CN110981951B - Preparation method for enriching immunoglobulin and lactoferrin in colostrum - Google Patents
Preparation method for enriching immunoglobulin and lactoferrin in colostrum Download PDFInfo
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- CN110981951B CN110981951B CN201911369456.4A CN201911369456A CN110981951B CN 110981951 B CN110981951 B CN 110981951B CN 201911369456 A CN201911369456 A CN 201911369456A CN 110981951 B CN110981951 B CN 110981951B
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- C07K14/79—Transferrins, e.g. lactoferrins, ovotransferrins
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Abstract
The invention discloses a preparation method for enriching immunoglobulin and lactoferrin in colostrum, which mainly comprises the following steps: 1) Separating and recovering milk fat by a centrifugal method; 2) Separating casein by combining a chymosin precipitation method and an acid precipitation method; 3) Centrifuging the solution to obtain supernatant as lactalbumin, and recovering the permeate after microfiltration and sterilization; 4) Concentrating whey with 50kDa ultrafiltration membrane to obtain double active components rich in immunoglobulin and lactoferrin. The invention adopts the combination of the rennin precipitation method and the acid precipitation method to separate the casein, can improve the yield of the whey protein, is beneficial to the subsequent enrichment of immunoglobulin and lactoferrin from the whey, takes colostrum as a raw material, has higher content of the immunoglobulin and the lactoferrin than normal milk, and can compound the obtained immunoglobulin and lactoferrin double-active components with active substances such as lysozyme, ginsenoside and the like to develop special diet food suitable for different people.
Description
Technical Field
The invention belongs to the technical field of functional food processing, relates to a preparation method for enriching immunoglobulin and lactoferrin in bovine colostrum, and discloses a novel preparation method.
Background
Colostrum is the milk secreted by female animals within 2-3 days after delivery, the content of protein, fat and sugar is 10-17 times of that of common milk, especially the content of immunoglobulin and lactoferrin is very high, the content of immunoglobulin is 40-150 times of that of common milk, and the content of lactoferrin is about 15 times of that of common milk, so the colostrum is known as the king of 21 century immunity. The colostrum product can be taken by weak middle-aged and elderly people, and has effects of improving autoimmune function and enhancing resistance.
The method comprises the steps of enriching immunoglobulin and lactoferrin double-active components from colostrum, preparing skim milk, separating casein from the skim milk to obtain whey protein, and enriching by taking the whey protein as a base material. The separation effect of casein will directly affect the yield of target protein, so how to increase the recovery rate of casein is the key to research on active protein separation. The current methods for separating casein are mainly acid precipitation, chymosin precipitation and fermentation agent combined chymosin precipitation. The acid precipitation method mainly adopts hydrochloric acid and lactic acid to adjust the pH value of the skim milk to about 4.6, the obtained clot is fine, a part of the clot can be lost into whey, and the casein yield is about 75 percent generally; the chymosin precipitation method is mainly characterized in that chymosin is added into milk to promote the casein precipitation, the obtained curd is large, but a part of whey can be wrapped, and the casein yield separated by the chymosin precipitation method is about 82 percent; the leavening agent is combined with the chymosin precipitation method, and the method mainly utilizes the leavening agent to ferment the skim milk, and then the chymosin is added to further promote the curd, and the yield of the casein is about 85 percent generally.
Disclosure of Invention
The invention aims to solve the technical problem of providing a preparation method for enriching immunoglobulin and lactoferrin in colostrum, wherein casein is separated by adopting a chymosin precipitation method and an acid precipitation method, the yield of whey protein can be improved, the subsequent enrichment of immunoglobulin and lactoferrin from whey is facilitated, the colostrum is used as a raw material, the content of immunoglobulin and lactoferrin is higher than that of normal milk, and the obtained immunoglobulin and lactoferrin double-active components can be compounded with active substances such as lysozyme, ginsenoside and the like.
The invention discloses a preparation method for enriching immunoglobulin and lactoferrin in colostrum, which is characterized by comprising the following steps:
(1) Separating butter fat: pasteurizing the primary emulsion, rapidly cooling to 30-35 deg.C, centrifuging at 3000-7000r/min with a milk fat separator, and removing fat;
(2) Separating casein by combining a chymosin precipitation method with an acid precipitation method: adding 0.01-0.2% (w/w) rennin into skim milk, reacting at 35-37 deg.C for 0.5-1 hr, adjusting pH of the emulsion to 4.6-4.8 with acid solution, centrifuging the solution at 7000-10000r/min to obtain casein precipitate and supernatant, which is whey protein, and recovering the supernatant;
(3) And (3) microfiltration sterilization: sterilizing the whey protein obtained in the step (2) by a microfiltration membrane of 0.1 mu m or 0.45 mu m, and taking the permeate for later use;
(4) And (3) active protein enrichment by ultrafiltration: and (3) carrying out ultrafiltration on the whey obtained in the step (3) by using a 50kDa ultrafiltration membrane to 30-50% of the original volume, recovering trapped fluid, adding ultrapure water to dilute the whey to the original volume, carrying out ultrafiltration again, repeating the process for three times, and carrying out freeze drying on the obtained sample solution for 24-48 hours to obtain the freeze-dried powder enriched with the immunoglobulin and lactoferrin double active components.
The recovery rate of the immune globulin and the lactoferrin can reach more than 92 percent, and the highest recovery rate of the enriched protein by the traditional membrane treatment method is about 85 percent.
The invention has the positive effects that:
adopting a chymosin precipitation method combined with an acid precipitation method to separate casein, firstly carrying out the first-stage curd by using chymosin, and then adjusting the pH value of the emulsion to about 4.6 by using an acid solution, wherein the method has better effect than the method of precipitating the casein by using single chymosin or an acid solution, and the casein recovery rate is more than 90%; compared with a starter combined with a chymosin precipitation method, the method saves the step of acidifying the starter, saves more time and has lower cost, and whey separated by the method contains higher content of active protein, so that the effect of enriching immunoglobulin and lactoferrin from raw fresh milk is better, the recovery rate of the immunoglobulin and lactoferrin can reach more than 92 percent, and the highest recovery rate of the protein enriched by the traditional membrane treatment method is about 85 percent; the chymosin precipitation method provided by the invention is combined with the acid precipitation method to have more advantages in the process of separating casein, provides favorable conditions for the subsequent step of enriching active protein, and the active components for enriching immunoglobulin and lactoferrin obtained by separation can be compounded with functional materials such as medicinal and edible raw materials, modified starch, oligosaccharide, probiotics and the like to develop and produce special diet food suitable for different crowds.
The specific implementation mode is as follows:
the present invention is further illustrated by the following examples, which do not limit the present invention in any way, and any modifications or changes that can be easily made by a person skilled in the art to the present invention will fall within the scope of the claims of the present invention without departing from the technical solution of the present invention.
Example 1
(1) Separating cream: 1000g of bovine colostrum is pasteurized and then rapidly cooled to 30 ℃, and is centrifuged by a milk fat separator at the rotating speed of 3000r/min to remove fat;
(2) Separating casein by combining a chymosin precipitation method with an acid precipitation method: adding 0.1g rennin into skim milk, reacting at 35 deg.C for 0.5 hr, adjusting pH of the emulsion to 4.8 with lactic acid solution, centrifuging the solution at 7000r/min to obtain casein precipitate and supernatant, which is whey protein, and recovering the supernatant;
(3) And (3) microfiltration sterilization: sterilizing the whey protein obtained in the step (2) by a 0.1-micron microfiltration membrane, and taking the permeate for later use;
(4) And (3) active protein enrichment by ultrafiltration: ultrafiltering the whey obtained in the step (3) by a 50kDa ultrafiltration membrane to 30% of the original volume, recovering trapped fluid, adding ultrapure water to dilute the whey to the original volume, and then carrying out ultrafiltration, repeating the process for three times, and freeze-drying the obtained sample solution for 24 hours to obtain freeze-dried powder enriched with the immunoglobulin and lactoferrin dual-active components;
(5) Comparative experiment: and (3) repeating the step (1) by taking 1000g of fresh bovine colostrum, changing the step (2) into adding 0.1g of rennin into skim milk, reacting for 0.5 hour at 35 ℃, centrifuging the solution at 7000r/min to obtain casein precipitate and supernatant, wherein the supernatant is whey protein, and recovering the supernatant for later use, and the following steps are the same as the steps (3) to (4), comparing the casein separating effect of the two methods and the recovery rate of the prepared medium immunoglobulin and lactoferrin enriched in the dual-active components of the immunoglobulin and the lactoferrin.
The experimental results are as follows:
under the condition of separating casein by adopting a chymosin precipitation method and an acid precipitation method, the recovery rate of the casein is 91 +/-0.1732%, the recovery rate of immunoglobulin in the enriched active ingredients is 90 +/-0.1011%, and the recovery rate of lactoferrin is 91 +/-0.1275%; the recovery rate of casein separated by using only rennin is 83 +/-0.1325%, the recovery rate of immunoglobulin in the enriched active ingredients is 80 +/-0.0523%, and the recovery rate of lactoferrin is 82 +/-0.1764%.
Example 2
(1) Separating cream: taking 2000g of sheep colostrum, pasteurizing, rapidly cooling to 32 ℃, and centrifuging at 5000r/min by using a milk fat separator to remove fat;
(2) Separating casein by combining a chymosin precipitation method with an acid precipitation method: adding 2g of rennin into skim milk, reacting for 0.8 hour at 36 ℃, adjusting the pH of the skim milk to 4.7 by using edible hydrochloric acid solution, centrifuging the solution at the rotating speed of 8000r/min to obtain casein precipitate and supernatant, wherein the supernatant is whey protein, and recovering the supernatant for later use;
(3) And (3) microfiltration sterilization: sterilizing the whey protein obtained in the step (2) by a 0.45-micrometer microfiltration membrane, and taking the permeate for later use;
(4) And (3) active protein enrichment by ultrafiltration: ultrafiltering the whey obtained in the step (3) by a 50kDa ultrafiltration membrane to 40% of the original volume, recovering trapped fluid, adding ultrapure water to dilute the whey to the original volume, and then carrying out ultrafiltration, repeating the process for three times, and freeze-drying the obtained sample solution for 36 hours to obtain freeze-dried powder enriched with the immunoglobulin and lactoferrin dual-active components;
(5) Comparative experiment: and (3) repeating the step (1) on 2000g of fresh sheep colostrum, changing the step (2) into the step of regulating the pH of the emulsion to 4.7 by using edible hydrochloric acid solution, centrifuging the solution at the rotating speed of 8000r/min to obtain casein precipitate and supernatant, wherein the supernatant is whey protein, and recovering the supernatant for later use, and comparing the casein separating effect of the two methods and the recovery rate of the prepared medium immunoglobulin and lactoferrin enriched with the dual-active components of immunoglobulin and lactoferrin with the same steps (3) - (4).
The experimental results are as follows: under the condition of separating casein by adopting a chymosin precipitation method and an acid precipitation method, the recovery rate of the casein is 92 +/-0.1652%, the recovery rate of immunoglobulin in the enriched active component is 90 +/-0.1720%, and the recovery rate of lactoferrin is 91 +/-0.1538%; the recovery rate of casein separated by using only rennet is 82 +/-0.1698%, the recovery rate of immunoglobulin in the enriched active ingredients is 80 +/-0.1592%, and the recovery rate of lactoferrin is 81 +/-0.0056%.
Example 3
(1) Separating cream: quickly cooling 3000g of bovine colostrum to 35 ℃ after pasteurization, and centrifuging at 7000r/min by using a milk fat separator to remove fat;
(2) Separating casein by combining a chymosin precipitation method with an acid precipitation method: adding 6g of rennin into skim milk, reacting for 1 hour at 37 ℃, adjusting the pH of the skim milk to 4.6 by using a tartaric acid solution, centrifuging the solution at the rotating speed of 10000r/min to obtain casein precipitate and supernatant, wherein the supernatant is whey protein, and recovering the supernatant for later use;
(3) And (3) microfiltration sterilization: sterilizing the whey protein obtained in the step (2) by a 0.1-micron microfiltration membrane, and taking the permeate for later use;
(4) And (3) active protein enrichment by ultrafiltration: ultrafiltering the whey obtained in the step (3) by a 50kDa ultrafiltration membrane to 50% of the original volume, recovering trapped fluid, adding ultrapure water to dilute the whey to the original volume, and then carrying out ultrafiltration, repeating the process for three times, and freeze-drying the obtained sample solution for 48 hours to obtain freeze-dried powder enriched with the immunoglobulin and lactoferrin dual-active components;
(5) Comparative experiment: and (3) repeating the step (1) by taking 3000g of fresh bovine colostrum, changing the step (2) into the step of regulating the pH of the emulsion to 4.6 by using a tartaric acid solution, centrifuging the solution at the rotation speed of 10000r/min to obtain casein precipitate and supernatant, wherein the supernatant is whey protein, and recovering the supernatant for later use, and comparing the casein separating effect of the two methods and the recovery rate of the prepared medium immunoglobulin and lactoferrin enriched with the dual-active components of the immunoglobulin and lactoferrin with the same steps (3) to (4).
The experimental results are as follows: under the condition of separating casein by adopting a chymosin precipitation method and an acid precipitation method, the recovery rate of the casein is 93 +/-0.0062%, the recovery rate of immunoglobulin in the enriched active ingredients is 91 +/-0.0532%, and the recovery rate of lactoferrin is 92 +/-0.1074%; the recovery rate of casein separated by using only rennin is 83 +/-0.1092%, the recovery rate of immunoglobulin in the enriched active ingredients is 82 +/-0.0002%, and the recovery rate of lactoferrin is 84 +/-0.0175%.
Claims (3)
1. A preparation method for enriching immunoglobulin and lactoferrin in colostrum is characterized by comprising the following steps:
(1) Separating butter fat: pasteurizing the primary emulsion, rapidly cooling to 30-35 deg.C, centrifuging at 3000-7000r/min with a milk fat separator, and removing fat;
(2) Separating casein by combining a chymosin precipitation method with an acid precipitation method: adding 0.01-0.2% (w/w) rennin into skim milk, reacting at 35-37 deg.C for 0.5-1 hr, adjusting pH of the emulsion to 4.6-4.8 with acid solution, centrifuging the solution at 7000-10000r/min to obtain casein precipitate and supernatant, which is whey protein, and recovering the supernatant;
(3) And (3) microfiltration sterilization: sterilizing the whey protein obtained in the step (2) by a microfiltration membrane with the diameter of 0.1 mu m or 0.45 mu m, and taking the permeate for later use;
(4) And (3) active protein enrichment by ultrafiltration: and (4) carrying out ultrafiltration on the whey obtained in the step (3) through a 50kDa ultrafiltration membrane to 30-50% of the original volume, recovering trapped fluid, adding ultrapure water to dilute the trapped fluid to the original volume, carrying out ultrafiltration again, repeating the process for three times, and carrying out freeze drying on the obtained sample solution for 24-48 hours to obtain the freeze-dried powder enriched with the double active components of the immunoglobulin and the lactoferrin.
2. The method of claim 1, wherein the chymosin of 2) is selected from the group consisting of chymosin of animal origin, chymosin of vegetable origin and chymosin of microbial origin.
3. The method of claim 1, wherein the acidic solution in 2) is selected from the group consisting of lactic acid, citric acid, acetic acid, tartaric acid and edible hydrochloric acid.
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1112394A (en) * | 1995-02-27 | 1995-11-29 | 湖北医科大学基础医学院科技开发部 | Preparation method for cow foremilk bio-active products |
| CN1629183A (en) * | 2004-09-03 | 2005-06-22 | 王秀英 | Active lactoprotein extracted from yak milk and its extraction method and use |
| CN1849903A (en) * | 2006-05-22 | 2006-10-25 | 江南大学 | Method for inductrialized separation of purified lactoferrins from cattle colostrum |
| CN101357941A (en) * | 2008-10-08 | 2009-02-04 | 青岛福仕奶业有限公司 | Heat resistant bovine colostrum immunoglobulin and preparation method |
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Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1112394A (en) * | 1995-02-27 | 1995-11-29 | 湖北医科大学基础医学院科技开发部 | Preparation method for cow foremilk bio-active products |
| CN1629183A (en) * | 2004-09-03 | 2005-06-22 | 王秀英 | Active lactoprotein extracted from yak milk and its extraction method and use |
| CN1849903A (en) * | 2006-05-22 | 2006-10-25 | 江南大学 | Method for inductrialized separation of purified lactoferrins from cattle colostrum |
| CN101357941A (en) * | 2008-10-08 | 2009-02-04 | 青岛福仕奶业有限公司 | Heat resistant bovine colostrum immunoglobulin and preparation method |
Non-Patent Citations (1)
| Title |
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| 马闯等.从牛初乳中提取酪蛋白、免疫球蛋白、乳白蛋白、乳铁蛋白生产工艺的研究.山东食品发酵.2004,(第undefined期),第6-7页. * |
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