A method for making Dendrobium leaf kombucha

Abstract

The present invention provides a method for preparing dendrobium leaf-kombucha, comprising crushing dendrobium leaves, enzymatically hydrolyzing them with cellulase and pectinase, mixing them with tea leaves and purified water, heating and boiling them, cooling them, filtering and removing the residue to obtain an extract, and fermenting them with a kombucha fungus solution to obtain kombucha. The resulting kombucha is simple and efficient to process, has a unique flavor, rich bubbles, and is rich in nutrients, making it suitable for industrial production.

Description

Technical Field

[0001] This invention belongs to the field of food processing technology, and in particular relates to a method for making Dendrobium leaf kombucha. Background Art

[0002] Kombucha is a fermented tea beverage made naturally from a symbiotic culture of bacteria and yeast and a fermentation substrate of sugar and tea. With a history of over two thousand years, it is also known as "kombucha" or "sea treasure." Kombucha contains three beneficial probiotics: yeast, acetic acid bacteria, and lactic acid bacteria. It also contains abundant digestive enzymes, organic acids, B vitamins, antioxidants, and trace minerals, which promote digestion, enhance the immune system, and have antioxidant and detoxifying effects. Currently, the traditional method of making kombucha uses black tea as the main ingredient. After filtering the tea, fermentation is carried out. This method produces products with undesirable flavors such as sourness, astringency, and pungent odors, resulting in inconsistent quality and poor consumer acceptance. Therefore, developing new kombucha flavors is an urgent problem to be solved.

[0003] Dendrobium officinale is a precious traditional Chinese medicine, a perennial herb belonging to the Dendrobium genus of the Orchidaceae family. Its dried stems are used medicinally, and due to its unique growing environment and medicinal effects, it is hailed as the first of the "Nine Immortal Herbs." With increasing emphasis on health and a renewed appreciation for traditional medicinal materials, the application and research of Dendrobium officinale are deepening. However, current utilization of Dendrobium officinale is mostly concentrated on the stems, with the leaves being underutilized. While Dendrobium officinale leaves are rich in polysaccharides, polyphenols, and other health-promoting components, they have a strong grassy aroma and high water content, leading to lower consumer acceptance. Therefore, utilizing kombucha fermentation to balance the flavor of Dendrobium officinale leaves, fully utilizing Dendrobium officinale resources, and expanding product categories are of great significance.

[0004] Chinese Patent Publication No. CN114794269A discloses a fruit-flavored kombucha beverage and its preparation method, comprising: inoculating kombucha culture into tea sugar water, fermenting and sterilizing to obtain a primary fermentation liquid; then adding pretreated fruit juice to the primary fermentation liquid, inoculating with lactic acid bacteria seed liquid, and fermenting to obtain a secondary fermentation liquid; finally inoculating with yeast seed liquid into the secondary fermentation liquid, and fermenting to obtain the fruit-flavored kombucha beverage. Although this method emphasizes the conversion of phenolic compounds into more free hydroxyl forms through stepwise fermentation, thereby enhancing their antioxidant activity and nutritional value, and the addition of fruit juice and lactic acid bacteria further enhances the aroma and color, the stepwise fermentation of fruit juice, lactic acid bacteria, and yeast in this method is complex and cumbersome, with a long production cycle, which is not conducive to centralized production.

[0005] Chinese Patent Publication No. CN116998566 A discloses a fucoidan-based kombucha nutrient beverage and its preparation method. The raw materials include water, fructooligosaccharides, lychee concentrate, selenium-enriched black fungus peptides, fucoidan, calcium lactate, pectin, vitamin C, sodium hyaluronate, and instant black tea powder. These raw materials are homogenized, filtered, and sterilized before being added to kombucha fermentation liquid for further homogenization and sterilization to obtain the product. Although this method significantly improves stability, flavor, taste, aftertaste, and anti-fatigue properties compared to existing technologies, the product requires a large amount of exogenous excipients, and the operation steps are complex, making it unsuitable for large-scale production. Summary of the Invention

[0006] To address the shortcomings of existing technologies, this invention provides a method for producing Dendrobium leaf-kombucha through research on the fermentation of Dendrobium officinale and kombucha. The method involves pre-processing Dendrobium officinale leaves, optimizing the ratio of Dendrobium officinale leaves to tea leaves, adding natural honey, and utilizing kombucha fermentation to improve the flavor of Dendrobium officinale products and increase the content of beneficial components. No additional excipients are added, thus better meeting people's demand for natural health products.

[0007] To achieve the purpose of the invention, the present invention adopts the following technical solution:

[0008] A method for preparing Dendrobium leaf kombucha includes the following steps:

[0009] S1: Select fresh Dendrobium leaves, free from disease and yellowing. Rub and wash them to remove dust and impurities. After draining the water, spread them out until there are no water stains on the surface of the leaves, and then perform high-temperature blanching.

[0010] S2: After the processed Dendrobium leaves are crushed, they are hydrolyzed by adding cellulase and pectinase.

[0011] S3: Mix the Dendrobium leaves and tea leaves treated in S2 with purified water in a certain proportion and heat to boiling. After cooling, filter to remove residue to obtain an extract. Add honey in a certain proportion to obtain a Dendrobium leaf-tea leaf-honey mixture.

[0012] S4: Prepare kombucha culture solution by activating kombucha bacteria to obtain kombucha culture solution;

[0013] S5: Fermentation: Add the kombucha culture obtained in S4 to the dendrobium leaf-tea leaf-honey mixture obtained in step S3, mix well, and then put it into a sterilized fermentation tank for static fermentation to obtain the kombucha.

[0014] Furthermore, the Dendrobium leaves described in S1 are stir-fried at a temperature of 180-220℃ for high-temperature blanching, and then immediately cooled after blanching for 3-5 minutes.

[0015] Furthermore, the Dendrobium leaves mentioned in S2 need to be crushed, ground, and homogenized. Then, 1-2% of the weight of Dendrobium leaves of cellulase and 2-3% of the weight of Dendrobium leaves of pectinase are added and mixed before enzymatic hydrolysis. The enzymatic hydrolysis temperature is 40-60℃ and the enzymatic hydrolysis time is 2-3 hours. After enzymatic hydrolysis, the mixture is sterilized for later use.

[0016] Furthermore, the cellulase added in S2 has an enzyme activity of 50–100 U / mg; the pectinase added has an enzyme activity of 400–800 U / mg.

[0017] Furthermore, the tea mentioned in S3 refers to one or more of the six major tea categories.

[0018] Furthermore, the honey described in S3 is natural floral nectar, including but not limited to jujube honey, osmanthus honey, lychee honey, and multifloral honey.

[0019] Furthermore, in S3, the mass ratio of Dendrobium leaves to tea leaves is 0.25 to 1:1; the mass-volume ratio of Dendrobium leaves to tea leaves to purified water is 1:100 to 200 g / mL; and the amount of honey added in S3 is 10 to 20 g per 100 mL of extract.

[0020] Further, the kombucha activation step described in S4 includes: inoculating kombucha into YPD liquid medium and pre-culturing it at 37°C and 200 r / min for 12 h as a seed culture; then inoculating it into shake flasks containing YPD liquid medium at an inoculation rate of 0.1~0.5% and culturing it at 37°C and 200 r / min for 24 h, with a bacterial concentration of not less than 1.0 × 10⁻⁶. 8 The resulting bacterial solution, obtained at CFU / mL, is called kombucha culture solution.

[0021] Furthermore, 5-10 mL of kombucha inoculum is added to every 100 mL of the Dendrobium leaf-tea-honey mixture described in S5.

[0022] Furthermore, the fermentation temperature described in S5 is 25–32°C; the fermentation time is 10–15 days.

[0023] Furthermore, the solution after fermentation as described in S5 is filtered to remove impurities, thus obtaining the Dendrobium leaf-kombucha.

[0024] Furthermore, the total sugar content of the Dendrobium leaf-kombucha is between 23.32 and 78.83 mg / mL; the pH value is below 4.0.

[0025] This invention provides a Dendrobium-kombucha fermented product prepared by the method described above.

[0026] This invention provides the Dendrobium-kombucha fermented product, which includes beverages and health products.

[0027] In the process of Dendrobium-kombucha fermentation, this invention utilizes kombucha to enhance the flavor and quality of Dendrobium officinale, preserves its effective components to the maximum extent, uses natural honey to provide a carbon source for the proliferation of fermentation bacteria while introducing floral flavor, and selects static fermentation which is conducive to the stability of tissue structure, which is also conducive to the stable fermentation of the thick bacterial film floating on the surface of the fermentation liquid.

[0028] The beneficial effects of this invention are reflected in:

[0029] 1. Utilizing kombucha fermentation of Dendrobium leaves to improve the flavor and quality of Dendrobium leaves and enhance their effective components;

[0030] 2. Dendrobium leaves are thick and leathery, which is not conducive to the release of beneficial components. Enzymatic hydrolysis by cellulase and pectinase can convert the insoluble cellulose of Dendrobium leaves into glucose, a carbon source that can be directly utilized by fermenting microorganisms, thereby improving the fermentation effect and promoting the rupture of Dendrobium leaf cell walls, thus increasing the release of beneficial components and enhancing health benefits.

[0031] 3. Using natural honey provides the carbon source for microbial growth, enhancing the flavor while increasing the beneficial health-promoting components of honey. This avoids the unfriendly carbon sources and monotonous flavor associated with processed foods.

[0032] 4. This invention expands the scope of utilization of Dendrobium officinale resources, avoids the waste of Dendrobium officinale leaf resources, and enhances the effective utilization of Dendrobium officinale leaves. Attached Figure Description

[0033] Figure 1 A graph showing the pH changes in Dendrobium leaf-kombucha;

[0034] Figure 2 A graph showing the changes in polyphenol content in Dendrobium leaf-kombucha;

[0035] Figure 3 A graph showing the change in total sugar content in Dendrobium leaf-kombucha;

[0036] Figure 4 Dendrobium leaf kombucha is made with different proportions of raw materials. Detailed Implementation

[0037] To explain in detail the technical content, objectives, and effects of the present invention, the following description is provided in conjunction with the embodiments and accompanying drawings.

[0038] Unless otherwise specified, the experimental methods used in the examples are conventional methods; unless otherwise specified, the materials and reagents used are commercially available.

[0039] Kombucha was activated by inoculating kombucha into YPD liquid medium and pre-culturing it at 37℃ and 200 r / min for 12 h to prepare a primary seed culture. Then, it was inoculated into shake flasks containing YPD liquid medium at an inoculation rate of 0.1~0.5% and cultured at 37℃ and 200 r / min for 24 h until the bacterial concentration was not less than 1.0×108 CFU / mL. The resulting bacterial culture was the kombucha culture. Example 1

[0040] Select fresh Dendrobium leaves, free from disease and yellowing, rub and wash them to remove dust and impurities, drain the water, and spread them out until the leaf surface is free of water stains. Then, stir-fry them at 180-220℃ for 3-5 minutes to kill the green, immediately cool them, grind them into a homogenous paste, take 100g, add 1g of cellulase and 2g of pectinase, mix, and then enzymatically hydrolyze at 40℃ for 2 hours before sterilization. Mix this paste with black tea at a mass ratio of 0.25:1, add 100 times the volume of purified water, soak and boil to obtain a mixed extract. After cooling to room temperature, filter to remove residue, and add osmanthus honey to obtain a Dendrobium leaf-tea-honey mixture. The amount of honey added is 10g per 100mL of the mixed extract. Add 5mL of kombucha culture solution per 100mL of the Dendrobium leaf-tea-honey mixture and mix well. Let it ferment in a sterilized fermentation tank at 25℃ for 10 days. After filtering to remove impurities, the kombucha is obtained.

[0041] The cellulase added has an enzyme activity of 50–100 U / mg; the pectinase added has an enzyme activity of 400–800 U / mg.

[0042] In Example 1, the black tea can be replaced by green tea, oolong tea, yellow tea, dark tea, white tea, or two or more of them. Example 2

[0043] Select fresh Dendrobium leaves, free from disease and yellowing, rub and wash them to remove dust and impurities, drain the water, and spread them out until the leaf surface is free of water stains. Then stir-fry them at 180-220℃ for 3-5 minutes to kill the green, immediately cool them, grind them into a homogenous paste, take 100g, add 2g of cellulase and 3g of pectinase, mix, and then enzymatically hydrolyze at 60℃ for 3 hours before sterilization. Mix this paste with green tea at a mass ratio of 0.5:1, add 150 times the volume of purified water, soak and boil to obtain a mixed extract. After cooling to room temperature, filter to remove residue, and add jujube honey to obtain a Dendrobium leaf-tea-honey mixture. The amount of honey added is 15g per 100mL of mixed extract. Add 7.5mL of kombucha culture solution per 100mL of Dendrobium leaf-tea-honey mixture and mix well. Let it ferment in a sterilized fermentation tank at 28℃ for 12 days. After filtering to remove impurities, the kombucha is obtained.

[0044] The cellulase added has an enzyme activity of 50–100 U / mg; the pectinase added has an enzyme activity of 400–800 U / mg.

[0045] In Example 2, the green tea can be replaced by black tea, oolong tea, yellow tea, dark tea, white tea, or two or more of them. Example 3

[0046] Select fresh Dendrobium leaves, free from disease and yellowing, rub and wash them to remove dust and impurities, drain the water, and spread them out until the leaves are free of water stains. Then, stir-fry them at 180-220℃ for 3-5 minutes to kill the green, immediately cool them, grind them into a homogenous paste, and take 100g of the paste. Add 2.5g of cellulase and 2.5g of pectinase, mix them, and then enzymatically hydrolyze them at 50℃ for 3 hours before sterilization. Mix the paste with black tea at a 1:1 mass ratio, add 200 times the volume of purified water, soak and boil to obtain a mixed extract. After cooling to room temperature and filtering to remove residue, add lychee honey to obtain a Dendrobium leaf-tea-honey mixture. The amount of honey added is 20g per 100mL of the mixed extract. Add 10mL of kombucha culture solution to each 100mL of the Dendrobium leaf-tea-honey mixture and mix well. Let it ferment in a sterilized fermentation tank at 32℃ for 15 days. After filtering to remove impurities, the kombucha is obtained.

[0047] The cellulase added has an enzyme activity of 50–100 U / mg; the pectinase added has an enzyme activity of 400–800 U / mg.

[0048] In Example 3, the black tea can be replaced by red tea, oolong tea, green tea, black tea, white tea, or two or more of them.

[0049] Comparative Example 1

[0050] Referring to Example 1, the difference is that the Dendrobium leaves were not subjected to high-temperature blanching.

[0051] Comparative Example 2

[0052] Refer to Example 2, except that honey is not added to the mixed extract.

[0053] Comparative Example 3

[0054] Referring to Example 3, the difference is that the mass ratio of Dendrobium leaves to tea leaves is 0.1:1.

[0055] Comparative Example 4

[0056] Referring to Example 3, the difference is that the mass ratio of Dendrobium leaves to tea leaves is 1.5:1.

[0057] The kombucha obtained in Examples 1-3 and Comparative Examples 1-4 were analyzed as follows.

[0058] Ten volunteer evaluators (5 males and 5 females, aged 20-32) were selected to conduct an overall sensory evaluation. Sensory evaluation indicators included: taste, color, aroma, and mouthfeel. Each indicator was scored from 1 to 5 points, with higher scores indicating better quality. The average sensory scores were used, and the results are shown in Table 1.

[0059] Table 1 Sensory Scores of Dendrobium Leaf-Kombucha with Different Treatments

[0060]

[0061] According to the sensory scoring table in Table 1, the kombucha obtained in Examples 1-3 all achieved high scores in taste, aroma, and color. Comparative Example 1's aroma score was affected by the high grassy aroma inherent in the Dendrobium leaves. Comparative Example 2 shows that the addition of honey significantly impacted the kombucha's taste, aroma, and acceptability; the lack of a direct energy source from honey resulted in insufficient fermentation, leading to a lower overall score. The scores in Comparative Examples 3-4 indicate that the ratio of Dendrobium leaves to tea leaves significantly affected the kombucha's taste, aroma, and color. Overall, the judges had a high level of acceptance for the Dendrobium leaf-kombucha.

[0062] pH determination: Take 5 mL of sample and measure the pH of the solution using a calibrated pH meter. The results are shown in the table below. Figure 1 pH is an important evaluation indicator for kombucha. pH changes due to fermentation are mainly affected by the content of added Dendrobium leaves and honey. The obtained Dendrobium leaf-kombucha all had a pH value below 4, and the lowest pH value reached 2.76 after adding 10g of white sugar.

[0063] Polyphenol content detection method: Take 1.0 mL of fermentation broth, add 2.5 mL of Folin-Ciocalteu reagent, shake well, add 2.5 mL of 15% Na₂CO₃ solution, and dilute to the mark with water, then shake well. Incubate at 40℃ for 60 min, allow to cool for 20 min, and measure the absorbance at 778 nm using a UV spectrophotometer. Establish a standard curve using gallic acid as a reference substance, and determine the polyphenol content using the Folin-Ciocalteu method. The results are as follows: Figure 2 As shown, the total amount of polyphenols obtained from Dendrobium leaf-kombucha is directly proportional to the proportion of Dendrobium leaf content added. It is a key component that constitutes the flavor and nutritional quality of Dendrobium leaf-kombucha, with polyphenol content ranging from 57.2 to 420.6 mg / L.

[0064] Total sugar content detection method: Add 500 μL of standard working solution and fermentation broth to 500 μL of 5% phenol solution, add 2.5 mL of concentrated sulfuric acid, mix well, react for 10-20 min, and measure the absorbance at 495 nm using a UV spectrophotometer. Establish a standard curve using glucose as the reference substance. The results are as follows: Figure 3As shown, Dendrobium leaves are rich in polysaccharides. The total sugar content of kombucha is directly proportional to the amount of Dendrobium leaves and honey added. Honey, as the main energy source for kombucha, is essential for successful fermentation. The total sugar content of the obtained Dendrobium leaf-kombucha ranged from 23.32 to 78.83 mg / mL.

[0065] The Dendrobium leaf-kombucha processed using the aforementioned technique Figure 4 Different proportions of raw materials result in different colors of Dendrobium leaf-kombucha. The kombucha is rich in bubbles and has the typical flavor and sensory characteristics of kombucha.

[0066] The above studies show that fermenting a system based on Dendrobium leaves, tea leaves, and sugar using kombucha microorganisms can improve its flavor, increase the content of its functional components, and enhance its nutritional value, demonstrating extremely broad development prospects and practical application potential.

Claims

1. A method for producing Dendrobium leaf-kombucha, characterized in that, The steps include: S1: Select fresh Dendrobium leaves, free from disease and yellowing. Rub and wash them to remove dust and impurities. After draining the water, spread them out until there are no water stains on the surface of the leaves, and then perform high-temperature blanching. S2: After the processed Dendrobium leaves are crushed, they are hydrolyzed by adding cellulase and pectinase. S3: Mix the Dendrobium leaves and tea leaves treated in S2 with purified water in a certain proportion and heat to boiling. After cooling, filter to remove residue to obtain an extract. Add honey in a certain proportion to obtain a Dendrobium leaf-tea leaf-honey mixture. S4: Prepare kombucha culture solution by activating kombucha bacteria. S5: Fermentation: Add the kombucha liquid obtained in S4 to the dendrobium leaf-tea-honey mixture obtained in step S3, mix well, and then put it into a sterilized fermentation tank for static fermentation to obtain the kombucha. The Dendrobium leaves described in S1 are stir-fried at a temperature of 180-220℃ for 3-5 minutes to kill the green, and then immediately shaken and spread out to cool. The Dendrobium leaves mentioned in S2 need to be crushed, ground, and homogenized. Add 1-2% of cellulase and 2-3% of pectinase by weight of Dendrobium leaves, mix, and then carry out enzymatic hydrolysis. The enzymatic hydrolysis temperature is 40-60℃, the enzymatic hydrolysis time is 2-3 hours, and the mixture is sterilized after enzymatic hydrolysis for later use. The cellulase activity described in S2 is 50–100 U / mg; the pectinase activity is 400–800 U / mg; the tea described in S3 is one or more of the six major tea categories; the honey described in S3 is natural nectar. The mass ratio of Dendrobium leaves to tea leaves in S3 is 0.25 to 1:1; the mass-volume ratio of Dendrobium leaves to tea leaves to purified water is 1:100 to 200 g / mL; the amount of honey added in S3 is 10 to 20 g per 100 mL of extract. The kombucha activation step described in S4 includes: inoculating kombucha into YPD liquid medium and pre-culturing it at 37°C and 200 rpm for 12 h as a seed culture; then inoculating it into shake flasks containing YPD liquid medium at an inoculation rate of 0.1–0.5% and culturing it at 37°C and 200 rpm for 24 h, with a bacterial concentration of not less than 1.0 × 10⁻⁶. 8 The resulting bacterial solution, with a concentration of CFU / mL, is the kombucha bacterial solution. The total sugar content of the Dendrobium leaf-kombucha ranges from 23.32 to 78.83 mg / mL; the pH value is below 4.

0. In step S5, 5-10 mL of kombucha inoculum is added to every 100 mL of the Dendrobium leaf-tea-honey mixture; the fermentation temperature is 25-32℃; the fermentation time is 10-15 days; and the solution after fermentation is filtered to remove impurities to obtain the Dendrobium leaf-kombucha.

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