CN103875906B - A kind of method of comprehensive utilization of the wheat embryo dregs of rice - Google Patents
A kind of method of comprehensive utilization of the wheat embryo dregs of rice Download PDFInfo
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Abstract
The invention discloses a kind of method of comprehensive utilization of the wheat embryo dregs of rice.The method with the wheat embryo dregs of rice for raw material, by steps such as twice flooding, enzymolysis, separation, acid precipitations, the products such as obtained glutathione, the homogeneous polypeptide of molecular weight, highly purified wheat plantule protein and animal feed, substantially increase the comprehensive utilization ratio of wheat embryo, solvent use amount is little, and not with an organic solvent, process stabilizing, easy and simple to handle, Standardization instrument, pollution-free, easy to utilize.
Description
Technical field
The present invention relates to technical field of biochemical industry, particularly relate to a kind of method of comprehensive utilization of the wheat embryo dregs of rice.
Background technology
Wheat embryo is the byproduct of wheat processing, the protein containing abundant, high-quality, fat, multivitamin and mineral matter.Owing to being rich in pigment and unrighted acid, be grinding into flour, Flour colour, long term storage performance, baking quality can be affected.China's wheat annual production is 100,000,000 tons, and the wheat embryo amount that can develop is up to 2,000,000 tons.Wheat embryo resource is fully utilized, grain resource utilization rate can be improved, can great wealth be created simultaneously.
At present, deep development and utilize defatted wheat germ protein resource to become the study hotspot of contemporary functional food, as the U.S., Japan and other countries carry out Depth Study to grain resource, makes it rise in value more than 10 times by the comprehensive utilization of grain germ.Wheat plantule protein is a kind of protein resource of high-quality, and its protein content is about 30%, and containing 8 kinds of essential amino acids, account for 34.7% of total amino acid, especially lysine, histidine and methionine content are all higher, far away higher than rice and flour.Wheat plantule protein has been widely used in the fields such as nutriment, health products, remedy diet.Wheat plantule protein contains high-quality full price phytoprotein, and has the functional characters such as good retentiveness, emulsibility, foaming characteristic, not only can increase the local flavor of food but also can improve structure and the quality of food.Contain in wheat embryo and manyly have bioactive amino acid sequence, just can obtain activated peptide section with enzyme hydrolysis, these peptide sections have anti-oxidant, anti-ageing function of waiting for a long time, high nutritive value.Glutathione content in wheat embryo is up to 98 ~ 107mg/100g, and the content of glutathione is the important indicator weighing antioxidant ability of organism size.Therefore, comprehensive utilization wheat plantule protein, polypeptide resource, the utilization rate improving wheat embryo has important Social benefit and economic benefit.
The method of existing exploitation wheat embryo resource, is mainly made into common punching food drink, health food, food additives etc.As " Wheat plumule protein drink " patent that the patent No. is CN03126118.3, disclose a kind of method utilizing wheat plantule protein preparation of drinks, this beverage is made up of wheat embryo, emulsifing thickener, fumet, antioxidant, sweetener, wherein wheat embryo content is 5-25%, the method just directly utilizes wheat embryo, do not carry out the deep processing such as degreasing, protein extraction to it, resource utilization is low.And for example the patent No. is " combined preparation process of wheat plantule protein and derived product thereof " patent of CN200710078256.4, disclose a kind of preparation method of wheat embryo Related product, the method with drying wheat plumule for raw material, through degreasing, solvent extraction, be separated, the techniques such as enzyme hydrolysis, prepare containing wheat embryo alcohol soluble protein, wheat embryo is anti-oxidant, anti-senility liquid, the multiple products such as hydrolyzed wheat plumule peptide concentrate, but the method does not carry out purifying to natural polypeptides such as glutathione, utilization rate is low, use methyl alcohol, the organic solvents such as ethanol, and solvent load is large, can impact environment.
At present, in China, the research for wheat embryo mainly concentrates on wheat-germ oil and wheat plantule protein two aspects, wherein for the exploitation relative maturity of wheat-germ oil, and the wheat embryo dregs of rice major part after degreasing is used for animal feed, cause the serious waste of high-quality protein resource.
Summary of the invention
The object of the invention is to the method for comprehensive utilization proposing the high wheat embryo dregs of rice of a kind of comprehensive utilization ratio.The method with the wheat embryo dregs of rice for raw material, the products such as glutathione, the homogeneous polypeptide of molecular weight, highly purified wheat plantule protein and animal feed are obtained by steps such as twice flooding, separation, enzymolysis, acid precipitations, substantially increase the comprehensive utilization ratio of wheat embryo, solvent use amount is little, and not with an organic solvent, process stabilizing, easy and simple to handle, Standardization instrument, pollution-free, easy to utilize.
For reaching this object, the present invention by the following technical solutions:
A method of comprehensive utilization for the wheat embryo dregs of rice, the method comprises:
(1) first time extracts: the wheat embryo dregs of rice are added flooding, then Separation of Solid and Liquid;
(2) separating glutathione: make liquid that step (1) is separated by metal chelate affinity chromatography medium, collects and penetrates liquid; Carry out wash-out with elution buffer, collect eluent, and desalination, concentrated, dry is carried out to eluent, obtain glutathione;
(3) the homogeneous polypeptide of molecular weight is prepared: use trypsase to carry out enzyme digestion reaction to the liquid that penetrates collected in step (2); After enzyme digestion reaction, classification process is carried out to enzymolysis liquid, concentrated, dry the homogeneous polypeptide of molecular weight;
(4) second time is extracted: the solid that step (1) is separated is added flooding, then Separation of Solid and Liquid;
(5) wheat plantule protein is prepared: the liquid that step (4) is separated is carried out acid precipitation, then Separation of Solid and Liquid; Acid precipitation gained solid is redissolved, then through concentrated, desalination, dry wheat plantule protein;
(6) animal feed is prepared: the solid be separated step (4) and step (5) acid precipitation gained liquid merge, dry, obtain animal feed.
In above-mentioned method of comprehensive utilization, as preferably, in step (1), the w/v by g/ml of the described wheat embryo dregs of rice and water is 1:(4-10), preferred 1:(5-8), more preferably 1:(6-7);
In a particular embodiment, the w/v by g/ml of the described wheat embryo dregs of rice and water is 1:4,1:5,1:6,1:7,1:8,1:9,1:10.
Preferably, step (1) described extraction temperature is 25-85 DEG C, is preferably 30-80 DEG C, is more preferably 50-60 DEG C;
In a particular embodiment, step (1) described extraction temperature is 25 DEG C, 30 DEG C, 35 DEG C, 40 DEG C, 45 DEG C, 50 DEG C, 55 DEG C, 60 DEG C, 65 DEG C, 70 DEG C, 75 DEG C, 80 DEG C, 85 DEG C.
Preferably, step (1) described extraction time is 10-50 minute, is preferably 10-30 minute, is more preferably 20 minutes;
In a particular embodiment, step (1) described extraction time is 10 minutes, 20 minutes, 30 minutes, 40 minutes, 50 minutes.
As preferably, in step (2), described metal chelate affinity chromatography medium is chelating copper ions affinity chromatography medium;
Preferably, described metal chelate affinity chromatography medium, before loading, first balances it with level pad;
Further preferably, described level pad is 10-500mM, pH value is the phosphate buffer of 6.0-8.5, and preferably 20-200mM, pH value are the phosphate buffer of 6.5-7.5, be more preferably 50-100mM, pH value is the phosphate buffer of 7.0;
In a particular embodiment, the concentration of described phosphate buffer is 10mM, 20mM, 50mM, 100mM, 150mM, 200mM, 250mM, 300mM, 350mM, 400mM, 450mM, 500mM; Its pH value is 6.0,6.5,7.0,7.5,8.0,8.5.
As preferably, in step (2), described elution buffer is 10-500mM, pH value is 6.0-8.5, phosphate buffer containing 0.5-2M sodium chloride, be preferably that 20-200mM, pH value are 6.5-7.5, phosphate buffer containing 0.5-1.5M sodium chloride, be more preferably 50-100mM, pH value is 7.0, phosphate buffer containing 1M sodium chloride;
In a particular embodiment, the concentration of described phosphate buffer is 10mM, 20mM, 50mM, 100mM, 150mM, 200mM, 250mM, 300mM, 350mM, 400mM, 450mM, 500mM; Its pH value is 6.0,6.5,7.0,7.5,8.0,8.5; The content of sodium chloride is wherein 0.5M, 1M, 1.5M, 2.0M.
As preferably, in step (3), before enzyme digestion reaction, the pH value penetrating liquid of collecting in first regulating step (2) to 6.5-8.5, be preferably 7.5-8.2, be more preferably 8.0;
Preferably, by mass, described tryptic consumption be penetrate liquid total protein quality 0.5-1.5%, be preferably 0.5-1%, be more preferably 0.75%;
Preferably, described enzyme digestion reaction temperature is 30-45 DEG C, is preferably 35-40 DEG C, is more preferably 37 DEG C;
Preferably, the described enzyme digestion reaction time is 1-5 hour, is preferably 2-4 hour, is more preferably 3 hours.
As preferably, in step (3), after enzyme digestion reaction, first the pH value of enzymolysis liquid is adjusted to 6.5-7.5, preferably 7.0, then classification process is carried out to enzymolysis liquid; Further preferably, milipore filter is used to carry out classification process.
As preferably, in step (4), the w/v by g/ml of the solid that described step (1) is separated and water is 1:(3-8), preferred 1:(4-6), more preferably 1:5;
In a particular embodiment, the w/v by g/ml of the solid that is separated of described step (1) and water is 1:3,1:4,1:5,1:6,1:7,1:8.
Preferably, step (4) described extraction temperature is 25-85 DEG C, is preferably 30-80 DEG C, is more preferably 50-60 DEG C;
In a particular embodiment, step (4) described extraction temperature is 25 DEG C, 30 DEG C, 35 DEG C, 40 DEG C, 45 DEG C, 50 DEG C, 55 DEG C, 60 DEG C, 65 DEG C, 70 DEG C, 75 DEG C, 80 DEG C, 85 DEG C.
Preferably, step (4) described extraction time is 20-70 minute, is preferably 30-60 minute, is more preferably 40 minutes;
In a particular embodiment, step (4) described extraction time is 20 minutes, 30 minutes, 40 minutes, 50 minutes, 60 minutes, 70 minutes.
As preferably, in step (5), described acid precipitation is specially: the liquid pH value that step (4) is separated is adjusted to 3.6-4.6, is preferably 4.0-4.2, be more preferably 4.0 and precipitate;
In a particular embodiment, the liquid pH value that step (4) is separated is adjusted to 3.6,3.8,4.0,4.2,4.4,4.6.
Preferably, use the phosphate buffer of 10-500mM, pH6.5-8.0, preferred 20-200mM, pH value are the phosphate buffer of 6.5-7.5, more preferably 50-100mM, pH value be 7.0 phosphate buffer acid precipitation gained solid is redissolved;
Further preferably, the w/v by g/ml of described acid precipitation gained solid and described phosphate buffer is 1:(10-50), preferably 1:(20-40), be more preferably 1:30.
As preferably, in step (6), after the solid be separated step (4) and step (5) acid precipitation gained liquid merge, adjust ph to 6.5-7.5, be preferably 7.0, spraying dry, obtains animal feed.
Most preferably, the method for comprehensive utilization of the wheat embryo dregs of rice of the present invention comprises the steps:
(1) first time extracts: be 1:(5-8 with water by the w/v of g/ml by the wheat embryo dregs of rice) mix, stirring and leaching 10-30 minute at 30-80 DEG C, rear centrifugation, collection supernatant and precipitation respectively;
(2) separating glutathione: balance chelating copper ions affinity column with level pad, then imports step (1) gained supernatant, collects and penetrates liquid; With elution buffer wash-out pillar, collect eluent, and desalination, concentrated, dry is carried out to eluent, obtain glutathione;
Described level pad is 10-500mM, pH value is the phosphate buffer of 6.0-8.5;
Described elution buffer is 10-500mM, pH value is 6.0-8.5, phosphate buffer containing 0.5-2M sodium chloride;
(3) the homogeneous polypeptide of molecular weight is prepared: the pH value penetrating liquid of collecting in step (2) is adjusted to 8.0, adds the trypsase of the 0.5-1% penetrating liquid total protein quality by mass, enzyme digestion reaction 1-5 hour at 37 DEG C; After having reacted, the pH value of enzymolysis liquid is adjusted to neutrality, then with milipore filter, classification process is carried out to enzymolysis liquid, obtain the homogeneous polypeptide of molecular weight finally by concentrated, dry;
(4) second time is extracted: be 1:(4-6 with water by the w/v of g/ml by step (1) gained precipitation) mix, stirring and leaching 30-60 minute at 30-80 DEG C, rear centrifugation, collection supernatant and precipitation respectively;
(5) wheat plantule protein is prepared: step (4) gained supernatant pH value is adjusted to 4.0-4.6 and carries out acid precipitation, centrifugation, collect supernatant and precipitation respectively; In gained precipitation, add 10-500mM, phosphate buffer that pH value is 6.0-8.5 redissolves, the w/v by g/ml of described precipitation and phosphate buffer is 1:(10-50); Then wheat plantule protein is obtained through ultrafiltration concentration, desalination, spraying dry;
(6) animal feed is prepared: step (4) gained precipitation and step (5) acid precipitation gained supernatant are merged, adjust ph is to neutral, and spraying dry obtains animal feed.
The method of comprehensive utilization of the wheat embryo dregs of rice provided by the invention has the following advantages:
(1) comprehensive utilization ratio is high: the method that the present invention adopts classification to be separated, extracted by first time and achieve low molecular weight polypeptide and albumen quick separating, first time extract isolate glutathione by metal chelate affinity chromatography medium, then by affinity chromatography medium penetrate liquid enzymolysis after prepare the homogeneous polypeptide products of molecular weight; Then the solids of sedimentation after being extracted first time carries out second time and extracts, and second time extract carries out the highly purified wheat plantule protein product of sour precipitation.
(2) solvent use amount is few, and not with an organic solvent; The total water consumption that two steps are extracted is no more than 12 times of the wheat embryo dregs of rice, and solvent for use is the aqueous solution of water or inorganic salts, and environmental friendliness is pollution-free.
(3) process stabilizing, easy and simple to handle, Standardization instrument, with low cost, easy to utilize.
The method of comprehensive utilization of the wheat embryo dregs of rice of the present invention can fully utilize wheat embryo resource, prepare the wheat germ polypeptide of high added value, protein product etc., the industries such as food, nutrition and health care, biochemistry, daily use chemicals can be widely used in, be conducive to the sustainable development of traditional food industry, be conducive to the competitiveness of raising industry.
Detailed description of the invention
Technical scheme of the present invention is further illustrated below by detailed description of the invention.It will be appreciated by those skilled in the art that these embodiments are only for illustration of the present invention, its scope do not limited the present invention in any way.
Embodiment 1
1) taking wheat embryo dregs of rice 10kg is positioned in agitator tank, and add water 50L, stirring and leaching 10 minutes at 30 DEG C, has extracted rear 10000g centrifugal 15 minutes, collects supernatant and precipitation respectively, obtains supernatant 38.3L, precipitation 19.2kg.
2) 1L chelating copper ions agarose affinity chromatography medium dress post is got, with the phosphate buffer of 3L20mM pH7.0, affinity column is balanced, import in affinity column by the supernatant collected in step 1), collect and penetrate liquid 45.6L, the total protein concentration penetrating liquid is 6.3g/L; Then carry out wash-out with 20mM pH8.5 containing the phosphate buffer of 1M sodium chloride, collect eluent.Desalination, concentrated, dry is carried out to eluent, obtains glutathione product 5.6g.
3) by step 2) in collect the liquid that penetrates be placed in reactor, adjust pH to 8.0, add 1.4g trypsase, enzymolysis is stirred 1 hour at 37 DEG C, enzymolysis liquid adjusts pH to neutral, the milipore filter of molecular cut off 10kDa, 5kDa, 3kDa, 1kDa is used to carry out classification process successively, concentrated, spraying dry obtains the polypeptide 125.4g that molecular weight is greater than polypeptide 7.6g, the polypeptide 15.4g of molecular weight 10-5kDa of 10kDa, the polypeptide 20.1g of molecular weight 5-3kDa, the polypeptide 27.4g of molecular weight 5-1kDa, molecular weight are less than 1kDa respectively.
4) be positioned in container by the 19.2kg collected in step 1) precipitation and 76.8L water, stirring and leaching 30 minutes at 30 DEG C, has extracted rear 10000g centrifugal 15 minutes, collects supernatant and precipitation respectively.
5) supernatant collected in step 4) imported in the heavy still of acid, adjust pH 4.0 to precipitate, after precipitation half an hour, centrifugal 15 minutes of 10000g, collects supernatant and precipitation respectively, is precipitated 86.1g.The phosphate buffer of 861ml20mM pH7.0 is added precipitation, redissolves, after ultrafiltration concentration desalination, spraying dry obtains wheat plantule protein product 83.2g.
6) supernatant that the precipitation of step 4) being collected and step 5) are collected merges, and adjusts pH to neutral, carries out spraying dry and obtain 18.7kg animal feed product.
Embodiment 2
1) taking wheat embryo dregs of rice 10kg is positioned in agitator tank, and add water 80L, stirring and leaching 30 minutes at 50 DEG C, has extracted rear 10000g centrifugal 15 minutes, collects supernatant and precipitation respectively, obtains supernatant 67.9L, precipitation 19.8kg.
2) 1L chelating copper ions agarose affinity chromatography medium dress post is got, with the phosphate buffer of 3L20mM pH7.0, affinity column is balanced, the supernatant collected in step 1) is imported in affinity column, collects and penetrate liquid 71.3L, penetrate the total protein concentration 4.7g/L of liquid; Then carry out wash-out with 20mM pH8.5 containing the phosphate buffer of 1M sodium chloride, collect eluent.Desalination, concentrated, dry is carried out to eluent, obtains glutathione product 8.3g.
3) by step 2) in collect the liquid that penetrates be placed in reactor, adjust pH to 8.0, add 1.7g trypsase, enzymolysis is stirred 1 hour at 37 DEG C, enzymolysis liquid adjusts pH to neutral, the milipore filter of molecular cut off 10kDa, 5kDa, 3kDa, 1kDa is used to carry out classification process successively, concentrated, spraying dry obtains the polypeptide 130.8g that molecular weight is greater than polypeptide 5.4g, the polypeptide 14.7g of molecular weight 10-5kDa of 10kDa, the polypeptide 18.7g of molecular weight 5-3kDa, the polypeptide 25.6g of molecular weight 5-1kDa, molecular weight are less than 1kDa respectively.
4) be positioned in container by the 19.8kg collected in step 1) precipitation and 118.8L water, stirring and leaching 60 minutes at 50 DEG C, has extracted rear 10000g centrifugal 15 minutes, collects supernatant and precipitation respectively.
5) supernatant collected in step 4) imported in the heavy still of acid, adjust pH 4.0 to precipitate, after precipitation half an hour, centrifugal 15 minutes of 10000g, collects supernatant and precipitation respectively, is precipitated 102.9g.The phosphate buffer of 1029ml20mM pH7.0 is added precipitation, redissolves, after ultrafiltration concentration desalination, spraying dry obtains wheat plantule protein product 100.8g.
6) supernatant that the precipitation of step 4) being collected and step 5) are collected merges, and adjusts pH to neutral, carries out spraying dry and obtain 18.4kg animal feed product.
Embodiment 3
1) taking wheat embryo dregs of rice 10kg is positioned in agitator tank, and add water 50L, stirring and leaching 10 minutes at 30 DEG C, has extracted rear 10000g centrifugal 15 minutes, collects supernatant and precipitation respectively, obtains supernatant 38.4L, precipitation 19.2kg.
2) 1L chelating copper ions agarose affinity chromatography medium dress post is got, with the phosphate buffer of 3L20mM pH7.0, affinity column is balanced, the supernatant collected in step 1) is imported in affinity column, collects and penetrate liquid 45.7L, penetrate the total protein concentration 6.3g/L of liquid; Then carry out wash-out with 20mM pH8.5 containing the phosphate buffer of 1M sodium chloride, collect eluent.Desalination, concentrated, dry is carried out to eluent, obtains glutathione product 5.6g.
3) by step 2) in collect the liquid that penetrates be placed in reactor, adjust pH to 8.0, add 2.8g trypsase, enzymolysis is stirred 1 hour at 37 DEG C, enzymolysis liquid adjusts pH to neutral, the milipore filter of molecular cut off 10kDa, 5kDa, 3kDa, 1kDa is used to carry out classification process successively, concentrated, spraying dry obtains the polypeptide 153.9g that molecular weight is greater than polypeptide 3.7g, the polypeptide 5.8g of molecular weight 10-5kDa of 10kDa, the polypeptide 10.4g of molecular weight 5-3kDa, the polypeptide 20.3g of molecular weight 5-1kDa, molecular weight are less than 1kDa respectively.
4) be positioned in container by the 19.2kg collected in step 1) precipitation and 115.2L water, stirring and leaching 60 minutes at 50 DEG C, has extracted rear 10000g centrifugal 15 minutes, collects supernatant and precipitation respectively.
5) supernatant collected in step 4) imported in the heavy still of acid, adjust pH 4.0 to precipitate, after precipitation half an hour, centrifugal 15 minutes of 10000g, collects supernatant and precipitation respectively, is precipitated 98.3g.The phosphate buffer of 4915ml20mM pH7.0 is added precipitation, redissolves, after ultrafiltration concentration desalination, spraying dry obtains wheat plantule protein product 97.9g.
6) supernatant that the precipitation of step 4) being collected and step 5) are collected merges, and adjusts pH to neutral, carries out spraying dry and obtain 18.6kg animal feed product.
Applicant states, the present invention illustrates technical scheme of the present invention by above-described embodiment, but the present invention is not limited to above-described embodiment, does not namely mean that the present invention must rely on above-described embodiment and could implement.Person of ordinary skill in the field should understand, any improvement in the present invention, to equivalence replacement and the interpolation of auxiliary element, the concrete way choice etc. of raw material selected by the present invention, all drops within protection scope of the present invention and open scope.
Claims (54)
1. a method of comprehensive utilization for the wheat embryo dregs of rice, is characterized in that, comprising:
(1) first time extracts: the wheat embryo dregs of rice are added flooding, then Separation of Solid and Liquid;
(2) separating glutathione: make liquid that step (1) is separated by metal chelate affinity chromatography medium, collects and penetrates liquid; Carry out wash-out with elution buffer, collect eluent, and desalination, concentrated, dry is carried out to eluent, obtain glutathione;
(3) the homogeneous polypeptide of molecular weight is prepared: use trypsase to carry out enzyme digestion reaction to the liquid that penetrates collected in step (2); After enzyme digestion reaction, classification process is carried out to enzymolysis liquid, concentrated, dry the homogeneous polypeptide of molecular weight;
(4) second time is extracted: the solid that step (1) is separated is added flooding, then Separation of Solid and Liquid;
(5) wheat plantule protein is prepared: the liquid that step (4) is separated is carried out acid precipitation, then Separation of Solid and Liquid; Acid precipitation gained solid is redissolved, then through concentrated, desalination, dry wheat plantule protein;
(6) animal feed is prepared: the solid be separated step (4) and step (5) acid precipitation gained liquid merge, dry, obtain animal feed.
2. method of comprehensive utilization according to claim 1, is characterized in that, in step (1), the w/v by g/ml of the described wheat embryo dregs of rice and water is 1:(4-10).
3. method of comprehensive utilization according to claim 2, is characterized in that, in step (1), the w/v by g/ml of the described wheat embryo dregs of rice and water is 1:(5-8).
4. method of comprehensive utilization according to claim 3, is characterized in that, in step (1), the w/v by g/ml of the preferred described wheat embryo dregs of rice and water is 1:(6-7).
5. according to the method for comprehensive utilization one of claim 1-4 Suo Shu, it is characterized in that, in step (1), described extraction temperature is 25-85 DEG C.
6. method of comprehensive utilization according to claim 5, is characterized in that, in step (1), described extraction temperature is 30-80 DEG C.
7. method of comprehensive utilization according to claim 6, is characterized in that, in step (1), described extraction temperature is 50-60 DEG C.
8. according to the method for comprehensive utilization one of claim 1-4 Suo Shu, it is characterized in that, in step (1), described extraction time is 10-50 minute.
9. method of comprehensive utilization according to claim 8, is characterized in that, in step (1), described extraction time is 10-30 minute.
10. method of comprehensive utilization according to claim 9, is characterized in that, in step (1), described extraction time is 20 minutes.
11., according to the method for comprehensive utilization one of claim 1-4 Suo Shu, is characterized in that, in step (2), described metal chelate affinity chromatography medium is chelating copper ions affinity chromatography medium.
12., according to the method for comprehensive utilization one of claim 1-4 Suo Shu, is characterized in that, in step (2), described metal chelate affinity chromatography medium, before loading, first balances it with level pad.
13. method of comprehensive utilization according to claim 12, is characterized in that, described level pad is 10-500mM, pH value is the phosphate buffer of 6.0-8.5.
14. method of comprehensive utilization according to claim 13, is characterized in that, described level pad is 20-200mM, pH value is the phosphate buffer of 6.5-7.5.
15. method of comprehensive utilization according to claim 14, is characterized in that, described level pad is 50-100mM, pH value is the phosphate buffer of 7.0.
16., according to the method for comprehensive utilization one of claim 1-4 Suo Shu, is characterized in that, in step (2), described elution buffer is 10-500mM, pH value is 6.0-8.5, phosphate buffer containing 0.5-2M sodium chloride.
17. method of comprehensive utilization according to claim 16, is characterized in that, in step (2), described elution buffer is 20-200mM, pH value is 6.5-7.5, phosphate buffer containing 0.5-1.5M sodium chloride.
18. method of comprehensive utilization according to claim 17, is characterized in that, in step (2), preferred described elution buffer is 50-100mM, pH value is 7.0, phosphate buffer containing 1M sodium chloride.
19., according to the method for comprehensive utilization one of claim 1-4 Suo Shu, is characterized in that, in step (3), before enzyme digestion reaction, the pH value penetrating liquid of collecting in first regulating step (2) is to 6.5-8.5.
20. method of comprehensive utilization according to claim 19, is characterized in that, in step (3), before enzyme digestion reaction, the pH value penetrating liquid of collecting in first regulating step (2) is to 7.5-8.2.
21. method of comprehensive utilization according to claim 20, is characterized in that, in step (3), before enzyme digestion reaction, and the pH value penetrating liquid of collecting in first regulating step (2) most 8.0.
22., according to the method for comprehensive utilization one of claim 1-4 Suo Shu, is characterized in that, in step (3), by mass, described tryptic consumption is the 0.5-1.5% penetrating liquid total protein quality.
23. method of comprehensive utilization according to claim 22, is characterized in that, in step (3), by mass, described tryptic consumption is the 0.5-1% penetrating liquid total protein quality.
24. method of comprehensive utilization according to claim 23, is characterized in that, in step (3), by mass, described tryptic consumption is penetrate liquid total protein quality 0.75%.
25., according to the method for comprehensive utilization one of claim 1-4 Suo Shu, is characterized in that, in step (3), described enzyme digestion reaction temperature is 30-45 DEG C.
26. method of comprehensive utilization according to claim 25, is characterized in that, in step (3), described enzyme digestion reaction temperature is 35-40 DEG C.
27. method of comprehensive utilization according to claim 26, is characterized in that, in step (3), described enzyme digestion reaction temperature is 37 DEG C.
28., according to the method for comprehensive utilization one of claim 1-4 Suo Shu, is characterized in that, in step (3), the described enzyme digestion reaction time is 1-5 hour.
29. method of comprehensive utilization according to claim 28, is characterized in that, in step (3), the described enzyme digestion reaction time is 2-4 hour.
30. method of comprehensive utilization according to claim 29, is characterized in that, in step (3), the described enzyme digestion reaction time is 3 hours.
31., according to the method for comprehensive utilization one of claim 1-4 Suo Shu, is characterized in that, in step (3), after enzyme digestion reaction, first the pH value of enzymolysis liquid are adjusted to 6.5-7.5, then carry out classification process to enzymolysis liquid.
32. method of comprehensive utilization according to claim 31, is characterized in that, in step (3), after enzyme digestion reaction, first the pH value of enzymolysis liquid are adjusted to 7.0, then carry out classification process to enzymolysis liquid.
33., according to the method for comprehensive utilization one of claim 1-4 Suo Shu, is characterized in that, in step (3), use milipore filter to carry out classification process.
34., according to the method for comprehensive utilization one of claim 1-4 Suo Shu, is characterized in that, in step (4), the w/v by g/ml of the solid that described step (1) is separated and water is 1:(3-8).
35. method of comprehensive utilization according to claim 34, is characterized in that, in step (4), the w/v by g/ml of the solid that described step (1) is separated and water is 1:(4-6).
36. method of comprehensive utilization according to claim 35, is characterized in that, in step (4), the w/v by g/ml of the solid that described step (1) is separated and water is 1:5.
37., according to the method for comprehensive utilization one of claim 1-4 Suo Shu, is characterized in that, in step (4), described extraction temperature is 25-85 DEG C.
38., according to method of comprehensive utilization according to claim 37, is characterized in that, in step (4), described extraction temperature is 30-80 DEG C.
39., according to method of comprehensive utilization according to claim 38, is characterized in that, in step (4), described extraction temperature is 50-60 DEG C.
40., according to the method for comprehensive utilization one of claim 1-4 Suo Shu, is characterized in that, in step (4), described extraction time is 20-70 minute.
41. method of comprehensive utilization according to claim 40, is characterized in that, in step (4), described extraction time is 30-60 minute.
42. method of comprehensive utilization according to claim 41, is characterized in that, in step (4), described extraction time is 40 minutes.
43., according to the method for comprehensive utilization one of claim 1-4 Suo Shu, is characterized in that, in step (5), described acid precipitation is specially: the liquid pH value that step (4) is separated is adjusted to 3.6-4.6 and precipitates.
44. method of comprehensive utilization according to claim 43, is characterized in that, in step (5), described acid precipitation is specially: the liquid pH value that step (4) is separated is adjusted to 4.0-4.2 and precipitates.
45. method of comprehensive utilization according to claim 44, is characterized in that, in step (5), described acid precipitation is specially: the liquid pH value that step (4) is separated is adjusted to 4.0 and precipitates.
46., according to the method for comprehensive utilization one of claim 1-4 Suo Shu, is characterized in that, in step (5), use the phosphate buffer of 10-500mM, pH6.5-8.0 to redissolve to acid precipitation gained solid.
47. method of comprehensive utilization according to claim 46, is characterized in that, in step (5), use the phosphate buffer of 20-200mM, pH6.5-7.5 to redissolve to acid precipitation gained solid.
48. method of comprehensive utilization according to claim 47, is characterized in that, in step (5), use the phosphate buffer of 50-100mM, pH7.0 to redissolve to acid precipitation gained solid.
49. method of comprehensive utilization according to claim 46, is characterized in that, the w/v by g/ml of described acid precipitation gained solid and described phosphate buffer is 1:(10-50).
50. method of comprehensive utilization according to claim 49, is characterized in that, the w/v by g/ml of described acid precipitation gained solid and described phosphate buffer is 1:(20-40).
51. method of comprehensive utilization according to claim 50, is characterized in that, the w/v by g/ml of described acid precipitation gained solid and described phosphate buffer is 1:30.
52., according to the method for comprehensive utilization one of claim 1-4 Suo Shu, is characterized in that, in step (6), after the solid be separated step (4) and step (5) acid precipitation gained liquid merge, adjust ph is to 6.5-7.5, and spraying dry, obtains animal feed.
53. method of comprehensive utilization according to claim 52, is characterized in that, in step (6), after the solid be separated step (4) and step (5) acid precipitation gained liquid merge, adjust ph to 7.0, spraying dry, obtains animal feed.
54. method of comprehensive utilization according to claim 1, is characterized in that, comprise the steps:
(1) first time extracts: be 1:(5-8 with water by the w/v of g/ml by the wheat embryo dregs of rice) mix, stirring and leaching 10-30 minute at 30-80 DEG C, rear centrifugation, collection supernatant and precipitation respectively;
(2) separating glutathione: balance chelating copper ions affinity column with level pad, then imports step (1) gained supernatant, collects and penetrates liquid; With elution buffer wash-out pillar, collect eluent, and desalination, concentrated, dry is carried out to eluent, obtain glutathione;
Described level pad is 10-500mM, pH value is the phosphate buffer of 6.0-8.5;
Described elution buffer is 10-500mM, pH value is 6.0-8.5, phosphate buffer containing 0.5-2M sodium chloride;
(3) the homogeneous polypeptide of molecular weight is prepared: the pH value penetrating liquid of collecting in step (2) is adjusted to 8.0, adds the trypsase of the 0.5-1% penetrating liquid total protein quality by mass, enzyme digestion reaction 1-5 hour at 37 DEG C; After having reacted, the pH value of enzymolysis liquid is adjusted to neutrality, then with milipore filter, classification process is carried out to enzymolysis liquid, obtain the homogeneous polypeptide of molecular weight finally by concentrated, dry;
(4) second time is extracted: be 1:(4-6 with water by the w/v of g/ml by step (1) gained precipitation) mix, stirring and leaching 30-60 minute at 30-80 DEG C, rear centrifugation, collection supernatant and precipitation respectively;
(5) wheat plantule protein is prepared: step (4) gained supernatant pH value is adjusted to 4.0-4.6 and carries out acid precipitation, centrifugation, collect supernatant and precipitation respectively; In gained precipitation, add 10-500mM, phosphate buffer that pH value is 6.0-8.5 redissolves, the w/v by g/ml of described precipitation and phosphate buffer is 1:(10-50); Then wheat plantule protein is obtained through ultrafiltration concentration, desalination, spraying dry;
(6) animal feed is prepared: step (4) gained precipitation and step (5) acid precipitation gained supernatant are merged, adjust ph is to neutral, and spraying dry obtains animal feed.
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| CN102406049A (en) * | 2011-10-25 | 2012-04-11 | 江苏江大源生态生物科技有限公司 | Joint preparation method of wheat germ oil and wheat germ protein |
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