JPH1017595A - Modified zein and its production - Google Patents

Abstract

PROBLEM TO BE SOLVED: To produce a new inexpensive modified zein, having water solubility, capable of retaining good physical properties of the zein and widely utilizable in the fields of medicines, healthy foods and foods by dissolving a protein zein in an ethanol solution and then hydrolyzing the zein with an acid under mild conditions. SOLUTION: This modified zein is obtained by passing a prolamine-based protein zein prepared from corn through a step of dissolving the zein in an ethanol solution and then hydrolyzing the zein with an acid under mild conditions. The modified zein has water solubility, emulsifiability, foaming properties and foam stability, is excellent in ability to reduce the surface tension and inexpensive and can be expected in wide utilization in the fields of medicines, healthy foods and foods. The modified zein is prepared by dissolving the protein zein in the ethanol solution, then partially hydrolyzing the zein with an acid at 0.01-3.2%. preferably 0.2-0.7% final concentration under temperature conditions of 40-80'C, preferably 50-60 deg.C for 4-24hr, preferably 4-6hr reactional time and subsequently desalting and drying the resultant solution.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD OF THE INVENTION

[0002] The present invention relates to a modified zein obtained from a prolamin-based protein obtained from corn and a method for producing the same.

More specifically, the zein is partially decomposed by an acid and solubilized in water while maintaining the original physical properties (particularly, molecular weight and film properties) of the zein as much as possible, so that solubility, emulsifying property, foaming property and the like are improved. The present invention relates to zein that has been improved in physical properties and enhanced in degradability by enzymes, thereby enabling its use in the pharmaceutical, health food and food fields.

[0004]

[Prior art]

[0005] Zein, a natural prolamin-based protein, is considered to be insoluble in water, poorly digestible, and low in nutrition (Nutrition and Food, 30 , 153 (197)
7)) Because it is not used as a food material,
It is used as a coating agent, coloring agent and adhesive.

For example, when zein is used as a coating agent for pharmaceuticals, it is used after being dissolved in a solvent such as an ethanol solution, but it is difficult to use.

For this reason, various zein reforming methods have been conventionally proposed.

[0008]

[Problems to be solved by the invention]

[0009] However, according to the conventional method, although the decomposition proceeds to make it water-soluble and easy to use, it loses the good physical properties of zein, such as film properties and emulsifying properties, and also has disadvantages such as high cost. is there.

Therefore, there has been a demand for a modified zein which has water solubility, does not impair the good physical properties of zein, is excellent in cost, and can be expected to be widely used in the fields of medicine, health foods and foods. .

[0011]

[Means for Solving the Problems]

The present inventors have conducted intensive studies to solve the above-mentioned problems. As a result, after dissolving zein with an ethanol solution, the acid concentration and various physical properties (molecular weight, film property, solubility, emulsifying property, Examination of the relationship between foaming properties, surface tension lowering ability, and digestibility by enzymes) revealed that zein had better physical properties (eg, film properties) by decomposing under milder conditions than those reported so far. The inventors have found that the physical properties (solubility, emulsifiability, etc.), which were disadvantages of zein, can be improved while leaving them, and completed the present invention.

That is, means for solving the problems of the present invention are as follows.

First, modified zein obtained by dissolving protein zein with an ethanol solution and then subjecting the zein to acid decomposition under mild conditions.

Second, after dissolving the protein zein with an ethanol solution, an acid having a final concentration of 0.01% to 3.2%, preferably 0.2% to 0.7% is used, and the temperature condition is adjusted. 40-80 ° C, preferably 50-60 ° C, reaction time 4
Modified zein obtained through a step of partially decomposing in 24 hours, preferably 4-6 hours.

Third, it has solubility in water, and
Modified zein with excellent emulsifying properties, foaming properties and surface tension.

Fourth, a modified zein having the same molecular weight distribution as protein zein and having the following characteristics (a) to (f): (A) High solubility in water. (B) Strong emulsifying properties. (C) It has high foaming properties and foam stability. (D) High surface tension lowering ability. (E) Degradability by proteolytic enzymes is high. (F) The film formability is equivalent.

Fifth, a method for producing modified zein, comprising dissolving protein zein with an ethanol solution and then subjecting the zein to acid decomposition under mild conditions.

Sixth, after dissolving the protein zein with an ethanol solution, an acid having a final concentration of 0.01% to 3.2%, preferably 0.2% to 0.7% is used, and the temperature condition is adjusted. 40-80 ° C, preferably 50-60 ° C, reaction time 4
A method for producing modified zein, which is produced through a step of partially decomposing in 24 hours, preferably 4-6 hours.

Seventh, the final concentration of the protein zein in an ethanol solution is 0.01% to 3.2%, preferably
0.2% to 0.7% acid is added and the temperature condition is 40 to 80
C., preferably at 50 to 60.degree. C. and a reaction time of 4 to 24 hours, preferably 4 to 6 hours, and then the resulting decomposition solution is desalted and then dried. Method for producing zein.

Eighth, the final concentration of the protein zein in an ethanol solution is 0.01% to 3.2%, preferably
0.2% to 0.7% acid is added and the temperature condition is 40 to 80
C., preferably at 50 to 60.degree. C. and a reaction time of 4 to 24 hours, preferably 4 to 6 hours. After partial decomposition, the obtained decomposition solution is centrifuged, the precipitate is washed with water and dried, and the supernatant is neutralized. A method for producing modified zein, comprising removing alcohol after drying, and drying.

[0022]

BEST MODE FOR CARRYING OUT THE INVENTION

As the zein used as a raw material for obtaining the modified zein according to the present invention, a commercially available zein powder or an alcohol extract (powder or extract) of corn gluten meal can be used.

Here, when alcohol is extracted from corn gluten meal, ethanol or isopropyl alcohol can be used.

A commercially available zein powder or an alcohol extract of corn gluten meal is used as a protein zein as a raw material,
When dissolving the protein zein with an ethanol solution, 5-kg of ethanol is added to 1 kg of the protein zein.
80 liters are added and dispersed, and then 10 liters of water is added to dissolve protein zein. At this time, the amount of ethanol is preferably 15 to 25 liters.

For the partial decomposition of zein after dissolving the protein zein with an ethanol solution, a final concentration of 0.01
% To 3.2% hydrochloric acid, preferably 0.2 to 3.2%.
More preferably, the concentration of hydrochloric acid is 0.7%.

As the acid used for the partial decomposition, general mineral acids and organic acids can be used.

The temperature conditions for the partial decomposition are 40 to 80.
° C, preferably 50 to 60 ° C.

The reaction time for partial decomposition is 4 to 2
4 hours, preferably 4 to 6 hours.

The removal of the acid from the decomposition solution obtained after the partial decomposition can be performed by ion exchange, dialysis, neutralization, washing with water, or the like.

The drying performed after the removal of the acid can be performed by air drying, warm air drying, spray drying, freeze drying, or the like.

The modified zein according to the present invention can be used as food additives as various food emulsifiers, detergents, raw materials for protein drinks, etc., and also in foods and the like that require emulsifying and foaming properties. Can also be used.

The modified zein according to the present invention is desirably used as a cleaning agent for those requiring non-toxicity such as foods and dishes.

When the modified zein according to the present invention is used as a raw material for protein drinks, the modified zein according to the present invention has excellent solubility in water and is resistant to heating. It is preferably used as a beverage that can be heat sterilized.

[0035]

【Example】

Hereinafter, the present invention will be described in more detail with reference to Examples and Test Examples.

[0037]

Embodiment 1

Freeze-dried corn gluten meal (FD)
-CGM) was defatted with ethanol: hexane (1: 1), extracted with 6 times the amount of ethanol by weight of FD-CGM at 60 ° C. for 6 hours, spray-dried, and dried. Zein (ZS-70E) for raw material in Examples
I got

To 30 g of this zein (ZS-70E), 600 ml of ethanol was added to disperse the mixture, and 300 ml of water was added to dissolve the mixture. Then, 90 ml of 20% hydrochloric acid was added to a final concentration of 1.82%. And reacted at 55 ° C. for 5 hours.

The solution obtained above was allowed to stand at 4 ° C. overnight, and then centrifuged to separate a precipitate P and a supernatant S.

The precipitate P was washed with water, pulverized, dissolved in 1% aqueous ammonia, centrifuged, and precipitated PP (DA-P
P) 3.83 g and supernatant PS.

The supernatant PS was adjusted to pH 5 with acetic acid, and then centrifuged to precipitate PSP (DA-PS) 10.
36 g were obtained.

On the other hand, the supernatant S was obtained by removing ethanol with a rotary evaporator to obtain a precipitate SP, dissolved by adding 1% aqueous ammonia, and centrifuged to obtain 5.59 g of the precipitate SPP (DA-SP). The supernatant was divided into SPS.

The supernatant SPS was adjusted to pH 5 with acetic acid, and then centrifuged to precipitate SPSP (DA-SS).
52 g were obtained.

[0045]

Embodiment 2

The same zein (ZS-70E) as in Example 1
To 5 g, 100 ml of ethanol was added and dispersed.
After adding 0 ml and dissolving, 15 ml of 20% constant boiling hydrochloric acid was added so that the final concentration was 1.82%, and 55 ml of 55% hydrochloric acid was added.
It was left at ℃ for 5 hours.

Thereafter, the precipitate obtained by dialysis against water was washed with water while crushing in a mortar, and filtered with Nutsche.
Air dried to obtain modified zein (TDA).

[0048]

Embodiment 3

The same zein (ZS-70E) as in Example 1
To 5 g, 100 ml of ethanol was added and dispersed.
After dissolving by adding 0 ml, 5 ml of 20% hydrochloric acid was added.
The mixture was added to a final concentration of 0.65%, and left at 55 ° C. for 5 hours.

Thereafter, the precipitate obtained by dialysis against water was washed with water while crushing in a mortar, and filtered with Nutsche.
Air-dried to obtain modified zein (3L-DA).

In this case, 5 g of zein (ZS-70E)
Was 3.4 g, and the yield was 68%.

Next, 3.0 ml of 20% hydrochloric acid was added.
In the same manner as above except that the amounts were changed to 1.5 ml and 0.15 ml, modified zein (5 L-DA) and modified zein (10 L
-DA) and modified zein (100 L-DA) were obtained.

In these cases, zein (ZS-70)
E) The yield (g) and yield (%) for 5 g were determined, respectively.

The results are shown in Table 1.

[0055]

[Table 1]

[0056]

[Test Example 1] [HPLC analysis]

According to this test example, the molecular weight distributions of the modified zein obtained in Examples 1 to 3 and the zein (ZS-70E) prepared in Example 1 as a control were determined by high performance liquid chromatography. Was measured as follows.

The modified zein (3L-DA), modified zein (5L-DA), modified zein (10L-DA), modified zein (100L-D) obtained in Examples 1 to 3 above.
A), modified zein (DA-PP), modified zein (DA-PP)
PS), modified zein (DA-SP), modified zein (DA
-SS), modified zein (TDA), and zein (ZS-70E) prepared in Example 1, 1 mg of each zein, 1 ml of 10 mM SDS-100 mM phosphate buffer (pH 6.8). And sonicated.

After that, high performance liquid chromatography (HPLC) through a 0.45 μm membrane filter
Was supplied with 10 μl.

In the high performance liquid chromatography, Bio-Sil SEC-250, 300 mm × 7.8 was used for the column.
mm (manufactured by Bio-Rad) and the mobile phase is 10 mM
SDS-100 mM phosphate buffer (pH 6.8) was used at a flow rate of 1.0 ml / min.
m was used.

FIG. 1 shows the results of analysis by high performance liquid chromatography.

[0062]

[Test Example 2] [Solubility test]

In this test example, the solubility of the modified zein obtained in Examples 1 to 3 and the zein (ZS-70E) prepared in Example 1 as a control were measured as follows.

500 mg of each modified zein obtained in Examples 1 to 3 was suspended in 10 ml of water, stirred for 30 minutes, and then bisected.

One of the two halves was adjusted to pH 4 with 1N sodium hydroxide solution,
The mixture was stirred for 30 minutes, a part was collected, centrifuged, and separated into a supernatant and a precipitate.

The separated supernatant was subjected to the Lowry method (J. Bio. Chem., 193 , 265 (195).
The protein concentration was measured using 1)) to determine the solubility.

The precipitate obtained by centrifugation was returned to a pH 4 solution, further adjusted to pH 5, stirred for 30 minutes, and a part was collected and centrifuged. As described above, the protein concentration was measured, and the solubility was determined.

Here, for pH 6 to pH 11, pH
As in the case of adjusting the pH to 5 after returning to the solution of 4,
After returning to the pH 4 solution, the pH was adjusted to each, the protein concentration was measured, and the solubility at each pH was determined.

The other of the two halves of pH 1 to 3 was adjusted to each pH using 1 N hydrochloric acid, and the protein concentration was measured to determine the solubility at each pH.

As a control, zein (ZS-70) was used.
Regarding E), the solubility at each pH was determined in the same manner as described above.

Each modified zein and zein (ZS-70E)
Is shown in Table 2.

[0072]

[Table 2]

[0073]

[Test Example 3] [Emulsification test]

According to this test example, the results obtained in Examples 1 to 3 were obtained.
Each modified zein and zein (ZS-70
E) The method of Pearce et al.
[J. Agric. Food Chem. ,26
(3), 716-723, 1978]
The emulsifiability was measured as follows.

10 mg of each modified zein and the same zein (ZS-70E) as in Example 1 as a control were dissolved in a phosphate buffer (pH 0.1, ionic strength: 0.1) of pH 8, and 6 ml of the solution was previously prepared. Take into a test tube containing 2 ml of soybean oil and at 25 ° C., Junke & Kunkel blender
And stirred at 25000 rpm for 1 minute.

Immediately after stirring, 100 μl was sampled from the center of the emulsified layer or from the center of the separated water layer when there was separated water, and 9.9 ml of 0.1% SDS-0.1 M sodium chloride (pH 7.0) was sampled. The mixture was mixed with the solution to prepare a sample for measuring emulsifiability.

Similarly, after 24 hours from the stirring, 100 μl was sampled from the center of the emulsified layer or from the center of the separated water layer when there was separated water, and 9.9 ml of 0.9 μl was sampled.
It was mixed with a 1% SDS-0.1 M sodium chloride (pH 7.0) solution to prepare a sample for measuring emulsifiability.

Here, the index of the emulsifying property is EAI (Emu
Using the LS activity value, immediately after the emulsification (EAI ₀ ) and after 24 hours (EAI ₀ )
_{24 hr} ), the EAI value was measured.

Further, after 24 hours, the emulsified sample
After heating at 80 ° C. for 30 minutes, the mixture was cooled, and the EAI ₈₀ ° C. was measured in the same manner as above.

For each measurement sample, the absorbance (A) at λ = 500 nm was measured using a spectrophotometer.

The EAI value was calculated by the following equation.

EAI = 2T / φC

In the above formula, T = 2.3 A / I, φ = the ratio of the oil layer to the total amount of the emulsified sample, 0.25, I = 0.01 m, the optical path length of the spectrophotometer cell, and C is the value before emulsification. The protein concentration of the prepared protein solution indicates 0.1.

Further, the emulsion stability was determined by the following equation.

[0085] · ΔEAI% = (EAI _max -EAI
₈₀ ° C) × 100 / EAI _max

In the above equation, EAI _max means EAI ₀
And EAI _{24 hr} , the larger one, and the smaller the ΔEAI value, the better.

Each modified zein and zein as a control (Z
Table 3 shows the measured values of emulsifiability and emulsification stability at 25 ° C and pH 8 for S-70E).

[0088]

[Table 3]

According to Table 3, each modified zein obtained in Examples 1 to 3 was more excellent in emulsifying property than zein (ZS-70E).

[0090]

[Test Example 4] [Foamability test]

According to this test example, each modified zein obtained in Examples 1 to 3 and zein (ZS-70
E), Slack et al. [J. Foo
dProc. Pres 10 , 81-88], and the foamability was measured as follows.

25 mg of each modified zein and the same zein (ZS-70E) as in Example 1 as a control were suspended in 24.5 ml of water, adjusted to pH 8 with an aqueous sodium hydroxide solution, and stirred for 30 minutes. did.

After stirring, the total amount was adjusted to 25 ml with water, and 10 ml (Vs) was sampled.

Using a foaming property measuring apparatus shown in FIG. 2, nitrogen was flowed at a flow rate of 30 ml / min for 80 seconds while maintaining the temperature at 25 ° C. with a water jacket, and the volume (Vf) of the generated foam was measured. .

Further, the volume (Vd) of the liquid discharged from the foam was measured after 2 minutes and after 4 minutes and 30 seconds after flowing nitrogen for 80 seconds.

The foaming property (% OVERRUN)
Was calculated by the following equation.

Foaming property (% OVERRUN) = ｛(V
_{f -V S) / V S}} × 100

In addition, foam stability (% DRAINAGE)
Was calculated by the following equation.

Foam stability (% DRAINAGE) =
_{(V d / V S) ×} 100

For each of the modified zein and zein (ZS-70E) obtained above at 25 ° C. and pH 8,
Table 4 shows the foamability and foam stability.

[0101]

[Table 4]

According to Table 4, the modified zein (DA-PP)
Other than the above, the foaming properties and foam stability were superior to zein (ZS-70E).

[0103]

[Test Example 5] [Surface tension test]

According to this test example, the surface tension of each of the modified zein obtained in Examples 1 to 3 and the control was measured by the Dunui method as follows.

Each of the modified zeins was dissolved in a sodium hydroxide aqueous solution having a pH of 8 so as to have a protein concentration of 0.1 W / V%, and the surface tension of each solution was measured at 25 ° C. according to the Dunuy method.

For control, zein (ZS-70) was used.
E) was dissolved in a sodium hydroxide aqueous solution having a pH of 8 so that the protein concentration was 0.1 W / V%, and the surface tension at 25 ° C. was measured according to the Dunui's method.

For reference, the surface tension of pure water at 25 ° C. was measured in the same manner.

Table 5 shows the results.

[0109]

[Table 5]

[0110]

[Test Example 6] [Digestibility test]

According to this test example, each of the modified zein obtained in Examples 1 to 3 and the zein (ZS-70E) prepared in Example 1 as a control were used as follows. (Amano Pharmaceutical Co., Ltd.), digestibility was measured by using the digestive enzymes pepsin and chymotrypsin according to the method of Nuya et al. (Nutrition and Food, 32 , 291 (1979)).

That is, modified zein (TDA) and zein (ZS-70E) were added to a 100 mM phosphate buffer (pH
8.0) to 10 mg / ml.
Pro leather was added at a ratio of 1/100 to the substrate.

The modified zein (TDA) and zein (ZS-70E) were added to 100 mM Clark Labs buffer (pH 2.0) for 2 hours.
After dissolving each to a concentration of mg / ml, pepsin was added at a ratio of 1/100 to the substrate.

Further, each modified zein and zein (ZS-
70E) in 100 mM phosphate buffer (pH 8.0)
Were dispersed at a concentration of 2 mg / ml, and chymotrypsin was added at a ratio of 1/100 to the substrate.

In any of the above cases, after the addition, sampling was performed over time while incubating at 37 ° C.
After adding and mixing the same amount of 20% trichloroacetic acid (TCA), No. 6. Then, the filtrate was directly used or diluted 10 times with water, and the absorbance at λ = 280 nm was measured.

The absorbance of the sample immediately after the addition of the enzyme was subtracted as a blank.

FIG. 3 shows the results of a digestion test using a prolaser.
FIG. 4 shows the results of the digestion test with pepsin, and FIGS. 5 to 7 show the results of the digestion test with chymotrypsin.

[0118]

[Test Example 7] [Film property test]

In this test example, the film properties of the modified zein obtained in Examples 1 to 3 and the zein (ZS-70E) prepared in Example 1 as a control were examined.

That is, each modified zein obtained in Examples 1 to 3 was dissolved in 10 ml of a 70% ethanol solution, spread on a petri dish, and allowed to stand at 60 ° C. for 12 hours. Observed with the naked eye.

Similarly, 500 mg of control zein (ZS
-70E) was dissolved in 10 ml of a 70% ethanol solution, spread on a petri dish, allowed to stand at 60 ° C for 12 hours, and then visually observed for film formation.

The results are shown in Table 6.

[0123]

[Table 6]

[0124]

【The invention's effect】

The modified zein according to the present invention has water solubility, does not impair the good physical properties of zein, is excellent in cost, and is expected to be widely used in the fields of medicine, health food and food.

[Brief description of the drawings]

FIG. 1 shows the results of analysis by high performance liquid chromatography.

FIG. 2 is a schematic diagram of a foaming property measuring device.

FIG. 3 shows the results of a digestion test using a prolaser.

FIG. 4 shows the results of a digestion test with pepsin.

FIG. 5 is a view showing the results of a chymotrypsin digestion test on TDA and the like.

FIG. 6 is a diagram showing the results of a chymotrypsin digestion test on DA-PP and the like.

FIG. 7 shows the results of a digestion test with chymotrypsin on 3L-DA and the like.

Claims (8)

[Claims] 1. Modified zein obtained by dissolving protein zein with an ethanol solution and then subjecting the zein to acid degradation under mild conditions. 2. After dissolving the protein zein with an ethanol solution, an acid having a final concentration of 0.01% to 3.2%, preferably 0.2% to 0.7% is used, and the temperature condition is 40 to 40%.
80 ° C, preferably 50-60 ° C, reaction time 4-24
Modified zein obtained through a step of partially decomposing in time, preferably 4 to 6 hours. 3. A modified zein which has solubility in water and is excellent in emulsifying property, foaming property and surface tension. 4. Modified zein having the same molecular weight distribution as protein zein and having the following characteristics (a) to (f). (A) High solubility in water. (B) Strong emulsifying properties. (C) It has high foaming properties and foam stability. (D) High surface tension lowering ability. (E) Degradability by proteolytic enzymes is high. (F) The film formability is equivalent. 5. A method for producing modified zein, comprising: dissolving protein zein with an ethanol solution, followed by acid degradation under mild conditions. 6. After dissolving the protein zein with an ethanol solution, an acid having a final concentration of 0.01% to 3.2%, preferably 0.2% to 0.7% is used, and the temperature condition is 40 to 40%.
80 ° C, preferably 50-60 ° C, reaction time 4-24
A method for producing modified zein, which is produced through a step of partially decomposing in 4 hours, preferably 4 to 6 hours. 7. An ethanol solution of protein zein,
Final concentration of 0.01% to 3.2%, preferably 0.2%
~ 0.7% acid is added, the temperature condition is 40 ~ 80 ° C, preferably 50 ~ 60 ° C, and the reaction time is 4 ~ 24 hours, preferably 4 ~ 6 hours. Is dried after desalting. 8. An ethanol solution of protein zein,
Final concentration of 0.01% to 3.2%, preferably 0.2%
~ 0.7% acid is added, the temperature condition is 40 ~ 80 ° C, preferably 50 ~ 60 ° C, and the reaction time is 4 ~ 24 hours, preferably 4 ~ 6 hours. , Centrifuging the precipitate, washing and drying the precipitate with water, neutralizing the supernatant, removing the alcohol, and drying.