Method for preparing golden camellia beverage through liquid fermentation
Through the liquid fermentation method, the seed bacteria liquid containing compost is connected to the tea soup, which solves the problem of slow fermentation rate and difficult to control process parameters during solid fermentation, and achieves efficient and safe production of Jinhua tea liquid beverages.
Patent Information
- Application Number
- CN202311749832.9
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2023-12-16
- Publication Date
- 2025-06-17
AI Technical Summary
In the prior art, solid fermentation and production of golden flower tea has problems such as slow fermentation rate, long cycle, difficult to detect and control process parameters, and fewer products.
By using the liquid fermentation method, the seed bacteria liquid containing compost was connected to the tea soup, and fermented in a fermentation tank, the fermentation temperature was controlled at 30-32℃, and the fermentation time was 3-5 days, and the Jinhua tea liquid beverage was obtained.
It improves the fermentation rate, reduces costs, achieves safe and clean production, is suitable for industrial production, and increases the economic value of tea waste resources.
Smart Images
Figure CN120154057A_ABST
Abstract
Description
Technical Field
[0001] The present invention relates to a method for producing Camellia nitidissima beverage by liquid fermentation. Background Art
[0002] With the gradual rapid development and improvement of the tea industry, the tea production has increased year by year, and the quantity of tea stalks and yellow tea leaves produced therefrom is extremely considerable. However, most tea stalks and yellow tea leaves have poor taste, weak aroma, and unfavorable proportion of internal substances, and are often sold at low prices or used as fertilizers to cover the tea tree rows, with low utilization rate. Eurotium cristarum is also known as "golden flower fungus" because it produces golden ascocarps during its growth and reproduction process, and it is the only secondary confidential protected strain in China's tea-making industry. Eurotium cristarum has the effects of conditioning the stomach, promoting digestion, reducing blood sugar, and reducing blood lipid on the human body, and this has been confirmed by clinical experiments of Chinese and foreign medical institutions and experts. Eurotium cristarum mainly exists in Fuzhuan tea among dark teas. Usually, people use Eurotium cristarum for the flower-blooming solid tea bricks, and take the quantity of golden flowers as an index to evaluate the quality, but its use for the fermentation of liquid tea is relatively lacking. With the exploration of the health care effects of Eurotium cristarum, the further development of its products has become a hot topic at present, but mostly solid fermentation is adopted, which has problems such as slow fermentation rate, long cycle, difficult detection and control of process parameters, and few products.
[0003] Based on the above problems, the applicant has conducted research on the above problems and produced this case. Summary of the Invention
[0004] The purpose of the present invention is to provide a method for producing Camellia nitidissima beverage by liquid fermentation with a higher fermentation rate and lower cost.
[0005] In order to achieve the above purpose, the present invention adopts the following technical scheme:
[0006] A method for producing Camellia nitidissima beverage by liquid fermentation, including a tea soup production process and a fermentation process. The tea soup production process is to add 25 - 30 L of water and 250 - 400 g of granulated sugar to every 500 - 600 g of tea stalks or yellow tea leaves, and boil in boiling water for 10 - 12 min to form a tea soup. The fermentation process is to inoculate the seed bacterial liquid containing a single strain of Eurotium cristarum into the tea soup according to 2% - 4% of the volume of the tea soup, ferment in a fermentation tank, the fermentation temperature is 30 - 32 °C, and the fermentation duration is 3 - 5 d to obtain a Camellia nitidissima liquid beverage.
[0007] As a preferred embodiment of the present invention, the seed bacterial liquid containing a single strain of Eurotium cristatum is prepared as follows: Step A, select the finished Wuyi rock tea Jin Guanyin with yellow small flowers stored naturally, add sterile physiological saline, and the mass ratio of the finished Wuyi rock tea Jin Guanyin to the sterile physiological saline is 1:10. After shaking for 1 h, take 200 μl and spread it on a sterilized PDA solid medium for inoculation, and place it in an incubator at 30 °C for culturing for 4 - 6 d. After the colonies appear, select single colonies for PDB liquid shaker culture. The culture temperature is 30 °C and the rotation speed is 150 r / min. After 4 d, take 100 μl and spread it evenly on the PDA solid medium again to grow colonies; Step B, purify the strain of Eurotium cristatum in Step A; Step C, inoculate the pure strain into a sterilized tea soup liquid medium. The tea soup liquid medium is formed by adding 50 ml - 100 ml of water and 0.5 g - 1 g of granulated sugar to every 1 g - 2 g of yellow tea leaves or tea stalks and boiling in boiling water for 10 - 12 min. The inoculation amount in the tea soup liquid medium is 2% - 4% of the volume of the tea soup liquid medium.
[0008] Adopting the technical solution of the present invention, the preparation method of the golden flower tea beverage by liquid fermentation with tea stalks and yellow tea leaves as raw materials not only has the health care effect of Eurotium cristatum, but also improves the economic value of tea waste resources and can reduce costs. Using this single strain for fermentation can not only avoid possible contamination by other miscellaneous bacteria and achieve safe and clean production, but also has a high production efficiency. Usually, only an inoculation amount of 2% - 4% v / v is required. Except for the sugar carbon source to increase sweetness and accelerate production, no other compounds need to be added to the fermentation broth, and fermentation can be completed in 3 - 5 d. The method is simple and easy to implement, more efficient and convenient to control, and is suitable for industrial production. In the embodiment, the strain is the natural Eurotium cristatum from the finished Wuyi rock tea Jin Guanyin, which is a single strain with strong adaptability, rather than the Eurotium cristatum produced by the natural "flowering" of traditional Fu brick tea. Compared with the strains reported in the past, which are greatly affected by temperature and humidity, this strain has low requirements for temperature and humidity (it can grow within the range of temperature 25 - 40 °C and humidity 60% - 90%, and the growth rate changes little). In addition, for the strains reported in the past, it usually takes 7 - 14 days to achieve full flowering during solid fermentation, while the present invention only requires 3 - 5 days. Description of the Drawings
[0009] Figure 1 It is the colony morphology identification diagram in the present invention.
[0010] Figure 2 It is the molecular identification diagram of Eurotium cristatum in the present invention.
[0011] Figure 3 It is the photo of the growth situation of the bacteria body during the 3 - day solid fermentation of tea stalks and yellow tea leaves in the present invention.
[0012] Figure 4These are comparison photos of the golden camellia beverage before and after fermentation in the present invention. DETAILED DESCRIPTION
[0013] In order to better understand the technical solution of the present invention, it is described in more detail below in conjunction with embodiments.
[0014] 1. Screening and identification of pure strains of Eurotium cristatum
[0015] 1.1 Isolation and identification of single strains
[0016] Small golden flowers were found on the naturally stored Wuyi rock tea Jin Guanyin, a finished product, and were identified. Add sterile physiological saline (solid-liquid ratio 1:10) to the finished Jin Guanyin tea, shake for 1 hour, take 200ul and spread it on the sterilized PDA solid culture medium, place it in a 30℃ incubator and culture it for about 4-6 days (days, the same below). After the colonies appear, select single colonies for PDB liquid shaker culture, temperature 30℃, speed 150r / min, take 100ul after 4 days and evenly spread it on the PDA solid culture medium. Morphological identification is carried out after the colonies are full. The colonies are round or oval, the main body is dark yellow, the color gradually darkens from the edge to the center, and the center is dark brown ( Figure 1 A,). Under electron microscope observation, the classic immature cleistothecia, mature cleistothecia and released spores of Eurotium cristatum can be seen ( Figure 1 B, C, D). Among them, Figure 1 In the figure, A is the appearance of the colony, B is the immature cleistothecia under an electron microscope, C is the mature cleistothecia under an electron microscope, D is the spore being released, and the arrows are the spores.
[0017] Single colonies were selected and DNA was extracted using a fungal DNA kit. The extracted DNA was used as a template and PCR amplified using universal primers ITS1 (5'-TCCGTAGGTGAACCTGCGG-3') / ITS4 (5'-TCCTCCGCTTATTGATATGC-3'). PCR amplification system: 20ul of the total reaction system contained 1ul of ITS1 and ITS4, 1ul of DNA template, 10ul of premix Taq, and 7ul of ddH2O. The amplification conditions were: 95℃ pre-denaturation for 5min, 95℃ denaturation for 50s, 55℃ annealing for 30s, 72℃ extension for 1min, 30 cycles, and finally 72℃ extension for 7min. After the PCR product was detected by 1% agarose gel electrophoresis, the PCR product with the target band, about 500bp, ( Figure 2 A) was sent to Shanghai Shenggong Biotechnology Co., Ltd. for sequencing. The sequencing results were compared with the NCBI database by BLAST and a phylogenetic tree was constructed ( Figure 2B). The result shows that it is in the same branch as Eurotium cristarum with a confidence level of 100%. Combining the morphological observation results, this strain was identified as Eurotium cristarum and named LGT. Figure 2 In A, PCR agarose gel electrophoresis (M: DL2000 DNA Marker; 1: blank control; 2: PCR product band).
[0018] 1.2 Purification of Eurotium cristarum
[0019] The identified Eurotium cristarum was purified by the streak plate method. A small amount of the strain was picked and streaked on a PDA solid medium, incubated at 30 °C in a constant temperature incubator, and repeated three times to obtain pure strains, which were stored at 4 °C for later use.
[0020] 2. Making beverages by liquid fermentation of tea stems and yellow tea leaves
[0021] 2.1 Preparation of the fermentation broth of Eurotium cristarum and verification of solid fermentation
[0022] The pure strain was inoculated into a sterilized tea broth liquid medium and cultured in a shaker at 30 °C and 150 r / min for 4 - 5 days to obtain the subsequent fermentation seed broth. The composition of the tea broth liquid medium was prepared by adding 50 - 100 ml of water and 0.5 - 1 g of granulated sugar to 1 - 2 g of yellow tea leaves or tea stems and boiling in boiling water for 10 - 12 min. The inoculation amount in the tea broth liquid medium was 2% - 4% of the volume of the tea broth liquid medium.
[0023] The production of solid golden flower tea stems and golden flower yellow tea leaves was tested with this seed broth: The tea stems and yellow tea leaves were put into conical flasks and autoclaved in an autoclave (121 °C, 15 min). After cooling, the seed broth was inoculated into the tea stems and yellow tea leaves (1 g of sample: 0.5 mL of seed broth), shaken well and placed for 3 - 4 days (the temperature can be within the range of 25 - 40 °C and the humidity can be within the range of 60% - 90%), and it can be found that the strain can cover the tea stems and yellow tea leaves, and the spore number can reach about 1*10 8 CFU / g, indicating that the activity of this strain is relatively strong. Figure 3 In A, it is the colony growth status diagram of tea stem fermentation on the 3rd day; in B, it is the colony growth status diagram of yellow tea leaf fermentation on the 3rd day.
[0024] This strain was also applied to various teas (white tea, green tea, oolong tea, black tea, etc.) and medicinal materials (golden-edged rose, longan, motherwort, etc.) for flower formation, and the same effect could be obtained, indicating that this strain has strong adaptability.
[0025] 2.2 Production of golden flower beverages
[0026] Take 500-600g tea stems or yellow slices, add 25-30L water and 250-400g white sugar, and boil in boiling water for 10-12min to obtain tea soup. Add the prepared Eurotium cristatum seed liquid into the sterile tea soup at 2%-4% of the volume of the tea soup, and place it in a 50L semi-closed fermentation tank for small-scale fermentation. The fermentation temperature is 30-32℃, the rotation speed is 75rpm, the pH is natural, and the fermentation time is 3-5d. A golden camellia liquid beverage can be obtained. Compared with before fermentation, the tea soup color becomes darker, the bitter taste disappears, and the taste is sweet and refreshing. Figure 4 A is the picture before fermentation. Figure 4 B is the picture after fermentation. Usually, on the 4th to 5th day of fermentation, the number of bacteria in the fermentation liquid is 2×10 8 cfu / mL, which meets the standards for probiotic beverages.
[0027] The protection scope of the present invention is not limited to this embodiment, and any person making similar changes thereto shall be deemed to be within the protection scope of the present invention.
Claims
1. A method for producing Camellia nitidissima beverage by liquid fermentation, characterized in that, It includes a tea soup production process and a fermentation process. In the tea soup production process, 25 - 30 L of water and 250 - 400 g of granulated sugar are added to every 500 - 600 g of tea stalks or yellow tea leaves, and they are boiled in boiling water for 10 - 12 min to form a tea soup. In the fermentation process, a seed bacterial solution containing a single strain of Eurotium cristatum is inoculated into the tea soup at 2% - 4% of the volume of the tea soup, and it is fermented in a fermentation tank at a fermentation temperature of 30 - 32 °C for 3 - 5 d to obtain a camellia flower liquid beverage.
2. The method for producing Camellia nitidissima beverage by liquid fermentation according to claim 1, characterized in that, The seed bacterial solution containing a single strain of Eurotium cristatum is prepared in the following way: Step A, select the finished Wuyi rock tea Jin Guanyin stored naturally with yellow small flowers, add sterile physiological saline, and the mass ratio of the finished Wuyi rock tea Jin Guanyin to the sterile physiological saline is 1:
10. After shaking for 1 h, take 200 μl and spread it for inoculation on a sterilized PDA solid medium, and place it in an incubator at 30 °C for 4 - 6 d. After the colonies appear, select single colonies for PDB liquid shaker culture. The culture temperature is 30 °C, and the rotation speed is 150 r / min. After 4 d, take 100 μl and spread it evenly on the PDA solid medium again to grow colonies; Step B, purify the strain of Eurotium cristatum in Step A; Step C, inoculate the pure strain into a sterilized tea soup liquid medium. The tea soup liquid medium is formed by adding 50 ml - 100 ml of water and 0.5 g - 1 g of granulated sugar to every 1 g - 2 g of yellow tea leaves or tea stalks and boiling them in boiling water for 10 - 12 min. The inoculation amount in the tea soup liquid medium is 2% - 4% of the volume of the tea soup liquid medium.