Preparation process of big-leaf black tea with sleep aiding effect

Abstract

The invention provides a preparation process of large-leaf black tea with a sleep aiding effect. The preparation process comprises the following steps: S1, fresh leaf picking and pretreatment; S2, food-grade magnesium oxide solution treatment; S3, neutralizing and cleaning; through precise control and synergistic effect of each step in the technological process, the content of theanine and gamma-aminobutyric acid in the tea leaves is remarkably increased, the content of caffeine in the tea leaves is effectively reduced, the big-leaf black tea with a good sleep-aiding effect is provided for the market, and further development of the big-leaf black tea in the sleep-aiding field is promoted.

Description

Technical Field

[0001] This invention relates to the field of tea preparation technology, specifically to a preparation process for large-leaf black tea with sleep-aiding effects. Background Technology

[0002] Traditional black tea, as one of the most consumed teas globally, derives its core benefits from the synergistic effects of its rich polyphenols, amino acids, and caffeine. Polyphenols, in particular, exert antioxidant effects by scavenging free radicals, thus slowing cell aging. Theanine can cross the blood-brain barrier, promoting alpha wave generation to soothe emotions. Caffeine, on the other hand, achieves its stimulating effect by antagonizing adenosine receptors; however, excessive intake can easily lead to side effects such as insomnia and palpitations, especially affecting metabolically sensitive individuals and nighttime tea drinkers. With increasing pressure in modern society and a rising incidence of sleep disorders, consumer demand for "nighttime functional beverages" has surged. Traditional tea, due to its caffeine content, has long been excluded from bedtime scenarios. The development of sleep-aiding black tea targets this market gap. The significance of this product lies in reshaping the tea consumption scenario, expanding "tea" from a daytime stimulant to a "nighttime health companion," both continuing the essence of China's millennia-old tea culture and aligning with the WHO's concept of "non-pharmacological intervention for chronic diseases." However, current black tea processing methods result in high caffeine content, severely hindering the development of black tea in the sleep-aid field. Summary of the Invention

[0003] In view of this, the present invention proposes a preparation process for large-leaf black tea with sleep-aiding effects to solve the above problems.

[0004] The technical solution of this invention is implemented as follows:

[0005] A process for preparing large-leaf black tea with sleep-aiding effects includes the following steps:

[0006] S1. Fresh leaf picking and pretreatment: Pick fresh leaves of one bud and two leaves or one bud and three leaves from large-leaf tea trees, and then subject the fresh leaves to low-temperature stress treatment.

[0007] S2. Food-grade magnesium oxide solution treatment: Immerse the pretreated tea leaves in a food-grade magnesium oxide solution at 33-38℃ for 15-20 minutes, while adding polyacrylamide-caffeine imprinted hydrogel microspheres during the soaking process.

[0008] S3. Neutralization and cleaning: Immerse the tea leaves treated with food-grade magnesium oxide solution in 0.1M sodium dihydrogen phosphate-citric acid buffer solution for 8-12 minutes, and then rinse 3 times with running water;

[0009] S4. Withering Enhancement: The treated tea leaves are placed in a withering trough for withering treatment;

[0010] S5. Rolling: Place the withered tea leaves into a rolling machine and roll them;

[0011] S6. Enzymatic hydrolysis: Papain is added to the rolled tea leaves, and the pH is adjusted to 5.5-6.0 with 0.1M citrate-sodium citrate buffer. Enzymatic hydrolysis is carried out at 35-40℃ for 1.5-2.0h. After enzymatic hydrolysis, the enzyme is inactivated by a pulsed electric field.

[0012] S7. Fermentation: The enzymatically hydrolyzed tea leaves are fermented in two stages. The first stage is carried out at 30-32℃ and 90-95% humidity; the second stage is carried out at 22-25℃ and 85-90% humidity. During the second stage of fermentation, an exogenous compound microbial agent is introduced.

[0013] S8. Drying: The fermented tea leaves are dried, then cooled to room temperature and vacuum-packed with nitrogen to obtain a large-leaf black tea with sleep-aiding effects.

[0014] Furthermore, the low-temperature stress treatment of S1 is as follows: fresh leaves are placed in an environment of 12-15℃ with humidity controlled at 80-90% for 3.5-4.5 hours.

[0015] Furthermore, in S2, the food-grade magnesium hydroxide solution has a mass concentration of 1.0-1.5% and a material-to-liquid ratio of tea leaves to the food-grade magnesium hydroxide solution of 1:6-8 (g / mL). The food-grade magnesium hydroxide solution also contains ascorbic acid, with a mass concentration of 1.5-2.0%. The amount of polyacrylamide-caffeine imprinted hydrogel microspheres added is 0.6-1.0% of the mass of the tea leaves.

[0016] Furthermore, the polyacrylamide-caffeine imprinted hydrogel microspheres were prepared by the following method: caffeine was added to deionized water at a ratio of 1:50-80 g / mL, and the mixture was magnetically stirred at 35-40℃ for 30-60 min to obtain a template solution; acrylamide was added to the template solution at a ratio of 1-2:10 g / mL, and the mixture was magnetically stirred at 35-40℃ for 30-60 min to obtain a pre-assembled solution; 2-5% (by mass) of acrylamide, 0.5-2.0% (by mass) of N,N'-methylenebisacrylamide, and ammonium persulfate were added sequentially to the pre-assembled solution, and the mixture was stirred evenly to obtain an aqueous phase; Span 80 was added to silicone oil at a ratio of 2-5:100 g / mL, and the mixture was stirred evenly to obtain an oil phase; the aqueous phase was then... Add the product dropwise to the oil phase at a volume ratio of 1.9-2.1:5.0, stir at 800-1200 rpm for 10-15 min, then purge with nitrogen for 10-15 min, then heat to 60-70℃ and react for 4-6 h. After the reaction is complete, cool to room temperature, allow to stand and separate into layers, collect the lower layer of hydrogel microspheres, wash with ethanol and deionized water 2-3 times each, add 0.1M acetic acid solution at a volume ratio of 1:10-20 to the hydrogel microspheres, shake at 150-200 rpm for 12-24 h to dissociate caffeine from the imprinted sites, then filter out the hydrogel microspheres, wash with deionized water 3-5 times, and dry at 38-42℃ for 15-25 h to obtain polyacrylamide-caffeine imprinted hydrogel microspheres.

[0017] Furthermore, in S4, the withering thickness is 10-15cm. In the early stage, the tea is treated at 26-29℃ and 70-80% humidity for 3.5-4.5h; in the middle stage, it is treated at 24-26℃ and 60-70% humidity for 6.0-8.0h; and in the later stage, it is treated at 23-25℃ and 58-62% humidity for 10.0-14.0h. During the withering process, the tea leaves are turned over every 2 hours.

[0018] Furthermore, the kneading in S5 is carried out according to the following procedure: First, the tea leaves are kneaded for 15-20 minutes under a pressure of 0.15-0.18 MPa and a kneading disc speed of 25-30 rpm. Then, the tea leaves are taken out and spread on a clean stainless steel tray, and left to stand for 5-10 minutes at 20-25℃ and 60-70% humidity. Then, the tea leaves are kneaded again for 25-30 minutes under a pressure of 0.22-0.25 MPa and a kneading disc speed of 25-30 rpm. After the second kneading, the tea leaves are taken out and any clumps of tea leaves are broken up and spread on a clean stainless steel tray, and left to stand for 8-12 minutes at 20-25℃ and 60-70% humidity.

[0019] Furthermore, in S6, the amount of papain added is 0.2-0.3% of the tea mass, the papain activity is ≥800U / mg, and the electric field strength, pulse number, and temperature are 22-30kV / cm, 180-250, and 25-35℃ when the enzyme is inactivated by pulsed electric field.

[0020] Furthermore, the first stage of fermentation lasts 5.5-6.5 hours, with a leaf thickness of 9-11 cm; the second stage of fermentation lasts 8-12 hours, with a leaf thickness of 14-16 cm. The exogenous compound microbial agent in the second stage consists of Lactobacillus plantarum and Bacillus subtilis in a 1:1 mass ratio, and the amount of the exogenous compound microbial agent added is 0.2-0.4% of the tea mass. During the first and second stages of fermentation, the mixture is turned over once every 2 hours.

[0021] Furthermore, the S8 uses microwave-far-infrared drying to dry the tea leaves. First, it is microwaved at 600-800W for 2-3 minutes and then transferred to a far-infrared dryer to dry at 60-70℃ for 35-45 minutes, during which it is turned over once every 5 minutes.

[0022] Compared with the prior art, the beneficial effects of the present invention are:

[0023] (1) This application involves soaking tea leaves in a food-grade magnesium oxide solution. The magnesium oxide hydrolyzes to form magnesium hydroxide, creating a weakly alkaline environment for the tea leaves. This alkaline environment promotes the dissolution of caffeine in the tea leaves, and the magnesium hydroxide in the solution... 2+ It coordinates with the carbonyl oxygen of caffeine to form a soluble complex, which can be adsorbed by microspheres, aiding in the removal of caffeine from tea. Furthermore, Mg... 2+ It can enhance enzyme activity, thereby promoting the synthesis of γ-aminobutyric acid and improving the sleep-aiding effect of black tea.

[0024] (2) The polyacrylamide-caffeine imprinted hydrogel microspheres added in this application specifically capture caffeine molecules through π-π stacking and hydrogen bonding via caffeine imprinted holes on the polyacrylamide backbone, significantly reducing the content of free caffeine in tea.

[0025] (3) This application utilizes papain to efficiently hydrolyze tea protein, releasing theanine and thus significantly increasing the theanine content. Furthermore, the protein hydrolysis provides a substrate for the synthesis of γ-aminobutyric acid (GABA), which helps to increase the GABA content.

[0026] (4) The exogenous compound microbial agent of this application contains *Lactobacillus plantarum*, which can convert caffeine into theobromine or theophylline through demethylation, thereby reducing the caffeine content. *Bacillus subtilis*, in particular, converts glutamic acid into γ-aminobutyric acid (GABA) through glutamate decarboxylase, and is the main source of GABA in the fermentation process.

[0027] (5) This application significantly increases the content of theanine and γ-aminobutyric acid in tea leaves and effectively reduces the content of caffeine in tea leaves by precisely controlling and synergistically controlling each step of the process, thus providing the market with large-leaf black tea with good sleep-aiding effects and helping the further development of large-leaf black tea in the field of sleep aid. Detailed Implementation

[0028] To better understand the technical content of this invention, specific embodiments are provided below to further illustrate the invention.

[0029] Unless otherwise specified, the experimental methods used in the embodiments of this invention are all conventional methods.

[0030] Unless otherwise specified, all materials and reagents used in the embodiments of this invention are commercially available.

[0031] Example 1

[0032] A process for preparing large-leaf black tea with sleep-aiding effects includes the following steps:

[0033] S1. Fresh leaf picking and pretreatment: Pick one bud and two leaves from the large-leaf tea tree, and then subject the fresh leaves to low-temperature stress treatment. The specific low-temperature stress treatment is as follows: place the fresh leaves in an environment of 12℃ and control the humidity at 80% for 3.5 hours.

[0034] S2. Food-grade magnesium oxide solution treatment: The pretreated tea leaves were soaked in a food-grade magnesium oxide solution at 33℃ for 20 minutes. Polyacrylamide-caffeine imprinted hydrogel microspheres were added during soaking. The food-grade magnesium hydroxide solution had a mass concentration of 1.0%, and the ratio of tea leaves to the solution was 1:6 (g / mL). The solution also contained ascorbic acid, with a mass concentration of 1.5%. The amount of polyacrylamide-caffeine imprinted hydrogel microspheres added was 0.6% of the tea leaf mass. The polyacrylamide-caffeine imprinted hydrogel microspheres were prepared by the following method: caffeine was added to deionized water at a ratio of 1:500 (g / mL) and magnetically stirred at 35°C for 60 min to obtain a template solution; acrylamide was added to the template solution at a ratio of 1:10 (g / mL) and magnetically stirred at 35°C for 60 min to obtain a pre-assembled solution; 2% (w / w) of acrylamide, 0.5% (w / w) of N,N'-methylenebisacrylamide, and ammonium persulfate were added sequentially to the pre-assembled solution and mixed thoroughly to obtain an aqueous phase; Span 80 was added to silicone oil at a ratio of 2:100 (g / mL) and stirred thoroughly to obtain an oil phase. The aqueous phase was added dropwise to the oil phase at a volume ratio of 1.9:5.0, and stirred at 800 rpm for 15 min. Nitrogen gas was then introduced for 10 min, followed by heating to 60 °C and reacting for 6 h. After the reaction was completed, the mixture was cooled to room temperature, allowed to stand and separate into layers, and the lower layer of hydrogel microspheres was collected. The microspheres were washed twice with ethanol and deionized water, respectively. A 0.1 M acetic acid solution was added to the hydrogel microspheres at a volume ratio of 1:10, and the mixture was shaken at 150 rpm for 24 h to dissociate caffeine from the imprinted sites. The hydrogel microspheres were then filtered out, washed three times with deionized water, and dried at 38 °C for 25 h to obtain polyacrylamide-caffeine imprinted hydrogel microspheres.

[0035] S3. Neutralization and cleaning: Immerse the tea leaves treated with food-grade magnesium oxide solution in 0.1M sodium dihydrogen phosphate-citric acid buffer solution for 8 minutes, and then rinse 3 times with running water.

[0036] S4. Withering Intensification: The treated tea leaves are placed in a withering trough for withering treatment. The withering thickness is 10cm. In the early stage, the tea leaves are treated at 26℃ and 70% humidity for 3.5h. In the middle stage, the tea leaves are treated at 24℃ and 60% humidity for 6.0h. In the later stage, the tea leaves are treated at 23℃ and 58% humidity for 10.0h. The tea leaves are turned over every 2 hours during the withering period.

[0037] S5. Rolling: Place the withered tea leaves into a rolling machine and roll them according to the following procedure: First, roll for 20 minutes under a pressure of 0.15MPa and a rolling disc speed of 25rpm. Then, take out the tea leaves and spread them on a clean stainless steel tray. Let them stand for 5 minutes at 20℃ and 60% humidity. Then, roll for 30 minutes under a pressure of 0.22MPa and a rolling disc speed of 25rpm. After the second rolling, take out the tea leaves and break up any clumps of tea leaves. Spread them on a clean stainless steel tray and let them stand for 8 minutes at 20℃ and 60% humidity.

[0038] S6. Enzymatic hydrolysis: Papain was added to the rolled tea leaves, and the pH was adjusted to 5.5 with 0.1M citrate-sodium citrate buffer. Enzymatic hydrolysis was carried out at 35℃ for 2.0 h. After hydrolysis, the enzyme was inactivated by a pulsed electric field. The amount of papain added was 0.2% of the tea leaf mass, the papain activity was 800 U / mg, and the pulsed electric field inactivation was performed at an electric field strength of 22 kV / cm, a pulse number of 180, and a temperature of 25℃.

[0039] S7. Fermentation: The enzymatically hydrolyzed tea leaves are fermented in two stages. The first stage is carried out at 30℃ and 90% humidity; the second stage is carried out at 22℃ and 85% humidity. An exogenous compound microbial agent is introduced during the second stage fermentation. The first stage fermentation lasts 6.5 hours, with a leaf thickness of 11cm. The second stage fermentation lasts 12 hours, with a leaf thickness of 16cm. The exogenous compound microbial agent in the second stage consists of *Lactobacillus plantarum* and *Bacillus subtilis* in a 1:1 mass ratio, and the amount of exogenous compound microbial agent added is 0.2% of the tea leaf mass. The *Lactobacillus plantarum* viable count is ≥5.0 × 10⁻⁶. 9 CFU / g, Bacillus subtilis viable count ≥3.0×10⁻⁶ 9 CFU / g; Stir once every 2 hours during the first and second stages of fermentation.

[0040] S8. Drying: The fermented tea leaves undergo drying treatment using a microwave-far-infrared drying process. First, the tea leaves are microwaved at 600W for 3 minutes, then transferred to a far-infrared dryer and dried at 60℃ for 45 minutes, stirring every 5 minutes during the process. Afterward, the tea leaves are cooled to room temperature, filled with nitrogen, and vacuum-packed to obtain a large-leaf black tea with sleep-aiding effects.

[0041] Example 2

[0042] A process for preparing large-leaf black tea with sleep-aiding effects includes the following steps:

[0043] S1. Fresh leaf picking and pretreatment: Pick one bud and three leaves of large-leaf tea tree, and then subject the fresh leaves to low temperature stress treatment. The specific low temperature stress treatment is as follows: place the fresh leaves in an environment of 13.5℃ and control the humidity at 85% for 4.0h.

[0044] S2. Food-grade magnesium oxide solution treatment: The pretreated tea leaves were immersed in a food-grade magnesium oxide solution at 35℃ for 17.5 min. Polyacrylamide-caffeine imprinted hydrogel microspheres were added during immersion. The food-grade magnesium hydroxide solution had a mass concentration of 1.2%, and the ratio of tea leaves to the solution was 1:7 (g / mL). The solution also contained ascorbic acid, with a mass concentration of 1.8%. The amount of polyacrylamide-caffeine imprinted hydrogel microspheres added was 0.8% of the tea leaf mass. The polyacrylamide-caffeine imprinted hydrogel microspheres were prepared by the following method: caffeine was added to deionized water at a ratio of 1:65 (g / mL) and magnetically stirred at 38°C for 45 min to obtain a template solution; acrylamide was added to the template solution at a ratio of 1.5:10 (g / mL) and magnetically stirred at 38°C for 45 min to obtain a pre-assembled solution; 3.5% and 1.2% (by mass) of N,N'-methylenebisacrylamide and ammonium persulfate were added sequentially to the pre-assembled solution and mixed thoroughly to obtain an aqueous phase; Span 80 was added to silicone oil at a ratio of 3.5:100 (g / mL) and stirred thoroughly to obtain an oil phase. The aqueous phase was added dropwise to the oil phase at a volume ratio of 2.0:5.0, and the mixture was stirred at 1000 rpm for 12.5 min. Nitrogen gas was then introduced for 12.5 min, followed by heating to 65 °C and reacting for 5 h. After the reaction was completed, the mixture was cooled to room temperature, allowed to stand and separate into layers, and the lower layer of hydrogel microspheres was collected. The microspheres were washed three times each with ethanol and deionized water. A 0.1 M acetic acid solution was added to the hydrogel microspheres at a volume ratio of 1:15, and the mixture was shaken at 180 rpm for 18 h to dissociate caffeine from the imprinted sites. The hydrogel microspheres were then filtered out, washed four times with deionized water, and dried at 40 °C for 20 h to obtain polyacrylamide-caffeine imprinted hydrogel microspheres.

[0045] S3. Neutralization and cleaning: Immerse the tea leaves treated with food-grade magnesium oxide solution in 0.1M sodium dihydrogen phosphate-citric acid buffer solution for 10 minutes, and then rinse 3 times with running water.

[0046] S4. Withering Intensification: The treated tea leaves are placed in a withering trough for withering treatment. The withering thickness is 13cm. In the early stage, the tea leaves are treated at 27.5℃ and 75% humidity for 4.0h. In the middle stage, the tea leaves are treated at 25℃ and 65% humidity for 7.0h. In the later stage, the tea leaves are treated at 24℃ and 60% humidity for 12.0h. The tea leaves are turned over every 2 hours during the withering period.

[0047] S5. Rolling: Place the withered tea leaves into a rolling machine and roll them according to the following procedure: First, roll for 17.5 minutes under a pressure of 0.16 MPa and a rolling disc speed of 28 rpm. Then, remove the tea leaves and spread them on a clean stainless steel tray. Let them stand for 7.5 minutes at 22.5℃ and 65% humidity. Then, roll for 27.5 minutes under a pressure of 0.24 MPa and a rolling disc speed of 28 rpm. After the second rolling, remove the tea leaves and break up any clumps of tea leaves. Spread them on a clean stainless steel tray and let them stand for 10 minutes at 22.5℃ and 65% humidity.

[0048] S6. Enzymatic hydrolysis: Papain was added to the rolled tea leaves, and the pH was adjusted to 5.8 with 0.1M citrate-sodium citrate buffer. Enzymatic hydrolysis was carried out at 38℃ for 1.7h. After hydrolysis, the enzyme was inactivated by a pulsed electric field. The amount of papain added was 0.25% of the tea leaf mass, the papain activity was 900U / mg, and the pulsed electric field inactivation was performed at an electric field strength of 25kV / cm, a pulse number of 230, and a temperature of 30℃.

[0049] S7. Fermentation: The enzymatically hydrolyzed tea leaves are fermented in two stages. The first stage is carried out at 31℃ and 92% humidity; the second stage is carried out at 23.5℃ and 88% humidity. An exogenous compound microbial agent is introduced during the second stage fermentation. The first stage fermentation lasts 6.0 hours, with a leaf thickness of 10cm. The second stage fermentation lasts 10 hours, with a leaf thickness of 15cm. The exogenous compound microbial agent in the second stage consists of *Lactobacillus plantarum* and *Bacillus subtilis* in a 1:1 mass ratio, and the amount of exogenous compound microbial agent added is 0.3% of the tea leaf mass. The *Lactobacillus plantarum* viability count is ≥5.0 × 10⁻⁶. 9 CFU / g, Bacillus subtilis viable count ≥3.0×10⁻⁶ 9 CFU / g; Stir once every 2 hours during the first and second stages of fermentation.

[0050] S8. Drying: The fermented tea leaves are dried using a microwave-far-infrared drying process. First, the leaves are microwaved at 700W for 2.5 minutes, then transferred to a far-infrared dryer and dried at 65℃ for 40 minutes, stirring every 5 minutes during the process. After cooling to room temperature, the tea leaves are vacuum-packed with nitrogen to obtain a large-leaf black tea with sleep-aiding effects.

[0051] Example 3

[0052] A process for preparing large-leaf black tea with sleep-aiding effects includes the following steps:

[0053] S1. Fresh leaf picking and pretreatment: Pick one bud and three leaves of large-leaf tea tree, and then subject the fresh leaves to low temperature stress treatment. The specific low temperature stress treatment is as follows: place the fresh leaves in an environment of 15℃ and control the humidity at 90% for 4.5 hours.

[0054] S2. Food-grade magnesium oxide solution treatment: The pretreated tea leaves were immersed in a food-grade magnesium oxide solution at 38℃ for 15 minutes. Polyacrylamide-caffeine imprinted hydrogel microspheres were added during immersion. The food-grade magnesium hydroxide solution had a mass concentration of 1.5%, and the ratio of tea leaves to the solution was 1:8 (g / mL). The solution also contained ascorbic acid, with a mass concentration of 2.0%. The amount of polyacrylamide-caffeine imprinted hydrogel microspheres added was 1.0% of the tea leaf mass. The polyacrylamide-caffeine imprinted hydrogel microspheres were prepared by the following method: caffeine was added to deionized water at a ratio of 1:80 (g / mL) and magnetically stirred at 40°C for 30 min to obtain a template solution; acrylamide was added to the template solution at a ratio of 2:10 (g / mL) and magnetically stirred at 40°C for 30 min to obtain a pre-assembled solution; 5% (g / mL) of acrylamide, 2.0% (g / mL) of N,N'-methylenebisacrylamide, and ammonium persulfate were added sequentially to the pre-assembled solution and mixed thoroughly to obtain an aqueous phase; Span 80 was added to silicone oil at a ratio of 5:100 (g / mL) and stirred thoroughly to obtain an oil phase. The aqueous phase was added dropwise to the oil phase at a volume ratio of 2.1:5.0, and the mixture was stirred at 1200 rpm for 10 min. Nitrogen gas was then introduced for 15 min, followed by heating to 70 °C and reacting for 4 h. After the reaction was completed, the mixture was cooled to room temperature, allowed to stand and separate into layers, and the lower layer of hydrogel microspheres was collected. The microspheres were washed three times each with ethanol and deionized water. A 0.1 M acetic acid solution was added to the hydrogel microspheres at a volume ratio of 1:20, and the mixture was shaken at 200 rpm for 12 h to dissociate caffeine from the imprinted sites. The hydrogel microspheres were then filtered out, washed five times with deionized water, and dried at 42 °C for 15 h to obtain polyacrylamide-caffeine imprinted hydrogel microspheres.

[0055] S3. Neutralization and cleaning: Immerse the tea leaves treated with food-grade magnesium oxide solution in 0.1M sodium dihydrogen phosphate-citric acid buffer solution for 12 minutes, and then rinse 3 times with running water.

[0056] S4. Withering Intensification: The treated tea leaves are placed in a withering trough for withering treatment. The withering thickness is 10cm. In the early stage, the tea leaves are treated at 29℃ and 80% humidity for 3.5h. In the middle stage, the tea leaves are treated at 26℃ and 70% humidity for 6.0h. In the later stage, the tea leaves are treated at 25℃ and 62% humidity for 10.0h. The tea leaves are turned over every 2 hours during the withering process.

[0057] S5. Rolling: Place the withered tea leaves into a rolling machine and roll them according to the following procedure: First, roll for 15 minutes under a pressure of 0.18 MPa and a rolling disc speed of 30 rpm. Then, take out the tea leaves and spread them on a clean stainless steel tray. Let them stand for 5 minutes at 25°C and 70% humidity. Then, roll for 25 minutes under a pressure of 0.25 MPa and a rolling disc speed of 30 rpm. After the second rolling, take out the tea leaves and break up any clumps of tea leaves. Spread them on a clean stainless steel tray and let them stand for 8 minutes at 25°C and 70% humidity.

[0058] S6. Enzymatic hydrolysis: Papain was added to the rolled tea leaves, and the pH was adjusted to 6.0 with 0.1M citrate-sodium citrate buffer. Enzymatic hydrolysis was carried out at 40℃ for 1.5h. After hydrolysis, the enzyme was inactivated by a pulsed electric field. The amount of papain added was 0.3% of the tea leaf mass, the papain activity was 1000 U / mg, and the pulsed electric field inactivation was performed with an electric field strength of 30kV / cm, a pulse number of 250, and a temperature of 35℃.

[0059] S7. Fermentation: The enzymatically hydrolyzed tea leaves are fermented in two stages. The first stage is carried out at 32℃ and 95% humidity; the second stage is carried out at 25℃ and 90% humidity. An exogenous compound microbial agent is introduced during the second stage fermentation. The first stage fermentation lasts 5.5 hours, with a leaf thickness of 9cm. The second stage fermentation lasts 8 hours, with a leaf thickness of 14cm. The exogenous compound microbial agent in the second stage consists of *Lactobacillus plantarum* and *Bacillus subtilis* in a 1:1 mass ratio, and the amount of exogenous compound microbial agent added is 0.4% of the tea leaf mass. The *Lactobacillus plantarum* viable count is ≥5.0 × 10⁻⁶. 9 CFU / g, Bacillus subtilis viable count ≥3.0×10⁻⁶ 9 CFU / g; Stir once every 2 hours during the first and second stages of fermentation.

[0060] S8. Drying: The fermented tea leaves undergo drying treatment using a microwave-far-infrared drying process. First, the tea leaves are microwaved at 800W for 2 minutes, then transferred to a far-infrared dryer and dried at 70℃ for 35 minutes, stirring every 5 minutes during the process. Afterward, the tea leaves are cooled to room temperature, filled with nitrogen, and vacuum-packed to obtain a large-leaf black tea with sleep-aiding effects.

[0061] Comparative Example 1

[0062] Compared with Example 2, the difference in this comparative example is that step S2, i.e., the food-grade magnesium oxide solution treatment step, is not performed in the preparation process.

[0063] Comparative Example 2

[0064] Compared with Example 2, the difference in this comparative example is that polyacrylamide-caffeine imprinted hydrogel microspheres were not added during the soaking in food-grade magnesium oxide solution in step S2.

[0065] Comparative Example 3

[0066] Compared with Example 2, the difference in this comparative example is that step S6, i.e., the enzymatic hydrolysis step, is not performed in the preparation process.

[0067] Comparative Example 4

[0068] Compared with Example 2, the difference in this comparative example is that no exogenous compound microbial agent was introduced in the second stage of fermentation in step S7.

[0069] Comparative Example 5

[0070] The difference between this comparative example and Example 2 is that the large-leaf black tea used is commercially available large-leaf black tea.

[0071] Component testing

[0072] Take 2g of the large-leaf black tea prepared in Examples 1-3 and Comparative Examples 1-5 respectively, and steep them in 100mL of hot water at 90℃ for 5 minutes with the lid on. Remove the tea leaves and collect the tea liquid. Take equal amounts of the tea liquid to determine the content of caffeine, γ-aminobutyric acid (GABA), and theanine. Caffeine was determined according to the high-performance liquid chromatography method in GB / T 8312-2013 "Determination of Caffeine in Tea", GABA was determined according to NY / T 2890-2016 "Determination of GABA in Rice by High-Performance Liquid Chromatography", and theanine was determined according to GB / T 23193-2017 "Determination of Theanine in Tea by High-Performance Liquid Chromatography". The tests were repeated 5 times, and the average value was taken. The results are shown in Table 1.

[0073] Table 1

[0074]

[0075] As shown in Table 1, compared to commercially available black tea, Examples 1-3 exhibit significantly lower caffeine content and substantially higher γ-aminobutyric acid (GABA) and theanine content. Therefore, the black tea prepared by this invention has excellent sleep-aiding effects.

[0076] Comparing Comparative Example 1 and Example 2, in step S2 of Example 2, food-grade magnesium oxide hydrolyzes to generate magnesium hydroxide, creating a weakly alkaline environment for the tea leaves. This alkaline environment promotes the dissolution of caffeine in the tea leaves, and the magnesium in the solution... 2+The caffeine complex coordinates with the carbonyl oxygen to form a soluble complex, which can be adsorbed by the microspheres. Furthermore, the polyacrylamide-caffeine-imprinted hydrogel microspheres added in step S2 specifically capture caffeine molecules through π-π stacking and hydrogen bonding via caffeine-imprinted holes on the polyacrylamide backbone, significantly reducing the content of free caffeine in tea. Therefore, if step S2 is omitted, caffeine cannot be adsorbed, and the residual amount will increase significantly. Additionally, Mg2+... + It can enhance enzyme activity, thereby promoting the synthesis of γ-aminobutyric acid (GABA). Without soaking in food-grade magnesium oxide solution, the enzyme activity in tea leaves is reduced, resulting in a decrease in subsequent GABA synthesis. Although the alkaline environment of food-grade magnesium oxide solution will cause a decrease in theanine content, considering the overall comprehensive effect, this loss can be fully compensated for by the significant reduction in caffeine and the increase in GABA, enabling the large-leaf black tea of ​​this invention to achieve a qualitative breakthrough in sleep improvement.

[0077] By comparing Comparative Example 2 with Example 2, it can be seen that the lack of polyacrylamide-caffeine imprinted hydrogel microspheres leads to the failure of the caffeine-specific adsorption pathway, which greatly weakens the removal effect of caffeine from tea.

[0078] Comparing Comparative Example 3 with Example 2, Example 2 significantly increased theanine content by efficiently hydrolyzing tea protein with papain to release theanine. Furthermore, the protein hydrolysis provides a substrate for γ-aminobutyric acid (GABA) synthesis, contributing to an increase in GABA content. However, papain treatment promoting protein hydrolysis also increased the dissolution of residual caffeine in the tea, resulting in a slight increase in caffeine content. However, considering the overall effect, this increase is fully compensated by the significant increase in theanine and the partial increase in GABA.

[0079] Comparing Comparative Example 4 with Example 2, the *Lactobacillus plantarum* in the exogenous compound microbial agent of Example 2 can convert caffeine into theobromine or theophylline through demethylation, thereby reducing the caffeine content. Without the inoculation of *Lactobacillus plantarum*, the demethylation metabolism of caffeine is hindered, the residual amount increases, and the caffeine content in the brewed tea rises. *Bacillus subtilis* converts glutamate into γ-aminobutyric acid (GABA) through glutamate decarboxylase, which is the main source of GABA during fermentation. Without the inoculation of *Bacillus subtilis*, the conversion of glutamate to GABA is hindered, leading to a significant reduction in GABA synthesis. Without the inoculation of *Lactobacillus plantarum* and *Bacillus subtilis*, the decomposition effect of the microbial agent on theanine disappears, and the theanine content relatively increases due to reduced consumption. This invention significantly increases the synthesis of GABA by inoculating an exogenous compound microbial agent, while simultaneously reducing the caffeine and theanine content. The reduction in theanine content is adequately compensated for the significant increase in GABA synthesis, resulting in a large-leaf black tea with effective sleep-aiding effects.

[0080] Sleep aid effect experiment

[0081] Fifty healthy ICR mice, aged 7-8 weeks and weighing 18-22g, with equal numbers of males and females, were randomly divided into five groups: Example 1 group, Example 2 group, Example 3 group, commercially available group, and blank control group. Mice were acclimatized for one week before the experiment, with free access to food and water, an ambient temperature maintained at 22-24℃, and a photoperiod of 12L:12D. During the experiment, the mice were administered the drug (10mL / kg body weight) by gavage for 7 consecutive days. The blank control group was given an equal volume of distilled water by gavage, while the black tea group received the same volume of black tea infusion (2g of black tea, steeped in 100mL of 90℃ hot water for 5 minutes, then cooled to 24-26℃ in a water bath). Thirty minutes after the last administration, a subthreshold dose of sodium pentobarbital (30mg / kg) was injected intraperitoneally. The sleep latency and sleep duration of the mice were immediately observed and recorded. The sleep latency from the end of injection to the disappearance of the righting reflex (back touching the ground > 30s) was timed with a stopwatch. Sleep duration was defined as the time from the disappearance of the righting reflex to the first spontaneous reversion to resume (>1 minute was considered wakefulness), and the total duration was recorded. All experimental procedures were performed under the same environmental conditions. The average sleep latency and sleep duration of each group of mice were recorded in Table 2.

[0082] Table 2

[0083]

[0084] As can be seen from Table 2, the sleep latency of the 1-3 groups of the present invention is short and the sleep duration is long, indicating that the large-leaf black tea prepared by the present invention has a real and effective sleep-aiding effect.

[0085] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A preparation process for large-leaf black tea with sleep-aiding effects, characterized in that, Includes the following steps: S1. Fresh leaf picking and pretreatment: Pick fresh leaves of one bud and two leaves or one bud and three leaves from large-leaf tea trees, and then subject the fresh leaves to low-temperature stress treatment. S2. Food-grade magnesium oxide solution treatment: Immerse the pretreated tea leaves in a food-grade magnesium oxide solution at 33-38℃ for 15-20 minutes, while adding polyacrylamide-caffeine imprinted hydrogel microspheres during the soaking process. S3. Neutralization and cleaning: Take out the tea leaves treated with food-grade magnesium oxide solution and soak them in 0.1M sodium dihydrogen phosphate-citric acid buffer solution for 8-12 minutes, then rinse them 3 times with running water; S4. Withering Enhancement: The treated tea leaves are placed in a withering trough for withering treatment; S5. Rolling: Place the withered tea leaves into a rolling machine and roll them; S6. Enzymatic hydrolysis: Papain is added to the rolled tea leaves, and the pH is adjusted to 5.5-6.0 with 0.1M citrate-sodium citrate buffer. Enzymatic hydrolysis is carried out at 35-40℃ for 1.5-2.0h. After enzymatic hydrolysis, the enzyme is inactivated by a pulsed electric field. S7. Fermentation: The enzymatically hydrolyzed tea leaves are fermented in two stages. The first stage is carried out at 30-32℃ and 90-95% humidity; the second stage is carried out at 22-25℃ and 85-90% humidity. During the second stage of fermentation, an exogenous compound microbial agent is introduced. S8. Drying: The fermented tea leaves are dried, then cooled to room temperature and vacuum-packed with nitrogen to obtain a large-leaf black tea with sleep-aiding effects.

2. The preparation process of a large-leaf black tea with sleep-aiding effects as described in claim 1, characterized in that, The low-temperature stress treatment of S1 is specifically as follows: placing fresh leaves in an environment of 12-15℃, controlling the humidity at 80-90%, and treating for 3.5-4.5 hours.

3. The preparation process of a large-leaf black tea with sleep-aiding effects as described in claim 1, characterized in that, The food-grade magnesium hydroxide solution in S2 has a mass concentration of 1.0-1.5% and a material-to-liquid ratio of tea leaves to the food-grade magnesium hydroxide solution is 1:6-8 (g / mL). The food-grade magnesium hydroxide solution also contains ascorbic acid, with a mass concentration of 1.5-2.0%. The amount of polyacrylamide-caffeine imprinted hydrogel microspheres added is 0.6-1.0% of the mass of the tea leaves.

4. The preparation process of a large-leaf black tea with sleep-aiding effects as described in claim 3, characterized in that, The polyacrylamide-caffeine imprinted hydrogel microspheres were prepared by the following method: caffeine was added to deionized water at a ratio of 1:50-80 g / mL, and the mixture was magnetically stirred at 35-40℃ for 30-60 min to obtain a template solution; acrylamide was added to the template solution at a ratio of 1-2:10 g / mL, and the mixture was magnetically stirred at 35-40℃ for 30-60 min to obtain a pre-assembled solution; 2-5% by mass of acrylamide, 0.5-2.0% by mass of N,N'-methylenebisacrylamide, and ammonium persulfate were added sequentially to the pre-assembled solution, and the mixture was stirred evenly to obtain an aqueous phase; Span 80 was added to silicone oil at a ratio of 2-5:100 g / mL, and the mixture was stirred evenly to obtain an oil phase; the aqueous phase was further prepared by volume... Add the mixture dropwise to the oil phase at a ratio of 1.9-2.1:5.0, stir at 800-1200 rpm for 10-15 min, then purge with nitrogen for 10-15 min, then heat to 60-70℃ and react for 4-6 h. After the reaction is complete, cool to room temperature, allow to stand and separate into layers, collect the lower layer of hydrogel microspheres, wash with ethanol and deionized water 2-3 times each, add 0.1M acetic acid solution to the hydrogel microspheres at a volume ratio of 1:10-20, shake at 150-200 rpm for 12-24 h to dissociate caffeine from the imprinted sites, then filter out the hydrogel microspheres, wash with deionized water 3-5 times, and dry at 38-42℃ for 15-25 h to obtain polyacrylamide-caffeine imprinted hydrogel microspheres.

5. The preparation process of a large-leaf black tea with sleep-aiding effects as described in claim 1, characterized in that, In the S4 process, the withering thickness is 10-15cm. The initial treatment is carried out at 26-29℃ and 70-80% humidity for 3.5-4.5 hours, the middle treatment is carried out at 24-26℃ and 60-70% humidity for 6.0-8.0 hours, and the final treatment is carried out at 23-25℃ and 58-62% humidity for 10.0-14.0 hours. The tea leaves are turned over every 2 hours during the withering process.

6. The preparation process of a large-leaf black tea with sleep-aiding effects as described in claim 1, characterized in that, The kneading in S5 is carried out according to the following scheme: First, the tea leaves are kneaded for 15-20 minutes under a pressure of 0.15-0.18 MPa and a kneading disc speed of 25-30 rpm. Then, the tea leaves are taken out and spread on a clean stainless steel tray, and left to stand for 5-10 minutes at 20-25℃ and 60-70% humidity. Then, the tea leaves are kneaded again for 25-30 minutes under a pressure of 0.22-0.25 MPa and a kneading disc speed of 25-30 rpm. After the second kneading, the tea leaves are taken out and the clumps of tea leaves are broken up and spread on a clean stainless steel tray, and left to stand for 8-12 minutes at 20-25℃ and 60-70% humidity.

7. The preparation process of a large-leaf black tea with sleep-aiding effects as described in claim 1, characterized in that, The amount of papain added in S6 is 0.2-0.3% of the tea mass, the papain activity is ≥800U / mg, and the electric field strength, pulse number, and temperature are 22-30kV / cm, 180-250, and 25-35℃ when the enzyme is inactivated by pulsed electric field.

8. The preparation process of a large-leaf black tea with sleep-aiding effects as described in claim 1, characterized in that, The first stage of fermentation lasts 5.5-6.5 hours, with the leaf thickness being 9-11 cm. The second stage of fermentation lasts 8-12 hours, with the leaf thickness being 14-16 cm. The exogenous compound microbial agent used in the second stage consists of Lactobacillus plantarum and Bacillus subtilis in a 1:1 mass ratio, and the amount of the exogenous compound microbial agent added is 0.2-0.4% of the tea mass. The tea leaves are turned over every 2 hours during both the first and second stages of fermentation.

9. The preparation process of a large-leaf black tea with sleep-aiding effects as described in claim 1, characterized in that, In the S8 process, microwave-far-infrared drying is used to dry the tea leaves. First, the leaves are treated with 600-800W microwave for 2-3 minutes and then transferred to a far-infrared dryer to dry at 60-70℃ for 35-45 minutes, during which they are turned over every 5 minutes.

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