JPH09110896A - Purification of component 3 of cow milk proteose peptone - Google Patents

Abstract

PROBLEM TO BE SOLVED: To purify the subject component 3 useful as e.g. a surfactant, emulsifier, by concentrating and dialyzing whey through an ultrafiltration membrane impermeable to the component 3 of cow milk proteose peptone but permeable to the other whey proteins. SOLUTION: Whey is concentrated and dialyzed using an ultrafiltration membrane 50000-500000 in molecular weight cut off impermeable to the component 3 of cow milk proteose peptone but permeable to the other whey proteins, the resultant concentrate is adjusted to pH 3.0-5.0 and then either heat-treated at 70-100 deg.C for 10-60min or heat-treated at higher temperatures for a shorter time so as to cause a denaturation equivalent to the case with the former heat treatment of effect aggregation and elimination of the remaining whey proteins other than the component 3 of the cow milk proteose peptone, thus easily and highly purifying the objective component 3 of the cow milk proteose peptone, having high surface activity and useful as e.g. a surfactant, emulsifier, stabilizer for foods, cosmetics, medicines, etc.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to a method for purifying component 3 of milk proteose peptone, and more specifically to a fraction containing protein 3 as a main component from a fraction generally called proteose peptone in the protein contained in milk. The present invention relates to a method for economically purifying whey from whey with high purity. The component 3 fraction obtained by the present invention has a high surface activity,
Surfactants or emulsifiers for foods, cosmetics and pharmaceuticals,
It can be used as a stabilizer.

[0002]

2. Description of the Related Art Proteose peptone in milk is a component of milk protein that does not precipitate even if whey is heated at 95 ° C. to 100 ° C. for 20 minutes and then adjusted to pH 4.6, but is precipitated with 12% trichloroacetic acid. Is defined as (Thomp
son, MP, Tarassuk, NP, Jenness, R., Lillevik,
HA, Ashworth, USand Rose, D .: Journal of Da
iry Science 48, 159 (1965)). This proteose peptone does not have a uniform chemical composition and is mainly composed of multiple protein fractions called components 3, 5 and 8 (Larson, BL, and GR Roll).
eri .: Journal of Dairy Science 38, 351 (1955)). After that, further components 3, 5, 8-slow, 8-fast
It is recognized that there are four types of ingredients (Kolar, CW, a
nd Brunner, JR, Journal of Dairy Science 53, 99
7 (1970)). Due to such heterogeneous constituents, the term proteose peptone is nowadays commonly used as a fraction name rather than as a substance name for a single constituent.

Of these, component 3 contains many components that have the same antigenicity as the glycoproteins that make up the fat globule membrane, and Kanno names this as lactophorin (Kan).
no, C .: Journal of Dairy Science 72, 883 (1989)).
Recently, the present inventors succeeded in determining the sequence of cDNA encoding the major glycoprotein that constitutes lactophorin (Tsukasaki, F., Motoshima, H., Kuriki, H.,
Minagawa, E. and Kanno, C. 24th In ternational Da
iry Congress: Melbourne, Australia, 18th-22nd Sept
ember, 1994, Brief communications and abstracts of
posters and invited papers, page 190, Gb21p, 1994)
, The amino acid sequence deduced from that sequence was determined by another investigator to determine the primary structure of the major glycoprotein of component 3 (Sorensen, E. S and Petersen, TE Journal.
of Dairy Research 60, 535 (1993)). That is, although the definitions of proteose peptone component 3 and lactophorin are slightly different, they are terms that indicate virtually the same component as a substance, so in the present invention, component 3 and lactophorin mean the same component. Used as.

The method for purifying Component 3 from the proteose peptone fraction of the present invention is treated as synonymous with the method for purifying lactophorin. We have already found that this lactophorin fraction has an extremely high surface activity and is highly useful as a natural surface-active agent (JP-A-4-1).
04830). Conventional fractionation methods for component 3 are described in the literature (eg Ng, WC, Brunner, JR And Rh.
ee KC Journal of Dairy Science 53,987 (1969)). However, the conventional method for purifying the fraction containing the component 3 as the main component from skim milk or whey requires a complicated process such as ammonium sulfate fractionation and is economically disadvantageous. There was a problem in using it as an emulsifier. Further, if a complicated purification method is used, impurities may be mixed in during the purification process, and there is a demand for the development of a simpler purification method for the component 3. The purification method by membrane treatment as in the present invention has not been reported yet. The reason is that the molecular weight of the major constituent protein of component 3 is about 27,000 under denaturation.
This is because it was thought that the fractionation was practically difficult because it was composed of glycoprotein having a molecular weight of 17,000 and 17,000, and there was not much difference in size from other mixed α-lactalbumin and β-lactoglobulin.

[0005]

The method for efficiently purifying component 3 or lactophorin has not been reported so far. It is an object of the present invention to develop a method for purifying a fraction particularly rich in component 3 from whey without using methods such as ammonium sulfate precipitation and column chromatography which have been conventionally reported. The whey used as a raw material in the present invention is not limited, and examples thereof include not only acidic whey and neutral whey, but also whey powder that is a powdered whey, whey protein concentrate (WP).
C), whey protein isolate (WPI) and the like can also be used. These are all commercially available and can be easily obtained. All of these contain the same three components as whey, and can be used as a raw material like whey if appropriately reduced with water or the like. Therefore, in this specification, "whey" is meant to include these unless otherwise specified.

[0006]

[Means for Solving the Problems] Even if milk or whey is heated in the range of pH 4 to 5 (about 90 ° C.) and the aggregates are removed by centrifugation, a fraction containing only proteose peptone is obtained. However, only the component 3 cannot be purified. However, as a result of diligent efforts, the present inventors simply concentrated whey after concentration with an ultrafiltration concentrator (a protein content of 0.05% (weight / volume) or more),
It was found that by heating at around 90 ° C., most of the proteins other than the component 3 were aggregated and removed, and the component 3 remained as the main component in the supernatant and could be purified. The present invention has been completed based on such findings.

That is, the present invention according to claim 1 concentrates and dialyzes whey by using an ultrafiltration membrane having a cut-off molecular weight that does not permeate the component 3 of milk proteose peptone but permeates other whey proteins. A method for purifying component 3 of milk proteose peptone, characterized in that the present invention according to claim 3 has a molecular weight cut-off of 1
After whey is concentrated and dialyzed using an ultrafiltration membrane of 10,000 to 1,000,000, the concentrated solution is heated at a temperature of 70 to 100 ° C for 10 minutes.
Milk proteose characterized by aggregating and removing whey proteins other than component 3 of milk proteose peptone by carrying out heat treatment at a higher temperature for a short period of time to cause heat denaturation for 60 minutes or equivalent. This is a purification method for peptone component 3.

[0008]

BEST MODE FOR CARRYING OUT THE INVENTION The present invention is a method for efficiently purifying milk proteose peptone component 3 from whey. Whey is obtained as a by-product of cheese manufacturing, casein manufacturing, etc., but whey (in a narrow sense)
There are two types of whey, acidic whey and sweet whey (or neutral whey). Acidic whey is produced mainly by the production of quark, cottage cheese, etc. and the production of casein, and is characterized by a low whey pH (usually pH 4-5). On the other hand, sweet whey is produced by the production of Gouda cheese, cheddar cheese, Camembert cheese, etc. and is characterized by a high whey pH (usually around pH 6). Any of these narrowly defined whey can be used in the method for purifying the component 3 of the present invention, and the above-mentioned whey powder, WPC, WPI and the like can also be used, and two or more of these can be used in combination. Is.

An ultrafiltration membrane is used for the concentration of whey. If a membrane having a molecular weight cut-off of 10,000 or more is used, the lactose and low molecular weight substances present in the whey can be effectively treated by performing membrane dialysis. High purity component 3
Fractions can be obtained. The upper limit of the molecular weight cut-off is 1,000,000. When the molecular weight cutoff is higher than that, the component 3 also permeates, and concentration becomes difficult. Furthermore, by carrying out membrane dialysis using a membrane having a cut-off molecular weight of 50,000 or more, preferably 100,000 to 500,000, casein degradation products other than component 3 and major α-lactalbumin, β-lactoglobulin, etc. The coexisting protein component can be effectively removed, and the fraction mainly containing the component 3 can be accumulated on the concentrated liquid side. In this case, the heat treatment described below is not always necessary. When membrane dialysis is performed using a membrane with a molecular weight cutoff of 10,000 or more and 1,000,000 or less,
By heating the obtained concentrated solution, protein components other than the component 3 can be aggregated and removed. As a result, it is possible to refine the component 3 more effectively. The heating temperature is 70 ° C or higher, usually 70 to 100 ° C. The heat treatment time is usually 10
It is suitable that the heating temperature is 80 ° C. for 20 minutes or more, and 90 ° C. for 10 minutes or more. However, since the purpose of this heat treatment is to heat-denature and agglomerate the protein, even heat treatment at a higher temperature for a shorter time, for example, heat treatment at 121 ° C. for about several seconds causes heat denaturation equivalent to this. Good. Examples of this method include a heating method using a plate heat exchanger. In addition, it is desirable to adjust the pH to 3 to 10, preferably 3.0 or more and less than 5.0 during the heat treatment. The same result can be obtained by performing the pH adjustment after heating and then removing the aggregates by centrifugation or the like. Further, concentration is carried out so that the protein concentration of the concentrated solution becomes 0.05 (weight / volume) or more.

The main component of Component 3 has a molecular weight of about 30,000 under denaturation and a molecular weight of 1 on gel filtration.
It has been revealed that it is about 30,000. However, it is considered that the whey has a molecular weight of 300,000 or more in the existing form in whey. Actually, the molecular weight cutoff is 50
With 3 million or 1 million membranes, component 3
Is accumulated on the concentrate side.

[0011]

Next, the present invention will be described by way of examples, which should not be construed as limiting the present invention. Example 1 <Identification of Component 3> Acidic whey produced in cheese production, for example, quark production, was concentrated using a membrane having a molecular weight cut off of 50,000, and the concentrate was subjected to polyacrylamide gel electrophoresis (PAGE) under non-denaturing conditions. Then, the presence of Component 3 was confirmed by staining with Coomassie Brilliant Blue R250. The gel concentration was 12.5%. FIG. 1 shows the electrophoretic photograph. It should be noted that electrophoresis was performed using literature (Davis, BJ Ann. NY Acad. Sci. 121, 404).
(1964)). The target component 3 can be easily confirmed at a position with relatively low mobility by this migration method. This fraction is named lactophorin by Kanno, as described above. The polyacrylamide gel electrophoresis of the present invention was carried out under these conditions.

Example 2 <Selective Concentration of Component 3 by Ultrafiltration> An example of concentration of Component 3 by ultrafiltration from acidic whey produced in quark production will be shown. This acidic whey has a pH
It was 4.23. This is further ion-exchange resin method p
A pH-adjusted whey in which H was replaced with 8.30 was prepared. The results of concentrating these two types of pH whey with a flat membrane type ultrafiltration device are shown in FIG. The experimental apparatus was a UF-LMS type UF module manufactured by Toyo Soda Co., Ltd., and a membrane (UF-300PS) having a molecular weight cutoff of 300,000 was used. 200 ml of each sample was passed through at a flow rate of 2.5 ml / min to obtain 40 ml of concentrated liquid and 160 ml of permeated liquid.

The resulting permeate was freeze-dried. After adding 60 ml of water, the concentrate was concentrated to 40 ml and freeze-dried. The obtained freeze-dried products were compared by electrophoresis (Fig. 2). As a result, it was clarified that the component 3 was concentrated in the concentrated liquid and the band of the component 3 was not seen on the permeated liquid side in both whey without pH adjustment and whey after pH adjustment. In addition, α having a high mobility other than that of the component 3
It was revealed that whey proteins such as lactalbumin and β-lactoglobulin are present in both the concentrate and the permeate. From these results, when whey is concentrated with an ultrafiltration membrane, α-lactalbumin and β-lactoglobulin can be permeated, and component 3 cannot be permeated. It is easy to understand that it can be concentrated to In FIG. 2, lane 1 is an undiluted acidic whey solution (pH 4.2), lane 2 is a concentrated solution obtained by concentrating the acidic whey with an ultrafiltration device, and lane 3 is an ultrafiltration of the acidic whey. Permeate when concentrated with a filtration device, lane 4 is acidic whey (pH 8.3) whose pH is adjusted by an ion exchange method, lane 5 is a concentrated liquid when the pH adjusted acidic whey is concentrated with an ultrafiltration device, Lane 6 shows the permeate when the pH-adjusted acidic whey was concentrated by an ultrafiltration device. It can be seen that the band of component 3 is not seen in the permeate of lanes 3 and 6.

Example 3 <Effect of heating on recovery of component 3> As shown in Example 2, it was revealed that the component 3 can be efficiently and selectively concentrated by ultrafiltration.
However, when proteins other than component 3 containing α-lactalbumin and β-lactoglobulin as the main components are slightly concentrated or not concentrated particularly when membrane dialysis is performed using a membrane having a small cutoff molecular weight. However, it became an equilibrium state, and it was found that they were mixed in the concentrated liquid.
Therefore, in order to further enhance the purity of the component 3, after concentrating, heat treatment is performed at 90 ° C. for 30 minutes to denature the components other than the mixed component 3 and aggregate them, and then separate them by centrifugation. Then, it was examined to remove the components other than the component 3.

As a result, the molecular weight cutoff is 100,000 or 50.
After concentrating the acidic whey using a membrane,
When heated at 0 ° C. for 30 minutes and centrifuged, it was revealed that the fraction other than the component 3 can be efficiently removed by the heat treatment regardless of which membrane is used. The results obtained by electrophoresis are shown in FIG. In FIG. 3, in Lane 1, acidic whey was concentrated with a membrane having a molecular weight cutoff of 500,000 and then heated,
Supernatant after centrifugation, lane 2 is a non-heated sample similar to lane 1, lane 3 is acidic whey concentrated with a membrane having a molecular weight cutoff of 100,000 and not heated, lane 4 is The same sample as in lane 3 is heated and centrifuged to show the supernatant. It is apparent that the bands other than the component 3 are reduced by performing centrifugation after heating by these electrophoresis.

Example 4 <Effect of molecular weight cut-off of ultrafiltration membrane on recovery of component 3> As described above, the component 3 had a molecular weight of 27,000 and 17 when subjected to electrophoresis under denaturation.
It is known that 000 relatively small molecules are the main constituents, and that the gel filtration has a molecular weight of about 130,000, but it is considered that the larger molecular weight exists in the natural state of whey. , Molecular weight cutoff of 10,000 or more 1
When concentrated using ultrafiltration membranes up to 0,000,000,
It was found that component 3 accumulates on the concentrate side. In other words, since it was previously thought that the molecular weight of component 3 was relatively close to that of casein, α-lactalbumin, β-lactoglobulin, etc., it was thought that efficient fractionation with a membrane was impossible. In addition, as shown in the present Example, it was revealed that even in a membrane having a very large molecular weight cutoff of 50,000 to 500,000, Component 3 could be concentrated without permeating the membrane.

FIG. 4 shows a molecular weight cutoff of 10,000, 50,000, 100,000,
Heat treatment (at 90 to 95 ° C. for 10 to 30 minutes) obtained by concentrating using 500,000 or 1 million ultrafiltration membranes,
The result of analyzing the supernatant obtained by centrifugation by electrophoresis is shown. In the figure, lane 1 was concentrated on a membrane having a molecular weight cut-off of 10,000, then heated and centrifuged to obtain a supernatant, which was subjected to polyacrylamide gel electrophoresis. As a result, lane 2 was a membrane having a molecular weight cut-off of 50,000. As a result of polyacrylamide gel electrophoresis of the supernatant obtained by concentrating, heating and centrifuging, lane 3 was concentrated with a membrane having a molecular weight cut-off of 100,000.
As a result of polyacrylamide gel electrophoresis of the supernatant obtained by heating and centrifuging, lane 4 was concentrated with a membrane having a molecular weight cut-off of 500,000 and then heated and centrifuged to obtain a polyacrylamide gel of the supernatant. As a result of acrylamide gel electrophoresis, lane 5 shows the results of polyacrylamide gel electrophoresis of the supernatant obtained by concentrating with a membrane having a molecular weight cut-off of 1,000,000, followed by heating and centrifugation. As is clear from this result, a membrane having a molecular weight cut-off of 10,000 to 500,000 could be concentrated on the concentrate side, and the component 3 did not appear on the permeate side. However, in the membrane having a cut-off molecular weight of 1,000,000, the component 3 leaked to the permeate side but was concentrated to some extent. Therefore, in order to efficiently concentrate and purify the component 3 by the ultrafiltration method, if a membrane having a cut-off molecular weight that the component 3 cannot pass through (for example, a membrane of 50,000 to 500,000) is used, the efficiency can be increased by membrane dialysis. It is possible to reduce the amount of contaminants, and it is economical because the clogging of the membrane hardly occurs.

Example 5 <Effect of protein concentration during heating> Even if the acidic whey produced in quark production is heated before concentration, no aggregates or precipitates are formed, and α-lactalbumin is also obtained by centrifugation. And β
Almost no lactoglobulin can be removed. Therefore, in order to efficiently purify the component 3 by the ultrafiltration method, it is necessary to perform heat treatment after concentration by membrane dialysis. At this time, it was examined how much protein concentration should be used for heat treatment to remove components other than the component 3 component. The acidic whey concentrate obtained using a membrane with a molecular weight cut off of 500,000 was lyophilized. The protein concentration is 0.05, 0.1, 0.3, 0.5, 0.7, 1.0,
It was prepared by reducing with water to 2.0% (W / V).
The protein content was determined by the dye method using bovine serum albumin as a standard protein. Heat treatment of the concentrated solutions prepared to these concentrations at 95 ° C. for 20 minutes, and then
After centrifugation at 5000 rpm, the supernatant was analyzed by electrophoresis so that the amount of protein in each lane was almost the same. Results are shown in FIG. The protein concentration of the sample in each lane is as follows. Lane 1: concentrated solution before heating (control), Lane 2: 0.0
5% (W / V), Lane 3: 0.1% (W / V), Lane 4: 0.3% (W / V), Lane 5: 0.5% (W /
V), lane 6: 0.7% (W / V), lane 7: 1.
0% (W / V), lane 8: 2.0% (W / V) As is clear from the figure, when the concentrated solution having a protein concentration of 0.05% or more was heat-treated, β-lactoglobulin It turns out that it can be removed. It was also found that α-lactalbumin can be removed at a protein concentration of 0.1% or more.

Example 6 <Influence of pH of concentrate> In order to effectively remove components other than the components of component 3 to efficiently purify component 3, how to adjust the pH of the concentrate may be adjusted. To investigate this, the effect of pH of the concentrated solution was examined. That is, the acidic whey concentrate obtained using a membrane having a molecular weight cutoff of 500,000 was freeze-dried. The acidic whey concentrate became alkaline (pH 8.3) upon continued membrane dialysis against water. This was prepared with various buffer solutions so as to have a protein concentration of 0.5% (W / V), and these concentrated solutions were heat-treated by boiling (95 to 100 ° C.) for 20 minutes to give 15,000.
After centrifugation at rpm, the supernatant was adjusted to the same protein amount and then analyzed by electrophoresis. FIG. 6 shows the results. In the figure, the pH of the sample in each lane is as follows. Lane 1: pH 3, Lane 2: pH 4, Lane 3: pH
5, lane 4: pH 6, lane 5: pH 7, lane 6:
pH 8, lane 7: pH 9, lane 8: pH 10 As is clear from the figure, α-lactalbumin and β-lactoglobulin are considered to be removable at any pH value. Furthermore, upon closer observation, it can be seen that a high molecular weight glycoprotein having a mobility lower than that of the component 3 can be removed by heat treatment under acidic conditions. The pH of the supernatant after centrifugation is 3.0 or more and 5.0.
When it was less than 1, the transparency was high, but it is considered that when heat treatment was performed on the alkaline side, the supernatant became cloudy and the amount of denatured protein increased. Therefore, the pH during heating is 3.0
It is preferably not less than 5.0 and less than 5.0. The same result can be obtained by adjusting the pH after heating and then removing the aggregates by centrifugation or the like.

Example 7 <Influence of heating temperature> In order to efficiently purify the component 3, what temperature should be used for the heat treatment was examined. Therefore, the acidic whey concentrate obtained using a membrane having a molecular weight cutoff of 500,000 was freeze-dried. This was prepared to have a protein concentration of 0.5% (W / V) (HCl
The pH was adjusted to 4.0 with. ) Then add the concentrate to 4
After heat treatment at 0 ° C, 50 ° C, 60 ° C, 70 ° C, 80 ° C, 90 ° C or boiling (95 to 100 ° C) for 20 minutes, centrifuge at 15000 ppm, and then use the supernatant as the same amount of protein. After adjusting so as to be, it was analyzed by electrophoresis. FIG. 7 shows the result. As a control, the one that was not heat-treated was used. In the figure, the heating temperature of the sample in each lane is as follows. Lane 1: concentrated solution before heating (control): Lane 2:40
° C, lane 3:50 ° C, lane 4:60 ° C, lane 5:
70 ° C., lane 6: 80 ° C., lane 7: 90 ° C., lane 8: boiling (95 to 100 ° C.) As a result, it was considered that considerable removal was possible by heat treatment at 90 ° C. for 20 minutes. . However, it became clear that if the heat treatment temperature was 95 to 100 ° C., the components other than the component 3 could be removed more efficiently.

Example 8 <Influence of heating time> In order to efficiently purify the component 3, how long the heating treatment should be conducted was examined. The same concentrated liquid as in Example 7 was boiled (95 to 10).
Heated at 0 ° C.) for 1 to 60 minutes. Then 15,000
After centrifugation at rpm, the resulting supernatant was analyzed by electrophoresis so that the amount of protein in each lane was the same. The result is shown in FIG. The heating time of the samples in each lane in boiling (95 to 100 ° C.) was as follows: lane 1: unheated (0 minutes), lane 2: 1 minute, lane 3: 5 minutes, lane 4: 10 minutes, lane 5: 15 minutes. , Lane 6: 2
0 minutes, lane 7:25 minutes, lane 8:30 minutes, lane 9:40 minutes, lane 10:50 minutes, lane 1
1: 60 minutes. As is clear from the figure, 95-1
It can be seen that the component 3 can be efficiently purified by heating at 00 ° C. for 5 minutes or more. Judging this result and the result of Example 7 together, the heat treatment at 90 ° C. for about 10 minutes is appropriate. It should be noted that, of course, this heat treatment is for denaturing the protein by heat, so that denaturation equivalent to this can be induced. It is clear that the results are obtained.

Example 9 <Production of Component 3 Fraction from Acidic Whey> Based on the results of each of the above Examples, Component 3 was actually purified from whey. That is, 270 kg of acidic whey (pH 4.3) produced in quark production is heated to 50 to 55 ° C.
Warm it to an ultrafilter (α-Laval UFR1 /
2, follow fiber PM100, molecular weight cutoff 10
10,000), and membrane dialysis with water was performed to concentrate to a total amount of 36 kg. Next, reverse osmosis (R)
O) (DDS, HR95) to 5 kg and then heat-treated for 30 minutes by boiling (95-100 ° C) for 3000
The precipitate was removed by centrifugation at rpm for 15 minutes. The pH of the concentrated liquid during the heat treatment was adjusted to 8.3. The resulting supernatant is freeze-dried to give 64 component 3 fractions.
g was obtained.

Example 10 <Production of highly transparent component 3 fraction from acidic whey> Since the component 3 obtained in Example 9 becomes cloudy when dissolved in water, in this example, the highly transparent component 3 A production example of is shown. Acid whey 363k
g was heated to 50 to 55 ° C., concentrated with an ultrafiltration device (α-Laval UFR1 / 2, follow fiber PM500, molecular weight cutoff of 500,000), and subjected to membrane dialysis with water to give a total amount of 2
It was concentrated to 2 kg. Then reverse osmosis membrane (RO) (DD
After concentrating to 5 kg with S, HR95), the pH was adjusted to 4.0 using 6N hydrochloric acid with stirring. Then, heat treatment by boiling (95 to 100 ° C) for 30 minutes, 3000
The precipitate was removed by centrifugation at rpm for 15 minutes. The resulting supernatant is lyophilized to obtain highly transparent component 3
54 g of fractions were obtained.

Example 11 <Analysis of Component 3 Fraction Obtained in Examples 9 and 10> The result of polyacrylamide electrophoresis analysis of the Component 3 fraction produced in Examples 9 and 10 is shown in FIG. It was In the figure, lane 1 is the component 3 fraction obtained in Example 9, and lane 2 is the component 3 fraction obtained in Example 10.

Example 12 <Example of Purification of Component 3 from Neutral Whey> In the above examples, only the example of purification of Component 3 from acidic whey was shown, but in this example, the same method is used from neutral whey. Shows that component 3 can be purified. FIG. 10 shows the analysis results of polyacrylamide gel electrophoresis of component 3 fractions obtained by treating with neutral whey in the same manner as in Example 9. As is clear from the figure,
The purity of the obtained three component fractions was almost the same as that from the neutral whey and that from the acidic whey.

[0026]

According to the present invention, the fraction containing the component 3 in the whey as a main component is used with almost no reagent.
A practical method for highly purifying only by physical methods is provided. Component 3, namely lactoforin, has already been found to be very emulsifying,
The present invention makes it possible for the first time to produce it economically.

[Brief description of the drawings]

FIG. 1 is an electrophoretic photograph showing the results of Example 1.

FIG. 2 is an electrophoretic photograph showing the results of Example 2.

FIG. 3 is an electrophoretic photograph showing the results of Example 3.

FIG. 4 is an electrophoretic photograph showing the results of Example 4.

5 is an electrophoretic photograph showing the results of Example 5. FIG.

FIG. 6 is an electrophoretic photograph showing the results of Example 6.

FIG. 7 is an electrophoretic photograph showing the results of Example 7.

FIG. 8 is an electrophoretic photograph showing the results of Example 8.

9 is an electrophoretic photograph showing the result of Example 11. FIG.

FIG. 10 is an electrophoretic photograph showing the results of Example 12.

 ─────────────────────────────────────────────────── ─── Continuation of the front page (72) Inventor, Choemon Sugano 1023-2 Shimohiraide-cho, Utsunomiya-shi, Tochigi Prefecture (72) Inventor Fuji Shijo, 465-1 Waatsu, Hiroshima-cho, Sapporo-gun, Hokkaido Within Yotsuba Dairy Co., Ltd. Research Center

Claims (4)

[Claims] 1. A milk proteose peptone characterized in that whey is concentrated and dialyzed using an ultrafiltration membrane having a cut-off molecular weight that does not permeate component 3 of milk proteose peptone but permeates other whey proteins. Purification of component 3 of. 2. The purification method according to claim 1, wherein the ultrafiltration membrane has a molecular weight cutoff of 50,000 or more and 500,000 or less. 3. The whey is dialyzed using an ultrafiltration membrane having a molecular weight cut-off of 10,000 or more and 1,000,000 or less, and then the concentrated solution is added to 70-1.
Heat treatment at a temperature of 00 ° C. for 10 to 60 minutes, or heat treatment at a higher temperature for a short time to cause heat denaturation equivalent to this, to aggregate whey protein other than component 3 of milk proteose peptone, A method for purifying Component 3 of milk proteose peptone, characterized in that it is removed. 4. The purification method according to claim 3, wherein the whey protein other than the component 3 is aggregated after the pH is adjusted to 3.0 or more and less than 5.0 before or after the heat treatment.