CN111732566B - Method for extracting theaflavin - Google Patents
Method for extracting theaflavin Download PDFInfo
- Publication number
- CN111732566B CN111732566B CN202010772945.0A CN202010772945A CN111732566B CN 111732566 B CN111732566 B CN 111732566B CN 202010772945 A CN202010772945 A CN 202010772945A CN 111732566 B CN111732566 B CN 111732566B
- Authority
- CN
- China
- Prior art keywords
- theaflavin
- black tea
- ultrasonic treatment
- fermentation
- ethyl acetate
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D311/00—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings
- C07D311/02—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings ortho- or peri-condensed with carbocyclic rings or ring systems
- C07D311/04—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring
- C07D311/58—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring other than with oxygen or sulphur atoms in position 2 or 4
- C07D311/60—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring other than with oxygen or sulphur atoms in position 2 or 4 with aryl radicals attached in position 2
- C07D311/62—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring other than with oxygen or sulphur atoms in position 2 or 4 with aryl radicals attached in position 2 with oxygen atoms directly attached in position 3, e.g. anthocyanidins
Abstract
The invention belongs to the technical field of preparation of tea extracts, and particularly relates to a method for extracting theaflavin from tea, particularly black tea, and particularly provides a preparation method of an extract containing theaflavin-3, 3-digallate (TF 3).
Description
Technical Field
The invention belongs to the technical field of preparation of tea extracts, and particularly relates to a method for extracting theaflavin from tea, particularly black tea.
Background
Theaflavin is a golden yellow pigment present in black tea. In biochemistry, theaflavin is a substance with a polyphenol hydroxyl group having a theanophenol ketone structure, and is the first compound with a definite pharmacological action to be found from tea leaves. Theaflavins comprise from 0.5% to 2% by weight of the dry tea of black tea and are dependent on the method of processing of the black tea. The theaflavin plays a certain role in bright color and strong taste in the tea soup, and is an important quality index of the black tea. In recent years, because theaflavin has a plurality of potential functions related to human health, such as antivirus, antioxidation, cardiovascular disease prevention and treatment, blood fat reduction, cancer prevention and the like, the theaflavin has received wide attention at home and abroad and becomes a research hotspot of tea quality chemistry and functional components.
Theaflavins are compounds derived from enzymatic oxidative condensation of polyphenols and their derivatives. The theaflavins in black tea have various components, of which theaflavin (TF 1), theaflavin-3-gallate (TF 2A), theaflavin-3 '-gallate (TF 2B) and theaflavin-3, 3' -digallate (TF 3) are the four most important components, and in the existing black tea, TF3 accounts for about 20% -25% of the four components.
As early as 2005, researchers in Taiwan published a paper that suggests that theaflavin-3, 3' -digallate (TF 3) has a good activity against coronavirus and can effectively inhibit the activity of coronavirus 3C-like protease, thereby preventing the coronavirus from replicating in host cells. This is of great significance for the prevention and treatment of coronavirus infection. The content of theaflavin in common black tea is very low, the content of TF3 in theaflavin is not high, and the effect of resisting coronavirus in vivo can not be achieved by taking the theaflavin TF3 as daily black tea, so that the method has great value and significance for improving the content of theaflavin in the black tea and the proportion of TF3 in the theaflavin and efficiently extracting the theaflavin substances by taking the black tea as a raw material.
Disclosure of Invention
The invention provides a preparation method of an extract containing theaflavin-3, 3-digallate (TF 3), which comprises the following steps:
(1) crushing black tea to obtain crushed black tea;
(2) and (3) uniformly mixing the black tea pieces obtained in the step (1) with an ethanol water solution to prepare a mixed solution.
(3) And (3) carrying out ultrasonic treatment on the mixed solution in the step (2) to obtain an ultrasonic treatment solution, wherein the temperature of the ultrasonic treatment is 55-70 ℃.
(4) Cooling the ultrasonic treated liquid to room temperature, adding lactobacillus for static fermentation to obtain fermentation liquid.
(5) Filtering, sterilizing and deslagging the fermentation liquor to obtain fermentation supernatant;
(6) ester extraction the fermentation supernatant to obtain an extracted ester phase.
(7) Concentrating the ester phase of step (6), and freeze drying to remove the ester to obtain theaflavin extract containing theaflavin-3, 3-digallate.
Preferably, the ultrasonic treatment process in the step (3) adopts gradient stepwise heating, firstly heating to 55-65 ℃, preferably 55-60 ℃ for at least 20 minutes, and then heating to 65-70 ℃ for at least 10 minutes;
preferably, in the step (2), the crushed black tea is mixed with the ethanol water solution uniformly according to the proportion that 1000ml of ethanol water solution is added to each 100g of crushed black tea to prepare a mixed solution, wherein the volume concentration of ethanol in the ethanol water solution is 10-20%;
preferably, in the step (4), the power of the ultrasonic treatment may be 350-600W,
the emission frequency of the ultrasonic treatment is 20-30KHz,
the ultrasonic treatment time is 30-60 minutes;
preferably, in the step (5), the lactic acid bacteria comprise lactobacillus bulgaricus and/or streptococcus thermophilus;
preferably, in the step (5), the addition amount of the lactic acid bacteria is 0.5-1.5% by mass volume percent,
the fermentation temperature of the static fermentation is 20-28 ℃,
the fermentation time of the static fermentation is 18-24 h.
Preferably, the ester extraction is performed with ethyl acetate.
The invention also provides an extract containing theaflavin-3, 3' -digallate (TF 3) prepared by the preparation method.
On the basis of the common knowledge in the field, the above preferred conditions can be combined randomly to obtain the preferred embodiments of the invention.
The reagents and starting materials used in the present invention are commercially available.
The invention has the following effects:
(1) in the crushed tea mixed liquid extracted by ethanol, in order to prevent the final yield from being reduced by excessive oxidation of polyphenols such as theaflavin and catechin contained in tea leaves caused by overhigh temperature, the temperature is usually kept lower before fermentation, but the invention discovers that the yield can be greatly improved by properly heating the mixed liquid after adding the ethanol solution into the crushed tea leaves, particularly by heating the mixed liquid in sections for a period of time, probably because the proper high temperature increases the extraction efficiency of ethanol on the one hand, and on the other hand, after ensuring sufficient ethanol extraction, the temperature is further raised and the mixed liquid is kept for a period of time, a large part of ethanol is evaporated, which is beneficial to the subsequent fermentation of lactic acid bacteria, but the temperature cannot be overhigh and the time cannot be overlong, and the overhigh temperature and overlong processing time can cause the excessive oxidation of polyphenols such as theaflavin and catechin, significantly affects the amount of TF3 theaflavins in the final product;
in the first stage, 55-60 ℃ is preferred, and the heating time is more than 20 minutes, preferably 20-40 minutes.
In the second stage, 65 to 70 ℃ is preferred, and the heating time is more than 10 minutes, preferably 10 to 30 minutes.
(2) The invention adopts lactic acid bacteria for fermentation, and can obviously improve the content of TF3 in the fermentation product.
Detailed Description
The invention is further illustrated by the following examples, which are not intended to limit the scope of the invention. The experimental methods without specifying specific conditions in the following examples were selected according to the conventional methods and conditions, or according to the commercial instructions.
HPLC detection of theaflavin adopts C18 chromatographic column, and mobile phase is acetonitrile: ethyl acetate volume ratio 21: 3, taking theaflavin-3, 3' -digallate (TF 3) of sigma company as a standard product.
Example 1: extraction of theaflavin-3, 3' -digallate (TF 3) from black tea (lactic acid bacteria fermentation)
1) Crushing commercially available common loose black tea in a traditional Chinese medicine crusher, sieving with a 80-mesh sieve to obtain black tea crushed powder, and weighing 1000g of the black tea crushed powder for later use;
2) putting 1000g of the black tea crushed powder into a stainless steel barrel, adding 10 liters of ethanol water solution with the volume concentration of 15%, and stirring and mixing uniformly to prepare a mixed solution;
3) pouring the mixed solution into an ultrasonic reaction kettle, controlling the temperature in the kettle to be 60 ℃, adjusting the power of an ultrasonic transmitter to be 500W, setting the ultrasonic transmitting frequency to be 25KHz, starting an ultrasonic program, controlling the ultrasonic intensity to generate obvious eddy current on the mixture slurry, carrying out ultrasonic treatment for 30 minutes, then heating to 70 ℃, and continuing the ultrasonic treatment for 20 minutes to obtain an ultrasonic treatment solution;
4) cooling the ultrasonic treatment solution to room temperature, subpackaging in 1000ml glass triangular bottles, subpackaging 500ml treatment solution in each triangular bottle, adding 5g of commercially available lactic acid bacteria powder (containing lactobacillus bulgaricus and streptococcus thermophilus powder) produced by Jiangsu microbial science and technology limited into each triangular bottle according to the mass volume percentage of 1%, covering a breathable sealing film on each triangular bottle, tightening, placing on a horizontal rotary shaking table, setting the culture temperature to be 25 ℃, performing static fermentation culture for 20 hours, starting the shaking table to perform horizontal rotary shaking for 5 minutes at a constant shaking period of 120rmp every 3 hours, and then closing the shaking table to continue the static fermentation culture;
5) using a circulating pump to set the conveying pressure of the circulating pump to be 0.5MPa, enabling liquid-phase feed liquid to pass through a 0.45-micron microporous filter membrane, removing thalli and other solid-phase impurities, and collecting effluent liquid after microporous filtration to obtain fermentation supernatant;
6) transferring the fermented supernatant into an extraction reaction kettle, adding ethyl acetate, wherein the volume ratio of the fermented supernatant to the ethyl acetate is 1, stirring and extracting at room temperature in the reaction kettle, separating a water phase and an ester phase, keeping the ethyl acetate phase, extracting the water phase once again with the ethyl acetate by the same method, collecting the second ethyl acetate phase, and combining the second ethyl acetate phase with the previous ethyl acetate phase;
7) concentrating the extracted ethyl acetate phase by rotary evaporation at 55 deg.C to 5% (20 times) of the original volume to obtain theaflavin concentrated solution;
8) freeze drying the theaflavin concentrate. And freeze-drying to remove ethyl acetate to obtain theaflavin extract with high content of TF 3. Precooling a freeze dryer before freeze-drying, placing a stainless steel tray containing a theaflavin concentrated solution pre-frozen at-40 ℃ on a shelf when the temperature of the shelf of the freeze dryer is reduced to 4 ℃, setting the temperature of the shelf of the freeze dryer to-40 ℃, starting a vacuum pump, setting the vacuum degree to 15Pa, and continuously freeze-drying for 10 hours to obtain 42.4 g of theaflavin extract, wherein the content of theaflavin-3, 3' -digallate (TF 3) is 48.36 +/-1.07 percent through HPLC detection.
Example 2: extraction of theaflavin-3, 3' -digallate (TF 3) from black tea broke (yeast fermentation)
1) Crushing commercially available common loose black tea in a traditional Chinese medicine crusher, sieving with a 80-mesh sieve to obtain black tea crushed powder, and weighing 1000g of the black tea crushed powder for later use;
2) putting 1000g of the black tea crushed powder into a stainless steel barrel, adding 10 liters of ethanol water solution with the volume concentration of 15%, and stirring and mixing uniformly to prepare a mixed solution;
3) pouring the mixed solution into an ultrasonic reaction kettle, controlling the temperature in the kettle to be 60 ℃, adjusting the power of an ultrasonic transmitter to be 500W, setting the ultrasonic transmitting frequency to be 25KHz, starting an ultrasonic program, controlling the ultrasonic intensity to generate obvious eddy current on the mixture slurry, carrying out ultrasonic treatment for 30 minutes, then heating to 70 ℃, and continuing the ultrasonic treatment for 20 minutes to obtain an ultrasonic treatment solution;
4) cooling the ultrasonic treatment liquid to room temperature, subpackaging in 1000ml glass triangular bottles, subpackaging 500ml treatment liquid in each triangular bottle, adding 5g of commercially available saccharomyces cerevisiae (containing saccharomyces cerevisiae powder) produced by Angel yeast GmbH into each bottle according to the mass volume percentage of 1%, covering a breathable sealing film on each triangular bottle, tightening, placing on a horizontal rotary shaking table, setting the culture temperature to be 25 ℃, carrying out static fermentation culture for 20 hours, starting the horizontal rotary shaking table to carry out horizontal rotary shaking for 5 minutes at intervals of 3 hours, and then closing the shaking table to continue the static fermentation culture;
5) using a circulating pump to set the conveying pressure of the circulating pump to be 0.5MPa, enabling liquid-phase feed liquid to pass through a 0.45-micron microporous filter membrane, removing thalli and other solid-phase impurities, and collecting effluent liquid after microporous filtration to obtain fermentation supernatant;
6) transferring the fermented supernatant into an extraction reaction kettle, adding ethyl acetate, wherein the volume ratio of the fermented supernatant to the ethyl acetate is 1, stirring and extracting at room temperature in the reaction kettle, separating a water phase and an ester phase, keeping the ethyl acetate phase, extracting the water phase once again with the ethyl acetate by the same method, collecting the second ethyl acetate phase, and combining the second ethyl acetate phase with the previous ethyl acetate phase;
7) concentrating the extracted ethyl acetate phase by rotary evaporation at 55 deg.C to 5% (20 times) of the original volume to obtain theaflavin concentrated solution;
8) freeze drying the theaflavin concentrate. And freeze-drying to remove ethyl acetate to obtain theaflavin extract with high content of TF 3. Precooling a freeze dryer before freeze-drying, placing a stainless steel tray containing a theaflavin concentrated solution pre-frozen at-40 ℃ on a shelf when the temperature of the shelf of the freeze dryer is reduced to 4 ℃, setting the temperature of the shelf of the freeze dryer to-40 ℃, starting a vacuum pump, setting the vacuum degree to 15Pa, and continuously freeze-drying for 10 hours to obtain 43.5 g of a theaflavin extract, wherein the content of theaflavin-3, 3' -digallate (TF 3) is 41.39 +/-0.74 percent through HPLC detection.
Example 3: extraction of theaflavin-3, 3' -digallate (TF 3) from black tea broke (without gradient heating)
1) Crushing commercially available common loose black tea in a traditional Chinese medicine crusher, sieving with a 80-mesh sieve to obtain black tea crushed powder, and weighing 1000g of the black tea crushed powder for later use;
2) putting 1000g of the black tea crushed powder into a stainless steel barrel, adding 10 liters of ethanol water solution with the volume concentration of 15%, and stirring and mixing uniformly to prepare a mixed solution;
3) pouring the mixed solution into an ultrasonic reaction kettle, controlling the temperature in the kettle to be 60 ℃, adjusting the power of an ultrasonic transmitter to be 500W, setting the ultrasonic transmitting frequency to be 25KHz, starting an ultrasonic program, controlling the ultrasonic intensity to enable mixture slurry to generate obvious vortex, and performing ultrasonic treatment for 50 minutes to obtain an ultrasonic treatment solution;
4) cooling the ultrasonic treatment solution to room temperature, subpackaging in 1000ml glass triangular bottles, subpackaging 500ml treatment solution in each triangular bottle, adding 5g of commercially available lactic acid bacteria powder (containing lactobacillus bulgaricus and streptococcus thermophilus powder) produced by Jiangsu microbial science and technology limited into each triangular bottle according to the mass volume percentage of 1%, covering a breathable sealing film on each triangular bottle, tightening, placing on a horizontal rotary shaking table, setting the culture temperature to be 25 ℃, performing static fermentation culture for 20 hours, starting the shaking table to perform horizontal rotary shaking for 5 minutes at a constant shaking period of 120rmp every 3 hours, and then closing the shaking table to continue the static fermentation culture;
5) using a circulating pump to set the conveying pressure of the circulating pump to be 0.5MPa, enabling liquid-phase feed liquid to pass through a 0.45-micron microporous filter membrane, removing thalli and other solid-phase impurities, and collecting effluent liquid after microporous filtration to obtain fermentation supernatant;
6) transferring the fermented supernatant into an extraction reaction kettle, adding ethyl acetate, wherein the volume ratio of the fermented supernatant to the ethyl acetate is 1, stirring and extracting at room temperature in the reaction kettle, separating a water phase and an ester phase, keeping the ethyl acetate phase, extracting the water phase once again with the ethyl acetate by the same method, collecting the second ethyl acetate phase, and combining the second ethyl acetate phase with the previous ethyl acetate phase;
7) concentrating the extracted ethyl acetate phase by rotary evaporation at 55 deg.C to 5% (20 times) of the original volume to obtain theaflavin concentrated solution;
8) freeze drying the theaflavin concentrate. And freeze-drying to remove ethyl acetate to obtain theaflavin extract with high content of TF 3. Precooling a freeze dryer before freeze-drying, placing a stainless steel tray containing a theaflavin concentrated solution pre-frozen at-40 ℃ on a shelf when the temperature of the shelf of the freeze dryer is reduced to 4 ℃, setting the temperature of the shelf of the freeze dryer to-40 ℃, starting a vacuum pump, setting the vacuum degree to 15Pa, and continuously freeze-drying for 10 hours to obtain 37.6 g of theaflavin extract, wherein the content of theaflavin-3, 3' -digallate (TF 3) is 38.85 +/-1.55% by HPLC detection.
Example 4: extraction of theaflavin-3, 3' -digallate (TF 3) (high concentration ethanol extract) from black tea
1) Crushing commercially available common loose black tea in a traditional Chinese medicine crusher, sieving with a 80-mesh sieve to obtain black tea crushed powder, and weighing 1000g of the black tea crushed powder for later use;
2) putting 1000g of the black tea crushed powder into a stainless steel barrel, adding 10 liters of 25% ethanol water solution by volume, and stirring and mixing uniformly to prepare a mixed solution;
3) pouring the mixed solution into an ultrasonic reaction kettle, controlling the temperature in the kettle to be 70 ℃, adjusting the power of an ultrasonic transmitter to be 500W, setting the ultrasonic transmitting frequency to be 25KHz, starting an ultrasonic program, and controlling the ultrasonic intensity to enable the mixture slurry to generate obvious vortex. Carrying out ultrasonic treatment for 45 minutes to obtain an ultrasonic treatment solution;
4) cooling the ultrasonic treatment solution to room temperature, subpackaging in 1000ml glass triangular bottles, subpackaging 500ml treatment solution in each triangular bottle, adding 5g of commercially available lactic acid bacteria powder (containing lactobacillus bulgaricus and streptococcus thermophilus powder) produced by Jiangsu microbial science and technology limited into each triangular bottle according to the mass volume percentage of 1%, covering a breathable sealing film on each triangular bottle, tightening, placing on a horizontal rotary shaking table, setting the culture temperature to be 25 ℃, performing static fermentation culture for 20 hours, starting the shaking table to perform horizontal rotary shaking for 5 minutes at a constant shaking period of 120rmp every 3 hours, and then closing the shaking table to continue the static fermentation culture;
5) using a circulating pump to set the conveying pressure of the circulating pump to be 0.5MPa, enabling liquid-phase feed liquid to pass through a 0.45-micron microporous filter membrane, removing thalli and other solid-phase impurities, and collecting effluent liquid after microporous filtration to obtain fermentation supernatant;
6) transferring the fermented supernatant into an extraction reaction kettle, adding ethyl acetate, wherein the volume ratio of the fermented supernatant to the ethyl acetate is 1, stirring and extracting at room temperature in the reaction kettle, separating a water phase and an ester phase, keeping the ethyl acetate phase, extracting the water phase once again with the ethyl acetate by the same method, collecting the second ethyl acetate phase, and combining the second ethyl acetate phase with the previous ethyl acetate phase;
7) concentrating the extracted ethyl acetate phase by rotary evaporation at 55 deg.C to 5% (20 times) of the original volume to obtain theaflavin concentrated solution;
8) freeze drying the theaflavin concentrate. And freeze-drying to remove ethyl acetate to obtain theaflavin extract with high content of TF 3. Precooling a freeze dryer before freeze-drying, placing a stainless steel tray containing a theaflavin concentrated solution pre-frozen at-40 ℃ on a shelf when the temperature of the shelf of the freeze dryer is reduced to 4 ℃, setting the temperature of the shelf of the freeze dryer to-40 ℃, starting a vacuum pump, setting the vacuum degree to 15Pa, and continuously freeze-drying for 10 hours to obtain 42.4 g of a theaflavin extract, wherein the content of theaflavin-3, 3' -digallate (TF 3) is 39.76 +/-1.41 percent through HPLC detection.
Example 5: extraction of theaflavin-3, 3' -digallate (TF 3) from black broiled tea (high temperature ultrasound)
1) Crushing commercially available common loose black tea in a traditional Chinese medicine crusher, sieving with a 80-mesh sieve to obtain black tea crushed powder, and weighing 1000g of the black tea crushed powder for later use;
2) putting 1000g of the black tea crushed powder into a stainless steel barrel, adding 10 liters of ethanol water solution with the volume concentration of 15%, and stirring and mixing uniformly to prepare a mixed solution;
3) pouring the mixed solution into an ultrasonic reaction kettle, controlling the temperature in the kettle to be 60 ℃, adjusting the power of an ultrasonic transmitter to be 500W, setting the ultrasonic transmitting frequency to be 25KHz, starting an ultrasonic program, controlling the ultrasonic intensity to generate obvious eddy current on the mixture slurry, carrying out ultrasonic treatment for 30 minutes, then heating to 78 ℃, and continuing the ultrasonic treatment for 20 minutes to obtain an ultrasonic treatment solution;
4) cooling the ultrasonic treatment solution to room temperature, subpackaging in 1000ml glass triangular bottles, subpackaging 500ml treatment solution in each triangular bottle, adding 5g of commercially available lactic acid bacteria powder (containing Lactobacillus bulgaricus and Streptococcus thermophilus powder) produced by Jiangsu Microkang Biotechnology Co., Ltd according to the mass volume percentage of 1%, covering a breathable sealing film on each triangular bottle, tightening, placing on a horizontal rotary shaking table, setting the culture temperature to be 25 ℃, performing static fermentation culture for 20h, starting the shaking table to perform horizontal rotary shaking for 5min at the constant shaking of 120rmp every 3h, and then closing the shaking table to continue the static fermentation culture;
5) using a circulating pump to set the conveying pressure of the circulating pump to be 0.5MPa, enabling liquid-phase feed liquid to pass through a 0.45-micron microporous filter membrane, removing thalli and other solid-phase impurities, and collecting effluent liquid after microporous filtration to obtain fermentation supernatant;
6) transferring the fermented supernatant into an extraction reaction kettle, adding ethyl acetate, wherein the volume ratio of the fermented supernatant to the ethyl acetate is 1, stirring and extracting at room temperature in the reaction kettle, separating a water phase and an ester phase, keeping the ethyl acetate phase, extracting the water phase once again with the ethyl acetate by the same method, collecting the second ethyl acetate phase, and combining the second ethyl acetate phase with the previous ethyl acetate phase;
7) concentrating the extracted ethyl acetate phase by rotary evaporation at 55 deg.C to 5% (20 times) of the original volume to obtain theaflavin concentrated solution;
8) freeze drying the theaflavin concentrate. And freeze-drying to remove ethyl acetate to obtain theaflavin extract with high content of TF 3. Precooling a freeze dryer before freeze-drying, placing a stainless steel tray containing a theaflavin concentrated solution pre-frozen at-40 ℃ on a shelf when the temperature of the shelf of the freeze dryer is reduced to 4 ℃, setting the temperature of the shelf of the freeze dryer to-40 ℃, starting a vacuum pump, setting the vacuum degree to 15Pa, and continuously freeze-drying for 10 hours to obtain 44.3 g of theaflavin extract, wherein the content of theaflavin-3, 3' -digallate (TF 3) is 38.28 +/-0.98 percent through HPLC detection.
Example 6: extraction of theaflavin-3, 3' -digallate (TF 3) from black tea broke (Normal temperature ultrasound)
1) Crushing commercially available common loose black tea in a traditional Chinese medicine crusher, sieving with a 80-mesh sieve to obtain black tea crushed powder, and weighing 1000g of the black tea crushed powder for later use;
2) putting 1000g of the black tea crushed powder into a stainless steel barrel, adding 10 liters of ethanol water solution with the volume concentration of 15%, and stirring and mixing uniformly to prepare a mixed solution;
3) pouring the mixed solution into an ultrasonic reaction kettle, controlling the temperature in the kettle to be 25 ℃, adjusting the power of an ultrasonic transmitter to be 500W, setting the ultrasonic transmitting frequency to be 25KHz, starting an ultrasonic program, and controlling the ultrasonic intensity to enable the mixture slurry to generate obvious vortex. Carrying out ultrasonic treatment for 45 minutes to obtain an ultrasonic treatment solution;
4) cooling the ultrasonic treatment solution to room temperature, subpackaging in 1000ml glass triangular bottles, subpackaging 500ml treatment solution in each triangular bottle, adding 5g of commercially available lactic acid bacteria powder (containing lactobacillus bulgaricus and streptococcus thermophilus powder) produced by Jiangsu microbial science and technology limited into each triangular bottle according to the mass volume percentage of 1%, covering a breathable sealing film on each triangular bottle, tightening, placing on a horizontal rotary shaking table, setting the culture temperature to be 25 ℃, performing static fermentation culture for 20 hours, starting the shaking table to perform horizontal rotary shaking for 5 minutes at a constant shaking period of 120rmp every 3 hours, and then closing the shaking table to continue the static fermentation culture;
5) using a circulating pump to set the conveying pressure of the circulating pump to be 0.5MPa, enabling liquid-phase feed liquid to pass through a 0.45-micron microporous filter membrane, removing thalli and other solid-phase impurities, and collecting effluent liquid after microporous filtration to obtain fermentation supernatant;
6) transferring the fermented supernatant into an extraction reaction kettle, adding ethyl acetate, wherein the volume ratio of the fermented supernatant to the ethyl acetate is 1, stirring and extracting at room temperature in the reaction kettle, separating a water phase and an ester phase, keeping the ethyl acetate phase, extracting the water phase once again with the ethyl acetate by the same method, collecting the second ethyl acetate phase, and combining the second ethyl acetate phase with the previous ethyl acetate phase;
7) concentrating the extracted ethyl acetate phase by rotary evaporation at 55 deg.C to 5% (20 times) of the original volume to obtain theaflavin concentrated solution;
8) freeze drying the theaflavin concentrate. And freeze-drying to remove ethyl acetate to obtain theaflavin extract with high content of TF 3. Precooling a freeze dryer before freeze-drying, placing a stainless steel tray containing a theaflavin concentrated solution pre-frozen at-40 ℃ on a shelf when the temperature of the shelf of the freeze dryer is reduced to 4 ℃, setting the temperature of the shelf of the freeze dryer to-40 ℃, starting a vacuum pump, setting the vacuum degree to 15Pa, and continuously freeze-drying for 10 hours to obtain 37.1 g of theaflavin extract, wherein the content of theaflavin-3, 3' -digallate (TF 3) is 35.62 +/-1.04 percent through HPLC detection.
Claims (8)
1. A method for preparing an extract containing theaflavin-3, 3-digallate comprises the following steps:
(1) crushing black tea to obtain crushed black tea;
(2) uniformly mixing the black tea pieces obtained in the step (1) with an ethanol water solution to prepare a mixed solution;
(3) carrying out ultrasonic treatment on the mixed solution obtained in the step (2) to obtain an ultrasonic treatment solution, wherein the temperature of the ultrasonic treatment is 55-70 ℃;
(4) cooling the ultrasonic treatment liquid to room temperature, adding lactic acid bacteria for static fermentation to obtain fermentation liquid;
(5) filtering, sterilizing and deslagging the fermentation liquor to obtain fermentation supernatant;
(6) ester extracting the fermented supernatant to obtain an extracted ester phase;
(7) concentrating the ester phase of step (6), and freeze drying to remove the ester to obtain theaflavin extract containing theaflavin-3, 3-digallate.
2. The method of claim 1, wherein: in the step (2), the crushed black tea is mixed with the ethanol water solution uniformly according to the proportion that 1000ml of ethanol water solution is added into each 100g of crushed black tea to prepare a mixed solution, wherein the volume concentration of ethanol in the ethanol water solution is 10-20%.
3. The method of claim 1, wherein: the ultrasonic treatment process of the step (3) adopts gradient stepwise heating, and the gradient stepwise heating is firstly heated to 55-60 ℃ for at least 20 minutes and then heated to 65-70 ℃ for at least 10 minutes.
4. The method of claim 1, wherein: in the step (3), the power of the ultrasonic treatment is 350-600W, and the transmitting frequency of the ultrasonic treatment is 20-30 KHz.
5. The method of claim 1, wherein: in the step (4), the lactic acid bacteria comprise lactobacillus bulgaricus and/or streptococcus thermophilus.
6. The method of claim 1, wherein: in the step (4), the addition amount of the lactic acid bacteria is 0.5-1.5% by mass volume, the fermentation temperature of the static fermentation is 20-28 ℃, and the fermentation time of the static fermentation is 18-24 h.
7. The method of claim 1, wherein: in the step (6), the ester extraction is performed by using ethyl acetate.
8. The method of claim 3, wherein: the gradient heating step by step is first to 55-60 ℃ for 20-40 minutes, followed by 65-70 ℃ for 10-30 minutes.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010772945.0A CN111732566B (en) | 2020-08-04 | 2020-08-04 | Method for extracting theaflavin |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010772945.0A CN111732566B (en) | 2020-08-04 | 2020-08-04 | Method for extracting theaflavin |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN111732566A CN111732566A (en) | 2020-10-02 |
| CN111732566B true CN111732566B (en) | 2020-11-27 |
Family
ID=72657136
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202010772945.0A Active CN111732566B (en) | 2020-08-04 | 2020-08-04 | Method for extracting theaflavin |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN111732566B (en) |
Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1699358A (en) * | 2005-06-15 | 2005-11-23 | 海南群力药业有限公司 | Method for extracting theaflavin |
| WO2009014758A1 (en) * | 2007-07-26 | 2009-01-29 | Wellgen, Inc. | Process of making theaflavins enriched tea extract |
| CN101886097A (en) * | 2010-06-18 | 2010-11-17 | 江苏德和生物科技有限公司 | Preparation process for high purity theaflavin and application thereof in preparing anti-influenza virus medicines |
| US20130023583A1 (en) * | 2009-10-07 | 2013-01-24 | Jiangsu Dehe Bio-Tech Co., Ltd. | Theaflavin Compositions, Related Processes and Methods of Use |
| CN104413179A (en) * | 2013-08-19 | 2015-03-18 | 贵州湄江印象茶业有限责任公司 | Fuzhuan tea fermentation process |
| CN106432167A (en) * | 2016-07-19 | 2017-02-22 | 广东省农业科学院茶叶研究所 | Method for extracting theaflavin from black tea |
| CN110183412A (en) * | 2019-06-10 | 2019-08-30 | 宁德师范学院 | The method of theaflavin is extracted from black tea |
-
2020
- 2020-08-04 CN CN202010772945.0A patent/CN111732566B/en active Active
Patent Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1699358A (en) * | 2005-06-15 | 2005-11-23 | 海南群力药业有限公司 | Method for extracting theaflavin |
| WO2009014758A1 (en) * | 2007-07-26 | 2009-01-29 | Wellgen, Inc. | Process of making theaflavins enriched tea extract |
| US20130023583A1 (en) * | 2009-10-07 | 2013-01-24 | Jiangsu Dehe Bio-Tech Co., Ltd. | Theaflavin Compositions, Related Processes and Methods of Use |
| CN101886097A (en) * | 2010-06-18 | 2010-11-17 | 江苏德和生物科技有限公司 | Preparation process for high purity theaflavin and application thereof in preparing anti-influenza virus medicines |
| CN104413179A (en) * | 2013-08-19 | 2015-03-18 | 贵州湄江印象茶业有限责任公司 | Fuzhuan tea fermentation process |
| CN106432167A (en) * | 2016-07-19 | 2017-02-22 | 广东省农业科学院茶叶研究所 | Method for extracting theaflavin from black tea |
| CN110183412A (en) * | 2019-06-10 | 2019-08-30 | 宁德师范学院 | The method of theaflavin is extracted from black tea |
Non-Patent Citations (2)
| Title |
|---|
| Lactic acid bacterial fermentation modified phenolic composition in tea extracts and enhanced their antioxidant activity and cellular uptake of phenolic compounds following in vitro digestion;Danyue Zhao et al.;《Journal of Functional Foods》;20151119;第20卷;第182-194页 * |
| 茶黄素的制备、分析、分离及功能活性研究进展;王洪新等;《食品与生物技术学报》;20110131;第30卷(第1期);第12-19页 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN111732566A (en) | 2020-10-02 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN111920040B (en) | Peony fermentation product and preparation method thereof | |
| CN109769991B (en) | Production process for enzymatic synthesis of polyester catechin by using exogenous enzyme | |
| CN109486604A (en) | A kind of preparation method of Stauntonia latifolia fruit wine | |
| KR101439321B1 (en) | Functional pear fermented liquid using by-product of pear processing and its application method | |
| CN116064685B (en) | Preparation process and application of eurotium cristatum fermented edible traditional Chinese medicine | |
| CN102690858A (en) | Preparation method for grape seed proanthocyanidin extract | |
| CN110731436A (en) | A black tiger acetic acid beverage, and its preparation method and application | |
| CN106729520A (en) | A kind of preparation method of Ginger P.E | |
| CN106036610A (en) | Selenium-enriched blueberry fruit powder product and preparation method thereof | |
| CN112568437B (en) | Extraction method of selenium polysaccharide in selenium-rich cyclocarya paliurus, selenium polysaccharide obtained by extraction method and application of selenium polysaccharide | |
| CN103571221A (en) | Extracting and purifying method of red-brown pigment of chestnut shells | |
| CN109381389A (en) | A kind of persimmon leaf fermentation extracting solution and preparation method and application | |
| CN111732566B (en) | Method for extracting theaflavin | |
| CN105566512B (en) | A kind of extracting method of persimmon fruit pectin | |
| CN113186069A (en) | Lactobacillus strain microbial fermentation process | |
| CN108753558A (en) | A kind of preparation method of red yeast rice Olive vinegar | |
| CN102258099B (en) | Camellia sinensisvar assamica Pu'er tea theabrownin and application thereof | |
| WO2010085009A1 (en) | Method for producing black garlic fermented and matured using enzyme | |
| CN114794260A (en) | Wall-broken cold-brewing tea and preparation method thereof | |
| CN112707881B (en) | Preparation method of blueberry anthocyanin | |
| CN112457939A (en) | Tea rice wine and preparation method thereof | |
| WO2022156615A1 (en) | Preparation method for non-ester-type tea polyphenol rich in egc | |
| CN114045193A (en) | Preparation method of malus micromalus makino high-calcium beverage co-produced biochar | |
| CN113287699A (en) | Elaeagnus angustifolia enzyme and preparation process thereof | |
| TW202131795A (en) | Tea stalk containing gallic acid and methods of manufacturing the same |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |