Method for preparing boxthorn leaf Kangpu tea with high antioxidant activity by using artificial fungicide

Abstract

The invention discloses a method for preparing boxthorn leaf Kangpu tea with high antioxidant activity by using an artificial microbial agent, and belongs to the field of food processing. The boxthorn leaf kangpu tea is prepared through fermentation of the artificially synthesized microbial agent, the fermentation method is easy to operate and short in fermentation time, the antioxidant activity of the kangpu tea is improved, the flavor and the health-care effect of boxthorn leaves are reserved to the maximum extent, agricultural resources are fully utilized, the environment is protected, and economic benefits and social benefits are improved.

Description

Technical Field

[0001] The present invention relates to a method for preparing wolfberry leaf kombucha with high antioxidant activity using an artificial bacterial agent, belonging to the field of food processing. Background Art

[0002] Kombucha originated in China and is also known as black tea fungus and sea treasure tea. It is a traditional beverage with a sweet and sour taste fermented from sugar tea water by various microorganisms such as acetic acid bacteria, yeast, and lactic acid bacteria. It has a unique fresh, sweet, and sour taste, and the types of tea used mainly include green tea and black tea, etc. Kombucha is also known as black tea fungus. The kombucha fermentation broth is rich in beneficial components such as phenolic substances, gluconic acid, acetic acid, lactic acid, D-glucaric acid-1,4-lactone, vitamins, and minerals, and has multiple health care effects. In recent years, domestic and foreign research has shown that kombucha has the effects of regulating the body's physiological functions, improving the body's immunity, helping digestion, promoting metabolism, conditioning the stomach, eliminating fatigue, and preventing and treating arteriosclerosis, etc. It is a microbial fermentation product with great development potential.

[0003] Natural fermentation kombucha is a tea beverage fermented by various microorganisms such as yeast, acetic acid bacteria, and lactic acid bacteria. The fermentation of kombucha mainly consists of two stages: the yeast fermentation stage and the acetic acid bacteria fermentation stage, among which yeast and acetic acid bacteria are the dominant strains. Currently, the yeasts isolated from kombucha mainly include Saccharomyces, Schizosaccharomyces, Brettanomyces, Candida, Zygosaccharomyces, Pichia, and Torulaspora, etc., and the acetic acid bacteria mainly include Acetobacter, Acetobacter pasteurianus, Acetobacter nitrogenifigens, Gluconobacter oxydans, Gluconobacter cerinus, and Komagataeibacter, etc. In addition, some kombucha also contains a small amount of lactic acid bacteria, mainly Lactobacillus. Due to the large number of microorganisms in kombucha and the complex and difficult-to-control fermentation process, which has a great impact on the flavor and nutrition of kombucha, therefore, inventing an artificial synthetic bacterial agent and optimizing the addition ratio of bacterial strains are crucial to overcome this difficulty, enabling yeast, lactic acid bacteria, and acetic acid bacteria to better coordinate growth and fermentation, shortening the fermentation time, and improving the antioxidant activity of kombucha. At the same time, wolfberry leaves are used as the fermentation raw material for application. Summary of the Invention

[0004] Technical Problem:

[0005] To solve the above technical problems, the present invention has developed a method for preparing wolfberry leaf kombucha with high antioxidant activity using an artificial synthetic bacterial agent, which can effectively shorten the fermentation time and improve the antioxidant activity of kombucha.

[0006] Technical Solution:

[0007] The first object of the present invention is to provide a bacterial agent, which comprises Saccharomyces cerevisiae, Lactobacillus plantarum, and Acetobacter pasteurianus.

[0008] In one embodiment, the Saccharomyces cerevisiae is Saccharomyces cerevisiae microcapsules; the wall material of the microcapsules is sodium alginate and gum arabic.

[0009] In one embodiment, the method for preparing the microcapsules is to add 1×10 8 ~1×10 10 CFU / mL of Saccharomyces cerevisiae to a mixed solution containing 30 - 35 g / L of sodium alginate and 10 - 15 g / L of gum arabic, and obtain Saccharomyces cerevisiae microcapsules after drying.

[0010] In one embodiment, the Saccharomyces cerevisiae includes Saccharomyces cerevisiae ATCC9763; the Lactobacillus plantarum includes Lactobacillus plantarum ACCC11095; the Acetobacter pasteurianus includes Acetobacter pasteurianus Huniang 1.01.

[0011] In one embodiment, the drying includes but is not limited to spray drying; the conditions for spray drying are an inlet air temperature of 145 - 165 °C, an outlet air temperature of 65 - 75 °C, and a homogenization pressure of 40 - 50 MPa.

[0012] The second object of the present invention is to provide a method for preparing goji leaf kombucha with high antioxidant activity using an artificial synthetic bacterial agent, and the production process is to add the artificial synthetic bacterial agent during the fermentation stage of goji leaf kombucha.

[0013] In one embodiment, the method includes the following steps:

[0014] (1) Tea juice preparation: Boil the tea leaves to obtain tea juice;

[0015] (2) Fermentation: Inoculate the bacterial agent into the tea juice obtained in step (1), filter after fermentation, and collect the filtrate to obtain the kombucha.

[0016] Further, the Saccharomyces cerevisiae is Saccharomyces cerevisiae ATCC9763.

[0017] Further, the lactic acid bacteria is Lactobacillus plantarum ACCC11095.

[0018] Further, the acetic acid bacteria is Acetobacter pasteurianus Huniang 1.01.

[0019] Further, the tea leaves are goji leaf tea; the goji leaf tea is obtained by boiling goji leaves.

[0020] Further, the production process specifically includes the following steps:

[0021] (1) Preparation of wolfberry leaf tea: Add 10 - 15% (w / v) sucrose and 1 - 3% (w / v) black tea leaves to drinking water, boil for 10 minutes, filter the tea leaves, cool the tea soup to 30°C after sterilization (121°C, 20 min);

[0022] (2) Strain activation: Activate Saccharomyces cerevisiae, Lactobacillus plantarum, and Acetobacter pasteurianus respectively to obtain Saccharomyces cerevisiae seed liquid, Lactobacillus plantarum seed liquid, and Acetobacter pasteurianus seed liquid, and microencapsulate Saccharomyces cerevisiae;

[0023] (3) Fermentation stage: Inoculate Saccharomyces cerevisiae seed liquid, Lactobacillus plantarum seed liquid, and Acetobacter pasteurianus into the tea juice at an inoculation amount of 1 - 5% respectively, maintain the temperature at 28 - 32°C, ventilate at 90 - 110 rpm for 5 - 7 h. Appropriate ventilation can promote the early growth of the strains. After closing the ventilation, carry out aerobic static fermentation for 6 d;

[0024] (4) Preparation of wolfberry leaf kombucha: Centrifuge, filter, and sterilize the fermentation broth to obtain the wolfberry leaf kombucha.

[0025] The third object of the present invention is to provide the application of the bacterial agent in the preparation of kombucha.

[0026] Beneficial effects:

[0027] Compared with the prior art, it has the following advantages: Using an artificially synthesized bacterial agent to ferment and prepare wolfberry leaf kombucha, the fermentation method is simple to operate, the fermentation time is short, the antioxidant activity of kombucha is improved, the flavor and health care effects of wolfberry leaves are maximally retained, agricultural resources are fully utilized, the environment is protected, and economic and social benefits are improved. Specific embodiments

[0028] The following specific embodiments are used to further illustrate the present invention so that those skilled in the art can better understand the present invention and implement it, but the examples given are not intended to limit the present invention.

[0029] The Lactobacillus plantarum involved in the following examples is Lactobacillus plantarum ACCC11095, the Saccharomyces cerevisiae is Saccharomyces cerevisiae ATCC9763, and the acetic acid bacterium or Acetobacter pasteurianus is Huniang 1.01.

[0030] Example 1:

[0031] (1) Preparation of wolfberry leaf tea: Add 10% (w / v) sucrose and 1% (w / v) wolfberry leaves to drinking water, boil for 10 minutes, filter the tea leaves, cool the tea soup to 30°C after sterilization (121°C, 20 min);

[0032] (2) Activation culture

[0033] Activation of Lactobacillus plantarum ACCC11095: Using an inoculation loop, pick a small amount of colonies from the original strain slant (or ampoule) under sterile conditions, place them on an MRS plate medium for streaking, then incubate at 37°C for 24 h with static culture. After inspection for no contaminants, the growth of the bacteria is regular. Visually observed, the colonies are plump. After smear and staining inspection for no contaminants, pick a single colony and place it in an MRS liquid medium. Control the liquid medium filling volume to be 20% of the Erlenmeyer flask, then incubate at 37°C with static culture for 18 h until the biomass reaches OD 600 0.8 - 1.2.

[0034] Activation of Saccharomyces cerevisiae ATCC9763: Using an inoculation loop, pick a small amount of colonies from the original strain slant (or ampoule) under sterile conditions, place them on an MRS plate medium for streaking, then incubate at 30°C for 48 h with static culture. After inspection for no contaminants, the growth of the bacteria is regular. Visually observed, the colonies are plump. After smear and staining inspection for no contaminants, pick a single colony and place it in an MRS liquid medium. Control the liquid medium filling volume to be 20% of the Erlenmeyer flask, then incubate at 30°C with static culture for 48 h until the biomass reaches 10 6 -10 7 CFU / mL.

[0035] Activation of Acetobacter pasteurianus Huniang 1.01: Using an inoculation loop, pick a small amount of colonies from the original strain slant (or ampoule) under sterile conditions, place them on an MRS plate medium for streaking, then incubate at 30°C for 48 h with static culture. After inspection for no contaminants, the growth of the bacteria is regular. Visually observed, the colonies are plump. After smear and staining inspection for no contaminants, pick a single colony and place it in an MRS liquid medium. Control the liquid medium filling volume to be 20% of the Erlenmeyer flask, then incubate at 30°C for 48 h with shaking culture, set the shaking speed of the shaker to 200 rpm until the biomass reaches OD 600 0.8 - 1.2.

[0036] The seed liquid obtained from the activation culture is used for the subsequent fermentation stage.

[0037] (3) Preparation of Saccharomyces cerevisiae microcapsules

[0038] Prepare a solution containing 30 - 35 g / L sodium alginate and 10 - 15 g / L arabic gum, filter and sterilize it through a microporous membrane (pore size 0.22 μm) as the wall material solution. Then add 10 8 ~10 10 CFU / mL of Saccharomyces cerevisiae, stir for 10 min and then pass through a colloid mill, followed by spray drying to obtain the microencapsulated bacteria. The conditions for spray drying are an inlet air temperature of 145 - 165°C, an outlet air temperature of 65 - 75°C, and a homogenization pressure of 40 - 50 MPa.

[0039] (4) Fermentation stage

[0040] Inoculate Saccharomyces cerevisiae microcapsules (initial inoculation concentration 0.3 g / L), Lactobacillus plantarum (initial inoculation concentration OD 600 0.8), and Acetobacter pasteurianus (initial inoculation concentration OD 600 0.8) into the tea juice obtained in step (1), maintain the temperature at 28 - 32°C, ventilate at 90 - 110 rpm for 5 - 7 h. Appropriate ventilation can promote the early growth of the strains. After closing the ventilation, carry out aerobic static fermentation for 6 d.

[0041] Centrifuge, filter, and sterilize the fermentation broth to obtain the described wolfberry leaf kombucha.

[0042] Comparative Example 1:

[0043] The types of raw and auxiliary materials, addition amounts, addition sequences, and fermentation conditions at each stage used in Comparative Example 1 are the same as those in Example 1, except that Lactobacillus plantarum ACCC11095 is not added in step (4) of the comparative example.

[0044] Comparative Example 2:

[0045] The types of raw and auxiliary materials, addition amounts, addition sequences, and fermentation conditions at each stage used in Comparative Example 2 are the same as those in Example 1, except that in step (3) of the comparative example, Saccharomyces cerevisiae is not microencapsulated, and the Saccharomyces cerevisiae seed liquid is directly inoculated into the tea juice.

[0046] Comparative Example 3:

[0047] The types of raw and auxiliary materials, addition amounts, addition sequences, and fermentation conditions at each stage used in Comparative Example 3 are the same as those in Example 1, except that in step (3) of the comparative example, the wall materials of the microcapsules, sodium alginate and arabic gum, are replaced with gelatin of the same total amount.

[0048] Table 1 Analysis table of wolfberry leaf kombucha fermentation time and antioxidant activity results

[0049]

[0050] Note: The detection method for the antioxidant activity data of the present invention: Total antioxidant capacity (T-AOC) detection kit (ABTS method), Total antioxidant capacity (T-AOC) detection kit (FRAP method), Total antioxidant capacity (T-AOC) detection kit (DPPH method), and the detection sample standard is 0.10 g / mL. The fermentation end point is when the glucose in the kombucha is lower than 0.5 g / L.

[0051] From the above data, it can be seen that the contents of ABTS, FRAP, and DPPH antioxidant capacities of the wolfberry leaf kombucha prepared by the present invention are all higher than those of the comparative examples, the fermentation time is shorter than that of the comparative examples, and the fermentation rate is faster. Therefore, the wolfberry leaf kombucha obtained by fermenting with an artificially synthesized bacterium agent in the present invention has the advantages of short fermentation time and strong antioxidant activity.

[0052] Table 2 Analysis table of sensory evaluation (flavor) results of wolfberry leaf kombucha

[0053]

[0054] Table 3 Analysis table of sensory evaluation (taste) results of wolfberry leaf kombucha

[0055]

[0056] As can be seen from Table 2 and Table 3, the taste, aroma, etc. of the wolfberry leaf kombucha prepared by the present invention are better than those of the comparative examples. Specifically, the floral and fruity aromas are richer, the umami taste is more prominent, and the irritating taste and bitterness are reduced. Therefore, the wolfberry leaf kombucha obtained by the co-fermentation of lactic acid bacteria and yeast in the present invention has an obvious fruity aroma, a brisk aroma, a clean and refreshing taste, many nutritional components, a mellow flavor, and a sour and refreshing taste. It is a fermented tea product with both good taste and nutritional and health care effects.

[0057] Although the present invention has been disclosed above with preferred embodiments, it is not intended to limit the present invention. Anyone familiar with this technology can make various modifications and decorations without departing from the spirit and scope of the present invention. Therefore, the protection scope of the present invention should be defined by the claims.

Claims

1. A bacterial agent, characterized in that, The microbial agent includes Saccharomyces cerevisiae, Lactobacillus plantarum and Acetobacter pasteurianus.

2. The microbial agent according to claim 1, wherein The Saccharomyces cerevisiae is Saccharomyces cerevisiae microcapsule; the wall material of the microcapsule is sodium alginate and arabic gum.

3. The microbial agent according to claim 2, wherein The preparation method of the microcapsules is as follows: to a mixed solution containing 30 - 35 g / L of sodium alginate and 10 - 15 g / L of arabic gum, add Saccharomyces cerevisiae with a final concentration of 10 8 ~10 10 CFU / mL, and obtain Saccharomyces cerevisiae microcapsules after drying.

4. The microbial agent according to claim 3, characterized in that, The Saccharomyces cerevisiae includes Saccharomyces cerevisiae ATCC9763; the Lactobacillus plantarum includes Lactobacillus plantarum ACCC11095; the Acetobacter pasteurianus includes Acetobacter aceti Huniang 1.

01.

5. A method for preparing kombucha using the bacterial agent according to any one of claims 1 to 4, characterized in that, The method includes the following steps: (1) Tea juice preparation: Add tea leaves with a final concentration of 1-3% w / v and sugar with a concentration of 10-15% w / v to water, boil and then filter the tea leaves to collect the tea juice; (2) Fermentation: Inoculate the tea juice obtained in step (1) with the microbial agent according to any one of claims 1-4, filter after fermentation, and collect the filtrate to obtain the kombucha.

6. The method according to claim 5, wherein In step (2), Lactobacillus plantarum and Acetobacter pasteurianus with an initial OD 600 of 0.8 - 1.2 are inoculated into the tea juice, and 0.3 - 0.5 g / L of Saccharomyces cerevisiae microcapsules are inoculated; The preparation method of the Saccharomyces cerevisiae microcapsules is as follows: 1×10 8 -1×10 10 CFU / mL of Saccharomyces cerevisiae is added to a mixed solution containing 30 - 35 g / L of sodium alginate and 10 - 15 g / L of arabic gum, and after drying, Saccharomyces cerevisiae microcapsules are obtained.

7. The method according to claim 6, characterized in that, The tea leaves in step (1) include but are not limited to wolfberry tea leaves.

8. The method according to claim 7, wherein The conditions for the fermentation in step (2) are: 28-32 °C, and ferment for at least 6 days.

9. The method according to claim 8, wherein In step (2), ventilate for 5-7 h in the early stage of fermentation.

10. Use of the microbial agent according to any one of claims 1-4 in the preparation of kombucha.

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