JPH0494643A - Riboflavin enriched cheese and production thereof - Google Patents

Abstract

PURPOSE:To obtain the title cheese having enriched vitamin B2 and high nutritive value by using an emulsion prepared by dissolving a riboflavin in milk fat and adding an emulsifiable substance to the solution. CONSTITUTION:A riboflavin (preferably riboflavin tetrabutyrate) is dissolved in milk fat and an emulsifiable substance (preferably spherical film of milk fat) is added to the solution to form an emulsion, which is used to give the objective cheese.

Description

DETAILED DESCRIPTION OF THE INVENTION (Industrial Field of Application) The present invention relates to vitamin-enriched cheese, and more particularly to cheese enriched with riboflavins. The present invention also relates to an industrial manufacturing method for efficiently manufacturing this cheese.

(Prior Art) Riboflavin is a water-soluble vitamin also called vitamin B2, and as is clear from its other name, lactoflavin, it is contained in large amounts in milk.

However, the riboflavin (hereinafter also referred to as RF) content in cheese prepared from milk is very low, and although cheese is a balanced nutritional food, it lacks RF, which is a growth promoting factor.

In other words, approximately 80% of the total RF in milk is not trapped in the cheese curd and is eliminated in the cheese whey, and this is because most of the RF in milk is present in free form. .

As described above, in the prior art, the RF content in cheese is very low compared to milk, and there are no known examples of successful industrial production of cheese with a high RF content. Furthermore, there are no known examples of success in aggressively fortifying cheese with RF.

(Problems to be Solved by the Invention) In view of the above-mentioned state of the art, an object of the present invention is to newly produce cheese with an increased RF content.

(Means for Solving the Problems) The present invention was made to achieve the above object, and as a result of examination from various aspects, although various improvements were made to the cheese manufacturing process, RF was excluded from whey. However, they were unable to control the pests, so they changed their thinking and focused on actively strengthening cheese using RF.

Therefore, we conducted extensive research and focused on the existence of RF bound to proteins in milk.
Milk Fat Globule Membrane
, hereinafter also referred to as MFGM) to strengthen 'RF,
We decided to strengthen the cheese with RF.

Most of the flavins bound to MFGH are FAD (f
lavin adenine dinucleotide
e). However, MFGM has RF, FMN (fl
avin mononucleotide), FAD
was not bound by addition of either of these, and by equilibrium dialysis, 98
% has been removed. Furthermore, even when RF, FMN, or FAD was added to MFGM from which flavin had been removed in advance, no amount greater than the initial binding amount was bound. This indicates that the flavin binding ability of MFGM has reached saturation.

Therefore, instead of RF, riboflavin tetrabutyrate (RTB), which is lipophilic and contains butyric acid, which is approved as a food additive in Japan and is important for the flavor of cheese, is used.
) was selected.

First, by dissolving RTB in milk fat and preparing an emulsion with MFGM, RTB is added to milk fat shields.
succeeded in incorporating.

The present invention was completed based on the above new findings.

The present invention will be explained in detail below.

(1) Buttermilk fraction containing MFGM (477 μg/g P
The bound flavins of protein (rotein) are NaCQ and KCQ.
salt, ammonium sulfate, Triton X-100, Tween
20, was rarely dissociated by nonionic surfactants such as Non1det P-40, as well as relatively mild anionic surfactants such as deoxycholic acid and cholate. In contrast, strong surfactants such as urea, guanidine hydrochloride, and sodium dodecyl sulfate (SDS) caused 44%, 98.6%, and 9%, respectively.
7.5% of flavins were dissociated (Table 1).

Table 1. Survival rate of bound flavin by various treatments aCQ KCQ raa Guanidine-HCQ Triton X-100
cholate Sodj,um dodecyl 5ulfate10
2.0 97.0 55.9 1.4 91.1 92.1 91.5 82.0 105.0 2.5 63.6 71.4 51.4 26.4 62.2 68.7 60. 5 46.4 62.1 28.5 83.7 100.0 102.4 13.3 74.1 79.2 74.1 Separate, 3 96.2 75.6 Methanol/chloroform2:1. (
v/ν) 14.2 (2) Relationship between pH of milk fat globule membrane, casein, and whey fractions and dissociation of bound flavins Buttermilk fraction containing MFGM (349,3p g/
Flavin in g protein) has a pH of 4.0.
-10.0, there was almost no dissociation, but at pH 3,
5 and pH 3.0, 97% of flavins were dissociated. M
Naturally, even at pH 4.9, which is the isoelectric point of FGM, it was hardly dissociated.

As is clear from the above results, especially the results in Table 1 (1), the binding between flavin and protein in MFGM, casein, and whey fractions is not necessarily the same, but is basically hydrophobic, and in particular, that of MFGM is It has become clear that this is due to relatively strong hydrophobic bonds. In addition, as is clear from (2), MFGH-bound flavin is present at pH 4,
It was hardly affected by pH'H up to 0, but that of casein and whey fractions was affected by pH and was liberated. These things also apply to MFG when forming cheese curds.
This suggests that the flavin of M is less likely to be released.

(3) Preparation of milk fat globule film After separating fresh milk, place it on a cream separator for 37 minutes.
Washed three times with distilled water at 0.degree. C., the washed cream is cooled in a water bath and immediately churned in a metal churn. MFGM is collected by centrifugation at 4° C., 186,000×g for 40 minutes.

MFGM products containing 0.02% sodium azide are N
2. Store in a water bath in an atmosphere and use within one month.

Prior to use, MFGM was 0. It is used after being dispersed in OIM's sodium phosphate buffer (PH7,0).

The contents of solids, fat, and protein in the MFGM dispersion are as follows:
They were 6.25%, 44%, and 52%, respectively.

(4) Reconstruction of milk fat bulges Unless otherwise stated, 1.25 μg of riboflavin tetrabutyrate (RTB, Wako Pure Chemical Industries, Ltd.) was dissolved per 1 g of milk fat.

This milk fat 2.5g and 0. OIM phosphate buffer (pi(
From 80 mg of MFGM dispersed in
By the method of no (J, Food Sci, 54
．． 1534.1989), an emulsion was prepared. In addition, 1.25 g of RTB was dissolved per 1 g of milk fat, and 1 g of this milk fat was mixed with 0.01 M phosphate buffer (pH 7.0).
The emulsion prepared from 80mg of MFGM dispersed in
It was measured with Further, the subnatant fraction was subjected to isoelectric precipitation at pH 4.8, and the RTB content of the precipitated and non-precipitated fractions was measured.

The results are shown in Table 2.

A Crαl 858.0±39.4 76.3±2.213.2±2
．． 0 38.3±2.8 Subnatant Precipitate 179.3±8.9 15.9±0.587.4±21
．． 3 7.8±1.9 0±2.4 41.6±0.9 1.0±0.3 20.1±2.0 at pH4,8uppernatant 43.9±7.0 24
．． 5±3.4at pH 4, 8 bottle ratio 91.8±0.5 14.4±2.3 bottles The amount of protein used was 30.24+ag.

** RT of precipitate and supernatant at pH 4,8
The ratio of B is calculated as 5 ubnatant 100.

As is clear from the above results, insoluble orange-red RTB was also observed in the centrifuged precipitate fraction along with a small amount of protein, but it accounted for only 8% of the total. Approximately 7 of RTB
6% was recovered in the cream fraction and 72% of RTB in the subnatal fraction was precipitated at pH 4.8. Therefore, it was revealed that about 90% of RTB was bound to MFGM in addition to the cream fraction and minute fat globules.

Based on these facts, RTB is dissolved in milk fat and this is
It was confirmed that encapsulation with FGM can be used for cheese production.

(5) Ultrasonication of milk fat globule coating A certain amount of RTB dissolved in methanol (MeOH) (b
ag) into a test tube, and after removing the solvent, add 2 r of MFGM suspension dispersed in phosphate buffer (pH 7,0) to this.
mfl (MFGM50+ag) was added, and ultrasonication was performed at an output of 100 for a certain period of time (1-10 minutes) while cooling with ice. A 44% (w/w) sucrose solution of lll111 (specific gravity 1.20) was added to this and centrifuged at 5000 rpm for 30 minutes. The layer with >d 1.20 was collected and dried under reduced pressure, and then RTB was extracted from the residue with methanol and measured by HPLC.

In this specification, the particle size and flavin of the emulsion were determined in the following manner.

The particle size and specific surface area of the bean l'yo55 constant emulsion are LA-50.
Measurement was performed using a type 0 laser diffraction particle size distribution analyzer (Horiba, Ltd.).

Fractionation of flavins by HPLC Samples were heated in boiling water for 3 minutes, and pronase (1:
50. from St,riseus Ca
After digestion for 60 minutes with lbiochem), the supernatant (200 μQ) obtained by centrifugation was injected into an SP 8700 LC pump, a 5P4270 integrator, and a CAPCELL P
AW C18 column (5pm, 4.6X250+nm
) using MeOH/10 mM Na phosphate (pH
5,5; 30%MeOH-+8 win→86%Me
OH→5 win), 0.8m+2/win, 40℃
It was analyzed in However, for RTB analysis, MeOH and 10 m
A 95°C 5-mixture solution (v/v) of M Na phosphate (pH 5,5)
) (flow rate: 1.2 mQ/win).

(6) Binding of RTB to MFGM by ultrasonic treatment As a result of the ultrasonic treatment of the milk fat globule membrane described above, the results shown in Table 1 were obtained, but the amount of RTB bound to MFGH was as follows.

When ultrasonicated for 3 minutes at an output of 100 υ, it reached a nearly constant level, but was not affected by the ultrasonic wave output direction (20-1501j) or the MFGM concentration (10 JOmg).

Table 3 shows the amount of RTB bound per lipid, protein, and sample. In particular, R binding to MFGM
Different results were obtained regarding the amount of TB depending on the treatment method used during MFGM preparation. That is, the amount of RTB bound was greater in the freeze-dried sample (MFGM 5 ).

It was also thought that heating due to sterilization of the cream used to prepare MFGM increased the binding of RTH (MFGM 1.
2). These facts indicate that partially denatured MFGI
This indicated that the amount of TB bound was high.

In addition, a considerable amount of RTB can be bound to buttermilk, which is a byproduct of butter production, and in particular, the RTB binding rate of buttermilk that has been dialyzed to remove lactose is 66%.
%, it showed the highest value among the samples tested. Based on this, when the amount of binding to casein was investigated, casein also bound RTB equivalent to that of untreated MFGM.

Hatake 1 freeze-drying 32.4±0.8718.2±1.181
0.4±0.8152.9±3.42MFGN 2 lyophilized words, 2±1.9717.5±0.9511.6±0
．． 635111.1±3.42M) 13M3 Non-lyophilized 15.3±0.69 14.9±0.52 6.6±
0.23 33.0±1.19f14 Non-lyophilized 13
．． 3±1.39 13.3±1.38 6.6±0.6
9 33.2±3.45 MFGMS non-lyophilized 11.
9±0.66 11.3±0.36 5.8±0.18
28.9±0.91MFGM 5 Freeze-dried 15.4±
1.1414.2±1.74 7.3±0.8936.
4±4.45 butter milk 1rMr21.0±1.23
52.3±0.38 4.5±0.03 22.6±
0.14 putter milk dialysis 19.3±1.0875.6
±4.2413.3±0.7566.4±3.72 Casein 5.9±0.36 - 5.
7±0.25 28.4±1.23 Ultrasonic treatment: output 601, processing time 4 minutes, MFGM50*eg, RTB
The test was carried out using 1 mg/2wxQ 10mM phosphate buffer.

MFGM 1 and 2 are sterilizing cream (90℃, 30 seconds)
was concentrated using an IF membrane and freeze-dried.

MFGH3,4,5 were prepared by the method of Kanno et al.
It was used as a suspension, but some was lyophilized. From butter production, buttermilk was also dialyzed to remove lactose and other dialyzable substances and freeze-dried. The acid casein was neutralized and freeze-dried.

(7) Uptake of RTB by reconstitution of milk fat globules The emulsifying activity of reconstituted fat globules is determined by emulsification time of 1-10 minutes and milk fat content of 20% or more (10-60%) at the maximum speed of the Polytron homogenizer. It became almost constant (Table 4
).

However, emulsion stability decreased significantly at milk fat contents of 30% and above. Also, the average diameter of the particles of reconstituted milk fat balls increases with increasing fat content in the presence of a constant MFGH, with an average diameter of 19.5% at 65% milk fat content.
At 2 μm, it was about 7.7 times that at 10% (2.5 μm). On the other hand, the specific surface area of the particles decreased, contrary to the increase in the milk fat content and the average diameter of the particles, and the specific surface area at 60% milk fat content decreased to 1/6.2 of that at 10% milk fat content. .

On the other hand, the average diameter of the particles of milk fat balls reconstituted with +80 mg MFGM and 25% milk fat and their specific surface area were not affected by the emulsification time. 80mg MFGM
Approximately 3.1 mg of RTB could be incorporated into a 10-emulsion reconstituted with 2.5 g of milk fat (25%) and 2.5 g of milk fat (25%).

A, Milk fat content 2.53±009 5.44±0.12 6.75±0.31 7.58±008 9.48±008 11.82±068 16.00±016 B, Emulsification time 268. 3±6.5 143.1±2.3 118.1±1.2 103.2±0.4 82.0±0.4 66.7±2.9 52.6±0.6 3 6. 57±0.31 118.1±1.25
6.32±0.12 127.0±1.710
6.37±0.08 127.6±2.92
0 6.27±0.24 130.3±1.2
(n=3, standard deviation above the mean) 183.6±6.8 105.9±2.3 87.4±2.5 65.2±261 53.3±1.0 42.5±1.6 33.7±1.9 1.25 2.50 3.13 3.75 5.00 6.25 7.50 873.6±25.4 852.1±17.5 838.3±3663 847.5 ±33.5 A suspension of 80 mg of MFGM in a total volume of 10 mjl dispersed in 10 mM phosphate buffer (pH 7,0) and milk fat (A
: 1-6.5 g, B: 2.5 g) at 45°C for a certain period of time (A: 2.5 g) at the maximum speed of a Polytron homogenizer.
3 minutes, B: 3-20 minutes) Stirring and emulsification.

(8) Emulsifying function of RTB Milk fat containing 1,25B RTB per gram of milk fat and 8
When emulsifying with different milk fat contents in the presence of 0 mg of MFGM, no difference was observed in the emulsifying activity values, but 50
A significant difference was observed in the average particle diameter of emulsions obtained in the presence of % milk fat depending on the presence or absence of RTB (Table 5).

A particularly remarkable phenomenon was observed with milk fat content in phase transformation. That is, phase transformation due to milk fat content was observed at 60% when RTB was not included, whereas when RTB was
was observed in 70% of cases. This suggests that RTB acts as an emulsifying agent for MFGM. This is probably because the isoalloxazine ring of RTB is oriented toward the hydrophobic side of the core lipid, and the butyric acid attached to ribitol is oriented toward the MFGH side of the interface.

20 5.51±0.05 5.44±0.1
240 12.26±0.26 9.48±0.
0850 13.68±0.32 11.82±0
．． 6860 15.98±0.1
6792±19 728±27 705±26 Umbrella&47±18 853±17 850±32 809 Shiba RTB contains 1.25 mg/g of milk fat, and a certain amount of MFGM (80+ng) was used.

*60% when using RTB-free milk fat
Phase conversion occurred at milk fat content, but phase conversion in the presence of RTB occurred at 70%.

(9) Summary of the above results Various findings were obtained by the inventors as described above, and the summary of the results is as follows.

1) It was revealed that the bond between flavin and protein in MFGM, casein, and whey fractions is basically hydrophobic. The bound flavin of MFGH was not affected by pH up to pH 4.0, but was dissociated at pH 3.0.

2) RF, FMN, and FAD added to MFGH were not bound. Also, MFG from which flavin has been removed in advance
RF to M.

Even when FMN and FAD were added, the amount of binding did not exceed the initial binding amount.

3) Instead of RF, lipophilic RTB (1, 25 mg
) 2.5g of milk fat and 80mg of MFG dissolved in
The emulsion with a total volume of 10 mfl prepared with
It was possible to incorporate RTB of g into the core lipid of reconstituted milk fat shield. After centrifugation of the emulsion, the precipitate fraction contains insoluble RTB (8% of the total) along with a small amount of protein.
Approximately 76% of RTB was recovered in the cream fraction, and the rest was recovered in microfat globules and MFGM.

4) By ultrasonicating MFGM and RTB,
It was possible to bind 6-12++g of RTB per gram of MFGM and 13 gram of RTB per gram of buttermilk Ig.

5) RTB was found to act significantly as an emulsification aid, as the particle diameter of emulsions containing RTB became smaller when the fat content was high.

(10) Production of riboflavin-enriched cheese To carry out the present invention, fat (e.g. milk fat) in which riboflavin tetrabutyrate (RTB) is dissolved and milk fat globule membrane (MFGM) are used.
(lyophilized products also show good results) and/or emulsions prepared from ultrasonication thereof may be used in the cheese manufacturing process.

For example, in the case of natural cheese, the above-mentioned emulsion is used when adjusting raw milk and/or its components (for example, when appropriately mixing and adjusting part or all of milk, cream, skim milk, partially skimmed milk, and buttermilk). You can process it using

However, in the present invention, the emulsion treatment is not limited to the above-described preparation of raw milk, but includes a step of adding rennet and starter and slowly stirring to coagulate to form a curd, a step of cutting the same, and a step of stirring the cut curd. The emulsion treatment may be performed in at least one of the steps in natural cheese production, such as heating and removing whey, crushing and salting, pressurizing, and aging in a fermentation chamber.

In addition to using the above-mentioned natural cheeses as raw materials for processed cheese, at least one of the process cheese manufacturing processes, such as selecting and combining natural cheeses, trimming, crushing, heating and melting with the addition of molten salt, and emulsification, is used as a raw material. , the emulsion treatment described above may be performed.

In the present invention, cheese includes not only the above-mentioned natural cheese and processed cheese, but also cheese spreads, cheese foods, powdered cheese, and other cheese products. In order to manufacture these cheese products according to the present invention, in addition to using the above-mentioned riboflavin-enriched natural cheese or processed cheese as a raw material, it is sufficient to appropriately emulse them in the manufacturing process of these cheese products.

Emulsion processing is carried out by adding and mixing various emulsions obtained in the above-mentioned examples at the time of ingredient adjustment of raw milk itself and/or raw milk, selective mixing of raw milk, etc. The emulsion may be used in at least one step. If necessary, this emulsion may be used as part or all of the milk fat when milk fat is added and reduced after the raw milk is defatted or partially defatted.

By changing the amount of this emulsion used or by changing the amount of RTB mixed into this emulsion, it is possible to freely control the amount of riboflavin added to cheese.

1) Production of Gouda cheese fortified with riboflavin Add RTB prepared according to the method described in section (7) to raw milk.
After adding 1% of the containing emulsion and sterilizing it by the usual method,
Cooled.

To this, 1% of a cheese starter containing 5 treptococcus 1actis, Str, and cremoris as main components was added, and after 30 minutes, 0.015 rennet was added.
% was added little by little as a 2% saline solution at 30°C.

After stirring slowly for 10 minutes, the mixture was left to stand for 30 minutes.

After stirring the generated curd and pulverizing it to the size of azuki beans, drain 1/3 of the total amount of whey, then add boiling water of 80°C or higher and stir to bring the whey temperature to about 35°C, then add whey 1. 1/3 of the whey was discharged, and the whey temperature was further increased to about 40° C., and the remaining 1/3 of the whey was discharged.

Salt (0.2% of the raw milk) was dissolved in the whey on the top of the curd, and the curd was stirred to mix thoroughly and then squeezed. After heating, it was cut and filled into a mold.

After pressing this with a cheese press to remove moisture, the curd was immersed in water at 10° C. for 10 hours while still in the mold.

After taking out the curd, it was soaked in saline solution for 3 days and drained.

This was aged for 3 months in a fermentation chamber controlled at a temperature of 10-18°C and a humidity of 80-85% to obtain Gouda cheese rich in riboflavin.

2) Production of riboflavin-enriched processed cheese 40% of the riboflavin-enriched Gouda cheese prepared above and 60% of cheddar cheese aged for 6 months were cut and mixed, and then further crushed with a chopper.

This was placed in a melting pot, heated to 48 to 50°C, and when softened, the temperature was further raised to reabsorb the separated fat and homogenize it. Here, 2% emulsifier containing polymerized phosphate and sodium citrate, and 0% RTB-containing emulsion prepared according to the method described in section (7) above.
．． 5% was added and heated to 67-68°C.

Heating was continued for 10 to 15 minutes, and when the melted cheese became uniform, it was immediately filled and packaged to obtain processed cheese rich in riboflavin.

(Effects of the Invention) The present invention has made it possible for the first time to successfully prepare cheese enriched with riboflavin.

Therefore, it is not only possible to prepare cheese with a riboflavin content comparable to that of cow's milk, which was previously impossible to prepare, but also cheese enriched with even higher amounts of riboflavin. Cheese, which is rich in minerals and trace elements, was further fortified with vitamin B2, greatly increasing its nutritional value and making cheese increasingly resemble a complete food.

Claims (1)

[Claims] (1) Cheese enriched with riboflavins. (2) The cheese according to claim 1, wherein the riboflavin is riboflavin tetrabutyrate. (3) A method for producing cheese enriched with riboflavins, which comprises adding and dissolving riboflavins in milk fat, adding an emulsifying substance to form an emulsion, and using this emulsion. (4) The method for producing cheese enriched with riboflavins according to claim 3, wherein the riboflavins are riboflavin tetrabutyrate. (5) The method for producing cheese enriched with riboflavins according to claim 3, wherein the emulsifying substance is a milk fat globule membrane. (6) The method for producing cheese enriched with riboflavins according to claim 5, wherein the milk fat globule coating is freeze-dried.