High-purity sasanquasaponin and extraction method therefor

Abstract

Disclosed in the present invention are high-purity sasanquasaponin and an extraction method therefor. The extraction method comprises the following steps: crushing sasanqua seed meal or tea seed meal, subjecting the crushed sasanqua seed meal or tea seed meal to dynamic countercurrent extraction by using an aqueous lower alcohol solution, and collecting the filtrate; concentrating the filtrate to recover the alcohol until the content of the alcohol in the concentrated solution is less than 3 wt%, so as to obtain a concentrated solution A; diluting the concentrated solution A with water, treating the diluted solution with an ultrafiltration membrane, and concentrating the ultrafiltration membrane filtrate with a nanofiltration membrane to obtain a concentrated solution B; adding water to the concentrated solution to obtain a dissolved solution, decolorizing the dissolved solution, adsorbing the decolorized solution with a macroporous adsorption resin, and then eluting same with an ethanol solution to obtain an eluate; and concentrating, sterilizing, filtering and drying the eluate to obtain sasanquasaponin. The sasanquasaponin prepared in the present invention has a high purity, a low content of impurities, and a light color, and is widely used in industries such as high-end washing and care products and cosmetics. The preparation method is simple, feasible, and suitable for large-scale production.

Description

A high-purity camellia saponin and its extraction method

[0001] This application claims priority to Chinese Patent Application No. 202411841228.3, filed on December 13, 2024, the disclosure of which is incorporated herein by reference in its entirety. Technical Field

[0002] This invention relates to the field of bio-extraction technology, and in particular to a high-purity camellia saponin and its extraction method. Background Technology

[0003] Tea saponins, also known as tea saponins, are a class of glycoside compounds extracted from the seeds of Camellia oleifera, Camellia sinensis, or tea. They are high-performance natural surfactants that can be widely used in light industry, chemical industry, pesticides, feed, aquaculture, textiles, oil extraction, mining, building materials, and highway construction.

[0004] The camellia saponins prepared using existing technologies contain various impurities such as proteins, polysaccharides, flavonoids, polyphenols, organic acids, tannins, tannins, and pigments. These impurities affect the product's formulation advantages and performance stability, severely impacting the unique surface-active functions and antibacterial and anti-dandruff effects of camellia saponins, limiting their application to pesticide adjuvants, low-end detergents, oil extraction, and building materials. Refining and purifying camellia saponins requires further removal of impurities such as proteins, camellia polysaccharides, flavonoids, polyphenols, organic acids, tannins, tannins, and other pigments from the crude saponins, especially colored impurities. This is a key technological challenge in the preparation of high-purity camellia saponins. Existing refining and purification methods include flocculation, precipitation, resin adsorption, hydrogen peroxide oxidation, or repeated recrystallization. However, these methods either have poor impurity removal effects, are complex and cumbersome, difficult to control, result in significant camellia saponin loss, or introduce and generate new chemical pollutants that are difficult to completely remove, making them unsuitable for large-scale industrial production. Summary of the Invention

[0005] In view of this, the purpose of this invention is to provide a high-purity camellia oleifera saponin and its extraction method. The high-purity camellia oleifera saponin has high purity, low impurity content, and light color, and can be widely used in high-end personal care products, cosmetics, pharmaceuticals, and the food industry. The preparation method is simple and feasible, and suitable for large-scale industrial production.

[0006] In a first aspect, the present invention provides a method for extracting high-purity camellia saponins, comprising the following steps:

[0007] The camellia seed cake or tea seed cake is crushed, and the crushed camellia seed cake or tea seed cake is dynamically countercurrently extracted with an aqueous low alcohol solution, and the filtrate is collected.

[0008] The filtrate was concentrated to recover alcohol until the alcohol content in the concentrate was less than 3 wt%, thus obtaining concentrate A.

[0009] Concentrate A is diluted with water, the diluted solution is treated with an ultrafiltration membrane, and the permeate from the ultrafiltration membrane is concentrated with a nanofiltration membrane to obtain concentrate B.

[0010] Water is added to the concentrated solution B to obtain a solution. The solution is then decolorized, and the decolorized solution is adsorbed by a macroporous adsorption resin and then eluted with an ethanol solution to obtain an eluent.

[0011] The eluent was concentrated, sterilized, filtered, and dried to obtain high-purity camellia saponins.

[0012] Compared with existing technologies, the preparation method of this invention first involves pulverizing camellia seed cake or tea seed cake, then performing dynamic countercurrent extraction on the pulverized camellia seed cake or tea seed cake to obtain a filtrate. The filtrate is then concentrated, diluted with water, treated with an ultrafiltration membrane, and the permeate from the ultrafiltration membrane is concentrated via nanofiltration to effectively remove macromolecular substances and small inorganic salts. Next, water is added to the concentrated solution for dissolution, decolorization, resin adsorption of the decolorized solution, and ethanol elution, further improving the purity of camellia saponins. Finally, the elution solution is treated to obtain high-purity camellia saponins. This invention's method for preparing high-purity camellia saponins results in high-purity camellia saponins with low impurity content and light color, making them widely applicable in high-end personal care products, cosmetics, pharmaceuticals, and food industries. Furthermore, the preparation method is simple and feasible, suitable for large-scale industrial production.

[0013] Furthermore, the conditions for pulverizing camellia seed cake or tea seed cake are as follows: the feeding speed is 10-200 kg / h, and the frequency is 30-50 Hz; the particle size of the pulverized camellia seed cake or tea seed cake is 0.1-2 mm; when performing dynamic countercurrent extraction on the pulverized camellia seed cake or tea seed cake, the extraction temperature is 30-50℃, the extraction frequency is 30-50 Hz, and the extraction propulsion speed is 5-10 m / h; the aqueous lower alcohol solution includes lower alcohol, compound salt, and water, wherein the mass concentration of lower alcohol is 50-90%, and the mass concentration of compound salt is 0.1-0.5%; the compound salt includes one or more of sulfites, bisulfites, or metabisulfites.

[0014] Compared with the prior art, the preparation method of the present invention uses an aqueous lower alcohol solution containing 0.1-0.5 wt% sulfite, bisulfite or metabisulfite, instead of water or an alcohol solution without the above-mentioned compound salts in the prior art. This can reduce the leaching of oils, proteins and macromolecular tannins in the raw material extraction, make filtration easier, reduce the further oxidation of some phenolic acid compounds during the extraction process, reduce the increase of color-developing substances, prevent the liquid from darkening, and reduce the burden of product decolorization.

[0015] Furthermore, the method for concentrating and recovering alcohol from the filtrate is nanofiltration membrane concentration, wherein the nanofiltration membrane has a specification of 150-200 Da, the operating pressure of the nanofiltration membrane is 0.5-3.0 MPa, the temperature is 10-65℃, the pH value is 3.0-12.0, and the flow rate of the filtrate is 0.5-5.0 m³ / h.

[0016] Further, the Brix value of the diluted solution after dilution with water is 1-10%; when the diluted solution is treated with an ultrafiltration membrane, the ultrafiltration membrane has a specification of 15-20 kDa, the inlet membrane pressure is 0.1-1.0 MPa, the temperature is 10-65℃, the pH value is 3.0-12.0, and the flow rate of the diluted solution is 0.5-5.0 m³ / h, retaining macromolecules with a molecular weight greater than 20 kDa, while continuously adding water for dialysis until the Brix value of the permeate is ≤0.5%; when the permeate from the ultrafiltration membrane is concentrated with a nanofiltration membrane, the nanofiltration membrane has a specification of 400-600 Da, the operating pressure is 0.1-3.0 MPa, the temperature is 10-65℃, the pH value is 3.0-12.0, and the flow rate of the permeate from the ultrafiltration membrane is 0.5-5.0 m³ / h, until the Brix value of the concentrated solution is 25-30%, thus obtaining concentrated solution B.

[0017] Furthermore, the Brix value of the solution is 8-10%.

[0018] Furthermore, the decolorization method of the solution is as follows: the solution is decolorized by adsorption with macroporous adsorption resin or by ion exchange with anion exchange resin. When the solution is decolorized by adsorption with macroporous adsorption resin or by ion exchange with anion exchange resin, the loading rate of the solution is 0.5-1 BV / h, and the loading volume is 0.5-5.0 BV of the resin volume. After the decolorization by macroporous adsorption resin or anion exchange resin is completed, a decolorized solution is obtained. The macroporous adsorption resin is a polystyrene macroporous adsorption resin or an acrylate macroporous adsorption resin, and the anion exchange resin is a polystyrene skeleton or an acrylate skeleton anion exchange resin.

[0019] Furthermore, when the decolorizing solution is adsorbed by the macroporous adsorption resin, the loading rate of the decolorizing solution is 1-2 BV / h, and the loading amount is 0.5-5.0 BV of the resin volume; when the ethanol solution is desorbed, the loading rate of the ethanol solution is 0.5-1 BV / h, and the loading amount is 1.0-5.0 BV of the resin volume.

[0020] Furthermore, the method for concentrating the eluent is as follows: the eluent is concentrated using a membrane. When the solid content of the concentrate reaches 20-30%, it is transferred to a single-effect external circulation concentrator for further concentration until the Brix value reaches 40-50%. The membrane used for membrane concentration has a specification of 100-200 Da. When the single-effect external circulation concentrator continues to concentrate, the concentration vacuum degree is 0.05-0.095 MPa, and the concentration temperature is 50-85℃.

[0021] Furthermore, the sterilization temperature is 80-85℃, and the sterilization time is 30-35 minutes;

[0022] Furthermore, the drying method is spray drying, with a feed rate of 10-100 L / h, an inlet air temperature of 180-190℃, and an outlet air temperature of 85-90℃.

[0023] Secondly, the present invention provides a high-purity camellia saponin with a purity of 97% or higher, a protein content of ≤1.0wt%, a polyphenol content of ≤1.0g / kg, and an absorbance Abs <0.06 at a wavelength of 420-610nm.

[0024] Compared with the prior art, the beneficial effects of the high-purity camellia saponins of the present invention are the same as those of the camellia saponin preparation method described above, and will not be repeated here. Detailed Implementation

[0025] To make the technical problems to be solved, the technical solutions, and the beneficial effects of the present invention clearer, the present invention will be further described in detail below with reference to specific embodiments. It should be understood that the specific embodiments described herein are only for explaining the present invention and are not intended to limit the present invention.

[0026] This invention provides a method for extracting high-purity camellia saponins, comprising the following steps:

[0027] The camellia seed cake or tea seed cake is crushed, and the crushed camellia seed cake or tea seed cake is dynamically countercurrently extracted with an aqueous low alcohol solution, and the filtrate is collected.

[0028] The filtrate was concentrated to recover alcohol until the alcohol content in the concentrate was less than 3 wt%, thus obtaining concentrate A.

[0029] Concentrate A is diluted with water, the diluted solution is treated with an ultrafiltration membrane, and the permeate from the ultrafiltration membrane is concentrated with a nanofiltration membrane to obtain concentrate B.

[0030] Water was added to concentrate B to obtain a solution. The solution was decolorized, and the decolorized solution was adsorbed by macroporous adsorption resin and then eluted with ethanol solution to obtain an eluent.

[0031] The eluent was concentrated, sterilized, filtered, and dried to obtain high-purity camellia saponins.

[0032] The preparation method of this invention first involves pulverizing camellia seed cake or tea seed cake, then performing dynamic countercurrent extraction on the pulverized camellia seed cake or tea seed cake using an aqueous low-grade alcohol solution to obtain a filtrate, which fully extracts camellia saponins from the camellia seed cake or tea seed cake. The filtrate is then concentrated to remove alcohols. The resulting concentrate A is diluted with water and treated with an ultrafiltration membrane to effectively remove high-molecular-weight and large-molecule proteins, camellia polysaccharides, starch, and other macromolecules from the diluted solution. The ultrafiltration permeate is then concentrated using a nanofiltration membrane to obtain concentrate B, further removing small-molecule inorganic salts from the ultrafiltration permeate, resulting in a significantly improved purity of concentrate B. Concentrate B is then subjected to water dissolution, decolorization, resin adsorption, and ethanol elution to obtain an eluent, removing colored substances and impurities from the camellia saponins, further improving their purity. Finally, the eluent is further processed to obtain high-purity camellia saponins. The present invention provides a method for preparing high-purity camellia saponins, which results in high purity, low impurity content, and light color of the prepared high-purity camellia saponins. It can be widely used in high-end personal care products, cosmetics, pharmaceuticals, food and other industries. Moreover, the preparation method is simple and feasible, and is suitable for large-scale industrial production.

[0033] In some embodiments, the conditions for pulverizing camellia seed cake or tea seed cake are as follows: the feeding speed is 10-200 kg / h, and the frequency is 30-50 Hz; the particle size of the pulverized camellia seed cake or tea seed cake is 0.1-2 mm; when the pulverized camellia seed cake or tea seed cake is subjected to dynamic countercurrent extraction, the extraction temperature is 30-50℃, the extraction frequency is 30-50 Hz, and the extraction propulsion speed is 5-10 m / h; the aqueous lower alcohol solution includes lower alcohol, compound salt, and water, wherein the mass concentration of lower alcohol is 50-90%, and the mass concentration of compound salt is 0.1-0.5%, preferably 0.2-0.4%; the compound salt includes one or more of sulfites, bisulfites, or metabisulfites. The lower alcohol in the aqueous lower alcohol can be methanol, ethanol, n-propanol, isopropanol and n-butanol, etc.; sulfites, bisulfites or metabisulfites can be mixed in any mass ratio to finally meet the content requirements in the aqueous lower alcohol solution. Preferably, the mass ratio of sulfites, bisulfites and metabisulfites can be (1-2):(1-2):(2-3).

[0034] It should be noted that the frequency of grinding camellia seed cake or tea seed cake refers to the working frequency of the grinder.

[0035] It should be noted that when performing dynamic countercurrent extraction on crushed camellia seed cake or tea seed cake, the extraction temperature refers to the actual working temperature when the aqueous low alcohol solution comes into contact with the crushed camellia seed cake or tea seed cake; the extraction frequency refers to the operating frequency range of the motor of the extraction equipment; and the extraction propulsion speed refers to the linear velocity of the crushed camellia seed cake or tea seed cake moving forward in the extraction equipment.

[0036] Specifically, the feeding rate for crushing camellia seed cake or tea seed cake can be 10 kg / h, 50 kg / h, 100 kg / h, 160 kg / h, or 200 kg / h; the crushing frequency can be 30 Hz, 35 Hz, 40 Hz, 45 Hz, or 50 Hz; and the particle size of the crushed camellia seed cake or tea seed cake can be 0.1 mm, 0.6 mm, 1.0 mm, 1.6 mm, or 2 mm. The extraction temperature for pulverized camellia seed cake or tea seed cake can be 30℃, 33℃, 38℃, 45℃, or 50℃; the extraction frequency for pulverized camellia seed cake or tea seed cake can be 30Hz, 35Hz, 40Hz, 42Hz, or 50Hz; and the extraction propulsion speed for pulverized camellia seed cake or tea seed cake can be 5m / h, 8m / h, 8.5m / h, 9m / h, or 10m / h.

[0037] Under the combined effect of the above parameters and an aqueous low-alcohol solution, camellia seed cake or tea seed cake is pulverized and extracted, allowing for the full extraction of camellia saponins from the camellia seed cake or tea seed cake. The aqueous low-alcohol solution contains 0.1-0.5 wt% of one or more of sulfites, bisulfites, or metabisulfites, which can reduce the leaching of oils, proteins, and macromolecular tannins during the extraction process, facilitate filtration, reduce further oxidation of some phenolic acid compounds during extraction, reduce the increase of color-developing substances, prevent the solution from darkening, and reduce the burden of product decolorization.

[0038] In some embodiments, the method for concentrating the filtrate to recover alcohol is nanofiltration membrane concentration, wherein the nanofiltration membrane has a specification of 150-200 Da, a pressure of 0.5-3.0 MPa, a temperature of 10-65°C, a pH value of 3.0-12.0, and the flow rate of the filtrate is 0.5-5.0 m³ / h.

[0039] It should be noted that the pressure refers to the transmembrane operating pressure during nanofiltration membrane concentration of the filtrate; the temperature and pH value refer to the temperature and pH value of the filtrate.

[0040] Specifically, the nanofiltration membrane can have specifications of 150 Da, 165 Da, 185 Da, 190 Da, and 200 Da, pressures of 0.5 MPa, 0.9 MPa, 1.5 MPa, 2.5 MPa, and 3 MPa, temperatures of 10℃, 20℃, 35℃, 50℃, and 65℃, pH values ​​of 3, 4, 5, 9, and 12, and filtrate flow rates of 0.5 m³ / h, 1.6 m³ / h, 2.5 m³ / h, 3.8 m³ / h, and 5 m³ / h.

[0041] When the filtrate is concentrated using nanofiltration membranes, under the above-mentioned parameters, the filtrate can be sufficiently concentrated to remove the alcohol. In practice, the alcohol recovered after concentration can be subjected to distillation and recycled.

[0042] In some embodiments, after the concentrate A is diluted with water, the Brix value of the diluted solution is 1-10%. When the diluted solution is treated with an ultrafiltration membrane, the ultrafiltration membrane has a specification of 15-20 kDa, the inlet membrane pressure is 0.1-1.0 MPa, the temperature is 10-65℃, the pH value is 3.0-12.0, the flow rate of the diluted solution is 0.5-5.0 m³ / h, and large molecules with a molecular weight greater than 20 kDa are retained. At the same time, water is continuously added for dialyzing until the Brix value of the permeate is ≤0. The process ends at 5%; when the ultrafiltration membrane permeate is concentrated by a nanofiltration membrane with a specification of 400-600 Da, it can allow substances with a molecular weight less than 400 Da to pass through and retain substances with a molecular weight greater than 600 Da. The operating pressure is 0.1-3.0 MPa, the temperature is 10-65℃, and the pH value is 3.0-12.0. The flow rate of the ultrafiltration membrane permeate is 0.5-5.0 m³ / h. The process ends when the Brix value of the concentrate is 25-30%, yielding concentrate B.

[0043] It should be noted that the Brix value refers to the content of soluble solids in a liquid.

[0044] It should be noted that when the diluent is treated by an ultrafiltration membrane, the inlet membrane pressure of 0.1-1.0 MPa during operation means that the inlet pressure of the membrane separation equipment should be maintained at 0.1-1.0 MPa during operation; the temperature and pH value refer to the temperature and pH value of the diluent.

[0045] It should be noted that when the permeate from the ultrafiltration membrane is concentrated by the nanofiltration membrane, the temperature and pH value refer to the temperature and pH value of the permeate from the ultrafiltration membrane.

[0046] Specifically, after dilution of concentrate A with water, the Brix value can be 1%, 3%, 6%, 8%, or 10%; the specifications of the ultrafiltration membrane can be 15 kDa, 17 kDa, 18 kDa, 19 kDa, or 20 kDa; the inlet membrane pressure during operation can be 0.1 MPa, 0.3 MPa, 0.5 MPa, 0.8 MPa, or 1.0 MPa; the temperature can be 10℃, 25℃, 37℃, 50℃, or 65℃; the pH value can be 3, 5, 7, 9, or 12; the flow rate of the diluent can be 0.5 m³ / h, 1.6 m³ / h, 3.0 m³ / h, 3.8 m³ / h, or 5 m³ / h; and at the end of ultrafiltration treatment, the Brix value of the permeate can be 0.1%, 0.2%, 0.35%, 0.4%, or 0.5%. When the permeate from the ultrafiltration membrane is concentrated by the nanofiltration membrane, the nanofiltration membrane specifications can be 400Da, 460Da, 500Da, 550Da, and 600Da; the operating pressure can be 0.1MPa, 0.8MPa, 1.5MPa, 2.0MPa, and 3.0MPa; the temperature can be 10℃, 25℃, 37℃, 50℃, and 65℃; the pH value can be 3, 6, 8, 10, and 12; the flow rate of the permeate from the ultrafiltration membrane can be 0.5m³ / h, 1.6m³ / h, 3.0m³ / h, 3.8m³ / h, and 5m³ / h; and the Brix value of the concentrate B can be 25%, 26.5%, 27.5%, 28%, and 30%.

[0047] Under the above membrane specifications and parameters, high molecular weight and large molecular weight proteins, tea oil polysaccharides, starch, macromolecular substances and small molecular weight inorganic salts in the diluent are fully removed, thereby improving the purity of tea oil saponin in the diluent.

[0048] In some embodiments, the Brix value of the solution is 8-10%; the decolorization method of the solution is adsorption by macroporous adsorption resin or ion exchange decolorization by anion exchange resin; when the solution is adsorbed by macroporous adsorption resin or ion exchange decolorized by anion exchange resin, the loading rate of the solution is 0.5-1 BV / h, and the loading amount is 0.5-5.0 BV of the resin volume; after the macroporous adsorption resin or anion exchange resin decolorization is completed, a decolorized solution is obtained; the macroporous adsorption resin is a weakly polar to polar macroporous resin with a particle size of 250 μm-1.25 mm, and can be polystyrene macroporous adsorption resin or acrylate macroporous adsorption resin; the anion exchange resin is a weakly basic anion exchange resin with a particle size of 250 μm-2 mm, and can be polystyrene skeleton or acrylate skeleton anion exchange resin.

[0049] Specifically, the Brix value of the solution can be 8%, 8.3%, 9%, 9.5%, or 10%; the loading rate of the solution can be 0.5 BV / h, 0.65 BV / h, 0.7 BV / h, 0.9 BV / h, or 1.0 BV / h; the loading volume can be 0.5 BV, 0.9 BV, 2.5 BV, 3.8 BV, or 5.0 BV of the resin volume; the macroporous adsorption resin used for decolorizing the solution can be any one or more of ADS-7, DA201, DM100, LKS03, LKS11, MN100, MN270, SD331, SD333, T19, and XDA-8; and the anion exchange resin used for decolorizing the solution can be any one or more of LX600J, D941, D900, D318, D301, D285, D201, and H280.

[0050] When decolorizing the solution, under the above-mentioned parameter conditions, colored substances in the solution can be effectively removed, resulting in a purer decolorized solution. It should be noted that conventional techniques for decolorizing solutions can be used, as long as they achieve the desired decolorization. This invention preferably uses macroporous adsorption resin for adsorption decolorization or anion exchange resin for ion exchange decolorization. It should be understood that during the macroporous adsorption resin adsorption decolorization or anion exchange resin ion exchange decolorization process, the initial clear water flowing out after sample loading should be removed. Collection should begin when the sample solution flows out. After sample loading is complete, the solution can be washed with 1-5 wt% saline solution. The sample solution and washing solution are collected to obtain the decolorized solution. Therefore, when using macroporous adsorption resin for adsorption decolorization or anion exchange resin for ion exchange decolorization, after the decolorizing resin has completed decolorization, it is only necessary to rinse the decolorizing resin with saline solution. This replaces the complex flocculation, precipitation, and oxidation-reduction steps in existing technologies, making the process simple and suitable for industrial production. The simple decolorization process, combined with the previous removal of high-molecular-weight and large-molecule proteins, camellia polysaccharides, starch, macromolecules, and small-molecule inorganic salts from camellia seed cake or tea seed cake extract, further improves the purity of camellia saponins. After the solution has been decolorized, the resin used for decolorization can be regenerated and recycled. Specifically, it is first rinsed with 1-5wt% hydrochloric acid, then washed with water until neutral, and then rinsed with 2-5wt% sodium hydroxide alkaline solution, washed with water until neutral, and then it can be used directly.

[0051] In some embodiments, when the decolorizing solution is adsorbed by the macroporous adsorption resin, the loading rate of the decolorizing solution is 1-2 BV / h, and the loading amount is 0.5-5.0 BV of the resin volume; when the ethanol solution is desorbed, the loading rate of the ethanol solution is 0.5-1 BV / h, and the loading amount is 1.0-5.0 BV of the resin volume.

[0052] Specifically, the macroporous adsorption resin is a non-polar to weakly polar macroporous resin, which can be polystyrene macroporous adsorption resin or acrylate macroporous adsorption resin, with a particle size of 250μm-1.25mm and a water content of 55-75%. The macroporous adsorption resin model used can be one or more of the following: AB-8, ADS-5, D101, D3520, D4006, D4020, H103, HP20, HP2MG, HPD100, HPD600, HZ801, HZ803, LX20, LX60, LK1180, LK1200, LK1300, LK1400, LK1500, LKS06, SP825, T28, T81, T83, X-5, XAD-5, XAD-8, XAD-16, and XAD-1600.

[0053] The decolorizing solution was adsorbed onto a macroporous adsorption resin. Under the parameters described above, this effectively adsorbed the camellia oleifera saponin in the decolorizing solution, resulting in a greater extraction of camellia oleifera saponin. After the macroporous adsorption resin adsorption was complete, the decolorizing solution was eluted with ethanol. It should be understood that after the ethanol elution was completed, the macroporous adsorption resin could be further rinsed with water, and the ethanol eluent and water wash liquid were collected and combined to obtain the eluent solution. After the macroporous adsorption resin eluted and eluted, it could be recycled after rinsing the resin with water until no alcohol remained.

[0054] In some embodiments, the method for concentrating the eluent is as follows: the eluent is concentrated by membrane concentration, and when the solid content of the concentrate reaches 20-30%, it is transferred to a single-effect external circulation concentrator for further concentration until the Brix value reaches 40-50%; the membrane used for membrane concentration has a specification of 100-200 Da; when the single-effect external circulation concentrator continues to concentrate, the concentration vacuum degree is 0.05-0.095 MPa and the concentration temperature is 50-85℃.

[0055] In some embodiments, the sterilization temperature is 80-85°C and the sterilization time is 30-35 minutes.

[0056] In some embodiments, the drying method is spray drying, with a feed rate of 10-100 L / h, an inlet air temperature of 180-190°C, and an outlet air temperature of 85-90°C.

[0057] The high-purity camellia oleifera saponins prepared by the method of this invention have a purity of over 97%, a protein content ≤1.0 wt%, a polyphenol content ≤1.0 g / kg, and an absorbance (Abs) <0.06 at a wavelength of 420-610 nm. The methods for testing the purity, protein content, polyphenol content, and absorbance of the high-purity camellia oleifera saponins are as follows:

[0058] Test method for the purity of high-purity camellia oil saponins: Test according to the method in Appendix A of GB / T 41549-2022;

[0059] Test method for high-purity camellia oleifera saponin protein content: The test shall be conducted according to the first method of Kjeldahl nitrogen determination in GB5009.5-2016;

[0060] The content of high-purity camellia oil saponins and polyphenols was tested using the Folin-Ciocalteu reagent method, which specifically included the following steps:

[0061] S1. Preparation of phenol standard stock solution: Accurately weigh 20 mg of gallic acid standard substance (CAS No.: 149-91-7), dissolve it in water and dilute to 100 mL in a volumetric flask, shake well, and store the phenol standard stock solution away from light.

[0062] S2. Preparation of standard working solutions: Pipette 0, 1 mL, 2 mL, 3 mL, 5 mL and 10 mL of phenol standard stock solution into 10 mL volumetric flasks, dilute with water to the mark, and mix thoroughly. The resulting phenol concentrations (expressed as gallic acid concentrations) are 0, 20 mg / L, 40 mg / L, 60 mg / L, 100 mg / L and 200 mg / L respectively.

[0063] S3. Preparation of sample test solution: Accurately weigh 1.0000g of high-purity camellia saponins into a beaker, dissolve in 10mL of purified water, and transfer to a 25mL volumetric flask. Rinse the beaker with 10mL of purified water and transfer the solution to the volumetric flask as well. Add water to the mark and mix well. Transfer 1.0mL to a 100mL volumetric flask, add water to the mark, and mix well to obtain the sample test solution.

[0064] S4. Pipette 0.2 mL each of the standard working solution and the sample test solution of various concentrations into a series of 10 mL graduated test tubes. Add 1.8 mL of distilled water, mix well, then add 0.2 mL of Folin-Ciocalteu phenol reagent, mix thoroughly, and let the reaction proceed for 5 min. When the color in the test tube turns light green, add 2.0 mL of 7% sodium carbonate solution and 0.8 mL of distilled water, and shake well. Let it stand at room temperature for 90 min, then measure the absorbance at 750 nm using a 1 cm cuvette, with a reagent (water) blank as a reference, using a spectrophotometer. Plot a standard curve with absorbance as the ordinate and concentration as the abscissa. Calculate the concentration of gallic acid in the sample test solution from the standard working curve.

[0065] S5. The content X of total polyphenols (calculated as gallic acid) in high-purity camellia saponins is calculated according to the following formula (1), in units (g / kg):

[0066] X = (C × V × K) / (m × 1000) Equation (1)

[0067] In formula (1): C is the concentration of gallic acid measured in the sample solution, in mg / L;

[0068] V represents the constant volume, in mL;

[0069] K is the dilution factor of the sample;

[0070] m represents the accurate amount of sample weighed, in grams.

[0071] 1000 is the conversion factor for milliliters (mL) to liters (L);

[0072] The calculation result should be rounded to three significant figures.

[0073] S6. Under repeatability conditions, the absolute difference between two independent measurements shall not exceed 10% of the arithmetic mean, and the average of the two measurements shall be taken as the result.

[0074] The absorbance of high-purity camellia oil saponins was tested using ultraviolet-visible spectrophotometry, specifically including the following steps:

[0075] Accurately weigh 0.500 g of the high-purity camellia saponin to be tested into a 50 mL volumetric flask, add 30 mL of purified water to dissolve it completely, then add another 15 mL of purified water to dissolve it completely and shake well. After there are no insoluble substances, make up to 50 mL and shake well to obtain the sample solution. Test the absorbance of the sample solution at wavelengths of 420 nm, 480 nm, 510 nm, 520 nm, 540 nm, and 610 nm according to the general technical requirements and guidelines (General Rule 0401) of the Chinese Pharmacopoeia (2020 edition).

[0076] To better illustrate the technical solution of the present invention, the following specific embodiments are also provided. It should be understood that, unless otherwise specified, all raw materials used in the following embodiments are commercially available.

[0077] Example 1:

[0078] A method for extracting high-purity camellia saponins from camellia seed cake includes the following steps:

[0079] (1) Raw material preparation: Camellia seed cake is grade 1 grass-free cake, and other aspects meet the requirements of GB35131 standard and are commercially available.

[0080] (2) Crushing and Extraction: Weigh 1000 kg of raw camellia seed cake, feed it evenly with a scraper at a speed of 10 kg / h, and adjust the frequency to 30 Hz. After crushing in the crusher, the particle size of the camellia seed cake is 0.6 mm. It falls into one end of the dynamic countercurrent extraction unit. The other end of the extraction unit is fed with 50% aqueous ethanol (containing 0.2 wt% sodium bisulfite and sodium metabisulfite, with a mass ratio of 1:1) through a pipeline. Extraction is carried out at a flow rate of 1 m³ / h. The extraction temperature is 50℃, the propulsion speed of the spiral shaft of the dynamic countercurrent extraction unit is 8 m / h, and the frequency is 30 Hz. The extract flows out from the port pipeline filter into the temporary storage tank. The camellia seed cake extraction residue is pushed to the end of the unit, filtered by a grid, and dried before being used as feed.

[0081] (3) Nanofiltration membrane concentration: The filtrate is concentrated using a nanofiltration membrane to recover alcohol until the alcohol residue in the concentrate is 1%. The membrane module is a hollow fiber membrane NF95 with a specification of 150 Da. The concentration temperature is 20℃, the pressure is 0.9MPa, the pH value is 4, and the flow rate is 0.5m³ / h. The recovered alcohol can be distilled and recycled.

[0082] (4) Dilution: Dilute the concentrate with water. The Brix value of the diluted solution is 6%.

[0083] (5) Ultrafiltration and Nanofiltration Concentration: The diluted solution is ultrafiltered through an ultrafiltration membrane (NF01, 15 kDa). During operation, the inlet membrane pressure is maintained at 0.3 MPa, the temperature at 25°C, the pH at 3, and the flow rate at 0.5 m³ / h. Water is continuously added for dialysis to retain macromolecules with a molecular weight greater than 20 kDa until the Brix value of the permeate is 0.2%. The permeate is then collected. The permeate is then concentrated through a series of nanofiltration membranes (NF10, 400 Da). This nanofiltration membrane allows molecules with a molecular weight less than 400 Da to pass through while retaining molecules with a molecular weight greater than 600 Da. During operation, the pressure is 0.8 MPa, the temperature at 10°C, and the pH at 3. The flow rate of the permeate from the ultrafiltration membrane is 1.6 m³ / h, and the concentration is continued until the Brix value of the concentrate is 25%.

[0084] (6) Resin column decolorization: Dissolve the concentrate obtained in step (5) in deionized water. The Brix value of the solution is 8%. Decolorize the solution on a resin column LX600J with a loading rate of 1 BV / h and a loading volume of 5 BV of the resin volume. Discard the effluent water and start collecting the effluent. After the injection is completed, use 2 wt% saline solution (2.5 times the resin volume) to push out the residual liquid. Collect the effluent and the water head liquid together to obtain the decolorized solution.

[0085] (7) Macroporous resin adsorption: The decolorizing solution is injected into the macroporous adsorption resin HP20 for adsorption. The loading rate is 1 BV / h and the loading volume is 5 BV of the resin volume. After the injection is completed, the residual liquid is rinsed with 2-3 BV of resin volume of water. The eluent and washing liquid are discarded. The solution is eluted with 2 BV of resin volume of 85% ethanol at a elution rate of 0.8 BV / h. Then, the ethanol is topped with 2 times the resin volume of water. The eluent and the top water are combined to form the elution solution.

[0086] (8) Concentration: The eluent is concentrated by membrane. The membrane module is a hollow fiber membrane NF95 with a specification of 200 Da. Ethanol is recovered until there is no alcohol. When the Brix value of the concentrate is 22%, it is transferred to a single-effect external circulation concentrator for further concentration. The concentration vacuum is 0.05 MPa and the concentration temperature is 60°C. The concentration ends when the Brix value of the concentrate is 40%.

[0087] (9) Sterilization: Pasteurize the concentrate and keep it at 82°C for 35 minutes.

[0088] (10) Drying: The sterilized concentrate was filtered through a pipeline filter and then spray-dried. The feed rate was adjusted to 100 L / h, the inlet air temperature was 190℃, and the outlet air temperature was 85℃. The product was collected. The final product had a high-purity camellia saponin yield of 10.4%, an extraction rate of 78.8%, and a camellia saponin content of 98.5%. The purity, protein content, polyphenol content, and absorbance of the high-purity camellia saponin product were tested. The results are shown in Table 1.

[0089] Example 2

[0090] A method for extracting high-purity camellia saponins from camellia seed cake includes the following steps:

[0091] (1) Raw material preparation: Camellia seed cake is grade 1 grass-free cake, and other aspects meet the requirements of GB35131 standard and are commercially available.

[0092] (2) Crushing and Extraction: Weigh 1000 kg of raw camellia seed cake, feed it evenly with a scraper at a speed of 160 kg / h, and adjust the frequency to 45 Hz. After crushing in the crusher, the particle size of the camellia seed cake is 0.1 mm. It falls into one end of the dynamic countercurrent extraction unit. The other end of the extraction unit is fed with 70% aqueous methanol (the aqueous ethanol contains 0.4 wt% sodium bisulfite and sodium sulfite, with a mass ratio of sodium bisulfite to sodium sulfite of 1:1) through a pipeline, and the extraction is carried out at a flow rate of 1 m³ / h. The extraction temperature is 38℃, the propulsion speed of the spiral shaft of the dynamic countercurrent extraction unit is 8.5 m / h, and the frequency is 42 Hz. The extract flows out from the port pipeline filter into the temporary storage tank. The camellia seed cake extraction residue is pushed to the end of the unit, filtered by a grid, and dried before being used as feed.

[0093] (3) Nanofiltration membrane concentration: The filtrate is concentrated using a nanofiltration membrane to recover alcohol until the alcohol residue in the concentrate is 1.5%. The membrane module is a hollow fiber membrane NF95 with a specification of 165 Da. The concentration temperature is 35℃, the pressure is 1.5MPa, the pH value is 3, and the flow rate is 1.6m³ / h. The recovered alcohol can be distilled and recycled.

[0094] (4) Dilution: Dilute the concentrate with water, and the Brix value of the diluted solution is 1%.

[0095] (5) Ultrafiltration and Nanofiltration Concentration: The diluted solution is ultrafiltered through an ultrafiltration membrane (NF01, 18 kDa). During operation, the inlet membrane pressure is maintained at 0.1 MPa, the temperature at 37°C, the pH at 5, and the flow rate at 1.6 m³ / h. Simultaneously, water is continuously added for dialysis to remove macromolecules with a molecular weight greater than 20 kDa until the Brix value of the permeate is 0.35%. The permeate is then collected. The permeate is then concentrated through a series of nanofiltration membranes (NF10, 460 Da). This nanofiltration membrane allows molecules with a molecular weight less than 400 Da to pass through while removing molecules with a molecular weight greater than 600 Da. During operation, the pressure is 1.5 MPa, the temperature at 37°C, and the pH at 8. The flow rate of the permeate from the ultrafiltration membrane is 3 m³ / h, and the concentration is continued until the Brix value of the concentrate is 28%.

[0096] (6) Resin column decolorization: Dissolve the concentrate obtained in step (5) in deionized water. The Brix value of the solution is 9%. Decolorize the solution on a resin column D941. The loading speed is 0.65 BV / h and the loading volume is 3.8 BV of the resin volume. Discard the effluent water. Start collecting the effluent. After the injection is completed, use 2wt% saline solution with 2.5 times the resin volume to push out the residual liquid. Collect the effluent and the water head liquid together to obtain the decolorized solution.

[0097] (7) Macroporous resin adsorption: The decolorizing solution was injected into macroporous adsorption resin LK20 for adsorption. The loading rate was 1.5 BV / h, and the loading volume was 0.5 BV of the resin volume. After the injection was completed, the residual liquid was rinsed with 2-3 BV of resin volume of water. The eluent and washing liquid were discarded. The solution was eluted with 3.8 BV of resin volume of 85% ethanol at a elution rate of 0.5 BV / h. Then, the ethanol was topped with water at twice the resin volume. The eluent and the top water were combined to form the elution solution.

[0098] (8) Concentration: The eluent is concentrated by membrane. The membrane module is a hollow fiber membrane NF95 with a specification of 180 Da. Ethanol is recovered until there is no alcohol. When the Brix value of the concentrate is 20%, it is transferred to a single-effect external circulation concentrator for further concentration. The concentration vacuum is 0.08 MPa and the concentration temperature is 50°C. The concentration ends when the Brix value of the concentrate is 45%.

[0099] (9) Sterilization: Pasteurize the concentrate and keep it at 84°C for 32 minutes.

[0100] (10) Drying: The sterilized concentrate was filtered through a pipeline filter and then spray-dried. The feed rate was adjusted to 10 L / h, the inlet air temperature was 180℃, and the outlet air temperature was 87℃. The product was collected, and the final product had a high-purity camellia saponin yield of 10.9%, an extraction rate of 82.6%, and a camellia saponin content of 98.7%. The purity, protein content, polyphenol content, and absorbance of the high-purity camellia saponin product were tested, and the results are shown in Table 1.

[0101] Example 3

[0102] A method for extracting high-purity camellia saponins from camellia seed cake includes the following steps:

[0103] (1) Raw material preparation: Camellia seed cake is grade 1 grass-free cake, and other aspects meet the requirements of GB35131 standard and are commercially available.

[0104] (2) Crushing and Extraction: Weigh 1000 kg of raw camellia seed cake, feed it evenly with a scraper at a speed of 50 kg / h and a frequency of 50 Hz. After crushing in a crusher, the camellia seed cake particles are 1.0 mm in diameter and fall into one end of a dynamic countercurrent extraction unit. The other end of the extraction unit is fed with 75% aqueous methanol (containing 0.1 wt% sodium sulfite, sodium bisulfite, and sodium metabisulfite in an ethanol-water mixture, with a mass ratio of 1:2:3) at a flow rate of 1 m³ / h. The extraction temperature is 33℃, the propulsion speed of the spiral shaft of the dynamic countercurrent extraction unit is 9 m / h, and the frequency is 50 Hz. The extract is filtered through a port pipe and sent to a temporary storage tank. The camellia seed cake residue is pushed to the end of the unit, filtered by a grid, and dried before being used as feed.

[0105] (3) Nanofiltration membrane concentration: The filtrate is concentrated using a nanofiltration membrane to recover alcohol until the alcohol residue in the concentrate is 2.8%. The membrane module is a hollow fiber membrane NF95 with a specification of 190 Da. The concentration temperature is 10℃, the pressure is 0.5MPa, the pH value is 9, and the flow rate is 2.5m³ / h. The recovered alcohol can be distilled and recycled.

[0106] (4) Dilution: Dilute the concentrate with water. The Brix value of the diluted solution is 8%.

[0107] (5) Ultrafiltration and Nanofiltration Concentration: The diluted solution is ultrafiltered through an ultrafiltration membrane (NF01, 17 kDa). During operation, the inlet membrane pressure is maintained at 1 MPa, the temperature at 65°C, the pH at 7, and the flow rate at 3.0 m³ / h. Simultaneously, water is continuously added for dialysis to remove macromolecules with a molecular weight greater than 20 kDa until the Brix value of the permeate is 0.5%. The permeate is then collected. The permeate is then concentrated through a series of nanofiltration membranes (NF10, 550 Da). This nanofiltration membrane allows the permeate to pass through molecules with a molecular weight less than 400 Da while removing molecules with a molecular weight greater than 600 Da. During operation, the pressure is 2.0 MPa, the temperature at 50°C, and the pH at 6. The flow rate of the permeate from the ultrafiltration membrane is 3.8 m³ / h, and the concentration is continued until the Brix value of the concentrate is 30%.

[0108] (6) Resin column decolorization: Dissolve the concentrate obtained in step (5) in deionized water. The Brix value of the solution is 9.5%. Decolorize the solution on a D900 resin column. The loading speed is 0.7 BV / h and the loading volume is 2.5 BV of the resin volume. Discard the effluent water. Start collecting the effluent. After the injection is completed, use 2wt% saline solution with 2.5 times the resin volume to push out the residual liquid. Collect the effluent and the water head liquid together to obtain the decolorized solution.

[0109] (7) Macroporous resin adsorption: The decolorizing solution was injected into the macroporous adsorption resin AB-8 for adsorption. The loading rate was 1.8 BV / h, and the loading volume was 4.2 BV of the resin volume. After the injection was completed, the residual liquid was rinsed with 2-3 BV of resin volume of water. The eluent and washing liquid were discarded. The solution was eluted with 4.5 BV of resin volume of 85% ethanol at a rate of 0.7 BV / h. Then, the ethanol was topped with water at a rate of 2 times the resin volume. The eluent and the top water were combined to form the elution solution.

[0110] (8) Concentration: The eluent is concentrated by membrane. The membrane module is a hollow fiber membrane NF95 with a specification of 160 Da. Ethanol is recovered until there is no alcohol. When the Brix value of the concentrate is 26%, it is transferred to a single-effect external circulation concentrator for further concentration. The concentration vacuum is 0.06 MPa and the concentration temperature is 75°C. The concentration ends when the Brix value of the concentrate is 48%.

[0111] (9) Sterilization: Pasteurize the concentrate and keep it at 80°C for 30 minutes.

[0112] (10) Drying: The sterilized concentrate was filtered through a pipeline filter and then spray-dried. The feed rate was adjusted to 25 L / h, the inlet air temperature was 185℃, and the outlet air temperature was 88℃. The product was collected, and the final product had a high-purity camellia saponin yield of 11.1%, an extraction rate of 85.1%, and a camellia saponin content of 99.3%. The purity, protein content, polyphenol content, and absorbance of the high-purity camellia saponin product were tested, and the results are shown in Table 1.

[0113] Example 4

[0114] A method for extracting high-purity camellia saponins from camellia seed cake includes the following steps:

[0115] (1) Raw material preparation: Camellia seed cake is grade 1 grass-free cake, and other aspects meet the requirements of GB35131 standard and are commercially available.

[0116] (2) Crushing and Extraction: Weigh 1000 kg of raw camellia seed cake, feed it evenly with a scraper at a speed of 100 kg / h and a frequency of 40 Hz. After crushing in a crusher, the camellia seed cake particles are 2.0 mm in diameter and fall into one end of a dynamic countercurrent extraction unit. The other end of the extraction unit is fed with 75% aqueous methanol (containing 0.3 wt% sodium metabisulfite) through a pipeline at a flow rate of 1 m³ / h for extraction. The extraction temperature is 45℃, the propulsion speed of the spiral shaft of the dynamic countercurrent extraction unit is 10 m / h, and the frequency is 35 Hz. The extract is filtered through the port pipeline and sent to a temporary storage tank. The camellia seed cake extraction residue is pushed to the end of the unit, filtered by a grid, and dried before being used as feed.

[0117] (3) Nanofiltration membrane concentration: The filtrate is concentrated using a nanofiltration membrane to recover alcohol until the alcohol residue in the concentrate is 0.5%. The membrane module is a hollow fiber membrane NF95 with a specification of 200 Da. The concentration temperature is 50℃, the pressure is 2.5 MPa, the pH value is 5, and the flow rate is 3.8 m³ / h. The recovered alcohol can be distilled and recycled.

[0118] (4) Dilution: Dilute the concentrate with water. The Brix value of the diluted solution is 3%.

[0119] (5) Ultrafiltration and Nanofiltration Concentration: The diluted solution is ultrafiltered through an ultrafiltration membrane (NF01, 19 kDa). During operation, the inlet membrane pressure is maintained at 0.5 MPa, the temperature at 50°C, the pH at 9, and the flow rate at 3.8 m³ / h. Simultaneously, water is continuously added for dialysis to retain macromolecules with a molecular weight greater than 20 kDa until the Brix value of the permeate is 0.1%. The permeate is then collected. The permeate is then concentrated through a series of nanofiltration membranes (NF10, 500 Da). This nanofiltration membrane allows the permeate to pass through molecules with a molecular weight less than 400 Da while retaining molecules with a molecular weight greater than 600 Da. During operation, the pressure is 3.0 MPa, the temperature at 65°C, and the pH at 12. The flow rate of the permeate from the ultrafiltration membrane is 0.5 m³ / h, and the concentration is continued until the Brix value of the concentrate is 26.5%.

[0120] (6) Resin column decolorization: Dissolve the concentrate obtained in step (5) in deionized water. The Brix value of the solution is 8.3%. Decolorize the solution on a resin column SD331. The loading speed is 0.9 BV / h and the loading volume is 0.9 BV of the resin volume. Discard the effluent water and start collecting the effluent. After the injection is completed, use 2wt% saline solution with 2.5 times the resin volume to push out the residual liquid. Collect the effluent and the water head liquid together to obtain the decolorized solution.

[0121] (7) Macroporous resin adsorption: The decolorizing solution was injected into macroporous adsorption resin LX60 for adsorption. The loading rate was 1.5 BV / h, and the loading volume was 2.8 BV of the resin volume. After the injection was completed, the residual liquid was rinsed with 2-3 BV of resin volume of water. The eluent and washing liquid were discarded. The solution was eluted with 5 BV of resin volume of 85% ethanol at a elution rate of 0.9 BV / h. Then, the ethanol was topped with 2 times the resin volume of water. The eluent and the top water were combined to form the elution solution.

[0122] (8) Concentration: The eluent is concentrated by membrane. The membrane module is a hollow fiber membrane NF95 with a specification of 100 Da. Ethanol is recovered until there is no alcohol. When the Brix value of the concentrate is 30%, it is transferred to a single-effect external circulation concentrator for further concentration. The concentration vacuum is 0.075 MPa and the concentration temperature is 85°C. The concentration ends when the Brix value of the concentrate is 50%.

[0123] (9) Sterilization: Pasteurize the concentrate and keep it at 83°C for 31 min.

[0124] (10) Drying: The sterilized concentrate was filtered through a pipeline filter and then spray-dried. The feed rate was adjusted to 75 L / h, the inlet air temperature was 186℃, and the outlet air temperature was 90℃. The product was collected. The final product had a high-purity camellia saponin yield of 10.9%, an extraction rate of 83.5%, and a camellia saponin content of 98.8%. The purity, protein content, polyphenol content, and absorbance of the high-purity camellia saponin product were tested. The results are shown in Table 1.

[0125] Example 5

[0126] A method for extracting high-purity camellia saponins from camellia seed cake includes the following steps:

[0127] (1) Raw material preparation: Camellia seed cake is grade 1 grass-free cake, and other aspects meet the requirements of GB35131 standard and are commercially available.

[0128] (2) Crushing and Extraction: Weigh 1000 kg of raw camellia seed cake, feed it evenly with a scraper at a speed of 200 kg / h and a frequency of 45 Hz. After crushing in a crusher, the camellia seed cake particles are 1.6 mm in diameter and fall into one end of a dynamic countercurrent extraction unit. The other end of the extraction unit is fed with 90% aqueous ethanol (containing 0.5 wt% sodium sulfite) through a pipeline at a flow rate of 1 m³ / h. The extraction temperature is 30℃, the propulsion speed of the spiral shaft of the dynamic countercurrent extraction unit is 5 m / h, and the frequency is 40 Hz. The extract flows from the end pipe filter into a temporary storage tank. The camellia seed cake extraction residue is pushed to the end of the unit, filtered by a grid, and dried before being used as feed.

[0129] (3) Nanofiltration membrane concentration: The filtrate is concentrated using a nanofiltration membrane to recover alcohol until the alcohol residue in the concentrate is 2%. The membrane module is a hollow fiber membrane NF95 with a specification of 185 Da. The concentration temperature is 65℃, the pressure is 3MPa, the pH value is 12, and the flow rate is 5m³ / h. The recovered alcohol can be distilled and recycled.

[0130] (4) Dilution: Dilute the concentrate with water, and the Brix value of the diluted solution is 10%.

[0131] (5) Ultrafiltration and Nanofiltration Concentration: The diluted solution is ultrafiltered through an ultrafiltration membrane (NF01, 20 kDa). During operation, the inlet membrane pressure is maintained at 0.8 MPa, the temperature at 10°C, the pH at 12, and the flow rate at 5 m³ / h. Water is continuously added for dialysis to retain macromolecules with a molecular weight greater than 20 kDa until the Brix value of the permeate is 0.4%. The permeate is then collected. The permeate is then concentrated through a series of nanofiltration membranes (NF10, 600 Da). This nanofiltration membrane allows molecules with a molecular weight less than 400 Da to pass through while retaining molecules with a molecular weight greater than 600 Da. During operation, the pressure is 0.1 MPa, the temperature at 25°C, and the pH at 10. The flow rate of the permeate from the ultrafiltration membrane is 5 m³ / h, and the concentration is continued until the Brix value of the concentrate is 27.5%.

[0132] (6) Resin column decolorization: The concentrate obtained in step (5) is dissolved in deionized water with a Brix value of 10%. The solution is decolorized by a mixed resin of LX600J and DM100. The loading speed is 0.5 BV / h and the loading volume is 0.5 BV of the resin volume. The effluent is discarded and the liquid is collected from the outflow. After the injection is completed, the residual liquid is pushed out with 2wt% saline solution with 2.5 times the resin volume. The effluent and the water are collected together to obtain the decolorized solution.

[0133] (7) Macroporous resin adsorption: The decolorizing solution is injected into the macroporous adsorption resin AB-8 for adsorption. The loading rate is 2 BV / h and the loading volume is 1.6 BV of the resin volume. After the injection is completed, the residual liquid is rinsed with 2-3 BV of resin volume of water. The eluent and washing liquid are discarded. The solution is eluted with 1 BV of resin volume of 85% ethanol at a elution rate of 1 BV / h. Then, the ethanol is topped with 2 times the resin volume of water. The eluent and the top water are combined to form the elution solution.

[0134] (8) Concentration: The eluent is concentrated by membrane. The membrane module is a hollow fiber membrane NF95 with a specification of 130 Da. Ethanol is recovered until there is no alcohol. When the Brix value of the concentrate is 25%, it is transferred to a single-effect external circulation concentrator for further concentration. The concentration vacuum is 0.095 MPa and the concentration temperature is 58°C. The concentration ends when the Brix value of the concentrate is 43%.

[0135] (9) Sterilization: Pasteurize the concentrate and keep it at 85°C for 35 minutes.

[0136] (10) Drying: The sterilized concentrate was filtered through a pipeline filter and then spray-dried. The feed rate was adjusted to 50 L / h, the inlet air temperature was 182℃, and the outlet air temperature was 86℃. The product was collected, and the final product had a high-purity camellia saponin yield of 10.8%, an extraction rate of 81.7%, and a camellia saponin content of 97.9%. The purity, protein content, polyphenol content, and absorbance of the high-purity camellia saponin product were tested, and the results are shown in Table 1.

[0137] Example 6

[0138] The high-purity camellia saponins were extracted according to the method of Example 5. The only difference from Example 5 is that the aqueous ethanol used in step (2) contains sodium sulfite, sodium bisulfite and sodium metabisulfite, and the mass ratio of sodium sulfite, sodium bisulfite and sodium sulfite is 2:1:2.

[0139] The final yield of high-purity camellia oleifera saponin product was 11.3%, the extraction rate was 86.9%, and the camellia oleifera saponin content was 99.7%. The purity, protein content, polyphenol content, and absorbance of the high-purity camellia oleifera saponin product were measured, and the results are shown in Table 1.

[0140] Comparative Example 1

[0141] High-purity camellia saponins were extracted according to the method of Example 3. The only difference from Example 3 is that the aqueous ethanol used in step (2) does not contain sodium sulfite, sodium bisulfite, and sodium metabisulfite.

[0142] The final yield of high-purity camellia oleifera saponin product was 10.2%, the extraction rate was 72.4%, and the camellia oleifera saponin content was 91.6%. The purity, protein content, polyphenol content, and absorbance of the high-purity camellia oleifera saponin product were measured, and the results are shown in Table 1.

[0143] Comparative Example 2

[0144] The high-purity camellia saponins were extracted according to the method of Example 3. The only difference from Example 3 was that the aqueous ethanol used in step (2) did not contain sodium sulfite, sodium bisulfite and sodium metabisulfite, and it was not treated in steps (5)-(6). The diluted solution in step (4) was directly adsorbed onto the macroporous resin in step (7).

[0145] The final yield of high-purity camellia oleifera saponin product was 9.5%, the extraction rate was 60.9%, and the camellia oleifera saponin content was 83.2%. The purity, protein content, polyphenol content, and absorbance of the high-purity camellia oleifera saponin product were measured, and the results are shown in Table 1.

[0146] Table 1. Performance parameters of camellia saponin products prepared in the examples and comparative examples.

[0147]

[0148] It should be noted that polyphenol content cannot be detected when it is below 0.05 g / kg. Therefore, "not detected" in Table 1 refers to polyphenol content below 0.05 g / kg.

[0149] As can be seen from Examples 1-5, the extraction rate of camellia saponins from camellia seed cake using the method of the present invention is all above 80%, and the purity of the obtained camellia saponin products is high, all above 97%. A comparison of Examples 5 and 6 shows that when extracting camellia seed cake, the aqueous lower alcohol solution containing compound salts, compared with the use of only salts, resulted in higher purity of the final camellia saponins, lower protein and polyphenol content, and lower absorbance. A comparison of Examples 3 and Comparative Example 1 shows that when extracting camellia seed cake, the aqueous lower alcohol solution containing compound salts, compared with the use of no compound salts, resulted in higher purity of the final camellia saponins, lower protein and polyphenol content, and lower absorbance. A comparison of Examples 3 and Comparative Example 2 shows that when extracting camellia seed cake, the use of an aqueous lower alcohol solution containing compound salts, combined with steps (5)-(6) of Example 3, significantly improved the purity of the final camellia saponins under the synergistic effect of the above conditions, resulting in lower protein and polyphenol content and lower absorbance.

[0150] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art can still modify the technical solutions described in the foregoing embodiments or make equivalent substitutions for some of the technical features. 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 method for extracting high-purity camellia saponins, characterized in that, Includes the following steps: Camellia seed cake or tea seed cake is crushed, and the crushed camellia seed cake or tea seed cake is subjected to dynamic countercurrent extraction using an aqueous low-alcohol solution. The filtrate is collected. The aqueous low-alcohol solution comprises low-alcohol, compound salt, and water, wherein the mass concentration of low-alcohol is 50-90%, and the mass concentration of compound salt is 0.1-0.5%; the compound salt comprises one or more of sulfites, bisulfites, or metabisulfites. The filtrate is concentrated to recover alcohol until the alcohol content in the concentrate is less than 3 wt%, thus obtaining concentrate A; The concentrate A is diluted with water, the diluted solution is treated with an ultrafiltration membrane, and the permeate from the ultrafiltration membrane is concentrated with a nanofiltration membrane to obtain concentrate B. Water is added to the concentrated solution B to obtain a solution. The solution is then decolorized, and the decolorized solution is adsorbed by a macroporous adsorption resin and then eluted with an ethanol solution to obtain an eluent. The eluent was concentrated, sterilized, filtered, and dried to obtain high-purity camellia saponins.

2. The extraction method according to claim 1, characterized in that, The conditions for crushing camellia seed cake or tea seed cake are as follows: feeding speed is 10-200 kg / h, frequency is 30-50 Hz; the particle size of the crushed camellia seed cake or tea seed cake is 0.1-2 mm.

3. The extraction method according to claim 1, characterized in that, When performing dynamic countercurrent extraction on the crushed camellia seed cake or tea seed cake, the extraction temperature is 30-50℃, the extraction frequency is 30-50HZ, and the extraction propulsion speed is 5-10m / h.

4. The extraction method according to claim 1, characterized in that, The method for concentrating and recovering alcohol from the filtrate is nanofiltration membrane concentration, wherein the nanofiltration membrane has a specification of 150-200 Da, the operating pressure of the nanofiltration membrane is 0.5-3.0 MPa, the temperature is 10-65℃, the pH value is 3.0-12.0, and the flow rate of the filtrate is 0.5-5.0 m³ / h.

5. The extraction method according to claim 1, characterized in that, The Brix value of the diluted solution after diluting the concentrate A with water is 1-10%. When the diluent is treated by an ultrafiltration membrane, the ultrafiltration membrane has a specification of 15-20 kDa, the inlet pressure is 0.1-1.0 MPa, the temperature is 10-65℃, the pH value is 3.0-12.0, the flow rate of the diluent is 0.5-5.0 m³ / h, and large molecules with a molecular weight greater than 20 kDa are retained. At the same time, water is continuously added for dialyzing until the Brix value of the permeate is ≤0.5%.

6. The extraction method according to claim 1, characterized in that, When the permeate from the ultrafiltration membrane is concentrated by the nanofiltration membrane, the nanofiltration membrane has a specification of 400-600 Da, the operating pressure is 0.1-3.0 MPa, the temperature is 10-65℃, the pH value is 3.0-12.0, and the flow rate of the permeate from the ultrafiltration membrane is 0.5-5.0 m³ / h, until the Brix value of the concentrate is 25-30%, thus obtaining concentrate B.

7. The extraction method according to claim 1, characterized in that, The Brix value of the solution is 8-10%.

8. The extraction method according to claim 1, characterized in that, The decolorization method of the solution is as follows: the solution is decolorized by adsorption with macroporous adsorption resin or ion exchange with anion exchange resin; when the solution is decolorized by adsorption with macroporous adsorption resin or ion exchange with anion exchange resin, the loading rate of the solution is 0.5-1 BV / h, and the loading amount is 0.5-5.0 BV of the resin volume. After the decolorization by macroporous adsorption resin or anion exchange resin is completed, a decolorized solution is obtained.

9. The extraction method according to claim 1, characterized in that, The macroporous adsorption resin is a polystyrene macroporous adsorption resin or an acrylate macroporous adsorption resin, and the anion exchange resin is a polystyrene skeleton or an acrylate skeleton anion exchange resin.

10. The extraction method according to claim 8, characterized in that, When the decolorizing liquid is adsorbed by the macroporous adsorption resin, the loading rate of the decolorizing liquid is 1-2 BV / h, and the loading amount is 0.5-5.0 BV of the resin volume.

11. The extraction method according to claim 1, characterized in that, During the ethanol solution analysis, the ethanol solution loading rate is 0.5-1 BV / h, and the loading volume is 1.0-5.0 BV of the resin volume.

12. The extraction method according to claim 1, characterized in that, The method for concentrating the eluent is as follows: the eluent is concentrated using a membrane. When the solid content of the concentrate reaches 20-30%, it is transferred to a single-effect external circulation concentrator for further concentration until the Brix value reaches 40-50%. The membrane used for membrane concentration has a specification of 100-200 Da. When the single-effect external circulation concentrator continues to concentrate, the concentration vacuum degree is 0.05-0.095 MPa and the concentration temperature is 50-85℃.

13. The extraction method according to claim 1, characterized in that, The sterilization temperature is 80-85℃, and the sterilization time is 30-35 minutes.

14. The extraction method according to claim 1, characterized in that, The drying method is spray drying, wherein the feed rate of the spray drying is 10-100 L / h, the inlet air temperature is 180-190℃, and the outlet air temperature is 85-90℃.

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