A kind of tea fermentation production process

Abstract

The invention discloses a tea fermentation production process, belongs to the technical field of tea processing, and the tea fermentation production process provided by the invention comprises the following steps: picking fresh tea leaves of one bud and four to five leaves, spreading them under an environment of 25°C and 60% humidity for 6h, until the water content drops to 68% to 70%, spraying the composite bacterial liquid on the surface of the pretreated fresh leaves, placing the inoculated tea leaves in a fermentation bin of 40 to 45°C and 80 to 85% humidity, passing oxygen, fermenting for 10 to 12h, then adjusting to 50 to 54°C and 25 to 30% humidity, stacking thickness of 40 to 50cm, fermenting for 7 to 8h, using pulse microwave to inactivate microorganisms, and then drying to a moisture content of 4 to 5%, completing the tea fermentation production process. The present invention accurately regulates the molecular structure of tea leaves through a microbial-enzyme synergistic catalytic system, solves the technical problems of low polyphenol oxidation efficiency and insufficient aromatic substances, and simultaneously achieves shortened processing time and enhanced flavor.

Description

Technical Field

[0001] The invention belongs to the technical field of tea processing, and in particular relates to a tea fermentation production process. Background Art

[0002] Traditional yellow tea processing relies on the natural yellowing process, which slowly promotes the oxidation of polyphenols in a hot and humid environment to form the "three yellow" characteristics. However, this process has significant defects: Long processing cycle: Natural yellowing takes more than 140 hours, which is inefficient and difficult to mass produce; Low oxidation efficiency: polyphenols are not completely oxidized, resulting in residual bitterness and affecting the taste; Single product: Traditional processes lack precise control over the oxidation pathway, making it difficult to generate diverse aromatic substances (such as pyrazine-like caramel-scented substances).

[0003] Existing technologies attempt to improve efficiency by optimizing physical parameters (such as shortening the yellowing time and adjusting the stacking thickness), but the fundamental problem has not been solved. For example: Patent CN113142323A uses wet heat accumulation to make the yellowing process, but does not introduce microorganisms or enzyme catalysis, resulting in insufficient production of aromatic substances; "Huoshan Huangda Tea Mechanical Processing Technology" shortens the yellowing time to 60 minutes through mechanical temperature control, but does not involve directional catalysis at the molecular level, and the polyphenol oxidation pathway is single.

[0004] None of the above technologies have achieved the coordinated regulation of polyphenol oxidation and aromatic substance production, and lack the use of microbial-enzyme synergistic systems. Therefore, there is an urgent need for an innovative process that can regulate the molecular structure of tea, shorten the processing cycle, and enhance the flavor and functionality. Summary of the invention

[0005] The purpose of the present invention is to provide a tea fermentation production process, which can accurately regulate the molecular structure of tea through a microbial-enzyme synergistic catalytic system, solve the technical problems of low polyphenol oxidation efficiency and insufficient aromatic substances, and at the same time achieve shortened processing time and enhanced flavor.

[0006] The purpose of the present invention can be achieved through the following technical solutions: A tea fermentation production process comprises the following steps: Fresh leaf pretreatment: Pick fresh tea leaves with one bud and four to five leaves, and place them in an environment of 25℃ and 60% humidity for 6 hours until the moisture content drops to 68% to 70%; Inoculation of compound bacterial solution: spray the compound bacterial solution on the surface of pretreated fresh leaves; Staged wet heat-dry fermentation: Place the inoculated tea leaves in a fermentation chamber at 40-45°C and 80-85% humidity, introduce oxygen, ferment for 10-12 hours, then adjust to 50-54°C and 25-30% humidity, stack to a thickness of 40-50 cm, and ferment for 7-8 hours; Dynamic inactivation and drying: Pulse microwaves are used to inactivate microorganisms, followed by drying to a moisture content of 4-5% to complete the tea fermentation production process.

[0007] Furthermore, the concentration of the introduced oxygen is 45% to 50%.

[0008] Furthermore, the frequency of the pulse microwave is 2450 MHz and the power is 300 W.

[0009] Furthermore, the composition of the composite bacterial solution is: 0.5%~1.5%(w / w) freeze-dried white rot fungus powder; 0.1%-0.3% (w / w) Ginkgo biloba leaf extract; The balance was citrate buffer at pH 5.0.

[0010] Furthermore, the ginkgo leaf extract is prepared by the following steps: The leaves of ginkgo biloba are washed with clean water, dried, and then crushed into 40-60 meshes. The ginkgo biloba powder is mixed with a 50% ethanol solution by mass at a solid-liquid ratio of 1:10-15. The extract is heated to reflux, extracted, and filtered. The extract is collected, filtered, purified by resin adsorption, and eluted. The eluate is collected, and the eluate is concentrated, dried, rotary evaporated, and spray-dried to obtain a ginkgo biloba extract.

[0011] Furthermore, the resin is AB-8 type macroporous adsorption resin.

[0012] Furthermore, the inlet air temperature of the spray drying is 150-180°C, the outlet air temperature is 80-90°C, and the atomization pressure is 0.2-0.5 MPa.

[0013] Furthermore, the total flavonoid glycoside content in the ginkgo leaf extract is 24.1-25.0%.

[0014] Beneficial effects of the present invention: The present invention provides a tea fermentation production process. Specifically, the present invention achieves technical breakthroughs and synergistic efficiency through the organic combination of "microorganism-enzyme synergistic catalytic system" and "staged fermentation process". The specific effects are as follows: (1) Synergistic catalysis of composite bacterial solution: White rot fungus laccase: specifically catalyzes the oxidation of ortho-hydroxyl groups of polyphenols to ortho-quinone structures, significantly reduces the content of ester-type catechins (8.7±0.3 mg / g in Example 5 vs. 23.4±0.4 mg / g in Comparative Example 2), and reduces bitterness; Ginkgo biloba extract: acts as a coenzyme factor to stabilize laccase activity, while its flavonoid glycosides induce the generation of pyrazine precursors, significantly increasing the content of caramel aroma substances (385±13 μg / g in Example 5 vs. 154±8 μg / g in Comparative Example 1). At the same time, the combination of the two not only accelerates the oxidation of polyphenols (the oxidation rate of polyphenols in Example 5 reaches 82.3±1.2%), but also directs the synthesis of specific aromatic compounds (the ortho-quinone content in Example 5 reaches 450±15 μg / g), achieving a dual improvement in flavor and functionality.

[0015] (2) Staged wet heat-dry fermentation process; Hot and humid stage (40-45°C, 80-85% humidity): high humidity promotes laccase-substrate binding and accelerates polyphenol oxidation; dry and stuffy stage (50-54°C, 25-30% humidity): low humidity environment accelerates quinone polymerization to form stable golden pigment, while dehydrating and concentrating aromatic substances. Moreover, the temperature and humidity gradient changes in the two stages enable efficient connection between oxidation and polymerization reactions, and the quinone content is increased to above 420 μg / g. DETAILED DESCRIPTION

[0016] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention. Example 1

[0017] Preparation of Ginkgo Biloba Extract: 1. Raw material selection and pretreatment: 1. Raw material collection: Choose Ginkgo biloba leaves picked in autumn, when the flavonoid glycoside content is higher. It is preferred to use mature leaves with complete leaves and no diseases or insect pests.

[0018] 2. Preprocessing: Cleaning: Use clean water to wash away dust and impurities on the blade surface; Drying: Place in a ventilated drying oven at 40°C and dry to a moisture content of ≤8%. In this embodiment, the moisture content is dried to 5%.

[0019] Crushing: Use a plant crusher to crush the dried leaves into 40 mesh particles.

[0020] 2. Solvent Extraction 1. Extraction solvent: 70% ethanol (volume fraction) is used as the extraction solvent, which has the ability to dissolve both polar and non-polar components.

[0021] 2. Extract parameters: (1) Solid-liquid ratio: 1:10 (w / v, i.e., 1 kg of ginkgo leaf powder is added to 10 L of 50% ethanol solution); (2) Temperature: 55°C constant temperature water bath; (3) Time: Reflux extraction twice, 1 h each time.

[0022] 3. Operation steps: Mix the ginkgo leaf powder with 50% ethanol solution by mass, heat to the set temperature and keep reflux, filter after the first extraction, add fresh solvent to the residue for secondary extraction, combine the two extracts and set aside.

[0023] 3. Purification and concentration 1. Preliminary filtration: Use a Buchner funnel or centrifuge (4000 rpm, 15-30 min) to remove suspended particles in the crude extract.

[0024] 2.Macroporous resin purification: (1) Resin type: AB-8 macroporous adsorption resin; (2) Column loading and balancing: The resin column (diameter: height = 1:5) is rinsed with deionized water until there is no alcohol smell; (3) Sample loading and adsorption: The extract was passed through the resin column at a flow rate of 2 BV / h, and the flavonoid glycosides were adsorbed; (4) Elution: First, use 3 BV of deionized water to wash away impurities such as polysaccharides, and then use 5 BV of 70% ethanol to elute the target flavonoid glycosides; (5) Collect the eluate: Collect the ethanol elution portion and set aside.

[0025] 3. Concentrated drying: (1) Rotary evaporation: Concentrate under reduced pressure at 50°C until there is no alcohol smell to obtain a concentrated solution.

[0026] (2) Spray drying: air inlet temperature 150°C, air outlet temperature 80°C, atomization pressure 0.2 MPa. After drying, a light yellow powder is obtained.

[0027] 4. Determination of flavonoid glycoside content: HPLC method (refer to the 2020 edition of the Chinese Pharmacopoeia): 1. Chromatographic column: C18 column (4.6×250mm, 5μm); 2. Mobile phase: acetonitrile-0.4% phosphoric acid aqueous solution (25:75, v / v); 3. Detection wavelength: 360nm; 4. Standards: quercetin, kaempferol, isorhamnetin mixed reference substances; 5. Calculation: Total flavonoid glycosides content = quercetin + kaempferol + isorhamnetin; 6. Measurement results: The total flavonoid glycoside content in the Ginkgo biloba extract prepared in this example is 24.4%. Example 2

[0028] Preparation of Ginkgo Biloba Extract: 1. Raw material selection and pretreatment: 1. Raw material collection: Choose Ginkgo biloba leaves picked in autumn, when the flavonoid glycoside content is higher. It is preferred to use mature leaves with complete leaves and no diseases or insect pests.

[0029] 2. Preprocessing: Cleaning: Use clean water to wash away dust and impurities on the blade surface; Drying: Place in a ventilated drying oven at 40°C and dry to a moisture content of ≤8%. In this embodiment, the moisture content is dried to 4.5%.

[0030] Crushing: Use a plant crusher to crush the dried leaves into 40 mesh particles.

[0031] 2. Solvent Extraction 1. Extraction solvent: 70% ethanol (volume fraction) is used as the extraction solvent, which has the ability to dissolve both polar and non-polar components.

[0032] 2. Extract parameters: (1) Solid-liquid ratio: 1:10 (w / v, i.e., 1 kg of ginkgo leaf powder is added to 10 L of 50% ethanol solution); (2) Temperature: 55°C constant temperature water bath; (3) Time: Reflux extraction twice, 2 h each time.

[0033] 3. Operation steps: Mix the ginkgo leaf powder with 50% ethanol solution by mass, heat to the set temperature and keep reflux, filter after the first extraction, add fresh solvent to the residue for secondary extraction, combine the two extracts and set aside.

[0034] 3. Purification and concentration 1. Preliminary filtration: Use a Buchner funnel or centrifuge (4000 rpm, 30 min) to remove suspended particles in the crude extract.

[0035] 2. Macroporous resin purification: (1) Resin type: AB-8 macroporous adsorption resin; (2) Column loading and balancing: The resin column (diameter: height = 1:5) is rinsed with deionized water until there is no alcohol smell; (3) Sample loading and adsorption: The extract was passed through the resin column at a flow rate of 2 BV / h, and the flavonoid glycosides were adsorbed; (4) Elution: First, use 3 BV of deionized water to wash away impurities such as polysaccharides, and then use 5 BV of 70% ethanol to elute the target flavonoid glycosides; (5) Collect the eluate: Collect the ethanol elution portion and set aside.

[0036] 3. Concentrated drying: (1) Rotary evaporation: Concentrate under reduced pressure at 50°C until there is no alcohol smell to obtain a concentrated solution.

[0037] (2) Spray drying: air inlet temperature 170°C, air outlet temperature 85°C, atomization pressure 0.4 MPa. After drying, a light yellow powder is obtained.

[0038] 4. Determination of flavonoid glycoside content: HPLC method (refer to the 2020 edition of the Chinese Pharmacopoeia): 1. Chromatographic column: C18 column (4.6×250mm, 5μm); 2. Mobile phase: acetonitrile-0.4% phosphoric acid aqueous solution (25:75, v / v); 3. Detection wavelength: 360nm; 4. Standards: quercetin, kaempferol, isorhamnetin mixed reference substances; 5. Calculation: Total flavonoid glycosides content = quercetin + kaempferol + isorhamnetin; 6. Measurement results: The total flavonoid glycoside content in the Ginkgo biloba extract prepared in this example is 25.0%. Example 3

[0039] Preparation of Ginkgo Biloba Extract: 1. Raw material selection and pretreatment: 1. Raw material collection: Choose Ginkgo biloba leaves picked in autumn, when the flavonoid glycoside content is higher. It is preferred to use mature leaves with complete leaves and no diseases or insect pests.

[0040] 2. Preprocessing: Cleaning: Use clean water to wash away dust and impurities on the blade surface; Drying: Place in a ventilated drying oven at 50°C and dry to a moisture content of ≤8%. In this embodiment, the moisture content is dried to 6%.

[0041] Crushing: Use a plant crusher to crush the dried leaves into 60 mesh particles.

[0042] 2. Solvent Extraction 1. Extraction solvent: 70% ethanol (volume fraction) is used as the extraction solvent, which has the ability to dissolve both polar and non-polar components.

[0043] 2. Extract parameters: (1) Solid-liquid ratio: 1:15 (w / v, i.e., 1 kg of ginkgo leaf powder is added to 15 L of 50% ethanol solution); (2) Temperature: 60°C constant temperature water bath; (3) Time: Reflux extraction twice, 2 h each time.

[0044] 3. Operation steps: Mix the ginkgo leaf powder with 50% ethanol solution by mass, heat to the set temperature and keep reflux, filter after the first extraction, add fresh solvent to the residue for secondary extraction, combine the two extracts and set aside.

[0045] 3. Purification and concentration 1. Preliminary filtration: Use a Buchner funnel or centrifuge (4000 rpm, 30 min) to remove suspended particles in the crude extract.

[0046] 2.Macroporous resin purification: (1) Resin type: AB-8 macroporous adsorption resin; (2) Column loading and balancing: The resin column (diameter: height = 1:5) is rinsed with deionized water until there is no alcohol smell; (3) Sample loading and adsorption: The extract was passed through the resin column at a flow rate of 2 BV / h, and the flavonoid glycosides were adsorbed; (4) Elution: First, use 3 BV of deionized water to wash away impurities such as polysaccharides, and then use 5 BV of 70% ethanol to elute the target flavonoid glycosides; (5) Collect the eluate: Collect the ethanol elution portion and set aside.

[0047] 3. Concentrated drying: (1) Rotary evaporation: Concentrate under reduced pressure at 50°C until there is no alcohol smell to obtain a concentrated solution.

[0048] (2) Spray drying: air inlet temperature 180°C, air outlet temperature 90°C, atomization pressure 0.5 MPa. After drying, a light yellow powder is obtained.

[0049] 4. Determination of flavonoid glycoside content: HPLC method (refer to the 2020 edition of the Chinese Pharmacopoeia): 1. Chromatographic column: C18 column (4.6×250mm, 5μm); 2. Mobile phase: acetonitrile-0.4% phosphoric acid aqueous solution (25:75, v / v); 3. Detection wavelength: 360nm; 4. Standards: quercetin, kaempferol, isorhamnetin mixed reference substances; 5. Calculation: Total flavonoid glycosides content = quercetin + kaempferol + isorhamnetin; 6. Measurement results: The total flavonoid glycoside content in the Ginkgo biloba extract prepared in this example is 24.1%. Example 4

[0050] A tea fermentation production process comprises the following steps: Step 1: Fresh leaf pretreatment: Pick fresh tea leaves with one bud and four to five leaves, and place them in an environment of 25℃ and 60% humidity for 6 hours until the moisture content drops to 70%.

[0051] Step 2: Inoculation of compound bacterial solution: Spray the surface of the fresh leaves pretreated in step 1 with a composite bacterial solution containing the following ingredients: White rot fungus (Trametes versicolor) freeze-dried powder: 0.5% (w / w); Ginkgo biloba extract prepared in Example 1: 0.1% (w / w); The solvent was citrate buffer at pH 5.0.

[0052] Step 3: Staged wet heat-dry fermentation: The first stage (wet heat catalysis): the inoculated tea leaves are placed in a fermentation chamber at 40°C and 80% humidity, oxygen (concentration 45%) is introduced, and fermented for 10 hours; The second stage (dry oxidation): adjust to 50℃, humidity 25%, stacking thickness 40cm, fermentation 7h.

[0053] Step 4: Dynamic inactivation and drying: Microorganisms were inactivated by pulse microwaves (frequency 2450 MHz, power 300 W, intermittent treatment 3 times, 30 seconds each time), followed by drum drying at 80°C to a moisture content of 4%. Example 5

[0054] A tea fermentation production process comprises the following steps: Step 1: Fresh leaf pretreatment: Pick fresh tea leaves with one bud and four to five leaves, and place them in an environment of 25℃ and 60% humidity for 6 hours until the moisture content drops to 68%.

[0055] Step 2: Inoculation of compound bacterial solution: Spray the surface of the fresh leaves pretreated in step 1 with a composite bacterial solution containing the following ingredients: White rot fungus (Trametes versicolor) freeze-dried powder: 1.5% (w / w); Ginkgo biloba extract prepared in Example 2: 0.3% (w / w); The solvent was citrate buffer at pH 5.0.

[0056] Step 3: Staged wet heat-dry fermentation: The first stage (wet heat catalysis): the inoculated tea leaves are placed in a fermentation chamber at 40°C and 80% humidity, oxygen (concentration 50%) is introduced, and fermented for 12 hours; The second stage (dry oxidation): adjust to 50℃, humidity 25%, stacking thickness 40cm, fermentation 8h.

[0057] Step 4: Dynamic inactivation and drying: Microorganisms were inactivated by pulse microwaves (frequency 2450 MHz, power 300 W, intermittent treatment 3 times, 30 seconds each time), followed by drum drying at 80°C to a moisture content of 4%. Example 6

[0058] A tea fermentation production process comprises the following steps: Step 1: Fresh leaf pretreatment: Pick fresh tea leaves with one bud and four to five leaves, and spread them out in an environment of 25℃ and 60% humidity for 6 hours until the moisture content drops to 68% to 70%.

[0059] Step 2: Inoculation of compound bacterial solution: Spray the surface of the fresh leaves pretreated in step 1 with a composite bacterial solution containing the following ingredients: White rot fungus (Trametes versicolor) freeze-dried powder: 1.5% (w / w); Ginkgo biloba extract prepared in Example 3: 0.3% (w / w); The solvent was citrate buffer at pH 5.0.

[0060] Step 3: Staged wet heat-dry fermentation: The first stage (wet heat catalysis): the inoculated tea leaves are placed in a fermentation chamber at 45°C and 85% humidity, oxygen (concentration 50%) is introduced, and fermented for 12 hours; The second stage (dry oxidation): adjust to 54℃, humidity 30%, stacking thickness 50cm, fermentation 8h.

[0061] Step 4: Dynamic inactivation and drying: Microorganisms were inactivated by pulse microwaves (frequency 2450 MHz, power 300 W, intermittent treatment 3 times, 30 seconds each time), followed by drum drying at 80°C to a moisture content of 5%.

[0062] Comparative Example 1 Comparative Example 1 is the control group of Example 5, in which the ginkgo leaf extract prepared in Example 3 is removed from the composite bacterial solution in Example 5, and the rest remain unchanged. The specific process is as follows: A tea fermentation production process comprises the following steps: Step 1: Fresh leaf pretreatment: Pick fresh tea leaves with one bud and four to five leaves, and place them in an environment of 25℃ and 60% humidity for 6 hours until the moisture content drops to 68%.

[0063] Step 2: Inoculation of compound bacterial solution: Spray the surface of the fresh leaves pretreated in step 1 with a composite bacterial solution containing the following ingredients: White rot fungus (Trametes versicolor) freeze-dried powder: 1.5% (w / w); The solvent was citrate buffer at pH 5.0.

[0064] Step 3: Staged wet heat-dry fermentation: The first stage (wet heat catalysis): the inoculated tea leaves are placed in a fermentation chamber at 40°C and 80% humidity, oxygen (concentration 50%) is introduced, and fermented for 12 hours; The second stage (dry oxidation): adjust to 50℃, humidity 25%, stacking thickness 40cm, fermentation 8h.

[0065] Step 4: Dynamic inactivation and drying: Microorganisms were inactivated by pulse microwaves (frequency 2450 MHz, power 300 W, intermittent treatment 3 times, 30 seconds each time), followed by drum drying at 80°C to a moisture content of 4%.

[0066] Comparative Example 2 Comparative Example 2 is the control group of Example 5, in which the freeze-dried powder of white rot fungus (Trametes versicolor) in the composite bacterial liquid in Example 5 is removed, and the rest remain unchanged. The specific process is as follows: A tea fermentation production process comprises the following steps: Step 1: Fresh leaf pretreatment: Pick fresh tea leaves with one bud and four to five leaves, and place them in an environment of 25℃ and 60% humidity for 6 hours until the moisture content drops to 68%.

[0067] Step 2: Inoculation of compound bacterial solution: Spray the surface of the fresh leaves pretreated in step 1 with a composite bacterial solution containing the following ingredients: Ginkgo biloba extract prepared in Example 2: 0.3% (w / w); The solvent was citrate buffer at pH 5.0.

[0068] Step 3: Staged wet heat-dry fermentation: The first stage (wet heat catalysis): the inoculated tea leaves are placed in a fermentation chamber at 40°C and 80% humidity, oxygen (concentration 50%) is introduced, and fermented for 12 hours; The second stage (dry oxidation): adjust to 50℃, humidity 25%, stacking thickness 40cm, fermentation 8h.

[0069] Step 4: Dynamic inactivation and drying: Microorganisms were inactivated by pulse microwaves (frequency 2450 MHz, power 300 W, intermittent treatment 3 times, 30 seconds each time), followed by drum drying at 80°C to a moisture content of 4%.

[0070] Comparative Example 3 Comparative Example 3 is the control group of Example 5, in which the composite bacterial solution in Example 5 is removed, and the rest remain unchanged. The specific process is as follows: A tea fermentation production process comprises the following steps: Step 1: Fresh leaf pretreatment: Pick fresh tea leaves with one bud and four to five leaves, and place them in an environment of 25℃ and 60% humidity for 6 hours until the moisture content drops to 68%.

[0071] Step 2: Staged wet heat-dry fermentation: The first stage (wet heat catalysis): the inoculated tea leaves are placed in a fermentation chamber at 40°C and 80% humidity, oxygen (concentration 50%) is introduced, and fermented for 12 hours; The second stage (dry oxidation): adjust to 50℃, humidity 25%, stacking thickness 40cm, fermentation 8h.

[0072] Step 3: Dynamic inactivation and drying: Microorganisms were inactivated by pulse microwaves (frequency 2450 MHz, power 300 W, intermittent treatment 3 times, 30 seconds each time), followed by drum drying at 80°C to a moisture content of 4%.

[0073] Test Example 1 The tea leaves produced in Examples 4 to 6 and Comparative Examples 1 to 3 were subjected to performance tests. The performance test process is as follows, and the test results are shown in Table 1: 1. Test of ester catechin content: (1) Sample preparation: Take 5.0 g of tea leaves from Example 5 and Comparative Examples 1 to 3, grind them through a 60-mesh sieve, add 50 mL of 80% methanol solution, perform ultrasonic extraction for 30 min (40 kHz, 25° C.), centrifuge (8000 rpm, 10 min), and filter the supernatant through a 0.45 μm filter membrane for later use.

[0074] (2) HPLC detection conditions: Chromatographic column: Agilent ZORBAX SB-C18 (4.6×250 mm, 5 μm); Mobile phase: phase A (0.1% formic acid in water), phase B (acetonitrile); Gradient elution: 0-10 min (Phase B 10%→30%), 10-25 min (Phase B 30%→50%); Flow rate: 1.0 mL / min, detection wavelength: 280 nm, column temperature: 30 °C; (3) Standard curve: EGCG was used as the standard, 0.1-1.0 mg / mL, and the linear regression equation was y=12540x+28.6(R 2 =0.999).

[0075] 2. Test of caramel aroma content: (1) Headspace extraction: 2.0 g of tea leaves were placed in a 20 mL headspace bottle, 5 μL of internal standard (1,2-dichlorobenzene, 10 μg / mL) was added, equilibrated at 60 °C for 10 min, and inserted into a DVB / CAR / PDMS extraction fiber (Supelco) for adsorption for 40 min.

[0076] (2) GC-MS detection conditions: Instrument: Agilent 7890B-5977A; Chromatographic column: HP-5MS (30m×0.25mm×0.25μm); Program temperature rise: 40℃(2min)→5℃ / min→250℃(5min); Ionization mode: EI, 70eV, ion source temperature 230°C; Scan range: m / z 35-450; (3) Quantitative analysis: Using pyrazine and 2-acetylfuran as standard substances, the internal standard method was used to calculate the content of total caramel aroma substances.

[0077] 3. Polyphenol oxidation rate test: (1) Sample preparation: Take 5.0 g of fermented tea sample, crush it through a 60-mesh sieve, add 50 mL of 80% methanol solution, extract by ultrasonication for 30 min (40 kHz, 25°C), centrifuge (8000 rpm, 10 min), and take the supernatant and filter it through a 0.45 μm filter membrane for later use.

[0078] (2) Plotting of standard curve: Using gallic acid (GA) as the standard substance, prepare standard solutions with concentration gradients of 0.1, 0.2, 0.4, 0.6, 0.8, and 1.0 mg / mL. Take 1 mL of the standard solution, add 5 mL of Folin phenol reagent (diluted 10 times), mix well, let stand for 5 min, add 4 mL of 7.5% Na2CO3 solution, mix well, and react at room temperature in the dark for 60 min. Measure the absorbance at a wavelength of 765 nm and plot the standard curve (the linear regression equation is y=12540x+28.6, R 2 =0.999).

[0079] (3) Sample determination: Take 1 mL of sample extract and follow the above steps to determine the absorbance and calculate the polyphenol content based on the standard curve.

[0080] (4) Calculation of polyphenol oxidation rate: polyphenol oxidation rate (%) = (polyphenol content before fermentation - polyphenol content after fermentation) / polyphenol content before fermentation × 100%.

[0081] 4. Quinone content test (1) Sample preparation: Take 5.0 g of fermented tea sample, crush it through a 60-mesh sieve, add 50 mL of 80% methanol solution, extract it by ultrasonication for 30 min (40 kHz, 25°C), centrifuge it (8000 rpm, 10 min), and take the supernatant and filter it through a 0.45 μm filter membrane for later use.

[0082] (2) HPLC detection conditions: Chromatographic column: Agilent ZORBAX SB-C18 (4.6×250 mm, 5 μm).

[0083] Mobile phase: phase A (0.1% formic acid aqueous solution), phase B (acetonitrile).

[0084] Gradient elution: 0-10 min (phase B 10%→30%), 10-25 min (phase B 30%→50%).

[0085] Flow rate: 1.0 mL / min, detection wavelength: 420 nm, column temperature: 30 °C.

[0086] (3) Standard curve drawing: o-quinone was used as the standard substance, and standard solutions with concentration gradients of 0.1, 0.2, 0.4, 0.6, 0.8, and 1.0 mg / mL were prepared. 10 μL of standard solutions were injected, and the peak area was recorded to draw the standard curve (the linear regression equation was y = 24500x + 15.8, R 2 =0.998).

[0087] (4) Sample determination: Take 10 μL of sample extract and inject it, record the o-quinone peak area, and calculate the quinone content according to the standard curve.

[0088] 5. Sensory scoring test (1) Testing method: Sensory evaluation by professional tea tasters.

[0089] (2) Sample preparation: Take 5.0 g of fermented tea sample, brew it with boiling water at a ratio of 1:50 (w / v) for 5 min, filter the tea soup, and cool it to room temperature for later use.

[0090] (3) Evaluation criteria: Soup color: dark yellow with a hint of brown, clear and bright is preferred (full score: 3 points); Aroma: Strong and long-lasting aroma is preferred (full score: 3 points); Taste: mellow and sweet without bitterness (full score 4 points).

[0091] Evaluation process: 5 professional tea tasters independently score the tea, and the total score of soup color, flavor and taste is accumulated and the average score of the 5 professional tea tasters is taken.

[0092] Scoring criteria: 9-10 points: excellent quality, no obvious defects; 7-8 points: good quality, with slight defects; 5-6 points: average quality, with obvious defects; 3-4 points: poor quality, serious defects; 1-2 points: The quality is extremely poor and unacceptable.

[0093] Table 1 Test results

[0094] It should be noted that, in this article, terms such as "include", "comprises" or any other variations thereof are intended to cover non-exclusive inclusion, so that a process, method, article or equipment that includes a series of elements includes not only those elements, but also includes other elements not explicitly listed, or also includes elements inherent to such process, method, article or equipment.

[0095] Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions and variations may be made to the embodiments without departing from the principles and spirit of the invention, and the scope of the invention is defined by the appended claims and their equivalents.

Claims

1. A tea fermentation production process, characterized in that: The following steps are involved: Fresh leaf pretreatment: Pick fresh tea leaves with one bud and four to five leaves, and place them in an environment of 25℃ and 60% humidity for 6 hours until the moisture content drops to 68% to 70%; Inoculation of compound bacterial solution: spray the compound bacterial solution on the surface of pretreated fresh leaves; Staged wet heat-dry fermentation: Place the inoculated tea leaves in a fermentation chamber at 40-45°C and 80-85% humidity, introduce oxygen, ferment for 10-12 hours, then adjust to 50-54°C and 25-30% humidity, stack to a thickness of 40-50 cm, and ferment for 7-8 hours; Dynamic inactivation and drying: Pulse microwaves are used to inactivate microorganisms, followed by drying to a moisture content of 4-5% to complete the tea fermentation production process.

2. A tea fermentation production process according to claim 1, characterized in that: The concentration of the introduced oxygen is 45% to 50%.

3. A tea fermentation production process according to claim 1, characterized in that: The frequency of the pulse microwave is 2450 MHz, and the power is 300 W.

4. A tea fermentation production process according to claim 1, characterized in that: The composition of the composite bacterial solution: 0.5%~1.5%(w / w) freeze-dried white rot fungus powder; 0.1%-0.3% (w / w) Ginkgo biloba leaf extract; The balance was citrate buffer at pH 5.

0.

5. A tea fermentation production process according to claim 4, characterized in that: The ginkgo leaf extract is prepared by the following steps: The leaves of ginkgo biloba are washed with clean water, dried, and then crushed into 40-60 meshes. The ginkgo biloba powder is mixed with a 50% ethanol solution by mass at a solid-liquid ratio of 1:10-15. The extract is heated to reflux, extracted, and filtered. The extract is collected, filtered, purified by resin adsorption, and eluted. The eluate is collected, and the eluate is concentrated, dried, rotary evaporated, and spray-dried to obtain a ginkgo biloba extract.

6. A tea fermentation production process according to claim 5, characterized in that: The resin is AB-8 type macroporous adsorption resin.

7. A tea fermentation production process according to claim 5, characterized in that: The inlet air temperature of the spray drying is 150-180° C., the outlet air temperature is 80-90° C., and the atomization pressure is 0.2-0.5 MPa.

8. A tea fermentation production process according to claim 5, characterized in that: The total flavonoid glycoside content in the ginkgo leaf extract is 24.1-25.0%.

Images