METHOD FOR PRODUCING POLYSACCHARIDE-PROTEIN COMPLEX

Abstract

[Problem] The present invention addresses the problem of providing: a material which can generate fine beads in a sparkling beverage and can keep the fine beads in the beverage without adversely affecting the flavor of the beverage when added to the beverage; and a method for producing the material. [Solution] A method for producing a polysaccharide-protein complex, comprising a heating step of heating a polysaccharide containing an uronic acid as a constituent sugar in the presence of a protein under the conditions in which the pH value is 2 to 5 and the temperature is 80 to 180 C. inclusive, wherein a fraction having a molecular weight of 12000 or more makes up 30% by mass or less of the polysaccharide-protein complex and a fraction having a molecular weight of 500 or more and less than 12000 makes up 30% by mass or more of the polysaccharide-protein complex.

Claims

1. A method for producing a polysaccharide-protein complex, comprising heating a polysaccharide containing uronic acid as a constituent saccharide in a presence of a protein at a pH of 2 to 5 and a temperature of 80 C. or more and 180 C. or less, wherein the polysaccharide-protein complex has 30% by mass or less of a fraction having 12000 or more of molecular weight and 30% by mass or more of a fraction having 500 or more and less than 12000 of molecular weight.

2. The method according to claim 1, wherein the polysaccharide-protein complex has 25% by mass or less of a fraction having 12000 or more of molecular weight and 40% by mass or more of a fraction having 500 or more and less than 12000 of molecular weight.

3. The method according to claim 1, wherein a ratio of the polysaccharide to the protein in the polysaccharide-protein complex is from 10:1 to 1:100 on mass basis.

4. The method according to claim 1, wherein a ratio of the polysaccharide to the protein in the polysaccharide-protein complex is from 5:1 to 1:10 on mass basis.

5. The method according to claim 1, wherein the polysaccharide is derived from pea seed.

6. The method according to claim 1, wherein a pH in the heating is 2.5 to 3.5.

7. A method for stabilizing foam of an effervescent beverage, comprising: a step of heating a polysaccharide containing uronic acid as a constituent saccharide in a presence of a protein at a pH of 2 to 5 and a temperature of 80 C. or more and 180 C. or less to obtain a polysaccharide-protein complex having 30% by mass or less of a fraction having 12000 or more of molecular weight and 30% by mass or more of a fraction having 500 or more and less than 12000 of molecular weight; and a step of adding the polysaccharide-protein complex to an effervescent beverage raw material.

8. A method for producing an effervescent beverage, comprising: a step of heating a polysaccharide containing uronic acid as a constituent saccharide in a presence of a protein at a pH of 2 to 5 and a temperature of 80 C. or more and 180 C. or less to obtain a polysaccharide-protein complex having 30% by mass or less of a fraction having 12000 or more of molecular weight and 30% by mass or more of a fraction having 500 or more and less than 12000 of molecular weight; and a step of adding the polysaccharide-protein complex to an effervescent beverage raw material.

9. The method according to claim 8, wherein the effervescent beverage contains an alcohol.

10. A polysaccharide-protein complex, comprising a protein and a polysaccharide containing uronic acid as a constituent saccharide, and having a 30% by mass or less of a fraction having 12000 or more of molecular weight and 30% by mass or more of a fraction having 500 or more and less than 12000 of molecular weight, wherein the complex is heated.

11. The complex according to claim 10, wherein a ratio of the polysaccharide to the protein in the polysaccharide-protein complex is from 10:1 to 1:100 on mass basis.

12. The complex according to claim 10, wherein the polysaccharide is derived from pea seed.

13. A foam stabilizer comprising the complex according to claim 10 as an active component.

14. An effervescent beverage comprising the foam stabilizer according to claim 13.

15. The effervescent beverage according to claim 14, comprising an alcohol.

Description

EXAMPLES

[0072] Hereafter, embodiments of the present invention will be explained concretely with reference to examples, but the technical concept of the present invention is not limited by the examples. Part(s) and % referred to in the examples mean mass basis unless stated otherwise.

[0073] After dehulling 50 kg of pea seeds, 5-fold amount of water was added to the cotyledon part, and then the seeds were immersed for 24 hours. The cotyledon part was ground with a Homomixer (5,000 rpm, for 30 minutes), and then protein and starch were extracted with pH 9 maintained using 30% sodium hydroxide solution. The components such as protein, and starch dispersed in water were removed by using a centrifugal filter (1,000 rpm, for 5 minutes), and a fiber was recovered. Moreover, 5-fold amount of water was added to the fiber and was stirred by using a Homomixer (3,000 rpm, for 30 minutes), and then the fiber was recovered by using a centrifugal filter (1,000 rpm, for 5 minutes). This operation was repeated twice and the fiber obtained was freeze dried to obtain 10 kg of a pea seed treated material was obtained.

Comparative Example 1

[0074] Into 940 parts of water, 60 parts of the above-described pea seed treated material was dispersed, then the mixture was adjusted to pH 6 using hydrochloric acid, and then heated at 120 C. for 90 minutes to extract a polysaccharide. An insoluble fiber in the liquid was removed by centrifugal separation (5,000 rpm, for 30 minutes) and the supernatant was recovered. The supernatant was heated to 60 C., and then an amylase (Fungamyl 800L; produced by Novozymes A/S) with an amount corresponding to 0.1% by mass of the solid content was added at pH 6 and a starch was decomposed for 1 hour. The amylase was then deactivated by heating in boiling water for 15 minutes, and then the mixture was freeze dried to obtain powdery water-soluble pea polysaccharide A.

Comparative Example 2

[0075] Into 940 parts of water, 60 parts of the above-described pea seed treated material was dispersed, then the mixture was adjusted to pH 3 using hydrochloric acid, and then heated at 120 C. for 90 minutes to extract a polysaccharide. An insoluble fiber in the liquid was removed by centrifugal separation (5,000 rpm, for 30 minutes) and the supernatant was recovered. The supernatant was adjusted to pH 6 with an alkali, and then an amylase (Fungamyl 800L; produced by Novozymes A/S) with an amount corresponding to 0.1% by mass of the solid content was added at 60 C., and the starch was decomposed for 1 hour. The amylase was then deactivated by heating in boiling water for 15 minutes, and then the mixture was freeze dried to obtain powdery water-soluble pea polysaccharide B.

Example 1

[0076] Into 940 parts of water, 40 parts of the above-described pea seed treated material (40 parts) and 20 parts of soybean protein (FUJIPRO-R; produced by Fuji Oil Co., Ltd.) were dispersed, then the mixture was adjusted to pH 3 using hydrochloric acid, and then heated at 120 C. for 90 minutes to extract a polysaccharide and form a complex of the polysaccharide and the protein. An insoluble fiber in the liquid was removed by centrifugal separation (5,000 rpm, for 30 minutes) and the supernatant was recovered. The supernatant was adjusted to pH 6 with an aqueous sodium hydroxide solution, and then amylase (Fungamyl 800L; produced by Novozymes A/S) with an amount corresponding to 0.1% by mass of the solid content was added at 60 C., and the starch was decomposed for 1 hour. The amylase was then deactivated by heating the resulting solution in boiling water for 15 minutes, and then the mixture was freeze dried to obtain powdery polysaccharide-protein complex A.

Examples 2 to 5

[0077] Polysaccharide-protein complexes B, C, D and E were obtained in the same manner as Example 1 except that potassium caseinate (Tatua 500: produced by Tatua Co-operative Dairy Co., Ltd.), wheat protein (produced by Hokkoku Food Co., Ltd.), pea protein (NUTRALYS F85M: produced by Rocket Japan), or dried albumen (Meringue Powder: produced by Taisei Co., Ltd.) was used instead of the soybean protein.

Example 6

[0078] Into weter, 30 parts of the water-soluble pea polysaccharide prepared in Comparative Example 1 and 30 parts of soybean protein (FUJIPRO-R; produced by Fuji Oil Co., Ltd.) were dissolved or dispersed, then the mixture was adjusted to pH 3 using hydrochloric acid, and then heated at 120 C. for 60 minutes to form a complex of the polysaccharide and the protein. The insoluble matter generated was removed by centrifugal separation (5,000 rpm, for 30 minutes) and the supernatant was recovered. The supernatant was adjusted to pH 4.5 with an alkali, and then freeze dried to obtain powdery polysaccharide-protein complex F.

Comparative Example 3

[0079] Into water, 30 parts of the water-soluble pea polysaccharide prepared in Comparative Example 1 and 30 parts of a soybean protein (FUJIPRO-R; produced by Fuji Oil Co., Ltd.) were dissolved or dispersed, then the mixture was adjusted to pH 3 using hydrochloric acid, then the insoluble matter was removed by centrifugal separation (5,000 rpm, for 30 minutes) without heating, and then the supernatant was recovered. The supernatant was adjusted to pH 4.5 with an alkali, and then freeze dried to obtain powdery composition A containing polysaccharide and protein.

Comparative Example 4

[0080] Into 540 parts of water, 30 parts of soybean protein (FUJIPRO R: produced by Fuji Oil Co., Ltd.) was suspended, then the mixture was adjusted to pH 7, and then protease (protease A Amano SD: produced by Amano Enzyme, Inc.) with an amount of 0.4% relative to the protein was added to the mixture to react at 40 C. for 60 minutes. The insoluble matter generated was removed by centrifugal separation (5,000 rpm, for 30 minutes) and the supernatant was recovered and boiled at 100 C. for 15 minutes to deactivate the enzyme, and then freeze dried to obtain an enzymatically decomposed protein. The enzymatically decomposed protein and the water-soluble pea polysaccharide B prepared in Comparative Example 2 were mixed together in a powdery form in a ratio of 1:2 to obtain composition B containing polysaccharide and protein.

Comparative Examples 5 and 6

[0081] Compositions C and D each containing polysaccharide and protein were obtained in the same manner as Comparative Example 4 except that the protease used was changed to THERMOASE PC1OF (produced by Amano Enzyme Inc.) or Papain W-40 (produced by Amano Enzyme Inc.).

Experiment Example 1

[0082] For the complexes A to F and the compositions A to D, the molecular weight was determined using a gel filtration HPLC method. An aqueous solution of complex or composition prepared using 20 mM phosphate buffer solution (pH 7.2) was filtered using a 0.2 m filter. The resulting filtrate was loaded to Superdex peptide 7.5/300GL (produced by GE Healthcare) and was eluted at a rate of 0.5 ml/minute using the above-mentioned phosphate buffer solution. The detection of each complex or composition was performed by measuring an absorbance at 214 nm. A calibration curve was produced using cytochrome c (molecular weight: 12384), aprotinin (molecular weight: 6512), gastrin I (molecular weight: 2098), angiotensin II (molecular weight: 1046), and triglutamic acid (molecular weight: 405) as molecular weight markers, and the distribution of molecular weight was determined on the basis of the calibration curve. Results are shown in Table 1.

TABLE-US-00001 TABLE 1 Ratio of each fraction present 12000 or 500 or more Less than more in and less than 500 in molecular 12000 in molecular weight molecular weight Type of (% by weight (% by protein Protease mass) (% by mass) mass) Example 1 Complex A Soybean 22.1 43.0 34.9 Example 2 Complex B Casein 24.2 45.7 30.1 Example 3 Complex C Wheat 25.3 36.0 38.7 Example 4 Complex D Pea 22.8 43.5 33.7 Example 5 Complex E Albumen 25.0 40.5 34.5 Example 6 Complex F Soybean 20.5 41.2 38.3 Comparative Composition A Soybean 36.3 23.0 40.8 Example 3 Comparative Composition B Soybean Protease A 1.6 28.2 70.2 Example 4 Comparative Composition C Soybean THERMOASE 2.3 29.4 68.3 Example 5 PC-10F Comparative Composition D Soybean Papain W- 1.2 2.0 96.8 Example 6 40

[0083] The complexes A to F contained much fraction being 500 or more and less than 12000 in molecular weight, which is believed to contribute to foam stability. On the other hand, the composition A contained low ratio of fraction being 500 or more and less than 12000 in molecular weight, which contributes to foam stability, but contained high ratio of fraction being 12000 or more in molecular weight because the composition A was unheated and not subjected to decomposition by heating. The compositions B to D formed by enzymatically decomposing the protein by the protease contained excessively increased amounts of fraction being less than 500 in molecular weight.

Application Example 1

[0084] For the water-soluble pea polysaccharides A and B, the polysaccharide-protein complexes A to F, and the compositions A and B, the foam stability of an aqueous solution was measured with the method described below. The results are shown in Table 2.

[0085] The polysaccharide, complex, or composition was dissolved in water to prepare each 0.2% aqueous solutions. Each of 30 ml of the aqueous solutions was put into a 100 ml Nessler color comparison tube with glass stopper. Then the stopper was put on the color comparison tube so as not to allow the solution to leak. And then, the color comparison tube was shaken along the height direction of the tube at a rotation speed of 150 rpm for 1 minute. The color comparison tube was fixed to a tube stand and was left to settle, and the thickness of a foam layer was measured at times of 5 minutes or 60 minutes after the beginning of the settlement. Using the thickness of the foam layer measured after 60 minutes as the index of foam stability: when the thickness is 30 mm or more, this is expressed by ; when the thickness is 20 mm or more and less than 30 mm, this is expressed by ; when the thickness is 10 mm or more and less than 20 mm, this is expressed by ; and when the thickness is less than 10 mm, this is expressed by . When the evaluation is or , this is rated as acceptable.

TABLE-US-00002 TABLE 2 Foam stability of aqueous solution Thickness of foam layer (mm) After After Foam Type of 5 minute 60 minute sta- protein settlement settlement bility Comparative Polysaccharide 16 14 Example 1 A Comparative Polysaccharide 5 3 X Example 2 B Example 1 Complex A Soybean 49 45 Example 2 Complex B Casein 54 47 Example 3 Complex C Wheat 35 29 Example 4 Complex D Pea 51 44 Example 5 Complex E Albumen 50 37 Example 6 Complex F Soybean 46 43 Comparative Composition A Soybean 14 13 Example 3 Comparative Composition B Soybean 24 13 Example 4

[0086] In all of the examples using the complexes A to F, extremely high foam stabilities were exhibited. Especially, the complexes obtained using soybean protein, casein, pea protein, or albumen had excellent foam stabilities. In addition, it was revealed that the complex A obtained using treated pea seeds and protein as starting raw materials and the complex F obtained using an existing water-soluble pea polysaccharide and protein as starting raw materials had almost the same foam stability, and it was found that complexes with superior foam stabilities may be produced in either production method. As observed in the composition A, the foam stability was not improved well in the case of omitting heating as compared to the case of using a polysaccharide independently. It was shown that heating was important in order to obtain a complex with high foam stability.

Application Example 2

Non-Alcoholic Carbonated Soft Beverage

[0087] For the water-soluble pea polysaccharides A and B, the polysaccharide-protein complexes A to F, and the composition A, the foam stability was measured by the method described below assuming a non-alcoholic carbonated soft beverage. The results are shown in Table 3.

[0088] Into a 100 ml Nessler color comparison tube, 49 ml of carbonated water was put and 1 ml of 10% aqueous solution of polysaccharide, complex or composition was added, and then they were mixed gently in a manner which avoids foaming. Using an ultrasonic foaming apparatus (Sonic Hour: manufactured by TAKARA TOMY A.R.T.S), an ultrasonic pulse was applied five times from the bottom of the color comparison tube for foaming. The thickness of the foam layer being after 2 minutes from the foaming was measured, and the criteria of foam stability are as follows: when the thickness is 40 mm or more, this is expressed by ; when the thickness is 30 mm or more and less than 40 mm, this is expressed by ; when the thickness is 20 mm or more and less than 30 mm, this is expressed by ; and when the thickness is less than 20 mm, this is expressed by . When the evaluation is or , this is rated as acceptable.

[0089] Organoleptic evaluation was performed for each carbonated beverage, and the foam quality (fineness of foam) and the flavor were evaluated according to the following criteria. The foam quality was evaluated on the basis of visual observation and mouthfeel at tasting, and the flavor was evaluated by tasting.

(Foam Quality)

[0090] : Very fine [0091] : Moderately fine [0092] : Coarse

(Flavor)

[0093] : Very good [0094] : Good [0095] : Bad

TABLE-US-00003 TABLE 3 Foam stability, foam quality and flavor in non-alcoholic carbonated soft beverage Type of Foam Foam protein stability quality Flavor Comparative Polysaccharide A X Example 1 Comparative Polysaccharide B X X Example 2 Example 1 Complex A Soybean Example 2 Complex B Casein Example 3 Complex C Wheat Example 4 Complex D Pea Example 5 Complex E Albumen Example 6 Complex F Soybean Comparative Composition A Soybean X X Example 3

[0096] Also for a beverage containing carbon dioxide gas, the beverages of Examples to which the complexes A to F were added were provided with high foam stability. In Application Example 1, there were not so remarkable differences regarding foam quality and flavor, but as to the carbonated beverages, carbonated soft beverages with fine foam quality and good clean flavor were obtained by adding the complexes of Examples.

Application Example 3

Alcoholic Effervescent Beverage

[0097] For the water-soluble pea polysaccharides A and B, the polysaccharide-protein complexes A to F, and the composition A, the foam stability was measured by the method described below assuming an alcoholic effervescent beverage. The results are shown in Table 4.

[0098] Into a 100 ml Nessler color comparison tube, 49 ml of a commercially available beer-like beverage produced without using malt, which is categorized as other brewages (effervescent) (1) was put and 1 ml of 10% aqueous solution of polysaccharide, complex or composition was added, and then they were mixed gently in a manner which avoids foaming. Using an ultrasonic foaming apparatus (Sonic Hour: manufactured by TAKARA TOMY A.R.T.S), an ultrasonic pulse was applied five times from the bottom of the color comparison tube for foaming. The thickness of the foam layer being after 2 minutes from the foaming was measured, and the criteria of foam stability are as follows: when the thickness is 60 mm or more, this is expressed by ; when the thickness is 40 mm or more and less than 60 mm, this is expressed by ; when the thickness is 20 mm or more and less than 40 mm, this is expressed by ; and when the thickness is less than 20 mm, this is expressed by . When the evaluation is or , this is rated as acceptable. As a comparison, a sample prepared without adding anything was evaluated under the same conditions (Comparative Example 7).

[0099] Organoleptic evaluation was performed for each beverage, and the foam quality (fineness of foam) and the flavor were evaluated according to the following criteria. The foam quality was evaluated on the basis of visual observation and mouthfeel at tasting, and the flavor was evaluated by tasting.

(Foam Quality)

[0100] : Very fine [0101] : Moderately fine [0102] : Coarse

(Flavor)

[0103] : Very good [0104] : Good [0105] : Bad

TABLE-US-00004 TABLE 4 Foam stability, foam quality and flavor in alcoholic effervescent beverage Type of Foam Foam protein stability quality Flavor Comparative Polysaccharide A X Example 1 Comparative Polysaccharide B X X Example 2 Example 1 Complex A Soybean Example 2 Complex B Casein Example 3 Complex C Wheat Example 4 Complex D Pea Example 5 Complex E Albumen Example 6 Complex F Soybean Comparative Composition A Soybean X X Example 3 Comparative Not added X X Example 7

[0106] Foam stability, foam quality, and flavor of the effervescent beverages containing alcohol of the individual Examples and Comparative Examples exhibited almost the same tendency as those of Application Example 2. Astringency peculiar to a soybean protein was felt a little in the composition A which was not heated. Meanwhile, flavor was improved in the complexes A to F which were heated.