Preparation method of instant golden camellia composition without pesticide residues

Abstract

The invention relates to the technical field of extraction of natural active substances, in particular to a preparation method of an instant golden camellia composition without pesticide residues, which comprises the following steps: taking golden camellia leaves and flowers, and carrying out flash extraction twice at low temperature; performing composite column chromatography, eluting with pure water, and concentrating under reduced pressure; carrying out enzymolysis, inactivation, cooling and filtration; performing macroporous resin chromatography, and eluting with pure water; and performing low-temperature concentration, cold centrifugation, freeze drying and the like. According to the preparation method, all nutritional ingredients of the california burclover leaves and flowers are reserved through low-temperature water flash extraction, pesticide residues in golden camellia are effectively removed through composite cation and anion exchange resin, the effects of decolorization and impurity removal are achieved, the content and purity of effective ingredients are optimized through enzymolysis and overload elution of macroporous resin, only water is used in the whole process, and the preparation method is safe, environmentally friendly and suitable for industrial production. The camellia nitidissima extracted product obtained by the preparation method is good in water solubility and free of pesticide residues, has tea fragrance, retains nutritional ingredients of camellia nitidissima, and also has pure taste.

Description

Technical Field

[0001] The invention relates to the technical field of natural active substance extraction, and in particular to a method for preparing an instant camellia chrysantha composition free of pesticide residues. Background Art

[0002] Camellia chrysantha belongs to the Camellia genus, subgenus Camellia, group Camellia chrysantha.

[0003] Existing Camellia chrysantha products are mainly tea products. The preparation of the products is mostly water extraction, and then other substances are added to make products, such as health tea, white tea, health fruit wine, with low extraction rate and light taste. The extracted components are mainly single polyphenols, polysaccharides, etc., such as the method for extracting polyphenols from Camellia chrysantha tea disclosed in Publication (Announcement) No. CN105168593A, which only extracts polyphenols, and the method for extracting Camellia chrysantha polysaccharides from Camellia chrysantha tea disclosed in CN105175569A, which only extracts Camellia chrysantha polysaccharides. And most of them have not been removed by agricultural residues, and the water-soluble ones are not clear and transparent, which will limit their scope of use. In order to better apply the requirements of various fields and retain the effective ingredients of the original plants at the same time, it is necessary to develop a new Camellia chrysantha composition, taking into account the retention of Camellia chrysantha nutrients, and the product has good water solubility, no agricultural residues, and tea fragrance to make up for the deficiencies of the prior art. Summary of the invention

[0004] The object of the present invention is to provide a method for preparing an instant camellia chrysantha composition free of pesticide residues, the preparation method comprising the following steps:

[0005] (1) Take the raw materials of golden camellia tea and flowers, crush them, pass through 200 mesh, and perform low-temperature flash extraction twice, and combine the filtrates of the two extractions to obtain a total filtrate;

[0006] (2) passing the total filtrate through a composite column chromatography, eluting with pure water, collecting the effluent and the water washings, and concentrating under reduced pressure;

[0007] (3) enzymatic hydrolysis, inactivation, cooling, and filtration;

[0008] (4) passing through a macroporous resin for chromatography and eluting with pure water until the effluent is clear and transparent, and collecting the effluent and the pure water eluate;

[0009] (5) low temperature concentration, sealing and keeping at low temperature for 12 hours, centrifuging while cold, taking the supernatant, and freeze-drying to obtain the Camellia chrysantha composition.

[0010] The step (1) specifically comprises mixing golden camellia tea leaves and flowers in a mass ratio of 2-5:1-2, crushing, passing through 200 meshes, flash-extracting with 4-10 times amount of pure water at 6-12° C., extracting twice, combining the filtrates from the two extractions, and obtaining a total filtrate.

[0011] Further, the material is put into a flash extractor, and the solvent is used as the extraction solvent. 2-5 times the amount of solvent is added each time. The motor speed is 10000-15000R / min. The extraction is performed at room temperature for 2 minutes, and the extraction is performed twice, each time for one minute. After each extraction, there is a rest for 1 minute. The filtrate is separated, and the filtrates from the two extractions are combined to obtain the total filtrate.

[0012] The step (2) is specifically as follows: after the total filtrate is completed, elution is performed with pure water in an amount 1-3 times the mass of the resin, the effluent and the water washing liquid are collected and combined, and concentrated to 5-6% of the solid content by a reduced pressure concentration device. The total amount of the mixed resin and the weight ratio of the raw materials are 1:4-6.

[0013] The composite column in step (2) consists of an upper layer and a lower layer, the upper layer is an anion resin, the lower layer is a cationic resin, the mass ratio of the anion resin to the cationic resin is 1-2:1-3. The model of the anion resin is D201, D2006 or D918, and the manufacturer is Amicogen Biopharmaceutical Co., Ltd. The model of the cationic resin is D001, D113 or D111, and the manufacturer is Amicogen Biopharmaceutical Co., Ltd.

[0014] The step (3) specifically comprises adding 0.5-1‰ of the weight of the raw material pectin lyase, performing enzymolysis at 48-52°C for 2-3 hours, heating to greater than 98°C, maintaining for 4-8 minutes to inactivate, cooling the solution to a temperature of 35-40°C, filtering through a 200-mesh filter cloth, and obtaining a filtrate after enzymolysis.

[0015] The step (4) specifically comprises subjecting the filtrate after enzymatic hydrolysis to macroporous resin chromatography, and then eluting with 1-3 times the mass of the resin in pure water until the effluent is clear and transparent, and collecting the effluent and the pure water eluate.

[0016] Furthermore, the weight ratio of the macroporous resin to the raw material is 1:0.4-0.1, the model is D18, D285 or DM28, and the manufacturer is Amicogen Biopharmaceutical Co., Ltd.

[0017] The step (5) specifically comprises the following steps: using a low temperature concentrator to concentrate the effluent obtained in step (4) and the pure water eluate to 25-30 BRIX at a temperature below 60°C, sealing the effluent, and maintaining the effluent at 4-10°C for 12 hours; centrifuging the effluent at a temperature below 10°C while cooling the effluent, taking the supernatant, and freeze-drying the effluent to obtain the Camellia chrysantha composition.

[0018] The golden camellia composition contains 5-20% polyphenols, 5-35% flavonoids and 6-25% polysaccharides.

[0019] Compared with the prior art, the present invention has the following beneficial effects:

[0020] (1) The preparation method adopts low-temperature water flash extraction and short-time extraction, so that the extracted effective ingredients can be better preserved, while retaining all the nutrients of the leaves and flowers of Camellia chrysantha; the composite cation and anion exchange resin can effectively remove the pesticide residues in Camellia chrysantha, while taking into account the effects of decolorization and impurity removal, thereby increasing the content of effective ingredients; enzymatic hydrolysis plus overload elution of macroporous resin, utilizing the separation of effective ingredients and impurities, optimizes the content purity of the effective ingredients, and at the same time can effectively remove the remaining trace pesticide residues again, ensuring that the product is free of pesticide residues; maintaining low-temperature centrifugation can effectively improve the water solubility of the product, and freeze-drying is selected to maximize the original physical properties of the product, and the product meets the needs of the development of big health.

[0021] (2) The entire process only uses water, which is safe and environmentally friendly. The process is simple and has good adaptability, making it suitable for large-scale factory production.

[0022] (3) The preparation method obtains a golden camellia extract composition, which has good water solubility, no pesticide residues, tea fragrance, retains the nutritional components of golden camellia, and also takes into account the pure taste. The aqueous solution has a good color. This composition has been tested and found to contain 5-20% polyphenols, 5-35% flavonoids, and 6-25% polysaccharides, and can be widely used in various aspects, such as food, beverages, etc. DETAILED DESCRIPTION

[0023] The specific implementation of the present invention is further described below in conjunction with examples, but the implementation and protection of the present invention are not limited thereto. It should be noted that if there are processes that are not particularly described in detail below, they can be implemented or understood by those skilled in the art with reference to the prior art.

[0024] Example 1

[0025] Take 1000g of tea leaves and flowers mixed in a mass ratio of 5:1, crush them, and pass through 200 meshes. Use 6 times the amount of pure water at 6°C, put the material into a flash extractor, use solvent as the extraction solvent, add 3 times the amount of solvent each time, the motor speed is 10000R / min, extract for 2 minutes at room temperature, extract 2 times, each time for one minute, rest for 1 minute in the middle after extracting once, separate the filtrate, merge the filtrate extracted twice, and obtain the total filtrate. The total filtrate is passed through a composite column chromatography, and the composite column consists of an upper layer and a lower layer, the upper layer is an anion resin (Aimi Biopharmaceutical Co., Ltd., D918), and the lower layer is a cationic resin (Aimi Biopharmaceutical Co., Ltd., D111). The ratio of the two is 1:1, and the total amount of mixed resin is 200g. After entering, 400mL of pure water is used for elution, and the effluent and water washing liquid are collected and combined. Use a vacuum concentration device to concentrate to 5% solids, then add 0.5mL pectin lyase, and enzymolyze at 48°C for 2 hours. After the enzymolysis is completed, heat to 100°C and keep for 4 minutes to inactivate. When the liquid is cooled to 35°C, pass through a 200-mesh filter cloth. Then pass through a macroporous resin chromatography (Aimi Biopharmaceutical Co., Ltd., D18), with a resin dosage of 100g. After entering, elute with 200g of pure water until the elution is clear and transparent. Collect the effluent and pure water eluent, use a low-temperature concentrator (temperature <60°C) to concentrate to 25BRIX, seal well, keep at a low temperature of 10°C for 12h, and centrifuge while cold. Take the supernatant and freeze-dry directly to obtain a golden camellia composition. This composition has been tested and found to contain 10% polyphenols, 12% flavonoids, and 15% polysaccharides.

[0026] Example 2

[0027] Take 1000g of golden camellia tea and flower mixed in a mass ratio of 2:1, crush, and pass 200 meshes. Use 4 times the amount of pure water at 12°C, put the material into a flash extractor, use solvent as the extraction solvent, add 3 times the amount of solvent each time, motor speed 15000R / min, extract for 2 minutes at room temperature, extract 2 times, one minute each time, rest for 1 minute in the middle after extracting once, separate the filtrate, merge the filtrate extracted twice, and obtain the total filtrate. The total filtrate is passed through a composite column chromatography, the composite column consists of an upper layer and a lower layer, the upper layer is an anion resin (Aimi Biopharmaceutical Co., Ltd., D201), and the lower layer is a cationic resin (Aimi Biopharmaceutical Co., Ltd., D001). The ratio of the two is 2:3, and the total amount of mixed resin is 250g. After entering, 250mL of pure water is used for elution, the effluent and washing liquid are collected and combined, and the solid content is 6% with a reduced pressure concentration device. Add 1mL pectin lyase (Novozymes) and enzymolyze at 52°C for 2 hours. After the enzymolysis is completed, heat to 99°C and keep for 8 minutes to inactivate. When the liquid is cooled to 40°C, pass through a 200-mesh filter cloth. Then pass through a macroporous resin chromatography (Aimi Biopharmaceutical Co., Ltd., D285), with a resin dosage of 400g. After the addition, elute with 400g of pure water until the effluent is clear and transparent. Collect the effluent and pure water eluate, concentrate to 30BRIX using a low-temperature concentrator (temperature <60°C), seal well, keep at a low temperature of 4°C for 12h, and centrifuge while cold. Take the supernatant and freeze-dry directly to obtain a golden camellia composition. This composition has been tested and found to contain 12% polyphenols, 25% flavonoids, and 20% polysaccharides.

[0028] Example 3

[0029] Take 1000g of tea leaves and flowers mixed in a mass ratio of 1:1, crush them, and pass through 200 meshes. Use 10 times the amount of pure water at 10°C, put the material into a flash extractor, use solvent as the extraction solvent, add 3 times the amount of solvent each time, the motor speed is 12000R / min, extract for 2 minutes at room temperature, extract 2 times, each time for one minute, rest for 1 minute in the middle after extracting once, separate the filtrate, merge the filtrate extracted twice, and obtain the total filtrate. The total filtrate is passed through a composite column chromatography, and the composite column consists of an upper layer and a lower layer, the upper layer is an anion resin (Aimi Biopharmaceutical Co., Ltd., D2006), and the lower layer is a cationic resin (Aimi Biopharmaceutical Co., Ltd., D113). The ratio of the two is 2:1, and the total amount of mixed resin is 180g. After entering, elute with 540ml of pure water, and merge and collect the effluent and water washing liquid. Use a vacuum concentration device to concentrate to 6% solids, add 8mL pectin lyase (Novozymes), and enzymolyze at 50°C for 3 hours. After enzymolysis, heat to 99°C and keep for 6 minutes to inactivate. When the liquid cools to 38°C, pass through a 200-mesh filter cloth. Then pass through a macroporous resin chromatography (Aimi Biopharmaceutical Co., Ltd., DM28), with a resin dosage of 200g. After entering, elute with 600g of pure water until the effluent is clear and transparent. Collect the effluent and pure water eluate, use a low-temperature concentrator (temperature <60°C) to concentrate to 28BRIX, seal well, keep at a low temperature of 6°C for 12h, and centrifuge while cold. Take the supernatant and freeze-dry directly to obtain a golden camellia composition. This composition has been tested and found to contain 7% polyphenols, 30% flavonoids, and 25% polysaccharides.

[0030] The results prepared in Examples 1-3 were tested for pesticide residues according to the following test methods: (1) GB / T5009.19-2008 Method for determining multi-component residues of organochlorine pesticides in food, (2) GB / T5009.20-2003 Method for determining organophosphorus pesticide residues in food, (3) GB 23200.3-2016 Method for determining cyclohexanone herbicide residues in food by liquid chromatography-mass spectrometry / mass spectrometry, and (4) GB 23200.6-2016 Method for determining chlorpyrifos residues in food by liquid chromatography-mass spectrometry / mass spectrometry. No pesticide residues were detected.

[0031] The above embodiments are only preferred implementation modes of the present invention and are only used to explain the present invention rather than to limit the present invention. Any changes, substitutions, modifications, etc. made by those skilled in the art without departing from the spirit of the present invention should fall within the scope of protection of the present invention.

Claims

1. A method for preparing an instant pesticide-free Camellia chrysantha composition, characterized in that: The preparation method comprises the following steps: (1) Take the raw materials of golden camellia tea and flowers, crush them, pass through 200 mesh, and perform low-temperature flash extraction twice, and combine the filtrates of the two extractions to obtain a total filtrate; (2) passing the total filtrate through a composite column chromatography, eluting with pure water, collecting the effluent and the water washings, and concentrating under reduced pressure; (3) enzymatic hydrolysis, inactivation, cooling, and filtration; (4) passing through a macroporous resin for chromatography and eluting with pure water until the effluent is clear and transparent, and collecting the effluent and the pure water eluate; (5) low temperature concentration, sealing and keeping at low temperature for 12 hours, centrifuging while cold, taking the supernatant, and freeze-drying to obtain the Camellia chrysantha composition.

2. The preparation method according to claim 1, characterized in that: The step (1) specifically comprises mixing golden camellia tea leaves and flowers in a mass ratio of 2-5:1-2, crushing, passing through 200 meshes, flash-extracting with 4-10 times amount of pure water at 6-12° C., extracting twice, combining the filtrates from the two extractions, and obtaining a total filtrate.

3. The preparation method according to claim 2, characterized in that: Put the material into a flash extractor, use solvent as the extraction solvent, add 2-5 times the amount of solvent each time, the motor speed is 10000-15000R / min, extract for 2 minutes at room temperature, extract twice, each time for one minute, rest for 1 minute in between each extraction, separate the filtrate, combine the filtrates from the two extractions to obtain the total filtrate.

4. The preparation method according to claim 1, characterized in that: The step (2) is specifically as follows: after the total filtrate is completed, elution is performed with pure water in an amount 1-3 times the mass of the resin, the effluent and the water washing liquid are collected and combined, and concentrated to 5-6% of the solid content by a reduced pressure concentration device. The total amount of the mixed resin and the weight ratio of the raw materials are 1:4-6.

5. The preparation method according to claim 1, characterized in that: The composite column in step (2) consists of an upper layer and a lower layer, the upper layer is an anion resin, the lower layer is a cationic resin, the mass ratio of the anion resin to the cationic resin is 1-2:1-3. The model of the anion resin is D201, D2006 or D918, and the manufacturer is Amicogen Biopharmaceutical Co., Ltd. The model of the cationic resin is D001, D113 or D111, and the manufacturer is Amicogen Biopharmaceutical Co., Ltd.

6. The preparation method according to claim 1, characterized in that: The step (3) specifically comprises adding 0.5-1‰ of the weight of the raw material pectin lyase, performing enzymolysis at 48-52°C for 2-3 hours, heating to greater than 98°C, maintaining for 4-8 minutes to inactivate, cooling the solution to a temperature of 35-40°C, filtering through a 200-mesh filter cloth, and obtaining a filtrate after enzymolysis.

7. The preparation method according to claim 1, characterized in that: The step (4) specifically comprises subjecting the filtrate after enzymatic hydrolysis to macroporous resin chromatography, and then eluting with pure water in an amount of 1 to 3 times the weight of the resin until the effluent is clear and transparent, and collecting the effluent and the pure water eluate.

8. The preparation method according to claim 7, characterized in that: The weight ratio of the macroporous resin to the raw material is 1:0.4-0.1, the model is D18, D285 or DM28, and the manufacturer is Aimi Biopharmaceutical Co., Ltd.

9. The preparation method according to claim 1, characterized in that: The step (5) specifically comprises the following steps: using a low temperature concentrator to concentrate the effluent obtained in step (4) and the pure water eluate to 25-30 BRIX at a temperature below 60°C, sealing the effluent, and maintaining the effluent at 4-10°C for 12 hours; centrifuging the effluent at a temperature below 10°C while cooling the effluent, taking the supernatant, and freeze-drying the effluent to obtain the Camellia chrysantha composition.

10. The preparation method according to claim 1, characterized in that: The golden camellia composition contains 5-20% polyphenols, 5-35% flavonoids and 6-25% polysaccharides.