici.
4, according to claim 1 in organic medium biocatalysis prepare the method for corn protein peptide, it is characterized in that described proteolytic enzyme derives from the dirty or pancreas of animal stomach.
5, according to claim 1 in organic medium biocatalysis prepare the method for corn protein peptide, it is characterized in that described hunting speed is 200～300rpm, described temperature of reaction is 40～60 ℃.
6, according to claim 1 in organic medium biocatalysis prepare the method for corn protein peptide, it is characterized in that the described reaction mixture that is separated into removes immobilization proteinase after filtration, organic solvent is removed in evaporation, the corn protein peptide that resulting throw out can obtain having different molecular weight with the ultra-filtration membrane or the nanofiltration membrane fractional separation of different interceptions.
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| CN102174626B (en) * | 2011-01-12 | 2013-05-01 | 武汉百信食品有限公司 | Method for preparing corn peptide |
| CN103141667A (en) * | 2013-04-08 | 2013-06-12 | 黑龙江省轻工科学研究院 | Method for producing corn protein peptide used for feeding by spent grains liquid of corn alcohol |
| CN113016453A (en) * | 2021-03-03 | 2021-06-25 | 浙江东郁广陈果业有限公司 | Apple nursery weeding method |
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Effective date of registration: 20170608 Address after: 510635, C28, Room 501, 1 John King Road, Guangzhou, Guangdong, Tianhe District Patentee after: Guangzhou Huayuan Food Science & Technology Co., Ltd. Address before: 510640 Tianhe District, Guangdong, No. five road, No. 381, Patentee before: South China University of Technology |
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