KR100353886B1 - Manufacturing process of Cacao Polyphenol which has novel antioxidant activity by fermentation - Google Patents

Abstract

The invention cacao bean husk as fermentation relates to a process for the preparation of water, and more particularly, the superoxide (Superoxide, O ₂ ^-) generated by the cacao bean husk jaentin oxidase (Xanthine Oxidase) having antioxidant activity high scavenging ability for the By optimizing the fermentation conditions of cacao bean husks with higher antioxidant activity and establishing a cacao bean husk fermentation method, the development of a new natural antioxidant material, and recycling a huge amount of cacao beans It relates to a method for producing a cacao bean husk fermentation having an antioxidant activity can be improved economical.

Description

Manufacturing process of Cacao Polyphenol which has novel antioxidant activity by fermentation

The invention cacao bean husk as fermentation relates to a process for the preparation of water, and more particularly, the superoxide (Superoxide, O ₂ ^-) generated by the cacao bean husk jaentin oxidase (Xanthine Oxidase) having antioxidant activity high scavenging ability for the By optimizing the fermentation conditions of cacao bean husks with higher antioxidant activity and establishing a cacao bean husk fermentation method, the development of a new natural antioxidant material and recycling of a large amount of cacao beans It relates to a method for producing a cacao bean husk fermentation having an antioxidant activity that can be improved economic efficiency.

Cacao, used as a source of chocolate and cocoa, has the mystical meaning of the god food, Theobroma cacao L. The origin of cacao is estimated from Central America to the northern part of South America, and has been cultivated for a long time in BC and has been treated with corn as a very valuable food. The Azteca leaders of the 13th and 16th centuries in Central America reported that they took tonics with spices mixed with cocoa, which was made from ground cacao beans. Later, it became widespread in Europe, and finally, in 1875, a chocolate was developed by a young Swiss man. With this breakthrough invention, the production of cacao beans has increased dramatically, and the popularity of chocolate has increased all at once, and now it is becoming a representative product of the world regardless of age, sex, or east and west.

Cacao beans provided as a basic raw material of the chocolate is somewhat different depending on the origin, but is not only rich in dietary fiber, but also contains a large amount of polyphenols. Polyphenol component is a substance contained in a large amount of green tea and the like, caries suppression effect [Sakanaka, S., et al., Biosci & Biochem., 56, 592, 1992], hypertension suppression effect [Iwata, K., et al., Jpn. Soc. Nutr. Food Sci., 40, 469, 1987], prevents cardiovascular disease [Hertog, MGL, et al., Lancet, 342, 1007, 1993] and antioxidant activity [Ramar etnam, N., et al., Trends in Food Sci . Technol., 6, 75, 1945] have been found to have various physiological activities. Recently, polyphenols are extracted from plants containing a large amount of polyphenols such as green tea, oolong tea, apples, grapes, etc. It is used as a material for medicines.

On the other hand, cacao beans are shelled out during the production process of chocolate so that nib is used as a raw material of chocolate or cocoa, and shells (about 15% by weight of the total cacao beans) that are produced as by-products are almost discarded. In other words, the world's production of cacao beans is about 2.5 million tons, and about 400,000 tons of cocoa beans are discarded. According to the 1990 Customs Statistics Korea, about 600 tons of cacao beans It is known to be being discharged as waste. However, the cacao bean husk contains a large amount of dietary fibers such as cellulose and polyphenols compared to kernels, and a method for recycling them into resources has been devised.

Therefore, the present inventors use the cacao bean husk extract obtained as a by-product during the production of chocolate, the price is cheap, and the content of the cacao polyphenol is different depending on the manufacturing process [Graham, HN, Preventive Medicine, 21, 334-350 , 1992], Balentine, DA, et al., Critical Reviews in Food Science and Nutrition, 37, 693-704, 1997, Harvowy, M., et al., Critical Reviews in Plant Science, 16, 415- 480, 1997] is to fermented cacao bean husk fermented with high antioxidant activity by fermentation.

For the above reason, the present invention, in the preparation of the cacao bean husk fermented product, the study of the culture conditions, fermentation conditions, etc. of the cacao bean husk, as a result, an optimized medium that can increase the antioxidant activity of the cacao bean husk fermented product By finding out conditions and fermentation conditions, this invention was completed.

Accordingly, an object of the present invention is to provide a method for producing a cacao bean hull fermented product having an antioxidant activity that significantly improves the antioxidant activity and at the same time recycles an enormous amount of cacao bean husk.

The present invention provides a method for producing a cacao bean husk fermented from cacao bean husk,

Selected cacao beans with an average size of 2 mm or less 3-10 g, degreasing rice 3-5 g, soybean peptide 1-3 g, NaHPO ₄ 1-3 g, (NH4) ₂ HPO ₄ 0.1-1 g, Na ₂ CO ₃ 0.1-1 g Was dissolved in 100 ml of distilled water and sterilized at 120 ° C. for 10 minutes,

In the sterilized medium Bacillus rickenformis (Bacillus licheniformis,KCCM 10042) inoculated with 3 ~ 5 ㎖ of fermentation broth and incubated for 1 day at 25 ~ 40 ℃, and

When the incubation is terminated, heat-treated for 45 minutes at 98 ~ 99 ℃ and filtered using diatomaceous earth, cacao soybean hull filtrate is added to the cacao polyphenol of 1 ~ 3g prepared by the post-sterilization process at 95 ~ 97 ℃ It is characterized by a method of producing cacao bean husk fermented product having antioxidant activity.

Referring to the present invention in more detail as follows.

The present invention confirms that the cacao bean husk fermented product shows high scavenging ability against superoxide (O ₂ ⁻ ) produced by Xanthine Oxidase, thereby optimizing the fermentation conditions of cacao bean husk. The present invention relates to a method for producing cacao bean husk fermented product, which further enhances antioxidant activity.

Here, the xanthin oxidase enzyme reacts with xanthin to produce superoxide (O ₂ ⁻ ). The resulting superoxide (Superoxide, O ₂ ^-) are involved in the human body, such as aging or cancer.

The cacao bean husks used in the present invention show high scavenging ability against the excess oxide, which is produced by diformazan reaction at 725 nm with nitroblue-tetrazolium. It can confirm by the absorbance which shows a specific peak.

In particular, in the present invention, in order to enhance the antioxidant activity against the excess oxide, it is characterized in the preparation of cacao bean husk fermentation in a specific medium and fermentation conditions.

When explaining the cacao bean husk fermented product of the present invention having such an antioxidant activity in more detail.

As a first process, the present invention performs the sterilization of the medium containing cacao beans. In the medium composition according to the present invention, only the average size of the cacao bean husk used as the main raw material of chocolate or cocoa drink is 2 mm or less, and then the cacao bean husk 3-10 g, degreasing rice 3-5 g, soybean peptide 1-3 g, NaHPO ₄ A medium containing 1 to ₃ g, 0.1 to 1 g of (NH ₄ ) ₂ HPO _{4 and} 0.1 to 1 g of Na ₂ CO ₃ is characterized. Sterilization at 120 ° C. for 10 minutes is carried out with the above medium composition. At this time, if the content of the cacao bean husk in the medium is less than 3g, there is a problem that the antioxidant activity of the fermentation broth is weakened after the fermentation, if more than 10g, the workability is worsened, there is a difficulty in producing the product. In addition, the degreasing rice bran and soybean peptides are used to enhance the antioxidant activity, the degreasing rice bran is easy to oxidize when used in excess of 5g, there is a problem that the antioxidant activity is weakened when used less than 3g. In addition, the soybean peptide is expensive, the price becomes weaker when used in excess of 3g, the antioxidant activity is weakened when used in less than 1g.

Next to the above process, the present invention performs a process of obtaining a cacao bean husk fermentation broth by culturing a medium containing cacao bean husk.

The present invention is inoculated with 3 to 5 ml of Bacillus licheniformis ( KCCM 10042) fermentation broth to the medium, and then incubated for 1 day at a temperature of 25 to 40 ℃. In particular, in the present invention, the incubation temperature is very important. At this time, when the incubation temperature is less than 25 ℃ fermentation time is long, there is a problem that the price competitiveness is lowered, when the fermentation at 40 ℃ or higher high antioxidant No activity can be obtained.

Lastly, the present invention performs a process of preparing a cacao bean husk fermentation having an antioxidant activity by containing a predetermined amount of the cacao polyphenol component in the cultured cacao bean husk fermentation.

In the present invention, the cultured cacao bean husk fermentation broth is heat-treated at 98-99 ° C. for 45 minutes, and the culture solution is filtered using diatomaceous earth (120 #, 12% w / v). Then, 1 to 3 g of cacao polyphenol is added to the filtrate of the cacao bean husk fermentation broth, and then a cacao bean husk fermentation broth having an antioxidant activity can be obtained through a post sterilization process at 95 to 97 ° C. for 1 hour.

The scavenging activity ^on-cacao bean husk fermentation are the superoxide (Superoxide, O ₂₎ generated by it cocoa polyphenols is contained 10 to 15%, jaentin oxidase (Xanthin Oxidase) according to the invention as described above As 70 to 85%, it can be confirmed that it is excellent in antioxidant activity.

Hereinafter, the present invention will be described in more detail with reference to Examples, but the present invention is not limited by Examples.

Reference Example 1 Optimal Media Condition Selection Test

An optimal ratio selection test of cacao bean husks in the medium was carried out using the following method.

Among cacao beans, only those with an average size of 2 mm or less are selected and 3 to 10 g of cacao beans, 3 to 5 g of degreasing rice, 1 to 3 g of soybean peptides, 1 to 3 g of NaHPO ₄ , 0.1 to 1 g of (NH 4) ₂ HPO _4, and Na _2. The medium was prepared with a composition of 0.1 to 1 g of CO ₃ and then sterilized at 120 ° C. for 10 minutes.

On sterile medium Bacillus rickenformis (Bacillus licheniformis,KCCM 10042) The fermentation broth was inoculated with 3 to 5 ml, and then cultured at 25 to 40 ° C for 1 day. After the incubation is terminated, heat treatment is performed at 98-99 ° C. for 45 minutes, followed by filtration using diatomaceous earth. Then, 1 to 3 g of cacao polyphenol (prepared according to the manufacturing method of Korean Patent Application No. 99-25998) is added to the filtrate, and the cacao having antioxidant activity through a post sterilization process at 95 to 97 ° C. for 1 hour. Bean husk fermentation was obtained.

Then, the thus obtained cacao bean husk with superoxide generation by the jaentin oxidase (Xanthine Oxidase) as the test method to use the fermentation (Superoxide, O ₂ ^-) was measured scavenging activity and polyphenol contents, the result then Table 1 shows.

[Test Methods]

1) superoxide (Superoxide, O ₂ ^- scavenging activity measurement test on):

Superoxide (Superoxide, O ₂ ^-) scavenging activity measurement test was done using hwagwang sunyak Industry Co., Ltd. (Japan) products, SOD Test Kit for the Measurement are as follows:

Sample (檢體, S) are mixed and the sample 0.1 ㎖ and color development reagent 1.0 ㎖, blind (盲檢, B l) was added to distilled water and 0.1 ㎖ coloring reagent 1.0 ㎖. Sample blind (檢體盲檢, SB l ) are mixed and the sample and the blank solution 0.1 ㎖ 1.0 ㎖, reagent blind (試藥盲檢, B l -B l) was prepared with distilled water and mix 0.1 ㎖ and the blank solution 1.0 ㎖ . Specimens (S), blind (B l ), specimen blind (SB l ), reagent blind (B l -B l ) were reacted at 37 ° C. for 20 minutes, and after the reaction was completed, 560 nm was added by adding 2.0 ml of reaction stopper. Absorbance was measured at.

As a component of each reaction solution used in the test, a color developing reagent was prepared by mixing xanthin 0.40 mmole and NO ₂ -TB 0.24 mmole in 1 L of 0.1 M phosphate buffer solution (pH 8.0), and the blank solution was 0.1 M phosphate buffer. Solution (pH 8.0) was used. Xanthine Oxidase was used as the enzyme, and 69 mM SDS solution was used as the reaction stop solution.

In Equation 1 above:

Es is the absorbance of the sample, E_{B l} Silver blind absorbance, Es _{- B l} Is the absorbance of the specimen blind, E_{B l- B l} The absorbance of the silver reagent blind is shown.

2) Content measurement test of polyphenol:

The content of polyphenols contained in each sample was measured by slightly changing the AOAC method. First, 750 μl of distilled water was added to an Eppendorf tube, and then 100 μl of a sample solution diluted to an appropriate concentration was added thereto. 50 μl of Folin Denis reagent and 100 μl of saturated sodium carbonate solution were added thereto, mixed well with a Vortex mixer, and reacted at room temperature for 1 hour. After the reaction was completed, the absorbance at 725 nm was measured to determine the content of polyphenols against the previously prepared standard curve. At this time, the standard curve was prepared using a conventional method using Epicatechin (Epicatechin) as a standard material.

In Table 1, the more the content of cacao bean shells increase the content of polyphenol is increased, but the superoxide generated by jaentin oxidase (Superoxide, O ₂ ^-) scavenging for is found to be decreased. Therefore, in the present invention, the content of cacao bean husk was used in the range of 3 to 10 g.

Reference Example 2: Optimal Fermentation Temperature Selection Test

Using the optimum medium conditions selected in Reference Example 1, the fermentation temperature was prepared in the same manner as in Reference Example 1 except that the fermentation temperature was changed as shown in Table 2 below. Further, the reference example 1 and superoxide in the same manner (Superoxide, O ₂ ^-) was measured scavenging activity measurement test and polyphenol contents for the results are shown in Table 2 below.

From Table 2, it can be seen that the antioxidant activity according to the fermentation temperature of the medium is the most suitable 25 ~ 40 ℃.

Example 1

Only the average size of cacao beans is less than 2 mm, and cacao beans peel 7g, degreasing rice 4g, soybean peptide 4g, NaHPO₄2g, (NH4)₂HPO₄0.5 g and Na₂CO₃After mixing 0.5 g, distilled water was added to make the volume 100 mL to form a medium. After sterilization for 10 minutes at 120 ℃, the medium Bacillus rickenformis (Bacillus licheniformis,KCCM 10042) was inoculated and incubated at 25 to 40 ° C. for 1 day. After the incubation was terminated, heat treatment was performed at 98-99 ° C. for 45 minutes, followed by filtration using diatomaceous earth (120 #, 12% w / v). 2 g of cacao polyphenol was added to the filtrate to obtain a cacao bean husk fermented product having antioxidant activity through a post sterilization process at 95 to 97 ° C. for 1 hour.

Comparative Example 1

Oolong tea extract (trade name: Sun Oolong, Japan Suntory Co., Ltd.), which is a commercially available product, was used. Then, the superoxide (Superoxide, O ₂ ^-) generated by jaentin oxidase (Xanthine Oxidase) measured in the same manner as in Reference Example 1, the scavenging effect on the measured results are shown in Table 3 below.

Comparative Example 2

Bio-Col (Tongyang Fermentation, Japan), a commercially available product, was used. Then, the superoxide (Superoxide, O ₂ ^-) generated by jaentin oxidase (Xanthine Oxidase) measured in the same manner as in Reference Example 1, the scavenging effect on the measured results are shown in Table 3 below.

Comparative Example 3

Biozyme (Bioscience Research Institute, Japan), which is a commercially available product, was used, and superoxide (O ₂ ⁻ ) generated by Xanthine Oxidase was measured by the same measurement method as that of Reference Example 1. The scavenging ability was measured and the results are shown in Table 3 below.

As described above, the present invention fermented cacao bean husks with antioxidant activity to the optimum conditions to produce a cacao bean husk fermented to further enhance the antioxidant activity, to develop a new antioxidant material and at the same time a huge amount of cacao bean husks Can be recycled to improve economics.

Claims (3)

In the method for producing a cacao bean husk fermented from cacao bean husk, Selected cacao beans with average size of 2 mm or less 3-10 g, skim rice bran 3-5 g, soybean peptide 1-3 g, NaHPO ₄ 1-3 g, (NH4) ₂ HPO ₄ 0.1-1 g, Na ₂ CO ₃ 0.1-1 g Sterilize at 120 ° C for 10 minutes with a medium composition of In the sterilized medium Bacillus rickenformis (Bacillus licheniformis,KCCM 10042) inoculated with 3 ~ 5 ㎖ of fermentation broth and incubated for 1 day at 25 ~ 40 ℃, and When the incubation was terminated, heat-treated at 98-99 ° C. for 45 minutes and filtered using diatomaceous earth, followed by adding 1 to 3 g of cacao polyphenol to the cacao bean husk filtrate, followed by a post sterilization process at 95-97 ° C. Method for producing a cacao bean husk fermentation having an antioxidant activity, characterized in that. According to claim 1, wherein said cacao bean husk fermentation are the superoxide (Superoxide, O ₂ ^-) generated by jaentin oxidase (Xanthin Oxidase) having an antioxidant activity, characterized in that scavenging effect on this of 70 ~ 85% Method for preparing cacao bean husk fermented product. The method of claim 1, wherein the cacao bean husk fermented product has a polyphenol content of 10 to 15%, characterized in that the cacao bean husk fermented product having an antioxidant activity.