CN101019580B - Preparation process of high content immune globulin whey powder - Google Patents
Preparation process of high content immune globulin whey powder Download PDFInfo
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- CN101019580B CN101019580B CN2007100202713A CN200710020271A CN101019580B CN 101019580 B CN101019580 B CN 101019580B CN 2007100202713 A CN2007100202713 A CN 2007100202713A CN 200710020271 A CN200710020271 A CN 200710020271A CN 101019580 B CN101019580 B CN 101019580B
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Abstract
The preparation process of high content immune globulin whey powder includes the following steps: improving the heat stability of cow colosrum, defatting, precipitating casein, isocratic elution for ultrafiltering and concentrating immune globulin from whey, measuring the separation degree of the immune globulin, vacuum freeze drying immune globulin to obtain milk powder, and packing. Test shows that the high content immune globulin whey powder has effective immune globulin content over 50 % and can raise immunity and prevent diseases effectively.
Description
Technical field
The present invention relates to immunology and milk production technical field, particularly relate to the preparation method of high content immune globulin whey powder.
Background technology
Immunoglobulin (Ig) (immunoglobulin, be called for short Ig) be the mankind and higher mammal be subjected to produce in the body after the antigenic stimulus can with the interactional class protein of antigentic specificity, be called antibody again, being prevalent in mammiferous blood, tissue fluid, lymph liquid and the exotocrine, is an important component part of body immune system.Immunoglobulin (Ig) is the class protein of homogeneous least in all proteins, according to its physicochemical property especially immunological properties, it can be divided into five classes, i.e. IgG, IgA, IgM, in IgD and the IgE, ox body based on IgG.
The commodity development of colostrum and functional component thereof can be traced to 1958, and Finland Immuno.Dynamic.Inc and other companies begin physiological function, consumption and the security of joint study colostrum and component thereof.Before penicillin and other antibiotic occur, the U.S. always with colostrum as a kind of disease-resistant food.Along with development of biology, many physiological functions (large space between muscles unit etc., 1987 of colostrum have been it is found that afterwards; C enturyLabsinev spa tent5013569).At United States Medicine circle, clearly proposing colostrum is " the recovery from illness agent of natural mystery ".
Colostrum nutrition is similar to human milk, and immune factor content is higher than people colostrum, immunoglobulin content be people's colostrum 50-100 doubly, except that antiviral and external cause a disease former, also can promote profitable strain growth and nutrition digestion to absorb, raising immunity.The crowd that colostrum is suitable for: after infant, pregnant and lying-in women, older, diabetic, cardiovascular patient, the operation (avenging 2002 in vain) such as fire victim, sub-health state crowd, the gangs of liking to be beautiful.
Since the nineties, China has also developed some successively and has strengthened the food of bovine colostrum immunoglobulin IgG, as the babies ' formula milk powder of the interpolation IgG of Wanda Mountain, Heilungkiang food factory, Beijing milk company utilized the pasturising milk with IgG activity of colostrum production " to come the good fortune breast " and colostrum powder in 1998.In the existing milk production, IgG content level differs greatly in the colostrum product, generally all between 3%-20%, and whether has activity and still is worth discussion.
This measuring commercially available skimmed milk powder, the content of IgG in the various milk powder such as dried sweetened milk, whey powder, but found that the content that can't measure the IgG in them with simple immunodiffusion method, promptly they all do not contain IgG or content to little.
Summary of the invention
The preparation method who the purpose of this invention is to provide a kind of immune globulin whey powder goods of high-load effectively improves the immunologic function of human body, prophylactic generation.
For achieving the above object, the technical solution adopted in the present invention is:
A kind of preparation method of high content immune globulin whey powder comprises following steps:
(1) chooses the raw material colostrum;
(2) in above-mentioned colostrum, add heat resistanceheat resistant sex change material;
(3) with above-mentioned colostrum degreasing, again the colostrum after the degreasing is precipitated, obtain whey liquid one time;
(4) an above-mentioned whey liquid is concentrated, obtain second whey liquid;
(5) above-mentioned second whey liquid is carried out drying, promptly get whey powder;
Concentrated employing isocratic elution ultrafiltration concentration method in the step (4) concentrates, and is about to a described whey liquid and carries out ultrafiltration earlier and concentrate, and carries out isocratic elution again;
Drying in the step (5) is through following operation, first primary drying, freezes, and the redrying that distils is again resolved under vacuum condition at last.
Colostrum in described step (1) is the healthy cow back 72 hours endocrine milk of giving a birth.
Described colostrum is at-15 ℃--and preserve in 25 ℃, add and take out man-hour and at room temperature melt.
Described heat resistanceheat resistant sex change material is sucrose, phosphate, a kind of or its mixture that makes up arbitrarily in the citrate.
Degreasing in step (3) is carried out on high speed freezing centrifuge, and the rotating speed of this centrifuge is 8000-12000 rev/min, and temperature is 2 ℃～5 ℃, and centrifugation time is 10 minutes-20 minutes.
Being precipitated as in step (3) precipitates the casein in the colostrum after the degreasing, this precipitation comprises following operation: after the pH value of the colostrum after the degreasing is adjusted to 5-6, on high speed freezing centrifuge, finish the precipitation operation again, the rotating speed of this centrifuge is 8000-12000 rev/min, temperature is 2 ℃-5 ℃, and centrifugation time is 15 minutes-25 minutes.
Ultrafiltration in step (4) concentrates and finishes on the ultrafiltration instrument, and its process was divided into for two steps, at first selected the 40-60kDa film to carry out ultrafiltration, with the 5-15kDa film filter liquor was held back then.
Also be provided with the step that detects the separation purity of immunoglobulin (Ig) in the described second whey liquid in step (4) and step (5).
The separation purity of described detection immunoglobulin (Ig) detects with the polyacrylamide gel electrophoresis technology.
During freezing in step (5), solidification point is-30 ℃---40 ℃, freeze-off time is 1.5～2.5 hours.
Lyophilization to second whey liquid in step (5) is to carry out in freeze drying equipment, described lyophilization comprises following operation, at first freeze drying equipment is cooled to and maintains-35 ℃--between 40 ℃, hothouse to equipment vacuumizes again, make vacuum reach 40-50Pa, heating sublimation 6-7 hour then.
Parsing in step (5) is to carry out in thermal analysis apparatus, and the resolution temperature of this thermal analysis apparatus is 35 ℃～45 ℃, and the parsing time is 6.5～7.5 hours.
The present invention compared with prior art has following advantage:
1. the content of immunoglobulin (Ig) is significantly higher than other milk powder in the dairy products of the present invention, and therefore the function of stronger raising immunity is arranged.
2. in the method for the present invention,, guaranteed the high-load and the high-purity of Immunoglobulin IgG in the dairy products with isocratic elution ultrafiltration concentration method separating immune globulin IgG.
3. of the present invention practical, simple to operate, step is clear, can adapt to the needs of large batch of production
For showing the increase significantly function of immunity of high content immune globulin whey powder goods with the inventive method preparation, carry out zoopery, process is as follows:
Get 27 of 615 inbred strais female mices (available from Chinese Medical Sciences University zoopery center).After weighing, get the mouse of body weight, be divided into 9 groups at random, 3 every group, make between each group body weight difference not remarkable at 21-24 gram.Be put at random then on 9 positions, extract 3 groups at random out, extract 3 groups afterwards more at random out, organize (III) in contrast, can make the environment indifference like this for remaining 3 groups as injection group (II) as drinking group (I).Drink every of group mouse every day and drink 1ml whey powder solution (0.2 gram whey powder, the content of Immunoglobulin IgG is about 40%), injection group mouse lumbar injection every day 0.1ml whey powder solution, 3:00 drank and injected IgG whey powder solution to mouse every afternoon.Experimental period is 30 days.Experimental session, mouse ad lib and drinking-water.
Test after 30 days, begin the immunity test experiments immediately, comprise: mouse peripheral T lymphocyte enzyme labeling decoration method (influencing the experimental technique of cellular immune function), chicken red blood cell is made immunogenic hemolysin (pluck eyeball and get blood) determination method, totally four tests of weighing of peritoneal macrophage functional examination method (influencing the experimental technique of humoral immune function), immune organ (spleen).
Interpretation is as follows,
(1) to the mouse cell Immune Effects
The results are shown in Table 19, drink IgG to mouse after, the shared percentage of T lymphocyte obviously improves, the poor heteropole of injection group and control group is (P≤0.01) significantly, illustrates that the IgG in the oral colostrum can obviously promote the mouse cell immunologic function.Injection group and control group comparing difference be extremely significantly (P≤0.01) also, illustrate that injecting IgG also can promote the mouse cell immunologic function.
Table 19 IgC is to the influence of the thin percentage composition of mouse T lymph
Table19effcct?of?IgG?on?the?percetage?centent?of?T?lyunphocyte?of?mice
(2) to mouse body fluid encourage can influence
The results are shown in Table 20, drink IgG to mouse after, OD value obviously improves, and illustrates that serum hemolysin obviously improves, the poor heteropole of injection group and control group is (P≤0.01) significantly, illustrates that the IgG in the oral liquid colostrum can obviously promote the mouse humoral immune function.Injection group and control group comparing difference be (P≤O.01) also extremely significantly, notes be described: serve as a contrast IgG and also can promote the mouse humoral immune function.
Table 20 IgG is to the influence of the disorderly growing amount of mouse haemolysis
Table?20?effect?of?IgG?on?hemgelysin?of?mlce
(3) to the influence of functions of murine peritoneal macrophages
The results are shown in Table 21, after drinking IgG to mouse, microscopy finds that the Turnover of Mouse Peritoneal Macrophages phagocytic rate obviously improves, the poor heteropole of injection group and control group is (P≤0.01) significantly, illustrates that IgG in the oral colostrum can effectively promote the phagocytic function of Turnover of Mouse Peritoneal Macrophages.Injection group and control group comparing difference be extremely significantly (P≤0.01) also, illustrate and injects the phagocytic function that IgG also can promote Turnover of Mouse Peritoneal Macrophages.
Table 21IgG is to the influence of Turnover of Mouse Peritoneal Macrophages phagocytic function
Table?21?effect?of?IgG?on?phaging?function?of?abdominal?in?mice
(4) to the influence of mouse immune organ (spleen) weight
The results are shown in Table 22, drinking and inject IgG to mouse all increases the weight of the immune organ of mouse (spleen), drink group and compare significant difference (P≤0.05) with control group, but the injection group is compared difference not significantly (P〉0.05) with control group
The influence that table 22IgG is heavy to mice spleen
Table?22?effcct?of?IgG?on?spleen?weight?of?mice
Annotate: the injection group has a dead mouse in the experimentation.
The analysis reason is as follows:
1, because mouse carries out once abdominal cavity injection every day, bigger to the mouse stimulation, may be stress be excessive and dead.
When 2, mouse was bought, because condition is limited, indoor temperature did not reach 20 degree.
3, may be improper in the operating process, cause dead mouse.
(5) brief summary
After this experiment is fed mouse with high content immune globulin IgG whey powder, its T lymphocyte, OD value (can promote serum to hold sanguinin), Turnover of Mouse Peritoneal Macrophages phagocytic rate all obviously increase than the mouse that does not feed the IgG whey powder under equivalent environment, and but the immune organ (spleen) of mouse is also had to be increased DeGrain.This has proved fully that also the IgG whey powder has growing of promotion mouse, increases immune function of mice.Experimental result shows that T lymphocyte percentage, optical density abdomen macrophage phagocytic rate, the spleen of experiment II group tested III raising more significantly.The main component IgG of IgG whey powder can strengthen immunization.Because of it all contains the effective active composition that strengthens immunity function.Experiment is compared with control group for two groups in this experiment, and T lymphocyte percentage, optical density, peritoneal macrophage phagocytic rate all have utmost point significant difference (p＜0.01), illustrates that this material has the effect of the immunity of organism of raising.Simultaneously can promote the splenocyte hyperplasia, perhaps this is one of approach that improves human body immune function.
Specific embodiment
A detailed process for preparing the immune globulin whey powder of high-load according to the inventive method is described in detail in detail below:
(1) healthy cow is given a birth back 72 hours endocrine fresh milk places-17 ℃ of refrigerators to preserve, and adds to take out man-hour at room temperature slowly to melt, to avoid the colostrum loss of biological activity.
(2) in colostrum, add heat resistanceheat resistant sex change material-natrium citricum, sodium phosphate, sucrose wherein, the molar concentration of natrium citricum is 200mmol/l, the molar concentration of sodium phosphate is 100mmol/l, and the mass content of sucrose is 15%, adds heat resistanceheat resistant sex change material to improve the heat endurance of colostrum;
(3) with above-mentioned colostrum degreasing, whole skimming processes adopts high speed freezing centrifuge, at 4 ℃, and under 10000 rev/mins of situations centrifugal 15 minutes.Centrifugal back finds that the blush sediment appears in lower floor, consider because the sampling time early, orlop should be blood red born of the same parents and macromolecular substances.Then the colostrum after the degreasing is processed with precipitation casein wherein, earlier its pH value is transferred to 4.5, the rotating speed of centrifuge is 10000 rev/mins, and temperature is 4 ℃, and centrifugation time is 20 minutes, and casein just can centrifugal get off, and obtains whey liquid one time;
(4) an above-mentioned whey liquid is carried out ultrafiltration and concentrate, finish on the ultrafiltration instrument, its process was divided into for two steps, at first select the 40-60kDa film that whey liquid is carried out ultrafiltration, with the 5-15kDa film filter liquor is held back then,, obtained second whey night again to the trapped fluid isocratic elution;
(5) with the separation purity of discontinuous vertical tabular polyacrylamide gel electrophoresis technical testing isocratic elution method ultrafiltration concentrated back second whey immunoglobulin (Ig) in night, after measured, immunoglobulin content is 50% in the emulsion;
(6) the Immunoglobulin IgG emulsion is packed into drying bottle after freezing 2 hours under-35 ℃ the temperature, is cooled to hutch water device and maintain again between-40 ℃, again hothouse is vacuumized, make vacuum reach 45Pa, heat supply then is for emulsion covers the heat of sublimation, after 7 hours, most of water can be removed with the form of ice crystal, and then 40 ℃ of following condition of high vacuum degree desorbs are after 7 hours, and discharging is sealed, finished product packing, warehouse-in.
Claims (9)
1. the preparation method of a high content immune globulin whey powder comprises following steps:
(1) chooses the raw material colostrum;
(2) add heat resistanceheat resistant sex change material in above-mentioned colostrum, described heat resistanceheat resistant sex change material is sucrose, phosphate, a kind of or its mixture that makes up arbitrarily in the citrate;
(3) will be through the colostrum degreasing after step (2) processing, again the colostrum after the degreasing is precipitated, obtain whey liquid one time, wherein, described being precipitated as precipitated the casein in the colostrum after the degreasing, and this precipitation comprises following operation: after the pH value of the colostrum after the degreasing is adjusted to 5～6, on high speed freezing centrifuge, finish the precipitation operation again, the rotating speed of this centrifuge is 8000～12000 rev/mins, and temperature is 2 ℃～5 ℃, and centrifugation time is 15 minutes～25 minutes;
(4) an above-mentioned whey liquid is concentrated, obtain second whey liquid;
(5) above-mentioned second whey liquid is carried out drying, promptly get whey powder;
It is characterized in that:
Concentrated employing isocratic elution ultrafiltration concentration method in the step (4) concentrates, and is about to a described whey liquid and carries out ultrafiltration earlier and concentrate, and carries out isocratic elution again; Wherein, described ultrafiltration concentrates and finishes on the ultrafiltration instrument, and its process was divided into for two steps, at first selected 40～60kDa film to carry out ultrafiltration, with 5～15kDa film filter liquor was held back then;
Drying in the step (5) is through following operation, first primary drying, freezes, and the redrying that distils is again resolved under vacuum condition at last.
2. the preparation method of a kind of high content immune globulin whey powder according to claim 1 is characterized in that: the colostrum in described step (1) is the healthy cow back 72 hours endocrine milk of giving a birth.
3. the preparation method of a kind of high content immune globulin whey powder according to claim 2, it is characterized in that: described colostrum is preserved in-15 ℃～-25 ℃, adds to take out man-hour and at room temperature melt.
4. the preparation method of a kind of high content immune globulin whey powder according to claim 1, it is characterized in that: the degreasing in step (3) is carried out on high speed freezing centrifuge, and centrifugation time is 15 minutes～20 minutes.
5. the preparation method of a kind of high content immune globulin whey powder according to claim 1 is characterized in that: also be provided with the step that detects the separation purity of immunoglobulin (Ig) in the described second whey liquid in step (4) and step (5).
6. the preparation method of a kind of high content immune globulin whey powder according to claim 5, it is characterized in that: the separation purity of described detection immunoglobulin (Ig) detects with the polyacrylamide gel electrophoresis technology.
7. the preparation method of a kind of high content immune globulin whey powder according to claim 1 is characterized in that: during freezing in step (5), solidification point is-30 ℃～-40 ℃, and freeze-off time is 1.5～2.5 hours.
8. the preparation method of a kind of high content immune globulin whey powder according to claim 1, it is characterized in that: the lyophilization to second whey liquid in step (5) is to carry out in freeze drying equipment, described lyophilization comprises following operation, at first freeze drying equipment is cooled to and maintains between-35 ℃～-40 ℃, hothouse to equipment vacuumizes again, make vacuum reach 40～50Pa, heating sublimation is 6～7 hours then.
9. the preparation method of a kind of high content immune globulin whey powder according to claim 1, it is characterized in that: the parsing in step (5) is to carry out in thermal analysis apparatus, the resolution temperature of this thermal analysis apparatus is 35 ℃～45 ℃, and the parsing time is 6.5～7.5 hours.
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| CN101283704B (en) * | 2008-05-23 | 2010-09-29 | 中国人民解放军第三军医大学 | Germfree mice artificial milk and its preparation method |
| ITRM20120412A1 (en) * | 2012-08-13 | 2014-02-14 | Biontologia S R L Lab | METHOD FOR THE PREPARATION OF A SERCERAL PROTEIN CONCENTRATE. |
| CN102919365B (en) * | 2012-11-08 | 2014-03-26 | 光明乳业股份有限公司 | Liquid dairy product rich in immune globulin and preparation method thereof |
| CN106390116A (en) * | 2016-08-19 | 2017-02-15 | 湖北华冠中科生物药业有限公司 | Bovine colostrum immunoglobulin G preparation for treating diarrhea and preparation method thereof |
| CN112244093A (en) * | 2020-10-20 | 2021-01-22 | 应荣科 | Modulated milk powder containing IgG, alpha-lactalbumin and lactoferrin and preparation method thereof |
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Non-Patent Citations (3)
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| 李长彪,刘长江,贺殿方,岳喜庆.超滤法分离牛初乳中免疫球蛋白IgG.食品工艺科技26 7.2005,26(7),116-117. * |
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