KR102409571B1 - Method of manufacturing sweet flag vinegar using the barley nuruk and the sweet flag vinegar manufactured thereby - Google Patents

Abstract

The present invention relates to a method for producing iris vinegar prepared from barley koji and to iris vinegar prepared thereby, wherein the yeast is prepared by inoculating grains steamed with Rhizopus oryzae 82-7 spawn, the yeast, After soaked and steamed barley, soaked water and Saccharomyces cerevisiae ( Saccharomyces cerevisiae ) YM33 yeast seed is mixed and fermented to prepare barley wine ( Acetobacter pasteurianus ) A11-2 Calamus vinegar prepared by mixing seed vinegar prepared with acetic acid bacteria and fermenting it with acetic acid has a high total phenol content and has excellent antioxidant and antibacterial activity.

Description

Method of manufacturing sweet flag vinegar using the barley nuruk and the sweet flag vinegar manufactured thereby

The present invention relates to a method for producing iris vinegar prepared from barley yeast and to iris vinegar produced thereby.

Vinegar, a fermented food that has been used for a long time, is not only used as a seasoning during cooking, but also variously used as a food preservative and medicine. Vinegar is manufactured by fermenting grains, fruits, and alcoholic beverages rich in sugar or starch as main raw materials with microorganisms. It contains various organic acids, sugars, amino acids, esters, etc. Vinegar has a very strong sterilizing power, so it is widely used in processed foods, etc., and has a stress-relieving effect. Recently, a new functional fermented vinegar market such as fermented vinegar for health using local agricultural products is being formed, and it is also being developed in the form of beverages for the purpose of dieting, skin care, prevention of adult diseases, and recovery from fatigue.

In general, fermented vinegar is made by proliferating acetic acid bacteria, an aerobic bacillus that oxidizes alcohol to produce acetic acid, in the fermentation substrate. At this time, since alcohol as a fermentation source is essential for the growth of acetic acid bacteria, alcohol fermentation for the production of alcohol is preceded for the production of traditional vinegar. That is, it is manufactured through a two-step fermentation process in which sugar or starch-containing fruits and grains are used as raw materials, sugar is fermented into alcohol by yeast, and alcohol is oxidized to acetic acid by acetic acid bacteria.

On the other hand, irises are perennial plants belonging to the Araceae family, and there are two types of irises: Acorus calamus L. and Soland.臭菖), it is called water chandelier (水菖蒲). Commonly known as water iris, iris grows well in wetlands and grows gregarious near ponds and lakes, and also grows wild in Hubei, Honam, and Sacheon in China, Jeju Island, Hwanghae, Pyeongbuk, Hamnam, Gyeongbuk, and Daegu in Korea. The rhizomes of Korean iris are collected and dried, and are used for the purpose of directional dryness, insulation, and aging prevention. If you make a powder from the underground root and apply it to boils, it is effective to suck out the juices. Bathing in the iris bath on the Dano Festival is used in the private sector to eliminate morale. In Donguibogam, there is a record that mixing the root juice of the iris with glutinous rice to make alcohol and mourning it for mourning. have.

As a prior document related to such an iris, Korea Patent Application Laid-Open No. 10-2003-0066449 has excellent effects for skin whitening, skin wound healing and improvement, or skin aging prevention, and is used as a cosmetic raw material that is easy to manufacture and excellent in stability. Disclosed is a method and composition for preparing an iris extract, and Korean Patent Publication No. 10-2013-0013693 discloses an external composition for preventing and improving atopic dermatitis comprising an iris extract as an active ingredient. However, there is no disclosure about vinegar prepared using iris.

Accordingly, the present inventors have studied the method of preparing vinegar by selecting iris among medicinal materials through an investigation of the ancient literature. As a result, it is possible to prepare iris vinegar with a high content of total phenolic components and excellent antioxidant and antibacterial activity. The present invention was completed by establishing the conditions.

It is an object of the present invention to provide a method for preparing iris vinegar having a high total phenol content and excellent antioxidant and antibacterial activity, and iris vinegar prepared thereby.

In order to achieve the above object, the present invention

1) Rhizopus oryzae ( Rhizopus oryzae ) 82-7 step of inoculating the steamed grains to prepare yeast; 2) Mixing the yeast of step 1), soaked and steamed barley, soaked water and Saccharomyces cerevisiae YM33 yeast seed, and fermenting it to prepare barley wine; And 3) the barley wine of step 2), iris powder, Acetobacter pasteurianus ( Acetobacter pasteurianus ) It provides a method for producing iris vinegar comprising the step of mixing and fermenting the seed seed prepared using A11-2 acetic acid bacteria. .

In addition, the present invention provides iris vinegar prepared by the above method.

The present invention relates to a method for producing iris vinegar prepared from barley koji and to iris vinegar prepared thereby, wherein the yeast is prepared by inoculating grains steamed with Rhizopus oryzae 82-7 spawn, the yeast, After soaked and steamed barley, soaked water and Saccharomyces cerevisiae ( Saccharomyces cerevisiae ) YM33 yeast seed is mixed and fermented to prepare barley wine ( Acetobacter pasteurianus ) A11-2 Calamus vinegar prepared by mixing seed vinegar prepared with acetic acid bacteria and fermenting it with acetic acid has a high total phenol content and has excellent antioxidant and antibacterial activity.

1 is a view showing the appearance according to the fermentation period of yeast prepared with Aspergillus niger ( Aspergillus niger ) 72-1 strain and Rhizopus oryzae 82-7 strain.
Figure 2 is inoculated with Aspergillus niger 72-1 strain and Rhizopus oryzae 82-7 strain in rice grown in the form of entry and cultured, followed by fermentation period (Day 0, 1 The graph is the result of measuring α-amylase activity (A) and gluco-amylase activity according to the first, third, and fifth days).
Figure 3 is a form of entry into rice, barley and brown rice, Aspergillus niger ( Aspergillus niger ) 72-1 strain and Rhizopus oryzae ) After inoculating and culturing the 82-7 strain, the fermentation period (day 0) , 1st day, 3rd day, 5th day) is a graph comparing the α-amylase activity (A) and gluco-amylase activity.
Figure 4 is a control, Saccharomyces cerevisiae ( Saccharomyces cerevisiae ) without the addition of yeast to investigate the alcohol fermentation characteristics according to yeast It is a graph comparing the sugar content (A), alcohol content (B) and production yield (C) of barley wine prepared with YM30 or YM33 strains.
Figure 5 is Saccharomyces cerevisiae ( Saccharomyces cerevisiae ) It is a graph showing the sugar content (A), fermentation temperature, and alcohol content (B) by fermentation period of barley wine prepared with YM33 strain by fermentation temperature and fermentation period.
Figure 6 is Acetobacter pasteurianus ( Acetobacter pasteurianus ) pH (A), acidity (B) and alcohol content (C) ) is a graph of the results of comparison.
Figure 7 is Acetobacter pasteurianus ( Acetobacter pasteurianus ) pH (A), acidity (B) and alcohol content (C) by fermentation temperature and fermentation period of calamus vinegar prepared using A11-2 acetic acid bacteria added is a graph of the results of comparison.

Hereinafter, the present invention will be described in detail.

the present invention

1) Rhizopus oryzae ( Rhizopus oryzae ) 82-7 step of inoculating the steamed grains to prepare yeast; 2) Mixing the yeast of step 1), soaked and steamed barley, soaked water and Saccharomyces cerevisiae YM33 yeast seed, and fermenting it to prepare barley wine; And 3) the barley wine of step 2), iris powder, Acetobacter pasteurianus ( Acetobacter pasteurianus ) It provides a method for producing iris vinegar comprising the step of mixing and fermenting the seed seed prepared using A11-2 acetic acid bacteria. .

In the step of producing barley liquor in step 2), white rice (rice flour) is used together to grow barley liquor together with yeast, and a method for making barley liquor using barley together with yeast is disclosed only at the station of the station. In the case of barley alcohol, the fermentation period is shorter and the flavor is improved.

In the manufacturing method, the iris of step 3 may be used without limitation, such as cultivated or commercially available irises, and the iris may be at least one selected from the group consisting of leaves, flower roots, stems, branches, peels and seeds, , for example, may be a root (rhizome).

The grain in step 1) may be any one of barley, rice or brown rice, preferably barley.

The Rhizopus oryzae 82-7 spawn of step 1) is 0.01 to 10%, 0.01 to 5%, 0.01 to 1%, 0.01 to 0.5%, 0.05 to 10%, 0.05 to godubap in the steamed grain. to 5%, 0.05 to 1%, or 0.05 to 0.5%.

Fermentation in the step of preparing barley wine in step 2) may be carried out at 21 to 35 °C, 21 to 29 °C, 23 to 27 °C, or 24 to 26 °C, preferably at 25 °C.

Fermentation in the step of preparing barley wine in step 2) may be performed for 5 to 30 days, 5 to 20 days, 5 to 15 days, 7 to 11 days, or 8 to 10 days, preferably for 9 days. can be done

The fermentation of step 3) may be made at 24 to 36 °C, 25 to 35 °C, 27 to 33 °C, or 29 to 31 °C, preferably at 30 °C.

The fermentation of step 3) may be carried out for 11 to 20 days, 12 to 19 days, 13 to 18 days, or 14 to 17 days, preferably for 15 days.

Calamus vinegar prepared by the above method has a high total phenol content, excellent antioxidant activity, and excellent antibacterial activity against pathogenic bacteria.

The pathogenic bacteria may be Gram-negative bacteria, preferably Escherichia coli or Salmonella.

In a specific embodiment of the present invention, the present inventors selected Rhizopus oryzae 82-7 seed as a yeast fungus for the production of iris vinegar (see FIGS. 1 and 2), confirming that barley is suitable as a fermenting agent (see FIG. 3).

In addition, the present inventors selected Saccharomyces cerevisiae YM33 yeast starter as a yeast starter for the production of iris vinegar, and derived the optimal fermentation temperature and fermentation time in the process of producing barley wine (Fig. 4 and 5).

In addition, the present inventors selected Acetobacter pasteurianus A11-2 acetic acid bacteria as the acetic acid bacteria for the production of iris vinegar, and derived the optimal fermentation temperature and fermentation time in the process of preparing iris vinegar using this. (See FIGS. 6 and 7 ), as a result of evaluating the functionality of calamus vinegar according to the presence or absence of seeds prepared with selected yeast, yeast and acetic acid bacteria, Rhizopus oryzae ) Yeast produced with 82-7 seeds, saka Calamus vinegar prepared by using Saccharomyces cerevisiae YM33 yeast seed, Acetobacter pasteurianus A11-2 acetic acid bacteria had high total phenol content, excellent DPPH radical scavenging ability, and gram It was confirmed that the antibacterial activity against negative bacteria was excellent (see FIGS. 8 and 9).

In addition, the present invention provides iris vinegar prepared by the above manufacturing method.

The method for producing iris vinegar may have the same characteristics as described above. In one example, the method comprises the steps of 1) preparing yeast by inoculating grains that have been steamed with Rhizopus oryzae 82-7 spawn; 2) Mixing the yeast of step 1), soaked and steamed barley, soaked water and Saccharomyces cerevisiae YM33 yeast seed, and fermenting it to prepare barley wine; and 3) mixing and fermenting the seed seed prepared by using the barley wine of step 2), iris powder, and Acetobacter pasteurianus A11-2 acetic acid bacteria. .

The calamus vinegar has a high total phenol content, excellent DPPH radical scavenging ability, and excellent antibacterial activity against Gram-negative bacteria.

Hereinafter, the present invention will be described in detail by way of experimental examples.

However, the following experimental examples only illustrate the present invention, and the content of the present invention is not limited by the following experimental examples.

<Experimental Example 1> Selection of yeast bacteria for the production of iris vinegar

In order to select yeast bacteria suitable for the yeast production method, which is the first step in the production of iris vinegar, Aspergillus niger ( Aspergillus niger ) 72-1 or Rhizopus Duck jae ( Rizopus oryzae ) 82-7 strains were distributed from the Korean Agricultural Culture Collection (KACC) (Table 1), and the following experiments were performed using them.

strain name scientific name of the strain KACC accession number 72-1 Aspergillus niger 93335P 82-7 Rizopus oryzae 46960

1-1. Comparison of enzyme activity according to fermentation period of each rice koji prepared with 2 types of yeast bacteria

In order to examine the difference in enzyme activity according to the two types of yeast, Aspergillus niger (Potato Dextrose Agar) grown on 5% bran medium.Aspergillus niger) 72-1 or Rhizopus duck material (Rizopus oryzae) Inoculate 82-7 strains (10⁶, 1%) and cultured for 5 days, then inoculated 10% each into soaked and steamed rice, fermented at 35°C for 0, 1, 3, and 5 days, and then used to measure enzyme activity.

For enzyme activity measurement, 5 g of sodium chloride was dissolved in 10 g of the yeast sample prepared above in distilled water, 50 mL of 10 mM acetate buffer was added, and 50 mL of sodium chloride buffer made to 1 L with distilled water was added, and stirred at room temperature for 3 hours. and leached. Then, it was filtered, and the filtrate was used as a crude enzyme solution to quantitatively analyze the enzymatic activity of α-amylase and glucoamylase, which are involved in starch decomposition. Enzyme activity is an enzyme kit for analysis of alcoholic beverages produced by Japan's Kigo Network. Each enzyme (α-amylase, glucoamylase, Kikkoman, Nod, Japan) solution was measured at 400 nm using a spectrophotometer (UV-2450, Shimadzu, JP). It was measured quantitatively.

Alpha-amylase (α-amylase) activity was obtained from 2-chloro-4-nitrophenyl-6-azide-6-deoxy-β-maltopentaside (N3-G5-β-CNP) at 1 μmole of 2-chloro Formation of -4-nitrophenyl was defined as 1 U/g.

Yeast (U/g) = (Es-Eb) × 0.179 × Df × extraction rate

Es: 10 minutes enzymatic reaction, absorbance of the sample

Eb: Absorbance of blank before enzymatic reaction

Df: sample dilution factor

Gluco-amylase activity was defined as 1 U/g of 1-μm 4-nitrophenol (PNP) produced from 4-nitrophenyl-β-D-glucoside during enzymatic reaction at 37°C.

Yeast (U/g) = (Es-Eb) × 0.171 × Df

Es: 10 minutes enzymatic reaction, absorbance of the sample

Eb: Absorbance of blank before enzymatic reaction

Df: sample dilution factor

As a result of inoculating and culturing two kinds of yeast bacteria in rice steamed in the form of entry, as shown in FIG. 1, Aspergillus niger 72-1 yeast bacteria are brown yeast, whereas rhizopus Duck jae ( Rizopus oryzae ) 82-7 yeast was confirmed to represent a black color (Fig. 1).

As a result of comparing the enzyme activity according to the fermentation period of each rice koji prepared with two types of yeast, as shown in FIG. 2, in the case of alpha-amylase activity, Aspergillus niger 72-1 yeast was While activity increased by day 5, rhizopus Duck jae ( Rizopus oryzae ) 82-7 yeast increased until the 3rd day, then decreased on the 5th day (FIG. 2A). In the case of gluco-amylase activity, the highest activity was shown on the 3rd day, whereas both yeast bacteria were not detected on the 5th day. .

1-2. Comparison of enzyme activity according to fermentation period of each yeast prepared with two types of yeast in various fermenting agents

In order to compare the enzyme activity according to the fermentation period of each yeast prepared with two types of yeast in various fermenting agents Aspergillus niger grown on potato glucose agar medium (Potato Dextrose Agar) in 5% bran medium ( Aspergillus niger ) 72-1 or Rhizopus Duck jae ( Rizopus oryzae ) After inoculating the strain 82-7 (10 ⁶ , 1%) and culturing for 5 days, 10% each was inoculated into soaked rice, barley and brown rice, and fermented at 35° C. for 0, 1 and 3 days <Experimental Example 1- 1> was used to measure the enzyme activity of alpha-amylase and glucoamylase in the same way.

As a result of inoculating and culturing koji bacteria in various fermenting agents in the form of entry, as shown in FIG. 3 , in the case of alpha-amylase activity, both koji bacteria showed high activity on the third day of fermentation in the order of barley, brown rice, and rice. Middle Rhizopus Duck material ( Rizopus oryzae ) showed the highest activity when inoculated with barley 82-7 yeast bacteria and fermented, which was 73 compared to the test group inoculated with Aspergillus niger 72-1 yeast bacteria on barley % It was confirmed that high activity was shown (FIG. 3A). In the case of gluco-amylase activity, 72-1 koji bacteria showed high activity in the order of brown rice, rice, and barley, and 82-7 koji bacteria showed high activity in the order of rice, barley, and brown rice. , showed the highest activity, which was 376% higher than that of the test group inoculated with 82-7 yeast on brown rice (FIG. 3B). In the case of calamus vinegar, rhizopus has the highest alpha-amylase activity because barley is used to make alcohol. Duck jae ( Rizopus oryzae ) 82-7 was selected as a strain suitable for the yeast production method for producing iris vinegar.

<Experimental Example 2> Selection of yeast bacteria for production of iris vinegar

In the calamus vinegar manufacturing method, in order to select a suitable yeast group in the barley wine manufacturing step, Saccharomyces cerevisiae ( Saccharomyces cerevisiae ) The following experiments were performed using YM30 or YM33 yeast (Table 2).

strain name scientific name of the strain KACC accession number YM30 Saccharomyces cerevisiae doesn't exist YM33 Saccharomyces cerevisiae 48334

Specifically, sterilized water 128 mL to 80 g of steamed rice after soaking for the production of shumo (160%), and Saccharomyces cerevisiae ( Saccharomyces cerevisiae ) YM30 or YM33 yeast was inoculated and cultured at 30°C for 3 days by adding a yeast culture solution (5 mL, 10 ⁸ CFU/g, 6.25%) grown for 1 day. Then, to make barley liquor, soaked and steamed barley, boiled and cooled water, koji and shumo were mixed well. This was fermented at 25° C. for 6 days, centrifuged (8,000 rpm, 5 minutes), and the filtrate filtered through filter paper (No. 2) was analyzed by the following method.

For the sugar content analysis, a certain amount of the filtrate was measured at room temperature using a refractometer (Pocket Refractometer PAL-1, ATAGO Co.).

For alcohol analysis, 70 mL of the distillate was received using an alcohol distiller, 100 mL of distilled water was used, and then measured using a density meter (DMA 5000M, Anton Paar, Wundshuh, Austria).

In order to confirm the production yield, the amount of the filtered filtrate by centrifugation was measured and expressed as a percentage using Equation 1 below.

[Formula 1]

As a result, as shown in FIG. 4 , there was no change in sugar content depending on the fungal spawn, and the fermentation using YM30 showed a higher sugar content than fermentation using YM33 according to yeast ( FIG. 4A ). As for the alcohol content, alcohol was not produced in the experimental group inoculated with yeast inoculated with yeast, and in the case of the yeast-added experimental group, when YM33 yeast was used, the alcohol content was 10% or more, 41% higher than in the yeast-free group high levels (Fig. 4B). As for the production yield, the production yield of the experimental group using YM33 yeast was 10% higher than the yeast-free group ( FIG. 4C ). Through the above results, it was confirmed that YM33 produced a lot of alcohol by using sugar as a fermentation substrate.

<Experimental Example 3> Establishment of optimal fermentation temperature and fermentation period of the selected yeast

Saccharomyces cerevisiae selected in the <Experimental Example 2> In order to establish the optimum fermentation temperature and fermentation period of YM33 yeast, the following experiments were performed.

Specifically, to 80 g of steamed rice after soaking for the production of main hair, add 128 mL of sterile water (160%), inoculate yeast, and add yeast culture (5 mL, 10 ⁸ CFU/g, 6.25%) grown for 1 day. Incubated at 30°C for 3 days. Then, to prepare barley liquor, soaked and steamed barley, boiled and cooled water, koji, and barley were mixed well, fermented at 20, 25, and 30℃ for 0, 3, 6, and 9 days, respectively, and centrifuged (8,000 rpm, 5 minutes) and filtered with filter paper (No. 2). In order to confirm the fermentation temperature and optimum conditions for the fermentation period, sugar content and alcohol content were measured in the same manner as in <Experimental Example 2> using the filtered filtrate.

Saccharomyces cerevisiae Fragrance components according to the fermentation temperature of barley wine prepared using YM33 yeast were measured using GC-2010 Plus and GCMS-TQ 8030 (Shimazu, Tokyo, Japan). Put 500uL of vinegar stock solution 4 times diluted with distilled water, 1g of NaCl, and 1.5ml of tertiary distilled water (including IS) in a 20ml vial, stirred at 25℃ for 30 minutes, and then adsorbed to SPME for 5 minutes. injected. GC Column (DB-WAX column, 30m × 0.25 mm id, 0.25 um film thickness; J & W scientific, Santa Clara, Calif., USA), Injection Temp (250 ℃), Column flow (1 mL/min), Carrier Under the condition of gas (Helium), the oven temperature was maintained at 40 °C for 3 minutes, then raised to 90 °C at 5 °C per minute, then raised to 230 °C at 19 °C per minute and maintained for 5 minutes. MS conditions were performed under the conditions of Ion source Temp (230 ℃), Interface Temp (280 ℃), Detector voltage (0.1 kV), and Event time (0.03s, 15ev) in Q3 scan mode, and the results are shown in Table 3 below. indicated.

Compound Barley wine 20℃ Barley wine 25℃ Barley Sake 30℃ Acetic acid, ethyl ester 3.39 3.41 1.99 Ethanol 10.72 12.22 11.36 2-Methyl-1-propanol 1.09 1.29 0.77 3-methyl-1-Butanol, acetate 0.27 0.43 0.17 3-Methyl-1-butanol 7.73 8.72 5.75 4-Methyl-2-heptanone 0.07 0.10 0.07 l-Limonene 0.09 0.29 0.21 Octanal 0.07 0.16 0.07 2-Methyl-2-octanol 0.11 0.12 0.13 Nonanal 0.49 1.22 0.49 2-Ethylhexanol 2.13 1.68 2.03 Decanal 0.22 0.14 0.15 Methyl salicylate 3.64 5.10 5.00

To select the optimal fermentation temperature and period of the selected yeast YM33, iris was prepared, and as a result of quality analysis thereof, as shown in Table 3 and FIG. 5, the sugar content increased as fermentation progressed, and relatively from the 3rd day of fermentation at 30°C. high, and showed the highest sugar content on the 9th day of fermentation (FIG. 5A). The alcohol content, like the sugar content, increased as the fermentation progressed, and when fermented at 25 and 30 °C for 9 days, the alcohol content was relatively higher than when fermented at 20 °C (FIG. 5B).

Saccharomyces cerevisiae As for the aroma component according to the fermentation temperature of barley wine manufactured using YM33 yeast, ethanol, the main component of alcoholic fermentation, was 10.72~12.23%, the highest when fermented at 25℃, and 2-Methyl-1 with a sweet flavor -propanol, it was confirmed that the highest content of Octanal with fruit flavor and Nonanal with rose orange flavor was fermented at 25℃ (Table 3).

Through the above results, a fermentation temperature of 25° C. and a fermentation period of 9 days were established as the optimal conditions for alcoholic fermentation for the production of iris vinegar.

<Experimental Example 4> Confirmation of the effect of acetic acid bacteria for the production of iris vinegar

Acetobacter pasteurianus ( Acetobacter pasteurianus ) A11-2 In order to evaluate the quality of iris vinegar according to the presence or absence of a spawn prepared with acetic acid bacteria, Acetobacter pasteurianus A11-2 deposited under the accession number KACC 92203P was synthesized. Seed was prepared by inoculating the medium and culturing at 30° C. for 3 days, then increasing the amount by 10 times step-by-step for 5 days in Cheongju (6-8% alcohol) and culturing 3 times. The pH, total acidity, and alcohol content were compared as follows using the prepared sample to which seedling was added and the sample to which seed was not added.

The pH was measured using a pH meter (Orion 3 star, Thermo scientific Co.) of the iris vinegar sample solution. The alcohol content was measured by using a density meter (DMA 5000M, Anton Paar, Wundshuh, Austria) after receiving 70 mL of the distillate using an alcohol distiller, distilling it into 100 mL of distilled water. The total acidity was obtained by neutralizing titration (pH 8.3) using 0.1 N NaOH by taking 1 mL of calamus vinegar sample solution, and converting the amount of the consumed solution into acetic acid (%) using Equation 2 below.

[Formula 2]

Organic acid content in food = titration mL × NaOH titer × acetic acid (0.006) × dilution factor × 100/S (g)

As a result of evaluating the quality of the prepared samples with and without seedlings, as shown in FIG. 6 , the pH of the samples with seedlings was lower than that of the control without seedlings, and fermentation 10 days From tea, it showed a tendency to increase as the fermentation progressed (FIG. 6A). The total acidity of the sample with the addition of seedling was high, which was 118% higher than that of the control without seedling. The sample showed a decreasing trend (Fig. 6B). Through this, it can be seen that, in the case of the sample to which the seed seed was added, over-fermentation in which the acetic acid bacteria decomposes the acetic acid produced after all the alcohol components, which are the fermentation substrates, were used. The alcohol content was not detected on the 10th day of fermentation in the sample with the addition of seedling, but in the case of the control without seedling, it was detected on the 20th day of fermentation, confirming that the fermentation proceeded slowly (Fig. 6C), and higher on the 20th day of fermentation compared to the 15th day of fermentation By showing the alcohol content, it was confirmed that alcoholic fermentation proceeds due to grains and yeast added during vinegar production.

According to the above results, Calamus vinegar was selected to be manufactured using the steps of entry using 82-7 yeast, alcohol fermentation using YM33 yeast, and acetic acid fermentation using A11-2 acetic acid bacteria.

<Experimental Example 5> Establishment of optimal fermentation temperature and fermentation period of acetic acid bacteria

Acetobacter pasteurianus ( Acetobacter pasteurianus ) In order to establish the optimum fermentation temperature and fermentation period of A11-2 acetic acid bacteria, the following experiments were performed.

Specifically, after washing 200 g of rice, soaking it in water to save water, steaming the pulverized rice flour in the form of beksulgi, 300 g of iris root powder, 500 g of yeast, 1 L of barley liquor, 200 mL of seed vinegar prepared in <Experimental Example 4> (10%) After mixing, fermented for 0, 5, 10, and 15 days at 30 and 37 ° C., and filtered to prepare iris vinegar. Iris vinegar without acetic acid bacteria was prepared by mixing 100 g of iris root powder, 800 g of barley wine, and 100 mL of seed vinegar, fermenting it at 30°C for 0, 5, 10, and 15 days, and then filtering it. pH, total acidity, and alcohol content were analyzed as follows using each prepared iris vinegar, and fragrance components according to the starting temperature of iris vinegar prepared using acetic acid bacteria were analyzed.

Fragrance components according to the fermentation temperature of iris vinegar prepared using acetic acid bacteria were measured using GC-2010 Plus, GCMS-TQ 8030 (Shimazu, Tokyo, Japan). Put 500uL of iris vinegar stock solution 4 times diluted with distilled water, 1g of NaCl, and 1.5ml of tertiary distilled water (including IS) into a 20ml vial, stir at 25℃ for 30 minutes, and then adsorb to SPME for 5 minutes was injected. GC Column (DB-WAX column, 30m × 0.25 mm id, 0.25 um film thickness; J & W scientific, Santa Clara, Calif., USA), Injection Temp (250 ℃), Column flow (1 mL/min), Carrier Under the condition of gas (Helium), the oven temperature was maintained at 40 °C for 3 minutes, then raised to 90 °C at 5 °C per minute, and then again raised to 230 °C at 19 °C per minute and maintained for 5 minutes. MS conditions were conducted under the conditions of Ion source Temp (230 ℃), Interface Temp (280 ℃), Detector voltage (0.1 kV), and Event time (0.03s, 15ev) in Q3 scan mode, and the results are shown in Table 4 below. indicated.

compound solid fermentation liquid fermentation iris 30℃ iris 37℃ iris 30℃ 0 days 10 days 15th 0 days 10 days 15th 0 days 10 days 15th Acetic acid, ethyl ester 0.60 1.26 2.43 0.79 2.94 4.64 1.01 27.15 5.00 Ethanol 2.80 0.00 0.51 2.93 0.19 0.16 4.06 1.06 0.21 2-Methyl-1-propanol 0.20 0.00 0.03 0.25 0.00 0.03 0.34 0.20 0.09 3-methyl-1-Butanol, acetate 0.00 0.00 0.07 0.00 0.00 0.05 0.00 0.83 0.21 3-Methyl-1-butanol 1.62 0.39 0.28 1.57 0.34 0.23 2.44 1.66 0.89 l-Limonene 0.46 0.40 0.21 0.25 0.30 0.27 0.16 0.31 0.36 Octanal 0.22 0.15 0.17 0.14 0.19 0.13 0.21 0.17 0.36 Nonanal 1.09 0.72 0.86 0.54 0.74 0.51 0.95 0.92 1.43 2-Ethylhexanol 1.41 1.93 1.92 2.35 1.92 2.12 1.91 1.94 1.94 Decanal 0.28 0.19 0.38 0.36 0.31 0.16 0.52 0.27 0.09 Methyl salicylate 5.81 9.48 7.91 11.30 10.74 6.41 10.45 3.89 7.16

Acetobacter pasteurianus ( Acetobacter pasteurianus ) Experimental results for establishing the optimum fermentation temperature and fermentation period of A11-2 acetic acid bacteria As shown in Table 4 and FIG. 7 , the pH decreased as fermentation progressed, and on the 5th day of fermentation After decreasing from 4.34 to 3.58 to 3.79, the decrease was small, and there was no significant difference between the experimental groups on the 15th day of fermentation (FIG. 7A). In the case of total acidity, it increased rapidly on the 5th day of fermentation and increased or maintained as fermentation progressed. increased to show an acidity of 4% or more. In addition, in the case of vinegar fermented at 37° C., the acidity was 4.23%, which was lower than that of vinegar fermented at 30° C. (5.25 to 5.53%) ( FIG. 7B ). In the case of alcohol content, it was found to be non-detectable on the 5th day of fermentation, and it was observed that the alcohol content was not overfermented and the acidity continued to increase compared to the alcohol content consumed quickly. It is presumed that this is because it does not In addition, in the case of the experimental group prepared except for the solid, it was not detected on the 10th day of fermentation, which is considered to be due to the difference in the initial alcohol content (FIG. 7C). In the case of aroma components according to the fermentation temperature of calamus vinegar prepared using acetic acid bacteria, acetic acid showed the highest content on the 15th day of fermentation in all experimental groups, and ethanol showed a tendency to decrease. In particular, in the iris vinegar experimental group fermented at 30°C for 15 days, acetic acid was the highest at 5%, and the contents of Octanal and Decanal, which had a positive flavor, were the highest (Table 4).

According to the above results, a fermentation temperature of 30° C. and a fermentation period of 15 days were selected as the acetic acid fermentation conditions for producing iris vinegar.

<Experimental Example 6> Functional evaluation of iris vinegar according to the presence or absence of seeds prepared with selected yeast, yeast and acetic acid bacteria

5-1. Comparison of total phenol content and DPPH radical scavenging ability of Calamus vinegar according to the presence or absence of seeds made with selected yeast, yeast and acetic acid bacteria and the period of acetic acid fermentation

In order to compare the total polyphenol content and DPPH radical scavenging ability of Calamus vinegar according to the presence or absence of seeds made with selected yeast, yeast and acetic acid bacteria and the period of acetic acid fermentation, Aspergillus niger as yeast bacteria 72-1 or Rhizopus Duck jae ( Rizopus oryzae ) 82-7 strain, yeast as Saccharomyces cerevisiae ( Saccharomyces cerevisiae ) YM33 strain or yeast-free, acetic acid bacterium Acetobacter pasteurianus ( Acetobacter pasteurianus ) According to the conditions of A11-2 strain, Calamus vinegar was prepared in a total of 8 experimental groups according to the conditions of the strain A11-2, and the following experiments were performed. .

For the total phenol content, 1 mL of Folin-Ciocalteu phenol reagent 2N was added to 250 μL of each calamus vinegar sample solution, and then left in the dark for 3 minutes. Then, 0.1N Na ₂ CO ₃ 2 mL was added and left in a dark room for 60 minutes, and absorbance was measured with a 765 nm spectrophotometer. A standard curve was prepared using tannic acid as a standard material and then converted. For the DPPH radical scavenging rate (%), 0.8 mL of 1.5 mM DPPH was added to 0.2 mL of each calamus vinegar sample solution diluted to an appropriate amount, left in the dark for 30 minutes, and absorbance was measured with a 517 nm spectrophotometer. The results of comparing the total polyphenol content and DPPH radical scavenging ability of iris vinegar on the 10th day of acetic acid fermentation are shown in Table 5, and the results of comparing the total polyphenol content and DPPH radical scavenging ability of iris vinegar on the 15th day of acetic acid fermentation are shown in Table 6 it was

Starter combination total polyphenols
(mg TAE/mL)
Antioxidant (%) DPPH 82-7+ YM33 +A11-2 770.78 ^a 88.07 ^ab 82-7+ YM33 + Seedling X 730.08 ^b 87.90 ^a 82-7+ Yeast X +A11-2 658.76 ^d 86.43 ^b 82-7+ Yeast X + Seedling X 689.88 ^c 86.25 ^b

Starter combination total polyphenols
(mg TAE/mL) Antioxidant (%) DPPH 82-7+ YM33 +A11-2 870.13a 87.31a 82-7+ YM33 + Seedling X 783.00c 85.31ab 82-7+ Yeast X +A11-2 758.59d 85.55ab 82-7+ Yeast X + Seedling X 786.38b 84.14b

As a result, as shown in Table 5, Table 6 and FIG. 8, the total phenol content of iris vinegar showed the highest value on the 15th day of fermentation, and the total phenol content of iris vinegar to which seed was added was relatively high. appeared (FIG. 8A). In the case of DPPH radical scavenging activity, the experimental group without seedling decreased as fermentation progressed, but the experimental group with seedling addition did not show a difference according to the period (FIG. 8B).

5-2. Comparison of antibacterial activity according to the presence or absence of seeds made with selected yeast, yeast and acetic acid bacteria and the period of acetic acid fermentation

In order to compare the antibacterial activity according to the presence or absence of seeds made with selected yeast, yeast and acetic acid bacteria and the period of acetic acid fermentation, Aspergillus niger as yeast bacteria ( Aspergillus niger ) 72-1 or Rhizopus Duck jae ( Rizopus oryzae ) 82-7 strain, yeast as Saccharomyces cerevisiae ( Saccharomyces cerevisiae ) According to the conditions of YM33 strain or yeast-free, acetic acid bacterium Acetobacter pasteurianus A11-2 strain or without seedling-free, iris vinegar was prepared in a total of 8 experimental groups, and the acetic acid fermentation period was 10 The following experiment was performed for one or 15 days.

Specifically, to measure antibacterial activity (mm), one colony was collected from a total of two pathogens, Escherichia coli KCTC 1309 and Salmonella typhimurium ATCC 41028, inoculated into 50 mL of TSB medium, and cultured overnight at 30°C to prepare a test bacterial solution. did. Then, 100 μL each of the test bacteria solution was put into the TSA medium and sprayed. Then, a paper disc was placed on the top, and 50 μL of calamus vinegar sample solution was dropped each, followed by incubation at 30° C. for 18 hours. Activity was assessed by measuring the inhibition zone. The results of comparing the antibacterial activity of iris vinegar on the 10th day of acetic acid fermentation are shown in Table 7 below, and the results of comparing the antibacterial activity of iris vinegar on the 15th day of acetic acid fermentation are shown in Table 8 below.

Starter combination antibacterial activity (clear zone, mm) coli Salmonella 82-7+ YM33 +A11-2 4 4 82-7+ YM33 + Seedling X 0 0 82-7+ Yeast X +A11-2 3 3 82-7+ Yeast X + Seedling X 2.5 2

Starter combination Antibacterial activity (%) coli Salmonella 82-7+ YM33 +A11-2 One 3 82-7+ YM33 + Seedling X One 0 82-7+ Yeast X +A11-2 2 0 82-7+ Yeast X + Seedling X One 2.5

As a result, as shown in Tables 7, 8, and 9, the iris vinegar experimental group with the addition of seed seed showed relatively high antibacterial activity compared to the iris vinegar experimental group without the addition of seed seed, and in the case of gram-negative bacteria, the 10th day of fermentation showed high antibacterial activity. In the case of Gram-positive bacteria, some experimental groups showed high antibacterial activity on the 10th day, but they did not show the same trend. There was no difference according to yeast and yeast bacteria (FIG. 10).

Agricultural Genetic Resource Center, National Academy of Agricultural Sciences KACC92203P 20171023

Claims (14)

1) preparing yeast by inoculating grains that have been steamed with Rhizopus oryzae 82-7 spawn deposited under the accession number KACC 46960;
2) The yeast of step 1), soaked and steamed barley, soaked water, and Saccharomyces cerevisiae YM33 yeast seed deposited under the accession number KACC 48334 are mixed, and the barley liquor is fermented. manufacturing a; and
3) A step of mixing and fermenting the seed seed prepared using iris powder, Acetobacter pasteurianus A11-2 acetic acid bacteria deposited under the accession number KACC 92203P with the barley wine of step 2).
A method of producing iris vinegar comprising a.
According to claim 1,
The method for producing iris vinegar, characterized in that the iris powder in step 3) is iris root powder.
According to claim 1,
The grain of step 1) is barley, rice, or brown rice, characterized in that any one, calamus vinegar manufacturing method.
4. The method of claim 3,
The grain of step 1) is barley, characterized in that, calamus vinegar manufacturing method.
According to claim 1,
The Rhizopus oryzae 82-7 spawn of step 1) is characterized in that it is inoculated at a concentration of 0.01 to 10w/w% in the steamed grain. A method for producing iris vinegar.
According to claim 1,
The fermentation of step 2) is a method for producing calamus vinegar, characterized in that it is made at 21 to 35 ℃.
7. The method of claim 6,
The fermentation is a method of producing iris vinegar, characterized in that made at 23 to 27 ℃.
According to claim 1,
The fermentation of step 2) is characterized in that it is made for 5 to 20 days, the method for producing iris vinegar.
9. The method of claim 8,
The fermentation method for producing calamus vinegar, characterized in that it is made for 7 to 11 days.
According to claim 1,
The fermentation of step 3) is characterized in that made at 24 to 36 ℃, Calamus vinegar production method.
According to claim 1,
The fermentation of step 3) is characterized in that it is made for 11 to 19 days, the method for producing calamus vinegar.
According to claim 1,
The method for producing iris vinegar, characterized in that the iris vinegar has a higher total phenol content than iris vinegar without the addition of seed vinegar.
According to claim 1,
The method for producing iris vinegar, characterized in that the iris vinegar has antibacterial activity against Gram-negative bacteria.
Calamus vinegar prepared by the method of claim 1.

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